ANNUAL REPORT EDUCATION ACTIVITIES 336 STAFF 338

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3 PREFACE ANNUAL REPORT MARIO NEGRI INSTITUTE, MILAN DEPARTMENTS Department of Oncology.. 7 Department of Environmental Health Sciences Department of Neuroscience.. 69 Department of Cardiovascular Research Department of Molecular Biochemistry and Pharmacology Department of Epidemiology LABORATORIES AND CENTERS Laboratory of Regulatory Policies Laboratory for Mother and Child Health Centre of Computer Science Engineering. 226 Italian Cochrane Center The Catullo and Daniela Borgomainerio Center 236 Library NEGRI BERGAMO LABORATORIES DEPARTMENTS Department of Molecular Medicine Department of Biomedical Engineering. 264 Laboratory of Biology and Therapy of Metastasis ALDO and CELE DACCO CENTER DEPARTMENT Department of Renal Medicine. 282 LABORATORIES AND CENTERS Laboratory of Clinical Epidemiology. 306 Laboratory of Coordination of Diagnosis and Information on Rare Diseases A. and A. Valenti Health Economics Center, CESAV. 326 The Transplant Research Center. 334 EDUCATION ACTIVITIES 336 STAFF 338 All the staff of the Institute is listed on its website PUBLICATIONS A comprehensive list of the Institute s publications is available on the website Section Publications 1

4 Edited by Isabella Bordogna printed May

5 PREFACE This booklet provides a brief description of the research and training work done at the Mario Negri Institutes in Milan and Bergamo. It is divided by departments, and in some cases by single laboratories. Details of the results are provided in the text itself, so here we shall just draw a few general remarks. Compared to previous years, our research programs are increasingly spreading out to involve the collaboration among several laboratories within each department, and some are interdepartmental. We have slimmed down the number of research topics in order to focus more closely, to reach a critical mass of scientists and resources employed on each theme. The tendency nowadays is towards widespread use of molecular biology techniques, especially for studying the mechanism of action of drugs. In vitro studies are an essential basis for thorough investigations although a significant number of in vivo experiments are still needed as this is the only mean we have of validating in vitro findings and establishing models that resemble human diseases as closely as possible. This has led to a substantial increase in the use of transgenic animals. The Institute is still concentrating on its traditional research lines: oncology, neurosciences, cardiovascular and renal diseases, organ transplants with strong cell biology and molecular biochemistry connotations. Significant work has been carried out on the environment and health as a whole. Experimental, clinical and epidemiological research on rare diseases and orphan drugs is growing all the time. The Mario Negri Institute strives to develop a multifaceted approach to all these research themes, ranging from basic research, to pharmacokinetics, pharmacology, controlled clinical trials, epidemiological analysis and whenever possible the epidemiology of services. So far we have published more than articles on peer reviewed scientific journals. If research is to continue young scientists must be continually trained. Working in the laboratory not only gives them an outlet for their ideas, but enables them to obtain worthwhile qualifications by taking part in the Institute s training schemes, which are recognised by the Lombardy Region in Italy, or by working for a Ph.D. awarded by the Open University, UK. Training courses are also available on biomedical statistics, for general practitioners, family pediatricians, and clinical trial nurses. A vital part of the Institute s work involves providing information, at all levels. This is done, in particular, through the Rare Diseases Information Center, the Center for Information on Medicinal Drugs and the Website ( Scientific activities have been affected by the recent move to the new headquarters in via G. Masa 19 in Milan, where there is more space and new and modern technologies. These are difficult years for research in Italy, and the Institute is most grateful to all those public and private bodies, foundations and private citizens who generously support the work outlined here, with their steady flow of contributions. Our sincere thanks to all of them. Silvio Garattini Director 3

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7 Mario Negri INSTITUTE FOR PHARMACOLOGICAL RESEARCH Milan ANNUAL REPORT 2007 departments and laboratories 5

8 DEPARTMENT OF ONCOLOGY STAFF Chief Maurizio D INCALCI, M.D. Oncological Studies Office and Documentation Scientific Documentalist Stefania FILIPPESCHI, Chemist Laboratory of Cancer Pharmacology Head Biophysics Unit Head Flow Citometry Unit Head Cancer Clinical Pharmacology Unit Head Maurizio D INCALCI, M.D. Paolo UBEZIO, Phys.D. Eugenio ERBA, Biochem.D Massimo ZUCCHETTI, Chem.Pharm.D. Laboratory of Molecular Pharmacology Head DNA Repair Unit Head Massimo BROGGINI, Ph.D. Giovanna DAMIA, M.D. Laboratory of Biology and Therapy of Metastasis Head Tumor Angiogenesis Unit Head Molecular Cancer Therapeutics Unit Head Raffaella GIAVAZZI, Biol.Sci.D., Ph.D. Giulia TARABOLETTI, Biol.Sci.D. Maria Rosa BANI, Biol.Sci.D., Ph.D. 6

9 Laboratory for the development of new pharmacological strategies Head Valter TORRI, M.D. Laboratory of Clinical Trials Head Irene FLORIANI, Dr.Sci.Biol., Dr.Stat., Ph.D. Laboratory of Translational and Outcome Research in Oncology Head Gynecology Oncology Unit Head Giovanni APOLONE, M.D. Roldano FOSSATI, Dr.Med.Chir. CERP:Center for the Evaluation and Research on Pain Head Giovanni APOLONE, M.D. Laboratory of Medical Research and Consumer Involvement Head Paola MOSCONI, Biol.Sci.D. 7

10 CURRICULA VITAE Maurizio D'Incalci obtained his Medical Degree cum Laude from the University of Milan in After specializing in Pharmacology at the Mario Negri Institute of Milan and in Oncology at the University of Genoa, he worked in the Laboratory of Molecular Pharmacology of the National Cancer Institute in Bethesda, MD, USA. Since 1986 he has been chief of the Laboratory of Cancer Chemotherapy at the Mario Negri Institute and since 1996 he has become chief of the Department of Oncology at the Mario Negri Institute. He has been President of the Pharmacology and Molecular Mechanisms Group of the European Organization for Research and Treatment of Cancer (EORTC). From 1994 to 1997 he was Chairman of the New Drug Development Coordinating Committee and from 1997 to 2000 he was chairman of the Research Division of the EORTC. He has been member of the Board of the EORTC from April 2000 to From 1997 he is the Preclinical Coordinator of the Southern Europe New Drug Organization (SENDO) and since 2006 the Chairman of the New Agents Committee (NAC). He is on the editorial board of many international cancer-related scientific journals and since September 2000 he is Editor for Experimental Oncology of the European Journal of Cancer. Dr D'Incalci is author of more than 440 papers on cancer chemotherapy published in peer reviewed international journals, and of several chapters in books on cancer chemotherapy. Selected publications Grosso F., Jones R.L. Demetri G.D., Judson I.R., Blay J.Y., Le Cesne A., Sanfilippo R., Casieri P., Collini P., Dileo P., Spreafico C., Stacchiotti S., Tamborini E., Tercero J.C., Jimeno J., D Incalci M., Gronchi A., Fletcher J.A., Pilotti S., Casali P.G. Efficacy of Trabectedin (ET-743) in advanced pre-treated myxoid liposarcomas. Lancet Oncology, 2007; 8: Salvati E., Leonetti C., Rizzo A., Scarsella M., Mottolese M., Galati R., Sperduti I., Stevens M., D Incalci M., Blasco M., Chiorino G., Horard B., Gilson E., Zupi G., Biroccio A. Telomere damage promotes antitumoral activity of the G- quadruplex ligand RHPS4. J. Clin. Invest., 2007; 117: Zongaro S., de Stanchina E., Colombo T., D Incalci M., Giulotto E., Mondello C. Stepwise neoplastic transformation of a telomerase immortalized fibroblast cell line. Cancer Research, 2005; 65:(24): Allavena P., Signorelli M., Chieppa M., Erba E., Bianchi G., Marchesi F., Garbi A., Lissoni A., de Braud F., Jimeno J. and D Incalci M. Anti-inflammatory properties of the novel antitumor agent Yondelis (Trabectedin): inhibition of macrophage differentiation and cytokine production. Cancer Res., 2005; 65(7): Lupi M., Matera G., Branduardi D., D Incalci M., Ubezio P., Cytostatic and cytotoxic effects of topotecan decoded by a novel mathematical simulation approach, Cancer Research, 2004; 64: Gambacorti-Passerini C., Zucchetti M., Russo D., Frapolli R., Verga M., Bungaro S., Tornaghi L., Rossi F., Pioltelli P., Pogliani E., Corneo G., Alberti D., D Incalci M. Alpha 1 acid glycoprotein (AGP) binds to STI571 and substantially alters its pharmacokinetics in chronic myeloid leukemia patients. Clin. Cancer Res.,2005; 9: Tavecchio M., Natoli C., Ubezio P., Erba E., D Incalci M. Dynamics of cell cycle phase perturbations by Trabectedin (ET-743) in nucleotide excision repair (NER)-deficient and NER-proficient cells, unravelled by a novel mathematical simulation approach. Cell proliferation, 2007; 40: Giovanni Apolone, got his Medical degree in 1982 (Pavia, Italy) and his post-doctoral specializations in Internal Medicine in 1987 (Pavia, Italy) and Pharmacological Research (1992). He is Head of the Laboratory of Translational and Outcome Research. He is also Vice-President of the Ethics Committee of the European Institute of Oncology in Milan (Italy) and listed as National Expert at the European Agency for the Evaluation of Medicinal products (EMEA) in London (UK). His main fields of interest are: Methodological, ethical and regulatory aspects of clinical research, with special emphasis on oncology and the cancer pain. Health care evaluation with special emphasis on oncology; Development and validation of case-mix and patient-reported outcome measures; Education and health promotion research and programs. He has authored or co-authored over 200 publications. 8

11 Selected publications Fossati R, Apolone G, Negri E., Compagnoni A, La Vecchia C, Mangano S, Clivio L, Garattini S, A double-blind, placebo controlled, randomized trial of bupropion for smoking cessation in primary care, Arch Intern Med 2007; 67: Bertele' V, Banzi R, Capasso F, Tafuri G, Trotta F, Apolone G, Garattini S Haematological anticancer drugs in Europe: any added value at the time of approval?, Eur J Clin Pharmacol 2007; 63: Apolone G, Mangano S, Compagnoni A, Negri Emanuele, Mosconi P, Mannino S, Villa M, Zuccaro P, A multidisciplinary project to improve the quality of cancer pain management in Italy. Background, methods and preliminary results J Ambul Care Manage 2006; 29: Apolone G, La Vecchia C, Garattini S. Targeted kinase inhibitors in lung cancer: from EGFR to patients Eur J of Cancer, 2006; 42: Apolone G, Joppi R, Bertelè V & Garattini S. Ten years of marketing approvals of anti-cancer drugs in Europe. Regulatory policy and guidance documents need to find a balance between different pressures BJC 2005; 93: Gallus S, Colombo P, Apolone G, Zuccaro P, La Vecchia C. A tax to prevent the epidemic of lung cancer. Lancet 2005; 366: 288 Massimo Broggini followed the faculty of Science of the University of Milan, got the specialization in Biochemistry at Mario Negri Institute, and the PhD degree at the Open University, London,UK. He worked for a period in the laboratory of Molecular Pharmacology of the National Cancer Institute of Bethesda, Md, in From 1991 is the head of the Molecular Pharmacology Unit of the Mario Negri Institute and from 1999 of the Laboratory of Molecular Pharmacology of the same Institute. His main fields of interest are the study of the mechanism of action of new anticancer agents, the search of proteins and genes altered in human cancer and the study of oncosuppressor genes. He is member of the "Pharmacology and Molecular Mechanisms Group" of the European Organisation for the Research and Treatment of Cancer (EORTC) and of the American Association for Cancer Research. He is in the Editorial board of the European Journal of Cancer. He is author of more than 100 articles published in international journals. Principali pubblicazioni Zangrossi S, Marabese M, Broggini M, Giordano R, D'Erasmo M, Montelatici E, Intini D, Neri A, Pesce M, Rebulla P, Lazzari L. Oct-4 expression in adult human differentiated cells challenges its role as a pure stem cell marker. Stem Cells. 2007;25(7): Marrazzo E, Marchini S, Previdi S, Broggini M. Questioning the oncogenic role of DeltaNp73alpha in different cell lines expressing p53 or not. Cancer Biol Ther. 2006;5(7): Polato F, Codegoni A, Fruscio R, Perego P, Mangioni C, Saha S, Bardelli A, Broggini M. PRL-3 phosphatase is implicated in ovarian cancer growth. Clin Cancer Res : Maffucci T, Piccolo E, Cumashi A, Iezzi M, Riley AM, Saiardi A, Godage HY,Rossi C, Broggini M, Iacobelli S, Potter BV, Innocenti P, Falasca M. Inhibition of the phosphatidylinositol 3-kinase/Akt pathway by inositol pentakisphosphate results in antiangiogenic and antitumor effects. Cancer Res. 2005;65: Marabese M, Marchini S, Sabatino MA, Polato F, Vikhanskaya F, Marrazzo E, Riccardi E, Scanziani E, Broggini M. Effects of inducible overexpression of DNp73alpha on cancer cell growth and response to treatment in vitro and in vivo. Cell Death Differ. 2005;12: Sabatino MA, Colombo T, Geroni C, Marchini S, Broggini M. Enhancement of in vivo antitumor activity of classical anticancer agents by combination with the new, glutathione-interacting DNA minor groove-binder, brostallicin. Clin Cancer Res. 2003;9: Irene Floriani got her degree in Biological Sciences at the University of Milan in 1988, her degree in Biostatistics and Experimental Statistics at the University of Milan in 2003 and her phd in Life Sciences at Open University of London (UK) in After ten-year experience in pharmaceutical industry, in 2002 she became Head of the Biometry and Data Management Unit of the Laboratory of Clinical Research in Oncology and since 2006 she is Head of Laboratory of Clinical Trials. She is also member as bio-statistician of three Italian Ethics Committees. Her main fields of interest are: statistical aspects of methodology of clinical research with focus on Controlled Clinical Trials in Oncology; Systematic Overview of the medical literature and Methodological aspects of diagnostic test evaluation. Selected publications Cascinu S, Labianca R, Barone C, Santoro A, Carnaghi C, Cassano A, Beretta GD, Catalano V, Bertetto O, Barni S, Frontini L, Aitini E, Rota S, Torri V, Floriani I. Adjuvant treatment of high-risk, radically resected gastric cancer patients with 5-fluorouracil, leucovorin, cisplatin, and epidoxorubicin in a randomized controlled trial. J Natl Cancer Institute (2007); 99: Ruzzo A, Graziano F, Loupakis F, Rulli E, Canestrari E, Santini D, Catalano V, Ficarelli R, Maltese P, Bisonni R, Masi G, Schiavon G, Giordano P, Giustini L, Falcone A, Tonini G, Silva R, Mattioli R, Floriani I, and Magnani M. 9

12 Phamacogeneic profiling in patients with advanced colorectal cancer treated first-line FOLFOX-4 chemotherapy Annals of Oncology (2007); 25: Cazzaniga M, Mustacchi G, Pronzato P, De Matteis A, Di Costanzo F, Floriani I on behalf of the NORA Study Group Adjuvant treatment of early breast cancer: do the St Gallen recommendations influence clinical practice? Results from the NORA study. Annals of Oncology (2007); 18: Mandalà M, Reni M, Cascinu S, Barni S, Floriani I, Cereda S, Berardi R, Mosconi S, Torri V, Labianca R. Venous thromboembolism predicts poor prognosis in irresectable pancreatic cancer patients. Annals of Oncology (2007); 10: Mustacchi G, Cazzaniga ME, Pronzato P, De Matteis A, Di Costanzo F, Floriani I on Behalf of the NORA Study Group. Breast cancer in elderly women: a different reality? Results from the NORA study. Annals of Oncology (2007); 18: Gregorc V, Spreafico A, Floriani I, Colombo B, Ludovini V, Pistola L, Bellezza G, Vigano MG, Villa E, Corti A. Prognostic value of circulating chromogranin A and soluble tumor necrosis factor receptors in advanced nonsmall cell lung cancer. Cancer (2007); 110; Raffaella Giavazzi got her Biological Sciences degree (1979) at the University of Milan, and her Ph.D. in Pharmacology at the Mario Negri Institute of Milan (1984), followed by a specialization in pharmacology (1994) at the University of Milan. From 1981 to 1983 she was a Fellow in the Cancer Metastasis and Treatment Laboratory, NCI-FCRDC, Frederick, MD., and from 1983 to 1985 Assistant Professor at the Department of Cell Biology of M.D. Anderson Hospital and Tumour Institute, University of Texas System Cancer Centre in Houston (TX). Raffaella Giavazzi s research interests are in the field of tumour biology and pharmacology. Specifically, she is studying aspects related to the metastatic process and angiogenesis. She is involved in the preclinical evaluation of new therapeutic strategies against cancer focusing on the angiogenesis inhibitors and combination therapies. From 1986 to 1993 she was Head of the Cancer Metastasis Treatment Unit and since 1993 she has been the Head of the Laboratory of Biology and Treatment of Metastasis at Mario Negri Institute for Pharmacological Research. She is also adjutant Professor in Oncology, Medical School-University of Brescia, member of the Teaching Committee for the PhD course in Physiology-Pharmacology-Molecular and Cellular Tossicology-University of Siena, member of the Executive Committee at SENDO (South Europe New Drug Development Organization) and member of the Pezcoller Foundation Scientific Committee. She was consulting scientist for the NCI-Drug Therapeutics Program, USA ( ); She is a member of the American Association for Cancer Research (AACR), International Metastases Research Society, EORTC-Screening and Pharmacology Group, European Association for Cancer Research (EACR), Italian Cancer Society (SIC) of which she was President ( ). She is on the Editorial Board of the European Journal of Cancer, Journal of Clinical Experimental Metastasis, Journal Exp. Therapeutic & Oncology, The International Journal of Biological Markers, Current Cancer Therapy Reviews and Journal of Chemotherapy. She has published approximately 150 articles on peer reviewed journals and 34 book chapters. Selected publications Martinelli M., Bonezzi K., Riccardi E., Kuhn E., Frapolli R., Zucchetti M., Ryan A.J., Taraboletti G., Giavazzi R.: Sequence dependent antitumour efficacy of the vascular disrupting agent ZD6126 in combination with paclitaxel. British Journal of Cancer 97:888-94, Giavazzi R., Bani M.R.,Taraboletti G.: Tumor host interaction in the optimization of paclitaxel-based combination therapies with vascular targeting compounds. Cancer Metastasis Rev. 26:481 88, 2007 Naumova E., Ubezio P., Garofalo A., Borsotti P., Cassis L., Riccardi E., Scanziani E., Eccles S.A., Bani M.R., Giavazzi R.: The vascular targeting property of paclitaxel is enhanced by SU6668, a receptor tyrosine kinase inhibitor, causing apoptosis of endothelial cells and inhibition of angiogenesis. Clin. Cancer Research 12(6): , Bani M.R., Nicoletti M.I., Alkharouf N.W., Ghilardi C., Petersen D., Erba E., Sausville E.A., Liu E.T., Giavazzi R.: Gene expression correlating with response to paclitaxel in ovarian carcinoma xenografts. Molecular Cancer Therapeutics 3(2): , Belotti D., Paganoni P., Manenti L., Garofalo A., Marchini S., Taraboletti G., Giavazzi R.: Matrix Metalloproteinases (MMP9 and MMP2) Induce the Release of Vascular Endothelial Growth Factor (VEGF) by Ovarian Carcinoma Cells: Implications for Acites Formation. Cancer Res. 63: , Micheletti G., Poli M., Borsotti P., Martinelli M., Imberti B., Taraboletti G., Giavazzi R.: Vascular-targeting Activity of ZD6126, a Novel Tubulin-binding Agent. Cancer Res. 63: , Paola Mosconi got his Biological Science degree (Milan 1982) and the specialisation in Pharmacological Research (Milan 1984). Her main areas of interest are: development of strategies to involve patients- 10

13 consumer associations in health debate, and research projects; assessment of quality of life, translation and cultural adaptation of questionnaires for quality of life; studies to evaluate the type of information on diseases and treatments received by patients, mainly in cancer patients; set-up of websites targeted on consumers/patients studies to evaluate the consumers level of satisfaction with the health services and cure. Paola Mosconi has participated as a teacher, or coordinator, to the realization of training course on Methodological aspects of clinical research or Evaluation of quality of life for health care professionals and representatives of voluntary associations. Selected publications O Connel D, Mosconi P. An active role for patients in clinical research? Drug Development Research 67 (3): , Mosconi P, Colombo Cinzia, La Bianca R, Apolone G. Oncologists' opinions about research ethics committees in Italy: an update, Eur J Cancer Prev 2006; 15: Leone M A, Beghi E, Righini C, Apolone G, Mosconi P. Epilepsy and quality of life in adults: a review of instruments. Epilepsy Research 66: 23-44, Mosconi P, Poli P, Giolo A, Apolone G. How health consumers feel about clinical research: a questionnaire survey. European Journal of Public Health 15: , Mosconi P, Buchanan M, Kyriakides S, Fernandez-Marcos A, Horvatin J, O'Connell D, Zernik N, on behalf of EUROPA DONNA. EUROPA DONNA: has strength in its heterogeneity. European J Cancer 40: , Mosconi P, Apolone G. Fixed and dynamic health related quality of life measurements. J Headache Pain S31-S34, Domenighetti G, D Avanzo B, Egger M, Berrino F, Perneger T, Saracci R, Mosconi P. and M. Zwahlen.Women s perception of the benefits of mammography screening: population-based survey in four countries. Int J Epidem 32: , Valter Torri got his Medical degree in 1985 and the specialization in medical Oncology in 1989 at the University of Milano. Education: 1985: MD Degree with full honors cum Laude, University of Milano; 1988 Post-Doctoral Degree in Pharmacological Research, Mario Negri Institute, Milano; 1989 Post-Doctoral Degree in Medical Oncology, University of Milano; Research Fellow at the Biometric Research Branch of Cancer Treatment Evaluation Program, NCI, Bethesda, MD (USA) Areas of Interest: Statistical aspects of clinical research methodology with focus on Controlled Clinical Trials in Oncology; Systematic Overview of the medical literature; Methodological aspects of diagnostic test evaluation. Present Position: Head of Laboratory of Clinical Research In Oncology, Oncology Department, Mario Negri Institute, Milano. Chronology of Professional Appointments: : Clinical research Fellow in Internal Medicine at the University Hospital, University of Milan; : Research assistant at the Clinical Trial Unit of the Laboratory of Clinical Epidemiology, Mario Negri Institute for Pharmacological Research, Milano; : Research fellow at the Biometric Research Branch of Cancer Treatment Evaluation Program, NCI, Bethesda, MD (USA); 1994: Head of Biometric Unit of the Laboratory of Cancer Clinical Epidemiology, Oncology Department, Mario Negri Institute for Pharmacological Research, Milano, Italy; 1995 Vice Director of the Italian Cochrane Center; 2001: Head of Laboratory of Clinical Research In Oncology, Oncology Department, Mario Negri Institute, Milano. 2006: Head of Laboratory for the development of new pharmacological strategies, Oncology Department, Mario Negri Institute, Milano. Selected publications Mandalà M, Reni M, Cascinu S, Barni S, Floriani I, Cereda S, Berardi R, Mosconi S, Torri V, Labianca R. Venous thromboembolism predicts poor prognosis in irresectable pancreatic cancer patients. Ann Oncol. 2007;18(10): Malesci A, Laghi L, Bianchi P, Delconte G, Randolph A, Torri V, Carnaghi C, Doci R, Rosati R, Montorsi MRoncalli M, Gennari L, Santoro A. Reduced likelihood of metastases in patients with microsatellite-unstable colorectal cancer. Clin Cancer Res. 2007;13(13): Garassino MC, Hollander L, Torri V. Bevacizumab for non-small-cell lung cancer. N Engl J Med. 2007;356(13):1373 Cascinu S, Labianca R, Barone C, Santoro A, Carnaghi C, Cassano A, Beretta GD, Catalano V, Bertetto O, Barni S, Frontini L, Aitini E, Rota S, Torri V, FlorianiI; Italian Group for the Study of Digestive Tract Cancer, Adjuvant treatment of high-risk, radically resected gastric cancer patients with 5-fluorouracil, leucovorin, cisplatin, and epidoxorubicin in a randomised controlled trial. J Natl Cancer Inst. 2007;99(8): Graziano F, Kawakami K, Ruzzo A, Watanabe G, Santini D, Pizzagalli F, Bisonni R, Mari D, Floriani I, Catalano V, Silva R, Tonini G, Torri V, Giustini L, Magnani M Methylenetetrahydrofolate reductase 677C/T gene polymorphism, gastric cancer susceptibility and genomic DNA hypomethylation in an at-risk Italian population. Int J Cancer : Torri V: Clinical trials and data management In: Oxford textbook of oncology, 2nd. ed. Vol. 1. Oxford Univ. Press, Oxford; 2002 :

14 Maria Rosa Bani got her Biological Sciences degree at the University of Milan in 1998 attaining the Italian Government Qualification to practice as Biologist in She obtained the specialization in Pharmacological Research from the Department of Education of the Regional Government of Lombardia in 1991 and the specialization in Biomedical Research from the Department of Education of the Regional Government of Abruzzo in In 2005 she was awarded the degree of Doctor of Philosophy (PhD), Discipline of Life Sciences of the Open University Research School (UK). From 1991 to 1995 she was a Post Doctoral Fellow in the Cancer Research Division, Sunnybrook Health Science Centre, University of Toronto (Canada); from 2000 to 2001 she was Guest Scientist at the Advance Technology Centre, National Cancer Institute, National Institute of Health (USA). At the MarioNegri Institute for Pharmacological Research she was a Fellow Research Scientist in the Laboratory of Biology and Treatment of Metastasis, Bergamo from 1996 to 2002 and she became a Staff Research Scientist in In April 2004 she became Head of the Molecular Cancer Therapeutics Unit. Since 2004 she is the Scientific Manager of STROMA, an integrated project within the 6 th Framework Program of the European Commission [FP6 LSHC-CT , STROMA]. She is a member of the American Association for Cancer Research (AACR), Italian Cancer Society (SIC) and European Association for Cancer Research (EACR). Maria Rosa Bani research interests are in the field of cancer biology and molecular therapeutics. She is co-author of 32 peer reviewed publications, 2 book chapters and 55 proceedings of which 15 selected for oral presentation at international meetings. Selected publications Giavazzi R., Bani M.R.,Taraboletti G.: Tumor host interaction in the optimization of paclitaxel-based combination therapies with vascular targeting compounds. Cancer Metastasis Rev. 26:481 88, Bani M.R., Nicoletti M.I., Alkharouf N.W., Ghilardi C., Petersen D., Erba E., Sausville E.A., Liu E.T. and Giavazzi R. Gene expression correlating with response to paclitaxel in ovarian carcinoma xenografts. Molecular Cancer Therapeutics 3: , Vikhanskaya F.*, Bani M.R.*., Borsotti P., Ghilardi C., Ceruti R., Ghisleni G., Marabese M., Giavazzi R., Broggini M. & Taraboletti G. p73 overexpression increases VEGF and reduces thrombospondin-1 production: implication for tumor angiogenesis. Oncogene 20 : , Taraboletti G., Sonzogni L., Vergani V., Hosseini G., Ceruti R., Ghilardi C., Bastone A., Toschi E., Borsotti P., Scanziani E., Giavazzi R., Pepper M.S., Stetler-Stevenson W.G. & Bani M.R. Post-transcriptional stimulation of endothelial cell matrix metalloproteinases 2 and 1 by endothelioma cells. Experimental Cell Research 258 : , Alessandri G., Chirivi R.G.S., Fiorentini S., Dossi R., Bonardelli S., Giulini S.M., Zanetta G., Landoni F., Graziotti P.P., Turano A., Caruso A., Zardi L. Giavazzi R. & Bani M.R., Phenotypic and functional characteristics of tumor-derived microvascular endothelial cells. Clinical & Experimental Metastasis 17 : , Bani M.R., Rak J., Adachi D., Wiltshire R., Trent J.M., Kerbel R.S. & Ben-David Y. Multiple features of advanced melanoma recapitulated in tumorigenic variants of early stage (radial growth phase) human melanoma cell lines: evidence for a dominant phenotype. Cancer Research 5 : , Giovanna Damia obtained her Medical Degree cum Laude from the University of Milan in After specializing in Pharmacology at the Mario Negri Institute of Milan and in Oncology at the University of Milan, she worked as a post-doctoral fellow in the Laboratory of Experimental Immunology of the National Cancer Institute, Frederick, USA. She worked as a research fellow in the Laboratory of Cancer Chemotherapy at the Mario Negri Institute and since April 2003 she has become chief of the DNA Repair Unit at the Mario Negri Institute. From 1992 to1995 she has been consultant of the General Secretariat of the Progetto Finalizzato CNR "Applicazioni Cliniche della Ricerca Oncologica". Since September 2005 she is Deputy Editor for Experimental Oncology of the European Journal of Cancer. Her main fields of interest are: mechanism of action of anticancer drugs, cell cycle checkpoints and natural compounds. Selected publications Simone M, Erba E, Damia G, Vikhanskaya F, Di Francesco A M, Riccardi R, Baldeyrou B, Bailly C, Cuevas C, Sousa- Faro J M F, D'Incalci M. Variolin B and its derivate deoxy-variolin B: New marine natural compounds with cyclindependent kinase inhibitor activity. Eur J Cancer 2005; 41: Carrassa L, Broggini M, Erba E, Damia G. Chk1, but not Chk2, is involved in the cellular response to DNA damaging agents. Differential activity in cells expressing or not p53. Cell Cycle 2004; 3: Damia G, Broggini M. Improving the selectivity of cancer treatment by interfering with cell response pathways. Eur J Cancer 2004; 40: Damia G, Broggini M. Cell cycle checkpoint proteins and cellular response to treatment by anticancer agents. Cell Cycle 2004; 3:

15 Carrassa L, Broggini M, Vikhanskaya F, Damia G. Characterization of the 5' flanking region of the human chk1 gene. Identification of E2F1 functional sites. Cell Cycle 2003; 2: Damia G, Sanchez Y, Erba E, Broggini M. DNA damage induces p53-dependent down-regulation of hchk1. J Biol Chem 2001; 276: Eugenio Erba has obtained his Biological and Biochemmistry Analysis Degree at the University of Urbino. He worked as a research fellow in the Laboratory of Cancer Chemotherapy at the Mario Negri Institute and since 1984 he is head of the Flow Cytometry Unit in the Department of Oncology at the Mario Negri Institute of Milan. He has worked as a visiting fellow in the Department of Istochemistry and Cytochemistry of the University of Leiden, The Netherlands in Since 1997 he is Teacher of Post-Graduate Studies in Cytometry at the University of Milan and Co-ordinator and Teacher of Post- Graduate Studies in Cytometry for the Italian Cytometry Group. He has been President of the Italian Cytometry Group from 1999 to Since 2001 he is member of the Executive Board of the Italian Cytometry Group. Scientific areas of interest: studies on the mechanism of action of different compounds with provided antitumoral activity evaluating the mechanism of cell death and cell cycle phase perturbations induced on different human cancer cell lines by using flow cytometry. Co-ordinator of working-group in a quality control study on flow cytometric DNA content analysis in human tumors. Selected publications Paulis M., Bensi M., Orioli D., Mondello C., Mazzini G., D Incalci M., Falcioni C., Radaelli E., Erba E., Raimondi E., De Carli L. Transfer of a Human Chromosomal Vector from a Hamster Cell Line to a Mouse Embryonic Stem Cell Line. Stem Cell, 25: (2007) Tavecchio M., Natoli C., Ubezio P., Erba E., D Incalci M. Dynamics of cell cycle perturbations by trabectedin (ET- 743) in nucleotide excision repair (NER) deficient and NER-proficient cells, unravelled by a novel mathematical simulation approach. Cell Prolif., 40: (2007) Dolfini E., Roncoroni L., Dogliotti E., Sala G., Erba E., Sacchi N., Ghidoni R. Resveratrol impairs the formation of MDA- MB-231 multicellular tumor spheroids concomitant with ceramide accumulation. Cancer Lett 2007; 249: Tognon G., Bernasconi S., Celli N., Faircloth G.T. Cuevas C., Jimeno J., Erba E., D Incalci M. Induction of resistance to Aplidin in a human ovarian cancer cell line related to MDR expression. Cancer Biology and Therapy, 4(12): (2005). Minuzzo M., Ceribelli M., Pitarque-Martì M., Borrelli S., Erba E., di Silvio A., D Incalci M., Mantovani R. Selective effects of the anti-cancer drug Yondelis (ET-743) on cell-cycle promoters. Mol. Pharmacol., 68: (2005). Simone M., Erba E., Damia G., Vikhanskaya F., Di Francesco A.M., Riccardi R., Bailly C., Cuevas C., Fernandez Sousa-Faro J.M., D Incalci M. Variolin B and its derivate deoxy-variolin B: new marine natural compounds with a cyclin-dependent-kinase inhibitor activity. Eur. J. Cancer, 41: (2005). Allavena P., Signorelli M., Chieppa M., Erba E., Bianchi G., Marchesi F., Garbi A., Lissoni A., de Braud F., Jimeno J. and D Incalci M. Anti-inflammatory properties of the novel antitumor agent Yondelis (Trabectedin): inhibition of macrophage differentiation and cytokine production. Cancer REs., 65(7): (2005). Tognon G., Frapolli G., Zaffaroni M., Erba E., Zucchetti M., Faircloth G.T., M. D Incalci, Fetal bovine serum, but not human serum inhibits the in vitro cytotoxicity of ET-743 (Yondelis TM; trabectedin). An example of potential problems for extrapolation of active drug concentrations from in vitro studies. Cancer Chemother Pharmacol., 53(1): (2004). Erba E., Cavallaro E., Damia G., Mantovani R., Di Silvio A, Di Francesco A.M., Riccardi R., Cuevas C., Faircloth G.T., D Incalci M. The unique biological features of the marine product YondelisTM (ET-743, Trabectedin) are shared by its analog ET-637, which lacks the C ring. Oncology Research, 14: (2004). Roldano Fossati got his Medical Degree cum Laude from the University of Milan in 1980, his Post- Doctoral Degree in Endocrinolgy cum Laude from the University of Verona in 1983 and his Post- Doctoral Degree in Medical Statistics from the University of Milan in He has been consultant at the Mario Negri Institute since 1983 and, at present, he is head of the Gynecology and Oncology Unit of the Laboratory of Translational and Outcome Research. Areas of Interest: Statistical and methodologic aspects of clinical research with focus on Controlled Clinical Trials in Oncology; Systematic Overview of the medical literature. Selected publications F.E. Johnson, K.S. Virgo, R. Fossati. Follow-up for patients with colorectal cancer after curative-intent primary treatment. J Clin Oncol 2004; 22: Roberto Labianca, Roldano Fossati, Alberto Zaniboni, Valter Torri, Silvia Marsoni, Donato Nitti, Lamberto Boffi, Marco Scatizzi, Berardino Tardio, Nicola Mastrodonato, Stefano Banducci, Giampiero Consani, Gianfranco Pancera on behalf of ACOI/GIVIO/GISCAD investigators. Randomized Trial of Intraportal and/or Systemic Adjuvant Chemotherapy in Patients with Colon Carcinoma. J Natl Cancer Inst 2004; 96:750-8 P. Benedetti Panici, A. Maggioni, N. Hacker, F. Landoni, S. Ackermann, E. Campagnutta, K. Tamussino, R. Winter, A. Pellegrino, S. Greggi, R. Angioli, N. Manci, G. Scambia, T. Dell'Anna, R. Fossati, I. Floriani, R.S. Rossi, R. Grassi, G. 13

16 Favalli, F. Raspagliesi, D. Giannarelli, L. Martella, C. Mangioni. Systematic Aortic and Pelvic Lymphadenectomy versus Resection of Bulky Nodes Only in Optimally Debulked Advanced Ovarian Cancer: A Randomized Clinical Trial J Natl Cancer Inst 2005; 97:1-6 Buda A, Fossati R, Colombo N, Fei F, Floriani I, Gueli Alletti D, Katsaros D, Landoni F, Lissoni A, Calzoni C, Sartori E, Scollo P, Torri V, Zola P, Mangioni C. Randomized trial of neoadjuvant chemotherapy comparing paclitaxel, ifosfamide, and cisplatin with ifosfamide and cisplatin followed by radical surgery in patients with locally advanced squamous cell cervical carcinoma: The SNAP01 (Studio Neo-Adjuvante Por. J Clin Oncol 2005; 23: Maggioni A, Benedetti Panici P, Dell'anna T, Landoni F, Lissoni A, Pellegrino A, Rossi RS, Chiari S, Campagnutta E, Greggi S, Angioli R, Manci N, Calcagno M, Scambia G, Fossati R, Floriani I, Torri V, Grassi R, Mangioni C.Randomised study of systematic lymphadenectomy in patients with epithelial ovarian cancer macroscopically confined to the pelvis. Br J Cancer. 2006; 18;95(6): Maggi R, Lissoni A, Spina F, Melpignano M, Zola P, Favalli G, Colombo A, Fossati R. Adjuvant chemotherapy vs radiotherapy in high-risk endometrial carcinoma: results of a randomised trial. Br J Cancer. 2006; 7;95(3): Fruscio R, Colombo N, Lissoni AA, Garbi A, Fossati R, Ieda' N, Torri V, Mangioni C.A phase II randomised clinical trial comparing cisplatin, paclitaxel and ifosfamide with cisplatin, paclitaxel and epirubicin in newly diagnosed advanced epithelial ovarian cancer: long-term survival analysis. Br J Cancer. 2008; [Epub ahead of print] Marabese M, Marchini S, Marrazzo E, Mariani P, Cattaneo D, Fossati R, Compagnoni A, Signorelli M, Moll UM, Codegoni AM, Broggini M. Expression levels of p53 and p73 isoforms in stage I and stage III ovarian cancer. Eur J Cancer Jan;44(1): Marchini S, Marabese M, Marrazzo E, Mariani P, Cattaneo D, Fossati R, Compagnoni A, Fruscio R, Lissoni AA, Broggini M. {Delta}Np63 expression is associated with poor survival in ovarian cancer. Ann Oncol Nov 12; [Epub ahead of print] Fossati R, Apolone G, Negri E, Compagnoni A, La Vecchia C, Mangano S, Clivio L, Garattini S; for the General Practice Tobacco Cessation Investigators Group. A double-blind, placebo-controlled, randomized trial of bupropion for smoking cessation in primary care. Arch Intern Med. 2007; 10;167(16): Giulia Taraboletti got her degree cum laude in Biological Sciences at the University of Pavia (Pavia, Italy) in 1983, and the specialization in Pharmacological Research at the Mario Negri Institute, Milano, Italy in From 1986 to 1988 she was a post-doctoral fellow at the Laboratory of Pathology, NCI, NIH, Bethesda, MD, and from research scientist at Mario Negri Institute in Bergamo, Italy. Since 1995 she is Head of the Unit of Tumor Angiogenesis, at Mario Negri Institute, in Bergamo. Research interests include tumor angiogenesis, endogenous inhibitors of angiogenesis (thrombospondin- 1) and preclinical studies of antiangiogenic and vascular disrupting compounds, including tubulintargeting agents. She is member of Metatasis Research Society (MRS, Board of Directors), American Association for Cancer Research (AACR), European Association for Cancer Research (EACR), and the Italian Society of Oncology (SIC). She is on the editorial board of European Journal of Cancer. Selected publications Giavazzi R., Bani M.R.,Taraboletti G. Tumor host interaction in the optimization of paclitaxel-based combination therapies with vascular targeting compounds. Cancer Metastasis Rev, 26:481 88, Martinelli M., Bonezzi K., Riccardi E., Kuhn E., Frapolli R., Zucchetti M., Ryan A.J., Taraboletti G., Giavazzi R. Sequence dependent antitumour efficacy of the vascular disrupting agent ZD6126 in combination with paclitaxel. Br J Cancer 97:888-94, Margosio B., Marchetti D., Vergani V., Giavazzi R., Rusnati M., Presta M., and Taraboletti G. Thrombospondin-1 as a scavenger for matrix-associated fibroblast growth factor-2. Blood 102: , Taraboletti G, D Ascenzo S, Borsotti P, Giavazzi R, Pavan R, and Dolo V Shedding of MMP-2, MMP-9 and MT1- MMP as membrane vesicle-associated components by endothelial cells. Am J Pathol,160: , 2002 Taraboletti G. Micheletti G, Rieppi M, Poli M, Turatto M, Rossi C, Borsotti P, Roccabianca P, Scanziani E, Nicoletti MI, Bombardelli E, Morazzoni P, Riva A, and Giavazzi R. Antiangiogenic and antitumor activity of IDN 5390, a new taxane derivative. Clin Cancer Res. 8: , 2002 Taraboletti G., Morbidelli L., Donnini S., Parenti A., Granger H.J., Giavazzi R., and Ziche M.The heparin binding 25 kda fragment of thrombospondin-1 promotes angiogenesis and modulates gelatinase and TIMP-2 production in endothelial cells. FASEB J., 14: , Paolo Ubezio got his B.Sc. degree in Physics at the University of Milan, in 1982, and the specialisation in Pharmacological Research Specialist" at the Mario Negri Institute for Pharmacological Research in Main activities are: i) Study of cell-cycle mathematical models; ii) Development of flow cytometric methods; iii) Optimization of anticancer drug scheduling. Since 1991 is Head of the Unit of Biophysics at the Mario Negri Institute Selected publications Basse, B., Ubezio, P. (2007) A generalised age and phase structured model of human tumour cell populations both umperturbed and exposed to a range of cancer therapies. Bull. Math. Biol. 69:

17 Lupi, M., Matera, G., Natoli, C., Colombo, V., Ubezio, P. (2007) The Contribution of p53 in the Dynamics of Cell Cycle Response to DNA Damage Interpreted by a Mathematical Model. Cell Cycle 6: Spinelli, L., Torricelli, A., Ubezio, P., Basse, B. (2006) Modeling the balance between quiescence and cell death in normal and tumor cell populations Math Biosciences 202: Lupi, M., Matera, G., Branduardi, D., D'Incalci M. and Ubezio, P. (2004) Cytostatic and cytotoxic effects of topotecan decoded by a novel mathematical simulation approach. Cancer Res. 64: Matera, G., Lupi, M.,and Ubezio, P. (2004) Heterogeneous cell response to topotecan in a CFSE-based proliferation test. Cytometry 62A: Ubezio, P. (2004) Unraveling the complexity of cell cycle effects of anticancer drugs in cell populations.discrete and Continuous Dynamical Systems-Series B 4: Tomasoni, D., Lupi, M., Bekkal Brikci, F. and Ubezio, P. (2003) Timing the changes of cyclin E cell content in G1 in exponentially growing cells. Exp Cell Res.; 288: Montalenti, F., Sena, G., Cappella, P., and Ubezio, P. (1998) Simulating cancer-cell kinetics after drug treatment: Application to cisplatin on ovarian carcinoma. Phys. Rev. E, 57: Massimo Zucchetti obtained his Chem. Pharm. Degree from the University of Milan in After specializing in Pharmacology at the Mario Negri Institute of Milan (1988), he worked in the Laboratory of Clinical Pharmacology of Department of Oncology at San Giovanni Hospital, Bellinzona, Switzerland ( ). Since 1996 he has been chief of the Cancer Clinical Pharmacology Unit at the Mario Negri Institute. He is member of the Pharmacology and Molecular Mechanisms Group of the European Organization for Research and Treatment of Cancer (EORTC) from 1988 up to date. His main field of interest are: - Clinical pharmacology, phase I and Phase II studies - Analysis of drugs, pharmacokinetic and pharmacodynamic studies in humans in GCP and GLP conditions - Pharmacokinetic and metabolic studies in animals - Pharmacokinetic drug interaction Dr Zucchetti is author of more than 80 papers on pre-clinical and clinical cancer chemotherapy published in peer reviewed international journals. Selected publications Frapolli R., Marangon E., Zaffaroni M., Colombo T., Falcioni C., Bagnati R., Simone M., D Incalci M., Manzotti C., Fontana G., Morazzoni P., Zucchetti M. Pharmacokinetics and metabolism in mice of IDN 5390 (13-(N-Boc-3-ibutylisoserinoyl)-C-7,8-seco-10-deacetylbaccatin III), a new oral C-seco-taxane derivative with antiangiogenic property effective on paclitaxel-resistant tumors. Drug Metabolism and Disposition, 34(12): (2006). Rizzari C., Citterio M., Zucchetti M., Conter V., Chiesa R., Colombini A., Malguzzi S., D Incalci M. Pharmacological study on pegylated asparaginase used in front-line treatment of children with acute lymphoblastic leukemia. Hematologica, 91: (2006). Fruscio R., Lissoni A.A., Frapolli R., Corso S., Mangioni C., D Incalci M., Zucchetti M. Clindamycin-Paclitaxel pharmacokinetic interaction in ovarian cancer patients. Cancer Chemother. Pharmacol., 58(3): (2006). Gambacorti Passerini C, Zucchetti M, Russo D, Frapolli R, Verga M, Bungaro S, Tornaghi L, Rossi F, Pioltelli P, Pogliani E, Alberti D, Corneo G, D'Incalci M Alpha 1 acid glycoprotein binds to imatinib (STI571) and substantially alters its pharmacokinetics in chronic myeloid leukemia patients Clin Cancer Res 2003; 9: Pratesi G, Laccabue D, Lanzi C, Cassinelli G, Supino R, Zucchetti M, Frapolli R, D'Incalci M, Bombardelli E, Morazzoni P, Riva A, Zunino F IDN 5390: An oral taxane candidate for protracted treatment schedules Br J Cancer 2003; 88: Zaffaroni M, Frapolli R, Colombo T, Fruscio R, Bombardelli E, Morazzoni P, Riva A, D'Incalci M, Zucchetti M High-performance liquid chromatographic assay for the determination of the novel C-Seco-taxane derivative (IDN 5390) in mouse plasma. J Chromatogr B Analyt Technol Biomed Life Sci 2002; 780:

18 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The Oncology Department comprises three preclinical experimental laboratories (Laboratory of Cancer Pharmacology, Laboratory of Molecular Pharmacology and Laboratory of Biology and Therapy of Metastasis) and four laboratories dealing with clinical research and clinical trials (Laboratory for the Development of New Pharmacological Strategies, Laboratory of Clinical Trials, Laboratory of Translational and Outcome Research in Oncology and Laboratory for Medical Research and Consumer Involvement). The Oncology department hosts the coordination center of two networks of hospitals that carry on clinical research in gynecologic cancer (MaNGO: Mario Negri Gynecologic Oncology) and in cancer pain (CPOR-SG: Cancer Pain Outcome Research Study Group) and a center for cancer pain assessment and research (CERP:Center for the Evaluation and Research on Pain). In some cases research projects are carried out by single laboratories or research units, in other cases by collaborations between different laboratories of the Oncology Department or other departments, or other groups outside the Institute (see National and International Collaborations). Preclinical laboratories focus on the discovery and development of new antitumor and antimetastatic drugs and their new combinations; on tumor biology, not only to acquire new scientific knowledge, but particularly as a base for more selective therapeutic approaches and to identify and evaluate experimental models for discovering and studying new drugs or treatments. Clinical new drug development involves close participation in the activity of SENDO (South Europe New Drug Development Organization) and studies driven by the Laboratory of Cancer Pharmacology, the Laboratory of Molecular Pharmacology and the Laboratory of Biology and Therapy of Metastasis. The Laboratory for the Development of New Pharmacological Strategies, the Laboratory of Clinical Trials, the Laboratory of Translational and Outcome Research in Oncology and the Laboratory for Medical Research and Consumer Involvement are involved in the evaluation of the effects of new therapeutic modalities in phase I/II and in phase III comparative and effectiveness outcome studies. Outcome Research implies organizing trials to clarify the results of certain health care practices and interventions in clinical practice. Observational (surveys) and outcome research (effectiveness) studies are carried out, in collaboration with regional and national health authorities and other scientific associations. At the preclinical and clinical level there are studies of various human tumors, with particular emphasis on ovarian tumors and more recently on soft tissue sarcomas. FINDINGS/MAIN RESULTS At nanomolar concentrations, ET-743 (Yondelis, Trabectedin) affects the regulatory mechanisms of the transcription. Nucleotide excision repair deficient cells that are hypersensitive to UV rays and to other DNA damaging drugs are resistant to Trabectedin. Use of mathematical models of tumor growth and anticancer treatment to interpret experimental data and to manage the complexity of underlying biological phenomena. The theoretical relationship between proliferation, quiescence and cell loss leading to growth control of tumor cell populations was found. 16

19 Variolin and some of its derivatives induce rapid apoptosis in some tumor cell lines that have a low sensitivity to other anticancer drugs. Gene profiling analysis shows specific molecular signatures according to the histotype and prognosis of stage I ovarian carcinoma. The expression of a truncated form of p63 (DNp63) increases with the increased malignancy of ovarian cancer. Patients expressing high levels of DNp63 have a worst prognosis. DNp63 represents therefore a new potential target for selective therapies in this malignancy. A new and efficient system to selectively downregulate CHK1 in vivo has been developed. The system is suitable to test combinations of drugs in vivo and to study new checkpoints inhibitors. An anthracycline derivative, Nemorubicin, has a peculiar mechanism of action and is active against tumors resistant to drugs such as cisplatin. The combination of the two drugs is highly synergic. Embryo fibroblasts isolated from DRAGO KO mice show a reduced response to treatment with different anticancer agents.drago gene is particularly responsive to p73. Inositol pentaphosphate analogues interfere with the PI3-kinase-induced phosphorylation of akt and possess antitumor activity in vitro and in vivo. Human umbilical cord-derived stem cells express checkpoints proteins only in specific differentiation stages. It is likely that this is related to the different susceptibility of the cells. Inhibition of PLC gamma, through sirna technology, reduces the in vivo growth of tumors and reduces the formation of metastasis. Identification of genes preferentially expressed by tumor associated endothelial cells. VEGF released by cancer cells modulates the gene expression in the tumor microenvironment (stroma). The production of VEGF influence the response to paclitaxel of an ovarian carcinoma xenograft model; the anti-vegf antibody bevacizumab (Avastin ) is of benefit in improving therapeutic efficacy. VEGF produced by ovarian tumor cells stimulates host MMP9 expression; the anti-vegf antibody bevacizumab (Avastin ) inhibited MMP9 expression and abolished ovarian tumor invasion. A new antiangiogenic domain of thrombospondin-1 (an endogenous inhibitor of angiogenesis) that binds the angiogenic factor FGF-2 has been identified and characterized. New antineoplastic compounds directed against the tumor vasculature (vascular disrupting agents) have been selected. The sequence of drug administration determines the efficacy of combination treatments with tubulin-binding vascular disrupting agents and cytotoxic drugs. 17

20 The histone deacetylase inhibitor SAHA potentiates the cytotoxic effect of paclitaxel in human ovarian carcinoma cells resistant to paclitaxel. The effect is mediated by the acetylation of tubulin. The expression of protease-activated receptor-1 (PAR-1) correlates with the malignant phenotype of human melanomas and is accountable for their motility and invasise features ICON4, a, randomised trial of second-line chemotherapy in advanced ovarian cancer, coordinated by the Mario Negri Institute and by MRC, showed for the first time a reduction in mortality in favour of platinum and paclitaxel chemotherapy. The response to chemotherapy was a good surrogate endpoint of survival in patients with locally advanced cervix carcinoma. Adjuvant chemotherapy with the regimen vindesin, mitomycin C and cisplatin (MVP) did not improve survival of non small cell lung cancer (NSCLC) patients compared with surgery alone. The website of the project PartecipaSalute ( has a very innovative character in comparison with the other health Italian sites because introduces and develops with ad-hoc instruments the information transfer in an active way. The LYMPHADENECTOMY trial in advanced ovarian cancer: two decades of uncertainty resolved (editorial). This trial showed that systematic lymphadenctomy does not improve overall survival and these results will spare many patients the unduly toxicity of this surgical procedure. Bupropion more than doubled the odds of continuous abstinence from smoking from week 4 to 7 and from week 4 to 12 months in a way similar to that observed in academic studies. The adherence of GPs and participants to the protocol was excellent, making our findings robust and easy to generalize to the context of primary care. A randomised phase II trial showed that chemotherapic multidrug regimens containing either ifosfamide or epirubicin are effective in advanced ovarian cancer, though ifosfamide is more toxic. This trial had an exceedingly long follow up and showed a median overall survival noticeably higher than usually reported in similar clinical settings. Authors suggest that an aggressive therapeutic approach to this disease, with several lines of chemotherapies, could have benefitted these patients. A survey carried out in Italy that collected data in 110 clinical centers about 1801 patients showed that as many as 40% of patients with cancer enter the oncologic/palliative care programs while clearly undertreated mostly because of sub optimal use of morphine or other opiate drugs. Data collected at three-month follow up (1461 patients) allowed to describe the trend over time of many analgesic and palliative endpoints and to find the proportion of nonresponder to analgesic therapy (25-30%). Most of the drugs recently approved by the European Agency (EMEA) for hematologic malignancies had some information debt about their true risk-benefit profiles and, notwithstanding they are prescribed to thousands of patients. A survey of all the assessments carried out by EMEA over the last decade gave evidence that in as many as 2/3 of new drugs were approved by EMEA without a real therapeutic benefit over the old ones 18

21 A randomized trial of patients with high risk (stage IcG3, IIG3 with myometrial invasion >50%, and III) endometrial carcinoma showed the substantial equivalence between radiotherapy or chemotherapy as an adjuvant therapy after surgery. Although both radiotherapic and chemotherapic approaches are still unsatisfactory, since the risk of progression or death remains high, this encouraging evidence of clinical activity suggest a possible use of their concurrent or sequential use in an adjuvant setting. The estimates of the prevalence and impact of cancer pain in a large and representative sample of cancer patients (1800) recruited by several Italian centers (more than 120), with the evaluation of the actual proportion of cases that received a substantial analgesic under-treatment (about 25%), mainly attributable to a sub-optimal utilization of opiods. An evaluation of the activity of the EMEA over the last 10 years, has documented that most of the new anti-cancers drugs has actually received an approval on the basis of very preliminary findings: in 48% of case the approval was based on studies using surrogate endpoints, and in 40% of cases the design of the study was non-comparative and non-randomized. The activity of training and information organized with the associations of citizens & patients in the framework of the PartecipaSalute project has been finalized to the organization of the Parita task Participate to the research project with the associations. Parita is organised to discuss with the scientific community the grey areas of the medical assistance and clinical research identified from the patients and their associations, and to develop ad hoc protocols for future research programs. Development and validation of a new short questionnaire, the PGWBI-short version available to be used in large sample of citizens or patients. NATIONAL COLLABORATIONS ASR, Agenzia Sanitaria Regionale, Bologna AIFA, Agenzia Italiana del Farmaco (Roma) Assessorato Sanità, Regione Emilia Romagna Azienda Sanitaria Locale (Rimini) Azienda Sanitaria Unica Regionale, (Marche) Casa Sollievo della Sofferenza, San Giovanni Rotondo (IRCCS) CNPDS, Centro Nazionale per la prevenzione e Difesa Sociale, Milano CNR IGBE, Pavia CNPDS, Centro Nazionale per la prevenzione e Difesa Sociale, Milano CNR, Istituto di Chimica del Riconoscimento Molecolare, Milano Cochrane Collaboration EUROPA DONNA Federazione Italiana Società Scientifiche Fondazione LUVI, Milano Fondazione Nerina e Mario Mattioli Onlus, Milano Fondazione Salvatore Maugeri, Pavia Fondazione SmithKline (FSK), Milano Fondo Edo Tempia, Laboratorio di Farmacogenomica, Biella I.A.S.I., Roma Istituto Clinico Humanitas, Rozzano MI 19

22 Istituto Dermopatico dell'immacolata, Roma Istituto Ortopedico Galeazzi, Milano Istituti Ortopedici Rizzoli, Bologna Istituto dei Tumori di Milano Istituto Europeo di Oncologia (IEO), Milano Istituto di Fisica, Politecnico di Milano Istituto di Genetica Molecolare CNR, Sezione di Istochimica e Citometria, Pavia Istituto Nazionale per la Ricerca sul Cancro (IST), Genova Istituto Regina Elena, Roma Laboratorio Cell factory, Policlinico di Milano Ospedale San Gerardo, Monza, Milano Ospedale San Matteo, Pavia Unità di Tossicologia e Scienze Biomediche, ENEA Centro Ricerche, Roma Università Cattolica del Sacro Cuore, Roma Università di Bari Università di Brescia Università di Chieti Università di L Aquila Università di Milano Università di Modena e Reggio Emilia Università di Monza Università di Catania Università di Padova Università di Pisa Università di Siena Università La Sapienza, Roma Zadig, Agenzia di Giornalismo Scientifico INTERNATIONAL COLLABORATIONS ARCAGY (Association de Recherche sur les Cancers Gynécologiques), France Breakthrough Breast Cancer Center, Instutite of Cancer Reasearch, London, UK Cancer Biomarkers and Prevention Group, University of Leicester, UK Cancer Research UK, London, UK Cancerdegradome Consortium, IP 6th FP, EC EORTC, Bruxelles, Belgium EUROPA DONNA European Agency for the Evaluation of Medicinal Products (EMEA), London, UK European Regulatory Issues on Quality of life Assessment (ERIQA), Paris, France Executive Board of GCIG (Gynecologic Cancer Intergroup) Genome Institute of Singapore (GIS), Singapore German Cancer Research Center, Division of Toxicology and Cancer Risk Factors, Heidelberg, Germany Goteborg University, Lundberg Laboratory for Cancer Research, Goteborg, Sweden Helios Klinikum Erfurt GmbH, Institute of Pathology, Germany Istituto Oncologico della Svizzera Italiana Johns Hopkins University, USA Ludwig Institute for Cancer Research, London, UK National Cancer Center, Singapore 20

23 Stony Brook University, NY USA Massachusetts General Hospital and Harvard Medical School, USA MD Anderson Cancer Center, Houston, Texas, USA MRC, London, UK National Cancer Institute (NCI), Bethesda and Frederick, MD, USA Ospedale San Giovanni, Bellinzona, Svizzera Paterson Institute for Cancer Research, Manchester, UK Southern Europe New Drug Organization (SENDO), Milan, Italy Stroma Consortium, IP 6th FP, EC Swiss Federal Institute of Technology, Zurigo, Swizeland The Sackler Institute, University College London, UK Tumor Biology and Metastasis Institute of Cancer Research, Sutton, UK University College, London Medical School, London, UK University of Birmingham, UK University of Cincinnati, USA University of Crete Medical School, Greece University of Newcastle, UK University of Pau, Francia University of Wisconsin, Madison, WI, USA Kyoto University, Japan Weizmann Institute of Science, Israel EDITORIAL BOARD MEMBERSHIP Attualità in Senologia (Paola Mosconi) British Journal of Cancer (Maurizio D Incalci) Chemotherapy (Maurizio D Incalci) Clinical Experimental Metastasis (Raffaella Giavazzi) Current Opinion in Oncologic, Endocrine and Metabolic Drugs (Maurizio D Incalci) Current Cancer Therapy Reviews (Raffaella Giavazzi) European Journal of Cancer (Maurizio D Incalci, Giovanna Damia, Raffaella Giavazzi,Massimo Broggini and Giulia Taraboletti) Health and Quality of Life Outcomes (Giovanni Apolone, Paola Mosconi) Health and Quality of Life Outcomes (Giovanni Apolone, Paola Mosconi) International Journal of Biological Markers (Raffaella Giavazzi) International Journal for Quality in Health Care (Giovanni Apolone) Journal of Ambulatory Care and Management (Giovanni Apolone) Journal of B.U.ON. (Maurizio D Incalci) Journal of Chemotherapy (Raffaella Giavazzi) Journal of Experimental Therapeutics and Oncology (Raffaella Giavazzi) Journal of Medicine and the Person (Giovanni Apolone) Journal of Preventive Medicine and Hygiene (Giovanni Apolone) Molecular Cancer Therapeutics (Maurizio D Incalci) Oncology Research (Maurizio D Incalci) Tumori (Maurizio D Incalci, Raffaella Giavazzi) (Paola Mosconi) (Paola Mosconi) 21

24 PEER REVIEW ACTIVITIES American Journal of Pathology, Annals of Oncology, Anti-cancer Drugs, Biochemical Pharmacology, British Journal of Cancer, British Journal of Pharmacology, British Medical Journal, Cancer Chemotherapy and Pharmacology, Cancer Detection and Prevention, Cancer Letters, Cancer Research, Carcinogenesis, Chemico-Biological Interactions, Clinical & Experimental Metastasis, Clinical Cancer Research, Cytometry, European Journal of Cancer, European Journal of Immunology, Faseb Journal, Gynecologic Oncology, Health and Quality of Life Outcomes, Intensive Care Medicine, International Journal of Biological Markers, International Journal of Cancer, International Journal for Quality in Health Care, Journal of Ambulatory Care and Management, Journal of Biological Chemistry, Journal of Biological Markers, Journal of Cell Biochemistry, Journal of Clinical Oncology, Journal of Experimental Therapeutics and Oncology, Journal of Medicine and the Person, Journal of the National Cancer Institute, Journal of Neurology, Journal of Preventive Medicine and Hygiene, Journal of the National Cancer Institute, Leukaemia, Molecular Cancer Therapeutics, Nature Biotechnology, Nature Reviews, Oncology Research, PharmacoEconomics, Quality of Life Research, Science, Tumori NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Ethical Committee, Centro di Riferimento Oncologico, Aviano PN, Italy Ethical Committee, Ente Ospedaliero San Paolo, Milan, Italy Ethical Committee, Istituto Europeo di Oncologia, Milan, Italy Ethical Committee, Istituto Neurologico Carlo Besta, Milan, Italy Ethical Committee, Istituto Scientifico Eugenio Medea, Bosisio Parini, Lecco, Italy Ethical Committee, Ospedale San Gerardo, Monza, Milan, Italy Ethical Committee, Ospedale Sant Anna, Como, Italy Ethical Committee, Ospedale della Valtellina e Valchiavenna, Sondrio, Italy Ethical Committee, IRCCS MultiMedica, Sesto San Giovanni, Milan, Italy Executive Board of GCIG (Gynecologic Cancer Intergroup) Scientific Committee, Associazione Italiana Ematologia e Oncologia Pediatrica, Monza, Milan, Italy Scientific Committee, Pezcoller Foundation, Trento, Italy Technical-Scientific Commitee, Associazione Italiana per la Ricerca sul Cancro, Milan, Italy Board of Directors, Metastasis Research Society (MRS) Board of Directors, Società Italiana di Cancerologia (SIC) Board of Directors, Società Italiana di Citometria (GIC) Directional Council Areas-CCI National Advisory Board 8th World Congress of Psycho-Oncology Developmental Therapeutics Program, National Cancer Institute (NCI) Decision Network and Executive Committee, South Europe New Drug Organization (SENDO) Executive Board, Europa Donna External Scientific Committee, Angiotargeting Consortium EU Sixth Framework Programme, University of Bergen, Norvegia NHS R&D National Coordinating Centre for Health Technology Assessment, UK Scientific Committee, Swiss Cancer League University Medical School of Siena, Italy 22

25 EVENT ORGANIZATION Conference: International Conference on Vascular Targeted Therapies in Oncology. Mandelieu (France), October 4-6, Investigator Meeting Cancer Pain Istituto di Ricerche Farmacologiche Mario Negri, december 17, Milan PARTICIPATION IN EVENTS IN WHICH THE DEPARTMENT WAS INVOLVED Meeting: 28th Winter PAMM Meeting New vicinal diaryl-substituted imidazole derivatives with vascular disrupting activity. Berlin (Germany), 31 January - 3 February Conference: 4th International Conference on Tumor Microenvironment: Progression, Therapy & Prevention. Identification of novel markers of tumor vasculature. Florence (Italy), 6 March - 10 March Conference: XVI Convegno Nazionale M. Gioia - Le nuove frontiere del diritto e della medicina legale: quale errore nella medicina estrema? Identificazione delle responsabilità nel percorso tra ricerca preclinica e l applicazione clinica in ambito oncologico. Pisa (Italy), 25 May -26 May Conference: European Conferences on Biomedical Optics (ECBO). Study of antiangiogenic drugs by fluorescence imaging and spectroscopy of a contrast agent in mice. Munich (Germany), 17 June - 21 June Symposium: International Symposium on New Directions in Cancer Management Targeting the Tumor Vasculature. strategies for Combination Therapy. Buenos aires (Argentina), 6 June - 8 June Meeting: V Meeting NIBIT- Network Italiano per la Bioterapia del Tumori. Angiogenic Targets and Combination Therapies. Siena (Italy), 21 September Conference: XXV National Conference of Citometria La Citometria nell era delle Biotecnologie, Pontificia Università Lateranense, Rome, Città del Vaticano, 3-6 October Conference: International Conference on Vascular Targeted Therapies in Oncology. Pharmacological approaches with vascular Targeting Agents in Combinations with Chemiotherapy. Mandelieu (France), 4 October - 6 October Conference: International Conference on Vascular Targeted Therapies in Oncology. VEGF released by ovarian cancer cells stimulates organ specific MMP9 expression and invasion. Mandelieu (France), 4 October - 6 October

26 Conference: International Conference on Vascular Targeted Therapies in Oncology. Molecular characterization of tumor endothelium and identification of novel markes. Mandelieu (France), 4 October -6 October Conference: International Conference on Vascular Targeted Therapies in Oncology. Binding of thrombospondin-1 type III repeats to fibroblast grown factor-2 as an exploitable mechanism of angiogenesis inhibition. Mandelieu (France), 4 October - 6 October Conference: International Conference on Vascular Targeted Therapies in Oncology. New tubulin-binding imidazole derivatives as vascular disrupting agents. Mandelieu (France), 4 October - 6 October Conference: International Conference on Vascular Targeted Therapies in Oncology. Molecular profile of stroma derived from human ovarian carcinoma xenograft tumors equipped of different angiogenic phenotypes. Mandelieu (France), 4 October - 6 October Meeting: IX Congresso Nazionale di Oncologia Medica (AIOM). Endpoints Farmacologici nello sviluppo e nell applicazione clinica degli agenti antiangiogenici. Palermo (Italy),12 October - 15 October Conference: AACR/NCI/EORTC International Conference Molecular Targets and Cancer Therapeutics: Discovery, Biology and Clinical Applications. Influence of VEGF production on paclitaxel response in a model of ovarian carcinoma xenograft. S. Francisco (USA), 22 October - 26 October Conference: 4th International Conference on Thrombosis and Hemostasis Issues in Cancer. Protease-activated receptor-1 (PAR-1) expression correlates with a malignant phenotype in human melanoma. Bergamo (Italy), 26 October - 28 October Gynecologic Cancer Intergroup (GCIG), General Assembly October 28, 2007, Charité Ruine, Campus Charité Mitte, Berlin VISEAR II, Vienna Initiative to Save European Academic Research II Vienna, november 12, 2007 Congress: 24 Congresso della Società Italiana di Chemioterapia. Pharmacological endpoints in the development of angiogenesis inhibitors. Verona (Italy), 25 November - 28 November Meeting: 49th Annual Meeting of the Italian Cancer Society (SIC) Modulation of endothelial cell gene expression by angiogenic factors. Pordenone (Italy), 26 November - 29 November Meeting: 49th Annual Meeting of the Italian Cancer Society (SIC) Modulation of tubulin acetylation affects the sensitive of endothelial cells to the antimotility effects of paclitaxel. Pordenone (Italy), 26 November - 29 November Congress: Swiss Proteomic Society Large scale comparative proteomic study of accessible vascular proteins in mouse liver metastases and normal liver. Losanna (Switzerland), 3 December -5 December

27 GRANTS AND CONTRACTS Agenzia Italiana del Farmaco AIL Associazione Italiana contro le Leucemie, Padova Amgem SpA, Milan AIRC Associazione Italiana per la Ricerca sul Cancro ASL Padova ASL Provincia di Lodi Astra Zeneca SpA Astra Zeneca UK Azienda Sanitaria Locale - Rimini Azienda Sanitaria Unica Regionale - Marche Bracco Imaging SpA, Milan Centro Cochrane Italiano Chiesi Farmaceutici SpA CNPDS, Centro Nazionale per la prevenzione e Difesa Sociale, Milano CNR Consiglio Nazionale delle Ricerche CNR-MIUR Ministero Istruzione Università e Ricerca Compagnia di San Paolo, Torino CTI Cell Therapeutics, Inc. Cyclacel Ltd. Dompé Eli Lilly Italia SpA Elsevier Science Ltd. EORTC-European Organization for Research and Treatment of Cancer EOS SpA European Commission - 6th Framework Programme (Cancerdegradome, STROMA) FIRB-MIUR Fondo per gli Investimenti della Ricerca di Base-Ministero Istruzione Università e Ricerca FIRC Fondazione Italiana per la Ricerca sul Cancro Fondazione Cassa di Risparmio delle Province Lombarde Fondazione Lu.V.I. Fondazione Nerina e Mario Mattioli Onlus Fondo Edo Tempia Grunenthal, Milano GlaxoSmithKline, Verona Indena SpA Institut de Recherche Pierre Fabre Istituto Superiore di Sanità Italfarmaco Komen Italia Onlus Lottomatica Madaus Srl Medac Merck Sharp & Dome Ministero della Sanità NCI SAIC Frederick Nerviano Medical Science S.r.l. 25

28 Novartis Farma SpA Optigenex Inc. Pfizer Global Research and Development Pharma Mar, SA Pharminox Ltd, UKPoliclinico di Padova / C.O.R. PTC Pharma AG Regione Emilia Romagna Regione Veneto Sara Bet, Roma SENDO-Tech Srl Sigma-Tau SpA Università degli Studi di Padova SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Mandalà M, Reni M, Cascinu S, Barni S, Floriani I, Cereda S, Berardi R, Mosconi S, Torri V, Labianca R. Venous thromboembolism predicts poor prognosis in irresectable pancreatic cancer patients. Ann Oncol 2007; 18: Cascinu S, Labianca R, Barone C, Santoro A, Carnaghi C, Cassano A, Beretta GD, Catalano V, Bertetto O, Barni S, Frontini L, Aitini E, Rota S, Torri V, Floriani I; Italian Group for the Study of Digestive Tract Cancer, Pozzo C, Rimassa L, Mosconi S, Giordani P, Ardizzoia A, Foa P, Rabbi C, Chiara S, Gasparini G, Nardi M, Mansutti M, Arnoldi E, Piazza E, Cortesi E, Pucci F, Silva RR, Sobrero A, Ravaioli A. Adjuvant treatment of high-risk, radically resected gastric cancer patients with 5-fluorouracil, leucovorin, cisplatin, and epidoxorubicin in a randomized controlled trial. J Natl Cancer Inst 2007; 99: Gregorc V, Spreafico A, Floriani I, Colombo B, Ludovini V, Pistola L, Bellezza G, Vigano MG, Villa E, Corti A. Prognostic value of circulating chromogranin A and soluble tumor necrosis factor receptors in advanced nonsmall cell lung cancer. Cancer 2007; 110: Mustacchi G, Cazzaniga ME, Pronzato P, De Matteis A, Di Costanzo F, Floriani I on Behalf of the NORA Study Group. Breast cancer in elderly women: a different reality? Results from the NORA study. Ann Oncol 2007; 18: Mosconi P., Colombo C., Satolli R., Liberati A. PartecipaSalute, an Italian project to involve lay people, patients associations and scientific-medical representatives on the health debate. Health Expect 2007; 10: Gallus S., Zuccaro P., Colombo P., Apolone G., Pacifici R., Garattini S., Bosetti C., La Vecchia C. Smoking in Italy : Effects of a comprehensive National Tobacco Regulation. Prev Med 2007; 45: Fossati R., Apolone G., Negri E., Compagnoni A., La Vecchia C., Mangano S., Clivio L., Garatini S., General Practice Tobacco Cessation Investigators Group. Arch Intern Med 2007; 167: Bianchi R., Gilardini A., Rodriguez-Menendez V., Oggioni V., Canta A., Colombo T., De Michele G., Martone S., Sfacteria A., Piedemonte G., Grasso G., Beccaglia P., Ghezzi P., D Incalci M., Lauria G., Cavaletti G. Cisplatininduced peripheral neuropathy: neuroprotection by erythropoietin without affecting tumour growth. Eur J Cancer 2007; 43: Salvati E., Leonetti C., Rizzo A., Scarsella M., Mottolese M., Galati R., Sperduti I., Stevens M.F.G., D Incalci M., Blasco M., Chiorino G., Bauwens S., Horard B., Gilson E., Stopacciaro A., Zupi G., Biroccio A. Telomere damage induced by the G-quadruplex ligand RHPS4 has an antitumor effect. J Clin Invest 2007; 117: Tavecchio M., Natoli C., Ubezio P., Erba E., D'Incalci M. Dynamics of cell cycle phase perturbations by trabectedin (ET-743) in nucleotide excision repair (NER)-deficient and NER-proficient cells, unravelled by a novel mathematical simulation approach. Cell Prolif 2007; 40: Lupi M., Matera G., Natoli C., Colombo V., Ubezio P. The contribution of p53 in the dynamics of cell cycle response to DNA damage interpreted by a mathematical model. Cell Cycle 2007; 6:

29 Sessa C., Cresta S., Cerny T., Baselga J., Rota Caremoli E., Malossi A., Hess D., Trigo J., Zucchetti M., D'Incalci M., Zaniboni A., Capri G., Gatti B., Carminati P., Zanna C., Marsoni S., Gianni L. Concerted escalation of dose and dosing duration in a phase I study of the oral camptothecin gimatecan (ST1481) in patients with advanced solid tumors. Ann Oncol 2007; 18: D'Incalci M., Steward W.P., Gescher A.J. Modulation of response to cancer chemotherapeutic agents by diet constituents. Is the available evidence sufficiently robust for rational advice for patients? Cancer Treat Rev 2007; 33: Bagnati R., Bianchi G., Marangon E., Zuccato E., Fanelli R., Davoli E. Direct analysis of isopropylthioxanthone (ITX) in milk by high-performance liquid chromatography/tandem mass spectrometry. Rapid Commun Mass Spectrom 2007; 21: Cavaletti G., Gilardini A., Canta A., Rigamonti L., Rodriguez-Menendez V., Ceresa C., Marmiroli P., Bossi M., Oggioni N., D'Incalci M., De Coster R. Bortezomib-induced peripheral neurotoxicity: A neurophysiological and pathological study in the rat. Exp Neurol 2007; 204: Gordon M.O., Torri V., Miglior S., Floriani I., Miller J.P., Gao F., Adamsons F., Poli D., D'Agostino R.B., Kass M.A. Validated prediction model for the development for the development of primary open-angle glaucoma in individuals with ocular hypertension. Ophthalmology 2007; 114: Basse B., Ubezio P. A generalised age- and phase-structured model of human tumour cell populations both unperturbed and exposed to a range of cancer terapies. Bull Math Biol 2007; 69: Vikhanskaya F., Lee M.K., Mazzoletti M., Broggini M., Sabapathy K. Cancer-derived p53 mutants suppress p53- target gene expression-potential mechanism for gain of function of mutant p53. Nucleic Acids Res 2007; 35: D'Incalci M., Brunelli D., Marangon E., Simone M., Tavecchio M., Gescher A., Mantovani A. Modulation of gene transcription by natural products. A viable anticancer strategy? Current Pharmaceutical Design 2007; 13: Di Francesco A.M., Meco D., Torella A.R., Barone G., D'Incalci M., Pisano C., Carminati P., Riccardi R. The novel atypical retinoid ST1926 is active in ATRA resistant neuroblastoma cells acting by a different mechanism. Biochem Pharmacol 2007; 73: Bertele' V., Banzi R., Capasso F., Tafuri G., Trotta F., Apolone G., Garattini S. Haematological anticancer drugs in Europe: any added value at the time of approval? Eur J Clin Pharmacol 2007; 63: Grassi R., Lombardi G., Reginelli A., Capasso F., Romano F., Floriani I., Colacurci N. Coccygeal movement: Assessment with dynamic MRI. Eur J Radiol 2007; 61: Torri V., Floriani I., Poli D., European Glaucoma Prevention Study Group (EGPS). Central corneal thickness in the European Glaucoma Prevention Study. Ophthalmology 2007; 114: Gambacorti Passerini C., Tornaghi L, Marangon E., Franceschino A., Pogliani E., D'Incalci M., Zucchetti M. Imatinib concentrantrations in human milk. Blood 2007; 109: 1790 Ruzzo A., Graziano F., Loupakis F., Rulli E., Canestrari E., Santini D., Catalano V., Ficarelli R., Maltese P., Bisonni R., Masi G., Schiavon G., Giordani P., Giustini L., Falcone A., Tonini G., Silva R., Mattioli R., Floriani I., Magnani M. Pharmacogenetic profiling in patients with advanced colorectal cancer treated with first-line FOLFOX-4 chemotherapy. J Clin Oncol 2007; 25: European Glaucoma Prevention Study Group (EGPS), Torri V., Floriani I., Rulli E., Poli D. Predictive factors for open-angle glaucoma among patients with ocular hypertension in the European Glaucoma Prevention Study. Ophthalmology 2007; 114: 3-9. Grosso F., Jones R.L., Demetri G.D., Judson I.R., Blay J-Y., Le Cesne A., Sanfilippo R., Casieri P., Collini P., Dileo P., Spreafico C., Stacchiotti S., Tamborini E., Tercero J.C., Jimeno J., D'Incalci M., Gronchi A., Fletcher J.A., Pilotti S., Casali P.G. Efficacy of trabectedin (ecteinascidin-743) in advanced pretreated myxoid liposarcomas: a retrospective study. Lancet Oncol 2007; 8:

30 Zangrossi S., Marabese M., Broggini M., Giordano R., D'Erasmo M., Montelatici E., Intini D., Neri A., Pesce M., Rebulla P., Lazzari L. Oct-4 expression in adult human differentiated cells challenges its role as a pure stem cell marker. Stem Cells 2007; 25: Martinelli M., Bonezzi K., Riccardi E., Kuhn E., Frapolli R., Zucchetti M., Ryan A.J., Taraboletti G., Giavazzi R. Sequence dependent antitumor efficacy of the vascular disrupting agent ZD6126 in combination with paclitaxel Br J Cancer 2007; 97: Giavazzi R., Bani M.R., Taraboletti G. Tumor-host interaction in the optimization of paclitaxel-based combination therapies with vascular targeting compounds. Cancer Metastasis Rev 2007; 26: Marabese M., Vikhanskaya F., Broggini M. p73: A chiaroscuro gene in cancer. Eur J Cancer 2007; 43: Damia G., D'Incalci M. Targeting DNA repair as a promising approach in cancer therapy. Eur J Cancer 2007; 43: Dolfini E., Roncoroni L., Dogliotti E., Sala G., Erba E., Sacchi N., Ghidoni R. Resveratrol impairs the formation of MDA-MB-231 multicellular tumor spheroids concomitant with ceramide accumulation. Cancer Lett 2007; 249: Apolone G., Mosconi P. Techniques for assessing the quality of life with a particular emphasis on physical exercise. Springer-Verlag Italia, Milano, 2007; Mosconi P., Satolli R., Colombo C., Liberati A., Donati S., Mele A. Hormone replacement therapy and information: in Italy a Consensus Conference to help woman decision. BMJ Malesci A., Laghi L., Bianchi P., Delconte G., Randolph A., Torri V., Carnaghi C., Doci R., Rosati R., Montorsi M., Roncalli M., Gennari L., Santoro A. Reduced likelihood of metastases in patients with microsatellite-unstable colorectal cancer. Clin Cancer Res 2007; 13: Miglior S., Torri V., Zeyen T., Pfeiffer N., Vaz J.C., Adamsons I., European Glaucoma Prevention Study Group (EGPS) Intercurrent factors associated with the development of open-angle glaucoma in the European Glaucoma Prevention Study. Am J Ophthalmol 2007; 144: Ubezio P., Lupi M., Matera G. Antiproliferative activity of cisplatin detected by CFSE in p53-proficient and p53- deficient cells. Immunol Invest 2007; 36: Paulis M., Bensi M., Orioli D., Mondello C., Mazzini G., D Incalci M., Falcioni C., Radaelli E., Erba E., Raimondi E., De Carli L. Transfer of a Human Chromosomal Vector from a Hamster Cell Line to a Mouse Embryonic Stem Cell Line. Stem Cell, 2007; 25:

31 LAY PRESS SELECTION PUBLISHED IN 2007 Apolone G, Colombo Cinzia Il dolore non necessario Partecipa Salute 2007 Apolone G, Mosconi P Le agenzie regolatorie: FDA, EMEA e AIFA In : La dispensa di PartecipaSalute, 2007; Apolone G Il dolore nel paziente con cancro: attività in corso e primi risultati Cancer Aging 2007; 5: s10-s12 Mosconi P, Colombo Cinzia Quale informazione per la donna in menopausa sulla terapia ormonale sostitutiva? Rispondere attraverso una Conferenza di Consenso Newsletter Ospedali Riuniti di Bergamo, Number 11 - September 2007; 7 Colombo Cinzia, Mosconi Paola L associazionismo sta cambiando: dall assistenza alla nascita di un progetto In: La dispensa di PartecipaSalute, Mario Negri Institute 2007; Mosconi P, Colombo Cinzia Dalla parte dei cittadini: al via il progetto farmaci equivalenti: facciamo chiarezza Newsletter Ospedali Riuniti di Bergamo, Number 8 - June 2007; 7 Mosconi P, Colombo Cinzia, Satolli R, Liberati A. La ricerca clinica risponde ai bisogni dei pazienti? Risultati di un indagine di PartecipaSalute Ricerca & Pratica 2007;23: Mosconi P. Quando la medicina arriva prima della malattia. Consumers Magazine Ottobre 2007 pag. 5 Mosconi P. Esperienze internazionali di coinvolgimento di associazioni di pazienti nella ricerca Le Donne per le Donne, Centro Congressi Villa Olmo (CO), September 2007; Mosconi P. Discussione: il coinvolgimento del paziente nei sistemi sanitari In: Rischio clinico e sicurezza del paziente. Modelli e soluzioni nel contesto internazionale. Società editrice Il Mulino, Bologna, March 2007 Mosconi P. La Commissione oncologica nazionale e la rappresentanza dei pazienti: occasione persa? ; 24 Agoust Mosconi P, Colombo Cinzia, Satolli R, Liberati A. Il corso di PartecipaSalute 2007 allo specchio: i commenti di chi ha organizzato ; 2007; 12 June Mosconi P, Colombo Cinzia, Satolli R. Le associazioni di pazienti e la ricerca clinica: un indagine di PartecipaSalute ; 2007; 28 May 29

32 Mosconi P, Colombo Cinzia I tuoi diritti quando partecipi alla ricerca clinica ; 2 February Mosconi P, Colombo Cinzia Dalla parte dei cittadini: la ricerca clinica risponde ai bisogni dei pazienti? Dati presentati durante la seconda giornata internazionale della ricerca clinica Newsletter Ospedali Riuniti di Bergamo, Numero 7 - May 2007; 6 Mosconi P, Colombo Cinzia Clinical Trial Day 2007: seconda giornata internazionale della ricerca clinica ; 2007; 31 May Mosconi P, Apolone G Sei domande sui comitati etici ; 15 January 30

33 RESEARCH ACTIVITIES Laboratory of Cancer Pharmacology Mode of action of Ecteinascidins A project ongoing since several years is about the characterization of marine natural products possessing antitumor activity. In particular we carried on the studies on the effects of ET-743 in cells defective for some DNA repair mechanisms. Cells deficient for Homologous Recombination (HR) are very sensitive to the drug, while cells deficient for Non Homologous End-Joining (NHEJ) are only slightly more sensitive, but surprisingly cell lines defective for Nucleotide Excision Repair (NER) are less sensitive to ET-743. Flow cytometric analysis coupled to a software of computer simulation, developed in our laboratory, has demonstrated that NER defective cells showed, after ET-743 treatment, cell cycle perturbations different than those occurring in NER proficient cells, probably for the activation of different and more efficient repair mechanisms. We study also a functional evaluation of the DNA repair mechanisms by the cell capacity to recognize and repair double helix breaks with a recently introduced test that is very sensitive to detect the phosphorylation of histone H2AX. An in vitro study is ongoing with flow cytometry and immunofluorescence techniques to evaluate in different tumor cell lines the phosphorylation level of histone H2AX in relation to the distribution of the cells in the different phases of the cell cycle and the cytotoxic effect induced after treatment with ET-743. Studies are in progress on the mechanism of action of new ET-743 derivates compounds that have shown antitumoral activity on cell lines with different DNA repair mechanisms. A new project is the study of the selective action of ET-743 on mixoid lyposarcoma, a pathology representing 10% of all soft tissue sarcomas, trying to understand if the significative antitumor effect is due to a selective action of the compound on pathogenetic alterations characteristic of this pathology. In particular we are trying to evaluate how ET-743 interact with the transcriptional modifications of specific genes due to the translocation FUS-CHOP that characterizes mixoid sarcomas or those caused by the interaction host-tumor, modifying inflammatory and angiogenetic processes. Studies are in progress to obtain cell lines and xenografts of mixoid lyposarcomas exhibiting the same molecular features of the patients tumors. Combinations of natural products of marine origin with other anticancer drugs We have observed additive or synergistic activity of ET-743 combined with other anticancer drugs such as cisplatin, doxorubicin, campthotecin and inhibitors of telomerase. Flow Cytometric analysis of the DNA content in human ovarian cancer: clinical correlations Conflicting results have been published on the prognostic significance of DNA aneuploidy on advanced ovary carcinoma (stage III or IV). The Citometry unit has reported one of the largest studies of the scientific literature indicating that the aneuploidy in the advanced ovary carcinoma is not an independent prognostic factor. In a large number of cases of stage I and II ovary tumors DNA content and the percentage of cells in S phase of the cell cycle, has been measured with flow cytometry, demonstrating that in the early stages of the disease DNA content is a prognostic factor important for ovary tumor. 31

34 Analysis of cell cycle data and interactions of different drugs The Biophysics Unit is engaged in theoretical and methodological studies aimed at a critical evaluation of current techniques of investigation of drug effects on heterogeneous cell populations. Several computing tools have been produced to simulate the cell proliferation at different levels (from molecular interactions to in vivo growth of solid tumours) and the process of measure. Collaborations are ongoing with other research groups for design and data analysis of drug combination studies in vitro. In this field, a number of computer programs have been developed, allowing comparative data analysis with the most common models of drug interaction. Evaluation of the complexity of the response of cell populations to treatment with anticancer drugs This project of the Biophysics Unit addresses the issue of establishing a connection between the intracellular drug interactions and the resulting cell cycle perturbations. It starts from the singlecell level of investigation to reach the cell-population level where the relevant end points of treatment efficacy are evaluated by flow cytometry and growth inhibition/cytotoxicity assays. The model adopted for data analysis and interpretation is the result of the merging of two mathematical models. One model describes the cell cycle, exploiting the results of the theory of age-structured cell population dynamics. The second model describes the response to the drug's challenge, using distinct parameters ("effect descriptors") measuring either the strength of cell cycle arrest, damage repair or cell death in every phase (G1, S and G2M). In this way, it is possible to reach a quantitative interpretation of the experimental results, overcoming the current qualitative and partial approaches to this problem, which are unable to resolve the overlapping of cytostatic and cytotoxic effects, and to establish a connection with phase-related events. Applying this procedure we demonstrated complex but biologically consistent patterns of time and dose-dependence for each cell cycle effect descriptor, following a short treatment with melphalan on a reference cell line of ovarian carcinoma. These results add to the previously reported studies on topotecan, cisplatin and taxol. Eventually, this project will produce a database containing the values associated to the new effect descriptors, related to few compounds but rich of information about them, especially in the dose and time dependence of the effects. This database will be used to compare the treatment response of the most common drugs adopted in the ovarian carcinoma. Cell cycle dysregulation in erlotinib-based treatments decoded by flow cytometry and mathematical modelling Epidermal growth factor receptor (EGFR) inhibitors represent one of the most promising class of anticancer compounds, some of them, like erlotinib, are already used for clinical therapy. Nevertheless, so far, the research has focused on molecular interaction of these compounds, somewhat neglecting the study of the dynamics of cell cycle perturbations and underscoring the importance of this issue for the optimization of both single and multidrug therapies. In order to fill this gap, the Biophysics Unit will apply the multidisciplinary approach, exploiting cell cycle simulation tools, to the study of cell cycle perturbations induced by erlotinib as a representative EGFR inhibitor. Studies of the time- and dose-dependence of perturbations induced by treatments are ongoing for single erlotinib treatment or for erlotinib combined with gemcitabine, irinotecan and oxaliplatin. The appreciation and the quantification of these effects will provide an important contribution to the comprehension of the mode of action of erlotinib alone or in combination. Such perspectives are vital if the events are to be decoded and used in predictive models for the exploration of pharmacodynamic actions and in understanding the origins of treatment failure. 32

35 Anticancer Drug Effects Decoded by Time-Lapse Imaging, Flow Cytometry and Modelling We aim to use flow cytometric (cell-population based analysis) and time-lapse imaging (single cell lineage based analysis) techniques to generate data that will be used to predict drug responses in term of the two major determinant of cytostatic/cytotoxic actions of anticancer drugs: specific cell cycle perturbations (detecting accumulation or depletion of cells in G1, S and G2M phases) and the commitment to cell death (apoptosis). The response to a treatment with the anticancer drugs will be investigated in a human osteosarcoma cell line U2OS. engineered in order to express fluorescent reporter (a fusion cyclin B1-GFP protein), so that it is now possible to follow the cells through G1, S and G2M using time-lapse microscopy. The working hypothesis is that quantitative analysis of time-lapse microscopy data could be integrated with the information provided by flow cytometry - allowing for the first time a joint interpretation of both kind of experiments through a common mathematical model that simulate the underlying phenomena. The final goal is to provide new levels of understanding and simulation tools to the cancer research community. Timing the changes of the cellular content of specific proteins inside G1, in exponentially growing cells We developed a method for measuring the content of immunocytochemically detected proteins in individual cells progressing through G1 phase. The feasibility was demonstrated in the analysis of cyclin E levels. The sequence of G1 events is tracked in unaltered cycling conditions, in a cell line in the phase of balanced growth in vitro, to avoid the pitfalls of synchronization. The method is based on i) a bromodeoxyuridine (BrdUrd) pulse-and-chase experimental plan; ii) triparametric flow cytometric detection of DNA, BrdUrd and cyclin E; iii) data analysis supported by the basic mathematical theory of asynchronous growing populations with variable cell cycle phase durations. Establishing a CFSE-based method for a quantitative measure of cytostatic effects of drugs on tumor cell populations Carboxyfluorescein diacetate succinimidyl ester (CFSE) is currently used to investigate migration and proliferation of hematopoietic cells. Several technical problems had precluded until now its use in the studies of the antiproliferative activity of anticancer drugs. We analysed several critical steps of the procedure, providing the way to overcome potential pitfalls. The project was eventually successful and the previous limitations were overcome. The outcome was a new standardised procedure of cytometry and data-analysis allowing a measure of the dynamics of cell cycle blockades after drug treatment. The new method was applied in the study of the drug topotecan, measuring the variability of response to treatment in terms percentage of cells immediately blocked, divided once or more times and then blocked or unaffected by the treatment. Pharmacokinetic of new taxane derivates Preclinical studies have been done on new taxane derivatives, chemically and biologically different from the conventional ones. For three of these compounds we evaluated the bioavailability after oral administration and the kinetic and metabolic profile has been entirely characterized by applying analytical methods based on HPLC/MS/MS technique developed in the Cancer Clinical Pharmacology Unit. They showed biological activity even in tumors with low susceptibility to other taxanes (cerebral tumor), suggesting a potential clinical interest. According to the hypothesis that these drugs act through an antiangiogenic mechanism, put forward by the Laboratory of the Biology and Therapy of Metastasis, it would be important to 33

36 investigate prolonged chronic treatments and therefore we are investigating the pharmacokinetic properties after oral administration and after prolonged daily treatment. An interesting pk profile was found for the 14-β-hydroxy-10-deacetylbaccatin III derivative IDN He showed good bioavailability and high distribution in brain achieving high concentration in comparison to that of plasma, demonstrating high ability to cross the bloodbrain-barrier. These data make this compound of great potential interest for the therapy of Central Nervous System tumors and metastasis. Clinical pharmacokinetics of gimatecan A study in collaboration with SENDO has shown the clinical pharmacokinetics of a new derivative of camptothecin (Gimatecan) in patients with sarcoma under phase II investigation. Studies still in progress show that this compound is rapidly absorbed after oral administration and has a long half life, with consequent long exposure to the drug. Pharmacokinetic parameters will be correlated to the clinical data, i.e. toxicity and antitumor activity, to characterize the properties of this new drug. Other relevant clinical studies ST The clinical pharmacokinetic of ST1926 a new oral retinoid derivative is under investigation in women with ovarian cancer.in particular we studied the pharmacokinetics, the bioavailability of the drug and its metabolism during a Phase I study, providing for the first time data on the main plasma metabolite of ST 1926 (the glucuronide conjugate). Our data show variable absorption of the drug and high conversion to the glucuronide. Antitumoral activity and pharmacokinetic properties of new drugs and combinations The antitumor activity, pharmacokinetic properties and toxicity of novel anticancer drugs with specific targets (e.g. different kinase inhibitors), conventional anticancer drugs (camptothecin) and combinations is being investigated using rodent tumors and human tumor xenografts. Laboratory of Molecular Pharmacology G2 checkpoint and cell cycle A new system able to specifically inhibit CHK1 expression in vivo in nude mice transplanted with human tumors has been developed. The system has been proved to be able to reduce the expression only in cancer cells. This plasmid allows the expression of the sirna only after induction with tetracycline and is therefore a unique tool to determine the effect of CHK1 inhibition in human tumors growing in nude mice following treatment with anticancer agents. Characterization of new potential oncosuppressor genes DRAGO gene, identified and cloned in our laboratory is one of the most interesting projects of the group. The characterization of the response of KO mice for DRAGO to ionising radiation is similar to normal mice. The characterization of the transcriptional regulation of DRAGO indicated that the gene is not only a p53-responsive gene, but, inside the p53 family, p73 has a strong ability to induce the transcription. Drago therefore, represents a new p73 responsive gene. Molecular characterization of ovarian carcinoma The molecular characterization of stage I ovarian carcinomas has been further studied. The gene expression profile analysis has showed interesting results. It is possible to identify genes able to discriminate in ovarian cancer the different histotypes, suggesting that specific biological and molecular characteristics are responsible for the 34

37 differences in morphology and clinical behaviour. These studies can help in identifying new specific molecular targets for the different subclasses of ovarian cancer that have a different clinical outcome. It is possible to identify patients with higher probability of relapse because there are genes able to differentiate these two classes of patients. We have demonstrated that stage I borderline patients have a gene expression profile similar to that of grade 1 patients, while they are well distinguishable from grade 2 and grade 3 patients. This can have important clinical implications for the treatment of this particular subgroup of patients. Expression of p63 in ovarian tumors P63 belongs to the family of p53 and its role is particularly relevant in embryo development. Its role as tumor suppressor has not been clarified yet. The gene encoding for p63 has a complex organisation which generates, among the others, a truncated isoform (DNp63), lacking the transactivation domain, which could act as a dominant negative for either p63 or p53. We have analysed the expression of the entire form (TAp63) and of DNp63 in approximately 90 ovarian cancer patients at stage I (early) and 90 at stage III (advanced tumor). The levels of TAp63 are similar in the two groups of patients, while the levels of DNp63 were higher in stage III tumors than in stage I tumors. The ratios between DNp63 and TAp63 increased consequently with the increase in malignancy. As a result, patients with higher ratio between DNp63 and TAp63 have a worst probability to survive. This has been observed not only in the whole population, but the tendency has been observed also analysing separately the two groups of patients. Altogether these results suggest that DNp63 can be considered as a new target to inhibit in this malignancy. Inhibition of the signal mediated by PI3K/akt In ovarian carcinoma cells PI3K gene is often overexpressed or mutated and this kinase is then constitutively activated. The consequence is the presence of proteins involved in cell survival, like akt, constitutively phosphorylated and then active. This project evaluated the capacity of tetra and pentaphosphate inositols and of some analogues to inhibit akt recruitment to the membrane and its further phosphorylation. IP5 and some new synthetic molecules, were shown to induce a reduction of the phosphorylation of akt and therefore to reduce cell growth. The activity of these molecules is not restricted to ovarian cancer, but has been shown also for other tumors such as breast and prostate. The possibility to combine PI3K inhibitors and mtor ( a kinase downstream to PI3K and akt) inhibitors has been evaluated. It has in fact been shown that some mtor inhibitors induce an aberrant phosphorylation, and hence activation of akt, and the combination of these inhibitors can block these undesired effects. The in vitro results show that the combination has at least an additive effect and open the way to find out new treatment schedules to verify whether the sequence of combination can be important for the antiproliferative effects. Oncosuppressors p53 and p73 The p53 analogue, p73 is present in different isoforms derived from alternative splicing of the C-terminal. Among these isoforms there is one called DNp73 in which the transactivation domain in the N-terminal of the protein is absent. This DN form of p73 is an antagonist of p53. The DNp73 isoforms can be further processed through alternative splicing, to generate a number of isoforms with a not yet clarified biological activity. We have previously shown that the alpha form of DNp73 does not modify either the growth in vitro and in vivo of cancer cells or the response to treatment with anticancer agents. We have therefore generated clones derived from the human lung cancer derived cell line H1299 which express DNp73 beta following induction. These clones show, upon induction, interesting effects on cell growth suggesting that the presence of high levels of this isoform can have unpredicted effects. Since the data generated on the expression of p73 isoforms in cancer 35

38 patients indicate that the beta isoform of DNp73 is indeed present in human cancer, the results obtained have a particular relevance and will be further analysed to verify which are the effects linked to the overexpression of this isoform. Identification of a new proteolytic mechanism of activation of p73 Another member of the p53 family, p73, has been characterised in the laboratory for its ability to modify the growth and response to therapy of cancer cells. We have further studied this potential tumor suppressor by studying new possible mechanisms of activation. We have found an additional regulation mechanism for the p73-alpha form that occurs through proteolytic cleavage connected to the activity of the serine protease HtrA2. Following apoptotic stimuli, HtrA2 accumulates in the nucleus and cleaves p73alpha in the C-terminal portion, enabling the protein to increase its transactivation activity on the apoptotic gene bax but not on the cell-cycle regulator gene p21. In the presence of HtrA2, p73 is more prone to cause caspase activation and nuclei fragmentation: p73 needs HtrA2 to activate and enhance its apoptotic functions. This new relation between p73 and HtrA2 may help to understand the different behavior of the p73 protein in cell physiology and in the responses of cancer cells to chemotherapy. Mechanisms of action of new antitumor drugs The mechanism of action of a new anthracycline derivative, nemorubicin (methoxy morpholino doxorubicin) has been characterised. Nemorubicin presents a pattern of antitumor activity in vitro and in vivo different from that of doxorubicin. We have found that cell lines with defects in DNA repair mechanisms, and particularly in the nucleotide excision repair (NER), are resistant to the treatment with this molecule. It is interesting to note that the majority of the drugs interacting with DNA, like cisplatin, show exactly the opposite: defects in NER are associated with high sensitivity. Moreover, a possible resistance mechanism for these drugs, is associated with an increased activity of NER in some tumors. Nemorubicin resistant cells have instead a reduced NER activity and show a collateral sensitivity to cisplatin. These result represent the molecular rational for the combined treatment between cisplatin and nemorubicin and /or for the treatment of cisplatin resistant tumors with nemorubicin. Generation of new cellular systems for in vivo imaging We have generated new cell clones derived from human cancer cells growing in vitro, which stably express fluorescent or luminescent probes which can allow us to follow in vivo the growth of primary tumors and metastasis in mice. These systems generated in human ovarian, breast and prostate cancer cell lines, can be implanted in nude mice and the growth and response to therapy followed by either optical and luminescent imaging or microtac analysis. These systems will be particularly useful to study the antimetastatic potential of new drugs. Characterization of response of stem cells to damage The therapeutic use of stem cells is continuously increasing. This project aims to investigate the ability of stem cells isolated from umbilical cord to respond to stress induction with particular emphasis on their ability to activate checkpoint proteins. Stem cells isolated and maintained in vitro with specific cytokine cocktails able to induce partial differentiation, have shown a peculiar expression of proteins controlling the cell cycle. In particular, there is a specific time point in the differentiation process in which cell cycle checkpoints proteins are present at high levels. This could imply that this represents the time point in which these cells are more susceptible and must be protected from external damages. The characterization of these important molecular aspects will be further studied. 36

39 Identification of cancer stem cells from ovarian cancer This project is aimed at isolating and characterizing a possible cancer stem cell from ovarian cancers. There are increasing evidences supporting the idea that few important multipotent cancer cells, termed cancer stem cells, are among the most relevant cells to be killed in a tumor. Normally present as quiescent cells inside the tumors, they are able to rapidly generate dividing and growing cancer cells. The current hypothesis is that normally dividing cancer cells can be preferentially killed by chemotherapy while the cancer stem cells would be more difficult to kill and would be responsible for the relapse following treatment. The possibility to identify and characterize the cancer stem cell would theoretically open the way to the selection of new generation molecules able to preferentially kill these cells. Cancer stem cells have been already identified in different human tumors including breast and hematopoietic tumors. We will focus our attention on human ovarian cancer by the use of antibodies directed against surface marker proteins to identify such potentially relevant cancer sub populations. Determination of the impact of EGFR mutations in the activity of tyrosine kinase inhibitors in patients with NSCLC We have started a multicenter, three year project aimed at identifying those patients who can have the chance to respond to tyrosine kinase inhibitors. The study will define whether patients without mutations in the EGFR have more chance to respond to conventional therapy rather than to treatment with TKI. Our laboratory is involved in the detection of mutations of EGFR in patients with NSCLC either in the tumor tissue or in the blood of patients with a secondary aim of defining whether is possible to detect mutations in the blood circulating DNA which are representative for the tumor. DNA sequencing and scorpion arms-based PCR methods are used for these studies. Laboratory of Biology and Therapy of Metastasis Physiologic regulation of angiogenesis Angiogenesis, the formation of blood vessels from existing ones is a fundamental process in tumor progression. A delicate balance between pro- and antiangiogenic factors finely tunes this process. In the last years we have been interested in endogenous angiogenesis-regulatory factors. During 2007 we have continued the study of trombospondin-1 (TSP-1), an endogenous inhibitor of angiogenesis. TSP-1 directly binds to angiogenic factors, in particular FGF-2 (Fibroblast Growth Factor-2), inhibiting their bioavailability and activity. In particular, we are studying the structure/function relationship of the different active domains of TSP-1, with the main aim to identify the FGF-2 binding site and design antiangiogenic compounds based on the active sequence of TSP-1. We studied the involvement of matrix metalloproteinases (MMPs) in angiogenesis and tumor progression. In particular in 2007, we studied the cross-talk between MMPs and Vascular Endothelial Growth Factor (VEGF), a factor that stimulates angiogenesis and vessel permeability, during ovarian carcinoma progression. In addition we evaluated the possibility of using VEGF inhibitors to affect MMP-dependent ovarian tumor invasion. We are analyzing modifications in metastasis/invasion-related gene expression profile in stroma cells induced by tumor VEGF. Lymphangiogenesis in ovarian carcinoma Lymphatic spread in early ovarian cancer is a predictor of outcome with potential clinical value. With the aim to clarify the molecular mechanisms involved in the process of lymphangiogenesis in ovarian cancer, the expression of VEGFC, a factor that stimulates lymphangiogenesis, has 37

40 been analyzed in serum and ascites of mice bearing human ovarian carcinoma xenografts and correlated with lymphonodes infiltration by neoplastic cells. How the microenvironment affects endothelial cell gene expression It is fundamental to understand qualitative and functional differences between tumor and normal tissue endothelial cells (EC) and the molecular mechanisms that drive the angiogenic process. This could lead to the identification of selective markers of the vascular endothelium associated to pathologies and/or of target molecules for the development of new drugs. To this purpose we analysed the gene expression profile of endothelial cells isolated from ovarian carcinoma and adrenal glands exposed or not to an angiogenic/tumor environment reconstituted in vitro. We have found that: i) the angiogenic/tumor environment is indeed able to modulate EC gene expression; ii) few genes are preferentially expressed by tumor associated endothelial cells. Preliminary results suggest that few of the genes might be of interest as markers of tumor endothelium. Their expression is higher in endothelial cells from tumor specimens than from normal tissues and they are not expressed by tumor cells. The putative new tumor endothelial markers have been further validated as anti-cancer / vascular targets by in situ hybridization analysis of normal and tumor tissues. The putative targets (identified as mrna transcript) are being produced as recombinant proteins, with the aim to isolate antibodies, directed against the recombinant proteins, suitable for immunohistochemical analyses. Preclinical models: role of Vascular Endothelial Growth Factor (VEGF) on tumor growth, vascularization and response to therapy To study the role of VEGF induced angiogenesis in the resistance of cancer to chemotherapy we have generated a variant of the human ovarian carcinoma A2780/1A9 by stable transfection with the VEGF121 isoform (1A9-VS1) or the antisense (1A9-VAS3). 1A9-VS1 cells express VEGF mrna and the protein is secreted in the culture supernatant. 1A9-VS1 xenografts i) show dilated blood vessels when implanted sc ii) produce ascites when implanted orthotopically in the peritoneal cavity, iii) release human VEGF in the plasma, iv) the level of circulating VEGF correlates with tumor burden. We have found that the response to chemotherapy (e.g. paclitaxel) might differ in xenografts producing high levels of VEGF. The blockade of VEGF, by the administration of Avastin, greatly improved the antitumor activity by paclitaxel treatment of 1A9-VS1 (T/C=11% vs T/C=40%). Studies are ongoing to elucidate the mechanism responsible of these differences. The hypotesis being that by producing/releasing VEGF, the cancer cells modify the microenvironment of the growing tumor, and in doing so, influence the response to therapy. This model is therefore being used to study gene expression. Based on circulating VEGF levels and morphological analysis of the tumor vasculature, samples were chosen and tissue slices of the 1A9-VS1 and 1A9-VAS3 were xenografts microdissected (PALM Microlaser system, in collaboration with the Institute of Pathology at Helios Klinikum, Germany) in order to isolate the stroma compartment to be evaluated by mean of Affymetrix s GeneChip Arrays technology. The microarray hybridisation data were analyzed with the aid of specialized softwares (i.e. GeneSpring and Rosetta Resolver) and differentially expressed genes were identified using a one-way, modified and error-weighted Analysis of Variance (ANOVA) with a P-value cut-off of Specifically, we discovered that in the stroma of tumors 227 were upregulated and 167 were down-regulated in consequence of the VEGF produced by the cancer cells. Gene Ontology (GO) enquiry (using Expression Analysis Systematic Explorer EASE), identified over-represented categories of potentially biologically relevant genes in the stroma of 1A9VS1 (high VEGF) tumors. Structural molecule activity, cell organization and biogenesis, 38

41 and basement membrane were among the up-regulated categories, with EASE score lower than Vascular targeting agents Antineoplastic therapies directed against the tumor vascular system may be designed with two different strategies. Antiangiogenic therapy prevents the formation of new vessels, while vascular disrupting treatment aims to selectively destroy the already formed tumor vessels. In 2007 we have focused on the identification of molecules with vascular targeting properties. Among the anti-vascular compounds, we have studied the properties of tubulin binding agents (analogues of colchicines and combretastatins), which cause microtubules depolymerization, selective damage to tumor blood vessels and tumor necrosis in experimental models in vivo. In collaboration with Prof. Bellina at the Department of Chemistry, University of Pisa (Prof. Bellina and Prof. Rossi), we have screened classes of compounds with such properties. The lead compound/s will be further characterized for pharmacological and anti-vascular/antineoplastic properties. Antineoplastic combination Therapies The optimization of biological therapies against selective targets in combinations with chemotherapy is one of the interest of this laboratory. The main class of compounds under study are the angiogenesis inhibitors and vascular disrupting agents that are thought to achieve optimal therapeutic efficacy when combined with conventional therapies. We have shown that in combination treatments, the vascular disupring activity of the tubulin binding ZD6126 is blocked by pretreatment with paclitaxel, a tubulin binding drug with opposite mechanism of action. An optimized schedule of administration of the two agents not only avoids the interference among drugs, but greatly improves the antineoplastic efficacy of the combination in experimental models. Studies are undergoing to further otpimize schedules of treatment on the basis of the mechanism of action of the drugs given in combination. There is clear evidence that histone deacetylase inhibitors are a new potential class of anticancer agents; recent reports indicate a greater antitumor effect of these compounds combined with radiation and chemotherapy. We investigated the effect of suberoylanilide hydroxamic acid (SAHA) on the paclitaxel-sensitive 1A9 and -resistant 1A9PTX22 cells. SAHA induced a comparable growth inhibition at micromolar concentrations and in a dosedependent manner in both 1A9 and 1A9PTX22 cells. Inhibition of cell proliferation, evaluated by combining doses of SAHA and paclitaxel ranging from IC30 to IC0, produced a synergistic effect. These findings show that SAHA has an antitumor activity in ovarian cancer cells with different sensitivity to paclitaxel. Furthermore, SAHA seems to be capable to partially restore sensitivity to paclitaxel in the 1A9PTX22 resistant cells. The effect is not due to a cell cycle blokade or augmented apoptosis, but is instead associated to enhanced tubulin acetylation. High PAR-1 expression is associated to a malignant phenotype Protease-activated receptor-1 (PAR-1) over-expression has been associated to a variety of human cancers, and increasing evidence implicates PAR-1 as a contributor to human melanoma malignancy. We investigated human melanoma cells, isolated from lesions representing various stages of disease progression, for the expression of PAR-1 (in collaboration with Prof. Naldini at the University of Siena) and evaluated their migratory and metastatic capabilities. Cells from advanced stage melanomas expressed higher levels of PAR-1 than those from early stages. The metastatic capability showed by the melanoma cells which overexpressed PAR-1 allowed these cells to colonize the lungs in % of the mice. Accordingly, melanoma cells overexpressing PAR-1 had higher migrated (chemotaxis assay) and invading (invasion assaymatrigel) cell counts than those expressing low PAR-1. Migration and invasion were decreased by both, PAR-1 sirna and SCH9797 (PAR-1 specific inhibitor). 39

42 Laboratory for the development of new pharmacological strategies The laboratory was born out of the consideration that the advent of oncological drugs endowed with mechanisms of action different from those of traditional chemiotherapics, introduces new treatment opportunities. At the same time, new problems arise concerning the choice of the most appropriate and effective design for research into the clinical activity profile of these new treatments. The traditional paradigm where the choice of dose is based on the maximal tolerated toxicity, and the screening of therapeutic activity focus on tumor mass reduction, may not necessarily be suitable for the evaluation of new agents whose targets may include the extracellular compartment or specific molecular targets. The clinical development of non toxic anti tumor molecules requires a critical review of the existing models as well as of all the aspects relative to the conduction of clinical trials including: dose selection criteria, methods for determination and confirmation of pharmacological activity, and the validation of new technologies and laboratory methods. This is where the need for a profound integration of the clinical screening and the preclinical research lies. It is a prerequisite for the construction of the pharmacological rationale for the identification of the most interesting molecules, the choice of dose, the hypotheses of combination with other drugs, and of the most appropriate indicators of clinical activity. The acquisition of know how and the development and application of new designs for clinical activity studies, including the use of randomization, the introduction of groups of patients treated with placebo, and new discontinuation designs, proceed in parallel to the above. Another fundamental issue in laboratory research is the recognition that the genomic characterization of any single tumor may now play a more relevant role in drug development and treatment identification. This notwithstanding, numerous uncertainties remain regarding the role of biomarkers in drug development and in the implementation of genomic technologies in clinical trials. It is therefore necessary to improve the methodology and more biomarkers evaluation already in the early stages of research, thus shifting translational research from a simple process of correlation search to one producing knowledge regarding the predictive role of the clinical activity of the investigational treatments. Therefore, the primary focus of the laboratory is the optimization of the methods for evaluating the activity of cytotoxic drugs, but mostly for those therapies aimed at specific molecular targets, as well as the identification of factors predictive of therapeutic response. Laboratory of Clinical Trials The Laboratory of Clinical Trials is involved in the planning, coordination and analysis of randomized clinical trials in oncology, conducted in cooperation with a network of medical oncologists. Main covered research areas are gastric, colo-rectal, breast and lung cancer. Gastric cancer ITACAS Intergruppo Nazionale Adiuvante Gastrico study is a randomised, open-label, multicenter, trial aimed at assessing the role of adjuvant chemotherapy in the treatment of gastric cancer. It compares the efficacy and safety of a sequential treatment (campto plus flurouracil/leucovorin, followed by taxotere and cisplatin) versus flurouracil/leucovorin regimen, used as standard reference in patients with radically resected adenocarcinoma of the stomach or gastroesophageal junction. The study, sponsored by Mario Negri Institute, involves 40

43 11 oncological collaborative groups and is being conducted in more than 110 Italian experimental centers. From February 2005, more than 750 patients out of the planned 1100 have been enrolled and it is expected to conclude the recruitment in the first half of Lung cancer The epidermal growth factor (EGFR) is overexpressed in many solid tumors including non small cell lung cancer (NSCLC) and is associated with disease progression and poor prognosis. EGFR is therefore a promising target for anticancer therapy. The development of agents that target the EGFR signal transduction pathways has provided a novel class of therapeutic agents. Erlotinib, an orally active, selective EGFR tyrosine kinase inhibitor (TKI) recently approved in NSCLC pre-treated patients is with gefitinib the most studied agent of this class of compounds. Recently, several authors reported results of analyses on retrospective data suggesting a relationship between response to TKIs and specific mutations in the EGFR tyrosine kinase domain, increased EGFR and Her-2 gene copy number and to the expression of p-akt and poorer clinical outcomes in patients with K-ras mutant when treated with erlotinib and chemotherapy. All these data are suggesting that a "tailored therapy" based on individual molecular features may result in better responses and optimization of sources and costs. Evidences so far available are based on retrospective or "post hoc" analyses of trials and are drawn on samples, non providing adequately powered tests. Furthermore, ongoing studies assess possible prognostic/predictive factors only as explorative analyses. As a consequence, in those trials no formal hypothesis primarily focused on a differential effect of treatment based on selected specific patient subgroups has been addressed with an adequate power. TAILOR AIFA trial is aimed at comparing the efficacy in terms of overall survival of erlotinib vs. docetaxel given as second line therapy in pts with advanced NSCLC. In particular, the predictive value of K-ras mutation, EGFR protein expression and EGFR gene amplification in determining the effect of erlotinib as compared to chemotherapy will be assessed. The study, sponsored by Azienda Ospedaliera Fatebenefratelli e Oftalmico of Milan and supported by the Agenzia Italiana del Farmaco (AIFA), started in late 2007 and will enroll approximately 1500 patients in three years. Colon cancer A randomised, phase III clinical trial aimed at identifying the best therapeutic adjuvant strategy in radically resected colon cancer patients is starting. The study, sponsored by Fondazione Giscad per la Cura dei Tumori and supported by the Agenzia Italiana del Farmaco (AIFA), will assess the following two questions: 1) Optimal duration of FOLFOX-4 regiomen (3 vs 6 months) 2) Efficacy of the addition of Bevacizumab to FOLFOX-4 regimen (only in high risk stage III patients) For both questions, primary efficacy endpoint will be recurrence free survival. Breast cancer TOP (Trastuzumab Optimisation trial) study is aimed at increasing the knowledge on the efficacy of herceptin in the treatment of locally advanced or metastatic breast cancer patients. It includes two randomised, open label, phase III clinical trials: the first addresses the impact of a maintenace therapy with herceptin in patients previously treated with chemotherapy plus herceptin, the second is focused on the efficacy in term of overall survival of a second-line of chemotherapy plus herceptin versus chemotherapy alone in patients progressed to a first-line containing herceptin. The results of TOP study will allow to evaluate the cost/benefit ratio of herceptin treatment and to optimise the therapeutic effectiveness of such a drug, already 41

44 approved in Italy, but used without clear data supporting evidence of benefit in the considered setting. Head and neek This is a randomized multicenter open label phase 3 factorial trial evaluating the overall survival in patients with locally advanced squamous cell carcinoma of head and neck treated with locoregional treatment (radiotherapy plus concomitant chemotherapy or cetuximab) with or without neoadjuvant chemotherapy. Patients will be randomized to receive 3 cycles of neoadjuvant chemotherapy (TPF) followed by radiotherapy plus concomitant chemo or cetuximab, or radiotherapy plus concomitant chemo or cetuximab alone. The primary objectives of the study are to compare the overall survival between neoadjuvant and no neoadjuvant arm and to compare the toxicity between concomitant chemoradiotherapy and radiotherapy plus Cetuximab. The study will be conducted by a multidisciplinary team, composed by oncologists, radiotherapists and othorinolaryngologists and will enroll approximately 350 patients in Italian centers. Laboratory of Translational and Outcome Research in Oncology The Laboratory is mainly aimed at documenting, by using either Randomized or Outcome Research studies, the value of new diagnostic and therapeutic interventions in oncology, paying particular attention to two critical steps: the passage from early to late clinical research (from the activity to efficacy evaluation) and from phase III to clinical practice (from efficacy to effectiveness). In order to facilitate the research activities and optimize the outputs, the Laboratory hosts the Coordination Centers of two multi-disciplinary Groups (MANGO: Mario Negri Gynecologic Oncology and the CP-OR: Cancer Pain Outcome Research Study Groups). As from 2007 on, all the activities of research and training in the field of chronic pain has been coordinated by a dedicated center (CERP:Center for the Evaluation and Research on Pain). CERP The objective of this newly set up center is in line with the Mario Negri Institute s purpose: CERP is aimed at advancing the scientific knowledge of chronic pain and particularly the cancer pain and at improving the quality of palliative care. Activities will be concerning three fields: research, formation and information. It ll be used a multidisciplinary and multi-institutional approach with special care to pharmacologic theraphy. Multidiscipinary teams will be created to value any contribution from either physicians or patients and with the involvement of scientific societies and patients associations. Several clinical activities focusing on patients resistant to standard analgesic treatments are currently on going and are being conducted with both private and public funds. The collaborative group in clinical gynecologic oncology named MaNGO The Mario Negri Gynecologic Oncology group (MaNGO) is a new name for a collaborative group that has been active in clinical gynecologic oncology for several years. Infact, this group consolidated its network and logistics while running the ICONs studies which were conducted in very close partnership with researchers at the Medical Research Council, Clinical Trial Unit, UK. MaNGO was formally set up in May 2006 is mainly representative of the northern part of Italy, although there are important sites in the central and southern part of the country too. Participating centers are either general public and private hospitals or university clinics. One of MaNGO s main statutory objectives was to foster an active collaboration with the Gynecologic Cancer Intergroup (GCIG), a true International Forum that circulates the scientific proposals from fifteen collaborative groups through ten countries. Currently, two randomized clinical trials in ovarian cancer have already started recruitment, one sponsored by a French group 42

45 (CALYPSO study) and the other sponsored by EORTC. Other collaboration with Dutch collaborative groups has been activated to conduct randomized clinical trials in endometrial cancer (PORTEC3 study). In 2007, several meetings of the Technical-Scientific Committe were held while MaNGO affiliates were conveyed in two General Assemblies. Colorectal cancer The assessment of efficacy of screening for relapses of colorectal carcinoma has been debated for a long time, with controversial results. GILDA is an open label, international, randomised study comparing two different strategies of post surgical surveillance in colorectal cancer (Dukes B2-C stage): minimalist versus intensive. Primary endpoints of this trial are disease free survival (which is used to assess diagnostic anticipation of metastases), overall survival, health related quality of life, direct and indirect costs evaluation. At present, GILDA trial is the largest randomised study evaluating the efficacy of two follow-ups in colorectal carcinoma. The trial was closed to patient entry in September 2006 when a total of 1200 patients had been enrolled. Follow up of patients is ongoing and preliminary results in terms of diagnostic anticipation and pattern of relapses are expected in Smoking cessation Bupropion is an antidepressant with dopaminergic and noradrenergic activity and has shown clinical efficacy for smoking cessation in specialized clinics when combined with high levels of psycosocial support. Aims of this trial is to confirm the efficacy of bupropion in the setting of family medical practice, within a simplified strategy of counselling not provided by trained counsellors. The research design is the double blind, placebo controlled, randomised clinical trial. Eligible for inclusion are subjects who have smoked an average of 10 cigarettes or more per day for the past year. The length of treatment is 7 weeks with three clinic visits scheduled over this period. Clinic visits for follow up assessments will occur at 26 and 52 weeks. Primary outcome measures are continuous abstinence and point prevalence rate of abstinence (self report of abstinence during the seven days preceding assessment); changes in weight, blood pressure, heart rate and the incidence of adverse events will be recorded and analysed. The study has reached the required number of patients and has been closed. A network of 71 Italian general practitioners (GPs) enrolled 593 participants between April 2004 and May Forty-one percent of the group given bupropion were continuously abstinent from week 4 to 7 as compared with 22% of the placebo group (OR, 2.37; 95% CI, 1.60 to 3.53). The continuous abstinence rates from week 4 to 12 months were 25% in the bupropion group and 14% in the placebo group (OR, 2.13; 95% CI, 1.33 to 3.41). The adherence of GPs and participants to the protocol was excellent, making our findings robust and easy to generalize to the context of primary care. This study, moreover, highlighted the great research potential of Primary Care setting which, in Italy, has been overlooked for a long time because of legislative vacuum. The study was published in Archives Internal Medicine in September Lymphoangiogenesis in epithelial ovarian cancer: clinical aspects and experimental studies Epithelial ovarian carcinoma is the leading cause of death from gynecologic malignancies in the developed world. Lymphatic spread in early ovarian cancer is a predictor of outcome with potential clinical value. In fact, the presence of node metastases upstages the patients with ovarian cancer apparently limited to the pelvis to FIGO stage IIIc disease and these patients are appropriate candidates for postoperative chemotherapic treatments. Even in advanced disease the metastatic involvement of aortic and pelvic nodes is still a predictor of survival. This project is aimed at providing further insight into the lymphatic spread of ovarian cancer. Two approaches will be followed to study this pathogenetic aspect of ovarian cancer: a clinical 43

46 approach where the lymphoangiogenetic activity of primary ovarian cancer will be quantified and correlated with the nodal status of women who underwent systematic lymphadenectomy (task 1) and an experimental approach where a model of human ovarian cancer xenograft overexpressing VEGF C will be set up and used to study the mechanisms of lymphatic spread and possible therapeutic approaches (task 2). Ovarian cancer: clinical and translational studies During 2007, the Unit of Gynecology-Oncology has coordinated the participation of a selected network of Italian hospitals to two international randomized clinical trials. The CALYPSO trial was sponsored by ARCAGY, France and compared two chemotherapy regimens (carboplatintaxol vs carboplatin- pegylated liposomal doxorubicin ) in patients with epithelial ovarian cancer in late relapse. The TARCEVA trial was sponsored by EORTC and evaluated the impact of adding erlotinib for two years in patients with no evidence of progression after first line, platinum-base chemotherapy for ovarian cancer. Both studies reached their target sample size in 2007 well in advance to the anticipated date of recruitment closure, thanks to really succesfull international collaboration. Results will be available in about three years. The Gynecology-Oncology Unit has started collaboration with the University of Newcastle to collect histologic specimens of patients enrolled in ICON3 randomized trial. This research aims at analyzing the p53 gene to test whether we can predict how much a patient with ovarian cancer will benefit from chemotherapy by looking at the genetic make-up of the tumour. In particular, we wish to see if it would be possible to predict whether certain patients' cancer could be treated equally well with either one drug (carboplatin alone) or with the addition of a second chemotherapy drug (carboplatin and paclitaxel). Patients experience more side effects if they are treated with the combination of two drugs. If we can tell which patients will do just as well without the second drug, then we can make their treatment safer and more tolerable whilst still being effective. During 2007, the Gynecology-Oncology Unit has finalized the protocol of an observational study aimed to describe the therapeutic approaches used to manage the ovarian cancer recurrence that develop between 6 and 12 months from the end of a first line platinum-based chemotherapy. This study should give clues to optimize the choise of drug combination in this specific clinical setting. Endometrial cancer: clinical studies In 2007, the Gynecology Oncology Unit registered in the Italian Clinical Trial Registry the PORTEC 3 protocol. This is an international randomized phase III sponsored by the Dutch Cooperative Gynecologic Oncology Group and aimed at comparing concurrent chemoradiation and adjuvant chemotherapy with pelvic radiation alone in high risk and advanced stage Endometrial Carcinoma. In late 2007 this protocol has been approved by the first Local Ethics Committee and it will be submitted to the other Ethics Committees of the interested sites in early During 2007 an observational study was also designed and it is ready to launch within spring Objective of this study is to assess the value of trans-vaginal ultrasound imaging in predicting the myometrial infiltration of endometrial carcinoma. Should this diagnostic procedure show satisfactory concordance rate with final histopathological examination clinicians could use it to improve surgical planning. During the whole 2007 two important collaborations were considered and finally drawn up with the Sapienza University, Rome and the Nordic gynaecologic oncology group. Such partnerships aimed at sharing and merging the database of very similar randomised clinical trials that singularly had been unable to reach the appropriate sample size. Such prospective poolings of the data should allow to have enough power to detect clinically important differences between treatments (i.e. lymphadenectomy versus no-lymphadenectomy and chemoradiotherapy versus radiotherapy alone, in stage I and high risk endometrial cancer, respectively). 44

47 Outcome research project in cancer pain In the context of a wide multidisciplinary project (J. Ambulatory Care Manage 29: ,2006), a nationwide multicenter, prospective outcome research study was launched in Italy in 2006 to investigate the epidemiology of cancer pain, the pattern and quality of analgesic-drug therapy, and the evolution of health outcomes over time. In a large, prospective, cohort of advanced cancer patients reporting pain, investigators collected predictive and prognostic variables, information about type of care, as well as several patientreported-outcomes, such as pain, quality of life, satisfaction with analgesic care using standardized questionnaires and data collections forms. 110 centers recruited 1801 patients from February 2006 to March Subjects were monitored for 28 days and then with a simplified scheme for further 8 weeks. At inclusion, 50% had bone metastasis, 73% a level of pain classified as moderate-severe, 48% reported episodes of breakthrough pain, 49% were still on active anti-cancer treatments and 60% were already on treatment with strong opioids. When the Index of Pain Management was computed to provide a rough estimate of how pain was treated (Cleeland et al, NEJM 330: , 1994), up to 45% had negative values suggesting a possible analgesic under-treatment, with large variations according to a selected list of clinical variables. In the sub-sample of patients with a complete follow-up at 28 days (no.=1461), all pain and palliative outcomes did significantly improve on average, with variations according to case mix, type of treatments and type of recruiting centers. Outcomes based on worst and average pain intensity showed the higher effect size estimates (0.84 and 0.69) when compared to patients' satisfaction and quality of life (0.44, 0.35). Up to 26% of patients were classified as non-responders. This outcome research study carried out at national level produced data to help implement future educative and research activities in Italy. Other research activities During 2007, other activities on patient-oriented clinical translational research in oncology were carried out. The focus was on the improvement of transferring information from the pre-clinical to clinical and research setting, and from clinical research to clinical practice. Most of this work involves data-entry, storage and other computerized applications for the management of large and complex databases of biological and clinical data. In addition to the methodological and bio-informatics issues that are relevant in this area, particular attention was also given to issues related to the ethical and legal issues pertaining the collection, storage and utilization of biological samples from patients and citizens. Finally, we would like to mention a research project about fairness in Health Care, sponsored by Regione Lombardia and run in collaboration with the Centro Nazionale di Prevenzione e Difesa Sociale (CNPDS) and the Centro Studi e Ricerche Medicina Generale (CseRMEG). This project aims to identify social, i.e. not-clinical, determinants of a unfair access to the National Health Care system, to evaluate the measurability in real conditions of such determinants, gauge their impact on care profiles and propose interventions for possible improvements. Specific objective of this Project is thus the creation of a classification system of the social vulnerability. That would be a pragmatic tool that will allow social researchers to precisely locate a single subject inside the social continuum and relate such vectorial space to a specific risk of missed of inappropriate access to Health services. A pilot phase with about 100 subjects is currently on going and it will be followed in 2008 by a survey made by about 100 General Practitioners. 45

48 Laboratory of Medical Research and Consumer Involvement This Laboratory promotes activities of research in the field of involvement of citizens and patients and their associations in decision making in health and medical issues. Moreover the activities of this laboratory include: researches on information conveyed to patients on illness and treatment, implementation of web site on the topics of the health and the information ( strategy of involvement of groups of patients for the publication of educational material; research on the evaluation of the quality of the life and satisfaction with care through studies on ad hoc selected groups, and implementation on validated ad hoc questionnaires. Strategic alliance between consumers and the scientific community PartecipaSalute PartecipaSalute ("Participate in Health Care") developed with the support of Compagnia di San Paolo foundation - is a project initiated at September 2003 to foster a strategic alliance between patients groups and professional societies with the common goal of promoting better health and shared decision-making. The project s main aims are: - to increase patients associations involvement in the healthcare debate and in decision making processes, also in the research field; - to promote a partnership between patients associations and medical societies soliciting medical societies attention to patients need and perspectives as far as concern both the production of medical information (clinical studies), and the dissemination of information. In 2007 the second edition of the Partecipasalute training course was organized: participants were 31 patients associations representatives. In this occasion a manual was given to all participants, dealing with the main arguments developed during the course. Regarding the website ( a new form was designed, with new tools useful for readers; at the end of 2007 the website s visits reached a month. New exploratory projects were defined together with patients associations in the PARITA working group/area. One example is the pilot project discussed and developed with volunteers associations interested in severe brain injury. Study on the appropriateness and appraisal of effectiveness in oncological setting, follow-up A timely oncological diagnosis and the appropriate surgical therapies remain the bench mark of the primary therapy of many oncological diseases. Effective adjuvant therapies have allowed to improve the survival of the patients, and have moreover supplied the base in order to set up programs of surveillance (follow-up). In such programs, also in absence of good results on the efficacy and effectiveness, the follow-up it is practiced with the assumption that the anticipated discovery of one relapse allows the activation of effective therapies and therefore improve the prognosis, with benefits in terms of health, consumption of resources and costs. In some cases, as an example in the breast and colon cancer, the follow-up it is pursued in the clinical practice, also in presence of randomised clinical studies, meta-analyses, consensus conference, editorials and statements of scientific associations and societies that suggest that obvious and great benefits in leading many diagnostic examinations after the primary therapies do not exist. In other cases (like in the case of the melanoma and some gynaecological tumours), because the lack of valid information on follow-up, exist a wide variability of behaviours between centres, doctors and patients. The research project is proposed to build a network of researchers, cancer centers and regional associations of patients who, after reviewing the literature, share an analysis on practical followup and on the results obtained, to set lines for shared research optimize assets and the practice, even through the production of documents on how to address follow-up more appropriate. The 46

49 activities cover patients with carcinoma of the breast, colon-rectum, endometrium. During 2007 were organised a series of working meetings that led to the development of a protocol for working gynaecology cancer and two protocols for breast cancer, one of which to be conducted in collaboration with general practioners. The project has been endorsed by the Ministry of Health. Conference of Consent on the Hormonal Replacement Therapy At the end of 2006 a Conference of Consent on Hormonal Replacement Therapy has been sponsorized by a bank foundation. The project will be articulated with multidiscliplinary working groups and it will carry out to a consent document on a topic still extremely controversial for public health, in particular regarding the information that is supplied to citizens and patients. During the 2007 the process leading to the celebration of the consensus conference was fully activated: definition of the scientific technical committee and three working groups multidisciplinary (clinical, media, citizens) who worked through review of the literature and collection of ad hoc data for the development of material to be presented to the jury of the consensus conference (May 2008). Numerous meetings have been working for the development of procedures to give room for discussion and organization of the preparatory work. SNAP project - Smoke, Nutrition, Alcohol and Physical Activity SNAP is a campaign for the health addressed to all the population of the province of Como, with particular attention to the young people between 11 and 20 years. This project, for want of FSE - Frontier Science & Technology Research Foundation, Southern Europe, a foundation for the support to the independent search - in collaboration with Istituto Mario Negri, has been launched with the attempt to increase knowledge and to modify the opinions, the attitudes and the behaviors of the young people on the four topics examined, through the circulation of paper material, a website ad hoc and a public event. In order to estimate the effectiveness of the formation-information plan, a form on knowledge, opinions and behaviors of the young people between the 11 and 20 years will be proposed, before and after the event. Throughout the last year it has been prearranged the informative material and the evaluation form to submit to the population and has been set up the website of the project. Project generic drugs, USL Bergamo As part of the work of the collaborative project between ASL Bergamo, AO ospedali Riuniti of Bergamo, Istituto di Ricerche Farmacologiche Mario Negri and Regione Lombardia, as well as between the goals, was activated a research project on generic drugs. The project has been the development and organization of an initiative - given to citizens in the province of Bergamo and their representatives (groups, associations of patients and citizens), the Primary Care Doctors and Pharmacists - on the subject of generic drugs whose consumption in the province of Bergamo to 31 December 2006 was 28%. The project, proposed by Mario Negri in activities related to the project PartecipaSalute, had intended to increase awareness among citizens about drugs equivalent and encourage good use of medicines. An evaluation after the first-consumption of medicines equivalent assess the effectiveness of this intervention-training information. The project started in May with a survey on a representative sample of the population in the province of Bergamo to assess knowledge, opinions and attitudes towards drugs equivalent. In autumn was organised the "Equivalent Drug Week" with brochures and posters distributed at pharmacies, studies of General Parctitioners, clinics and facilities accredited districts. The "Week of Drug Equivalent" had its high point in the one-day open to the public entitled "Who's afraid of generic drug?". In collaboration with Dr Alessandro Nobili, Laboratory of assessement of quality of care and service for Elder. 47

50 A population based evaluation of an intervention to improve cancer pain management The purpose of the project, designed in 2005 on the basis of preliminary results of the activities carried out in the context of the Outcome Research project above summarized (see the part b: evaluation of the volume and quality appropriateness - of prescription of analgesic drugs in a large administrative data base ) is to evaluate the effect of a community-oriented multimodal intervention to change the quantity and quality of analgesic drugs for the treatment of cancer pain. The project, after the pilot phase carried out in a smaller geographic area (Cremona: 350,000 citizens) will be carried out in an Italian Region (Le Marche: citizens). The project, sponsored by he Italian Agency for Drug evaluation (AIFA), was started in 2006 with the creation of the integrated system (data-bases) that will be used to monitor the efefct of the intervention using pre-dfined indicators of opiods prescriptions, and the -assembling of relevant panels and working groups of experts (methodologists, clinicians, general practitioners, health care managers, representative of citizens and patients, etc). During the year we organised the operational phases of the project. Several organizational meetings aimed to define the methodology of the project and its feasibility in the Marche region, have been organised. In collaboration with Dr Giovanni Apolone, Laboratory of Research Translational and Outcome in Oncology. Quality of life projects No specific research projects have been carried out on quality of the life evaluation. However are on going the activities of support and coordination of other groups using the instruments of quality of life translated and validated by our research group, SF-36, SF-12, PGWBI. During the year it has been periodically up-to-date the specific website 48

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52 DEPARTMENT OF ENVIRONMENTAL HEALTH SCIENCES STAFF Head Roberto FANELLI, Biol.Sci.D. Laboratory of Analytical Biochemistry Head Chiara CHIABRANDO, Biol.Sci.D. Laboratory of Environmental Chemistry and Toxicology Head Emilio BENFENATI, Chem.D. Industrial and Environmental Health Unit Head Laboratory of Food Toxicology Marco LODI, Chemist Head Ettore ZUCCATO, M.D. Laboratory of Mass Spectrometry Head Enrico DAVOLI, Anim.Sci.D. Laboratory of Molecular Toxicology Head Protein and Gene Biomarkers Unit Head Luisa AIROLDI, Pharm.D. Roberta PASTORELLI, Biol.Sci.D Department s Units Environmental Pollutants Risk Assessment Unit Head Elena FATTORE, Biol.Sci.D Analytical Instrumentation Unit Head Renzo BAGNATI, Chem.D. 50

53 CURRICULA VITAE Roberto Fanelli, Head of the Environmental Health Sciences Department since 1997, Laboratory Head , Researcher , Research fellow at the Mario Negri Institute. Doctoral Degree in Biological Sciences (University of Milan, 1973), Assistant Professor in Biochemistry at Baylor College of Medicine (Houston, Texas). Member of the Commissione Consultiva Prodotti Fitosanitari (Ministero Salute), Member of the Scientific Panel on Contaminants in the Food Chain (European Food Safety Authority, ), Certified Italian Toxicologist. Member of the Comitato Scientifico Ente Risi. Research areas: Sources, diffusion, toxicology, human exposure and risk assessment of persistent environmental pollutants. Environmental risk of plant protection products. Development of analytical methods for identification and measurement of biomarkers in toxicology. Mechanisms of toxic action by proteomic techniques. Selected publications: 1. Pastorelli R, Carpi D, Campagna R, Airoldi L, Pohjanvirta R, Viluksela M, Hakansson H, Boutros P C, Moffat I D, Okey A B, Fanelli R. Differential expression profiling of the hepatic proteome in a rat model of dioxin resistance: correlation with genomic and transcriptomic analyses. Mol Cell Proteomics 2006; 5: Zuccato E, Chiabrando C, Castiglioni S, Calamari D, Bagnati R, Schiarea S, Fanelli R. Cocaine in surface waters: a new evidence-based tool to monitor community drug abuse. Environ Health 2005; 4: 14 ( 2005) 3. Pastorelli R, Carpi D, Airoldi L, Chiabrando C, Bagnati R, Fanelli R, Moverare S, Ohlsson C. Proteome analysis for the identification of in vivo estrogen-regulated proteins in bone. Proteomics 2005; 5: Zuccato E, Castiglioni S, Fanelli R. Identification of the pharmaceuticals for human use contaminating the Italian aquatic environment. J Hazard Mater 2005; 122: Airoldi L, Magagnotti C, Pastorelli R, Fanelli R. Enzyme polymorphisms influencing the metabolism of heterocyclic aromatic amines. J Chromatogr B Analyt Technol Biomed. Life Sci 2004; 802: Fattore E, Di Guardo A, Mariani G, Guzzi A, Benfenati E, Fanelli R. Polychlorinated dibenzo-p-dioxin and dibenzofurans in the air of Seveso, Italy, 26 years after the explosion. Environmental Science Technology 2003; 37: Luisa Airoldi, Head of the Molecular Toxicology Laboratory since 1994, Unit Head , Researcher , Technician at the Mario Negri Institute. Doctoral Degree in Pharmacy (University of Milan, 1975), Postdoctoral fellow at the Massachusetts Institute of Technology (Cambridge, MA, 1976) and at the Northwestern University Medical School (Chicago, Il, 1977), Researcher at the Yale University Medical School (New Haven, CT, ). Research areas: Proteomics in toxicology with particular interest on the study of proteome changes in tissues and biological fluids from animals and humans after exposure to toxic compounds; molecular epidemiology focused on the identification and measurement of biomarkers of exposure to environmental carcinogens and disease susceptibility; chemical carcinogenesis centered on the study of carcinogens mechanism of action. Selected publications: 1. Vineis P, Hoek G, Krzyzanowski M, Vigna-Taglianti F, Veglia F, Airoldi L, Overvad K, Raaschou-Nielsen O, Clavel- Chapelon F, Linseisen J, Boeing H, Trichopoulou A, Palli D, Krogh V, Tumino R, Panico S, Bueno-De-Mesquita HB, Peeters PH, Lund E E, Agudo A, Martinez C, Dorronsoro M, Barricarte A, Cirera L, Quiros JR, Berglund G, Manjer J, Forsberg B, Day NE, Key TJ, Kaaks R, Saracci R, Riboli E. Lung cancers attributable to environmental tobacco smoke and air pollution in non-smokers in different European countries: a prospective study. Environ Health ; 6:7. 2. Pastorelli R, Saletta F, Campagna R, Carpi D, Dell'Osta C, Schiarea S, Vineis P, Airoldi L, Matullo G Proteome characterization of a human urothelial cell line resistant to the bladder carcinogen 4-aminobiphenyl Proteome Sci : 6 3. Pastorelli R, Carpi D, Campagna R, Airoldi L, Pohjanvirta R, Viluksela M, Hakansson H, Boutros P C, Moffat I D, Okey A B, Fanelli R. Differential expression profiling of the hepatic proteome in a rat model of dioxin resistance: correlation with genomic and transcriptomic analyses. Mol Cell Proteomics 2006; 5: Airoldi L, Vineis P, Colombi A, Olgiati L, Dell'Osta C, Fanelli R, et al. 4-Aminobiphenyl-hemoglobin adducts and risk of smoking-related disease in never smokers and former smokers in the European Prospective Investigation into Cancer and Nutrition Prospective study. Cancer Epidemiol Biomarkers Prev 2005; 14: Pastorelli R, Carpi D, Airoldi L, Chiabrando C, Bagnati R, Fanelli R, Moverare S, Ohlsson C. Proteome analysis for the identification of in vivo estrogen-regulated proteins in bone. Proteomics 2005; 5: Magagnotti C, Pastorelli R, Pozzi S, Andreoni B, Fanelli R, Airoldi L. Genetic polymorphisms and modulation of 2-amino-1- methyl-6-phenylimidazo[4,5-b]pyridine (PhIP)-DNA adducts in human lymphocytes. Int J Cancer 2003; 107:

54 Emilio Benfenati, Head of the Laboratory of Environmental Chemistry and Toxicology since 1997, Unit Head , Researcher , Research fellow at the Mario Negri Institute. Researcher at Istituto Biochimico Italiano Doctoral Degree in Chemistry (University of Milan, 1979). Member of Commissione Consultiva Prodotti Fitosanitari (Ministero Salute ), Certified Italian Chemist. Resarch areas: Computer-based models for chemistry and toxicology; Molecular descriptors; QSAR; Toxicity prediction; Metabolism studies; Characterization and assessment of wastes, industrial effluents, emissions from landfill and incinerator; Integration of chemical analysis and eco-toxicological data; Chemical analysis of organic compounds by mass spectrometry. Principali pubblicazioni: 1. Lo Piparo E, Koehler K, Chana A, Benfenati E, Virtual screening for aryl hydrocarbon receptor binding prediction, J Med Chem 2006; 49: Casalegno M, Sello G, Benfenati E, Top-priority fragment QSAR approach in predicting pesticide aquatic toxicity, Chem Res Toxicol 2006; 19: Lo Piparo E, Fratev F, Lemke F, Mazzatorta P, Smiesko M, Fritz J I, Benfenati E, QSAR models for Daphnia magna toxicity prediction of benzoxazinone allelochemicals and their transformationproducts, J Agric Food Chem 2006; 54: Benfenati E. (Ed.), Quantitative Structure-Activity Relationships for Pesticide Regulatory Purposes, Elsevier, pp (2007). 4. Benigni R, Netzeva T, Benfenati E, Bossa C, Franke R, Helma C, Hulzebos E, Marchant C, Richard A, Woo Y-T, Yang C. The expanding role of predictive toxicology: An update on the (Q)SAR models for mutagens and carcinogens. J Environ Sci Health C 25: (2007). 5. Roncaglioni A, Benfenati E. In silico-aided prediction of biological properties of chemicals: oestrogen receptor-mediated effects. Chem Soc Rev 37: (2008). 6. Porcelli C, Boriani E, Roncaglioni, A, Chana A, Benfenati. Regulatory perspectives in the use and validation of QSAR. A case study: DEMETRA model for daphnia toxicity. Environ Sci Technol 42: (2008). Chiara Chiabrando, Head of the Analytical Biochemistry Laboratory since 1997, Unit Head , Researcher , Research fellow at the Mario Negri Institute. Doctoral degree in Biological Sciences (University of Milan, 1974), Postdoctoral fellow at the Baylor College of Medicine (Houston, Texas, ). Postgraduate degree in Pharmacological Research, Mario Negri Institute (1977). Research areas: Development and application of bio-analytical methods based on mass spectrometry in the fields of biochemistry, metabolism, clinical chemistry and pharmacology. Identification and characterization of proteins and peptides of biomedical interest by proteomic approaches and mass spectrometry. Proteomics in toxicology. Selected publications 1. Castiglioni S, Zuccato E, Crisci E, Chiabrando C, Fanelli R, Bagnati R. Identification and measurement of illicit drugs and their metabolites in urban wastewaters by liquid chromatography tandem mass spectrometry (HPLC-MS-MS). Anal Chem 2006, 78: Pastorelli R, Carpi D, Airoldi L, Chiabrando C, Bagnati R, Fanelli R, Moverare S, Ohlsson C. Proteome analysis for the identification of in vivo estrogen-regulated proteins in bone. Proteomics. 2005;5: Zuccato E, Chiabrando C, Castiglioni S, Calamari D, Bagnati R, Schiarea S, Fanelli R. Cocaine in surface waters: a new evidence-based tool to monitor community drug abuse. Environ Health. 2005;4: Chiabrando C, Avanzini F, Rivalta C, Colombo F, Fanelli R, Palumbo G, Roncaglioni MC; PPP Collaborative Group on the antioxidant effect of vitamin E. Long-term vitamin E supplementation fails to reduce lipid peroxidation in people at cardiovascular risk: analysis of underlying factors. Curr Control Trials Cardiovasc Med. 2002;3:5. 5. Chiabrando C, Rivalta C, Bagnati R, Valagussa A, Durand T, Guy A, Villa P, Rossi JC, Fanelli R. Identification of metabolites from type III F2-isoprostane diastereoisomers by mass spectrometry. J Lipid Res. 2002;43: Chiabrando C, Valagussa A, Rivalta C, Durand T, Guy A, Zuccato E, Villa P, Rossi JC, Fanelli R. Identification and measurement of endogenous beta-oxidation metabolites of 8-epi-Prostaglandin F2alpha.J Biol Chem Jan 15;274:

55 Enrico Davoli, Head of the Mass Spectrometry Laboratory since 1997, Unit Head , Researcher , Research Fellow at the Mario Negri Institute. Fellow at USDA, Beltville, MD Doctoral Degree in Animal Sciences (University of Milan, 1983), Postdoctoral fellow at the University of Nebraska (Lincoln, NE, 1987) and at the University of Colorado Health Sciences Center (Denver, CO, 1988). Postgraduate degree in Pharmacological Research, Mario Negri Institute (1988). Member of the American Association for Mass Spectrometry of the Environment and Safety Commission of IGQ and of the ETS (Emission Trading System) commission. Member of the National Biomass Research Center Scientific Committee. Research areas: Development of methodology, instrumentation and software for environmental research. Studies of urban air pollution and characterization of environmental odor annoyance. Selected Publications 1. Riservato M, Rolla A, Davoli E. An isotopic dilution approach for 1,3-butadiene tailpipe emissions and ambient air monitoring. Rapid Commun Mass Spectrom 2004; 18: Davoli, L. Cappellini, M. Moggi and R. Fanelli. Automated, high speed analysis of selected organic compounds in urban air by on line isotopic dilution cryofocussing GC/MS. J. Am. Soc. Mass Spectrometry. 5: E. Davoli, L. Cappellini, M. Moggi S. Ferrari and R. Fanelli. On-Line Monitoring of Benzene Air Concentrations While Driving in Traffic by Isotopic Dilution GC/MS. Int. Arch. Occup. Environ. Health 1996; 68: E. Davoli, L. Cappellini, R. Fanelli, M. Bonsignore, M. Gavinelli. On-Site Analysis of World War II Cylinders and Barrels with Unknown Contents. Field Anal. Chem. Technol. 2001; 5: E. Davoli, L. Gangai, P. Morselli, D. Tonelli, Characterisation of Odorant emissions from Landfills by SPME and GC/MS. Chemosphere 2003; 51: E. Zuccato, P. Grassi, E. Davoli, L. Valdicelli, D. Wood, G. Reitano, R. Fanelli. PCB concentrations in some foods from four European countries. Food Chem Toxicol 2008 ; 46 : R. Bagnati, G. Bianchi, E. Marangon, E. Zuccato, R. Fanelli, E. Davoli. Direct analysis of isopropylthioxanthone (ITX) in milk by high-performance liquid chromatography/tandem mass spectrometry. Rapid Commun Mass Spectrom 2007 ; 21 : Ettore Zuccato, Head of the Food Toxicology Laboratory since 2005, Unit Head , Researcher , Technician at the Mario Negri Institute. Doctoral degree in Medicine (University of Milan, 1986), Postdoctoral degree in Human Nutrition (1999), Postdoctoral fellow at the King s College School of Medicine (London, UK, ). Member of the ANSISA, EMEA expert, member of the Commissione Consultiva per i Prodotti Fitosanitari, and expert for the evaluation of plant protection products for registration within the EU. Research areas: Food safety, including the study of dietary chemical contaminants, safety assessment of GMO in human nutrition, food allergens and toxicants, emerging issues in food toxicology, risk perception and risk communication to the consumers, and evaluation of plant protection products for registration within the European Union. Environmental pollution by pharmaceuticals, and monitoring of illicit drugs in surface waters to estimate community drug abuse. Selected publications 1. Castiglioni S, Zuccato E, Crisci E, Chiabrando C, Fanelli R, Bagnati R. Identification and measurement of illicit drugs and their metabolites in urban wastewaters by liquid chromatography tandem mass spectrometry (HPLC-MS-MS). Anal Chem 2006, 78: Pomati F, Castiglioni S, Zuccato E, Fanelli R, Rossetti C and Calamari D. Effects of Environmental Contamination by Therapeutic Drugs on Human Embryonic Cells. Environ. Sci. Technol. 2006, 40, Zuccato E, Calamari D, Castiglioni S, Chiabrando C, Bagnati R, Fanelli R. Cocaine in surface water: a new evidence-based tool to monitor community drug abuse. Environmental Health: A Global Access Science Source 2005, 4:14 4. Calamari D, Zuccato E, Castiglioni S, Bagnati R, Fanelli R. Strategic survey of therapeutic drugs in the rivers Po and Lambro in northern Italy. Environ Sci Technol 2003; 37: Zuccato E, Calamari D, Natangelo M, Fanelli R. Presence of therapeutic drugs in the environment. Lancet 2000; 355: Zuccato E, Calvarese S, Mariani G, Mangiapan S, Grasso P, Guzzi A, Fanelli R. Level, sources and toxicity of polychlorinated biphenyls in the Italian diet. Chemosphere 1999; 38 (12):

56 Renzo Bagnati, Head of the Analytical Instrumentation Unit since 2005, Researcher , Research fellow at the Mario Negri Institute. Doctoral degree in Chemistry (University of Turin, 1985), Postgraduate degree in Pharmacological Research, Mario Negri Institute (1989). Research areas: Mass spectrometry applied to the analysis of biological and environmental relevant substances (proteins, peptides, hormones, pharmaceuticals, drugs of abuse, pesticides). Selected Publications 1. Castiglioni S, Zuccato E, Crisci E, Chiabrando C, Fanelli R, Bagnati R. Identification and measurement of illicit drugs and their metabolites in urban wastewater by liquid chromatography-tandem mass spectrometry. Anal Chem 2006; 78: Castiglioni S, Bagnati R, Fanelli R, Pomati F, Calamari D, Zuccato E. Removal of pharmaceuticals in sewage treatment plants in Italy. Environmental Science Technology 2006; 40: Zuccato E, Chiabrando C, Castiglioni S, Calamari D, Bagnati R, Schiarea S, Fanelli R. Cocaine in surface waters: a new evidence-based tool to monitor community drug abuse. Environ Health. 2005;4: Pastorelli R, Carpi D, Airoldi L, Chiabrando C, Bagnati R, Fanelli R, Moverare S, Ohlsson C. Proteome analysis for the identification of in vivo estrogen-regulated proteins in bone. Proteomics 2005; 5: Bagnati R, Ramazza V, Zucchi M, Simonella A, Leone F, Bellini A, Fanelli R. Analysis of dexamethasone and betamethasone in bovine urine by purification with an 'on-line' immunoaffinity chromatography-hplc system and determination by GC-MS. Anal Biochem 1996; 235: Davoli E, Fanelli R, Bagnati R. Purification and analysis of drug residues in urine samples by on-line immunoaffinity chromatography/high-performance liquid chromatography/continuous-flow fast atom bombardment mass spectrometry. Anal Chem 1993; 65: Elena Fattore, Head of the Environmental Pollutants Risk Assessment Unit since 2005, Researcher , Research fellow at the Mario Negri Institute. Doctoral Degree in Biological Sciences (University of Milan, 1991), Postgraduate degree in Pharmacological Research, Mario Negri Institute (1994), Postdoctoral fellow at the National Institute of Environmental Medicine, Karolinska Institutet, Stockholm ( ). Member of the Group of Experts of the Italian Presidency of the Council of Ministers for the Surveillance of Exposure to Endocrine Disrupters. Research areas: Environmental chemistry, toxicology, assessment of human exposure and risk from environmental pollutants with emphasis on dioxins and dioxin-like compounds. Selected publications 1. Carubelli G, Fanelli R, Mariani G, Nichetti S, Crosa G, Calamari D, Fattore E. PCB contamination in farmed and wild sea bass (Dicentrarchus labrax L.) from a coastal wetland area in central Italy. Chemosphere 2007 ; 68 : Fattore E, Fanelli R, Turrini A, Di Domenico A. Current dietary exposure to polychlorodibenzo-p-dioxins, polychlorodibenzofurans, and dioxin-like polychlorobiphenyls in Italy. Mol Nutr Food Res 2006; 50: Fattore E, Guardo A, Mariani G, Guzzi A, Benfenati E, Fanelli R. Polychlorinated Dibenzo-p-Dioxins and Dibenzofurans in Air of Seveso, Italy, 26 years after the accident. Environ Sci Technol 2003; 37: Fattore E, Fanelli R, La Vecchia C. Persistent organic pollutants in food: Public health and implications. J Epidemiol Community Health 2002; 56: Fattore E, Trossvik C, Håkansson H. Relative potency values derived from hepatic vitamin A reduction in male and female Sprague-Dawley rats following subchronic dietary exposure to individual polychlorinated dibenzo-p-dioxin and dibenzofuran congeners, and a mixture thereof. Toxicol Appl Pharmacl 2000; 165: Fattore E, Benfenati E, Mariani G, Fanelli R, Evers E H. Pattern and Sources of Polychlorinated dibenzo-p-dioxins and Dibenzofurans in Sediment from Venice Lagoon. Environ Sci Technol 1997; 31:

57 Marco Lodi, Head of the Industrial and Environmental Unit since 2002, Consultant at the Mario Negri Institute. General Certificate of Education in Industrial Chemistry (Milan, 1974). Member of AIDII (Italian Industrial Hygiene Association), certified by ACGIH (American Conference of Governmental Industrial Hygienist). Research areas: Emission sources, environmental diffusion, toxicology, human exposure and risk assessment of persistent environmental pollutants. Environmental risk of chemical pollution products. Development of sampling methods for environmental toxic compounds. Selected publications 1. Benfenati E, Azimonti G, Auteri D, Lodi M Environmental and ecological toxicology: computational risk assessment. Computational toxicology. Risk assessment for pharmaceutical and environmental chemicals John Wiley, Hoboken, NJ, 2007; Grosso M, Cernuschi S, Palini E, Lodi M, Mariani G. PCDD/Fs release during normal and transient operation of a full scale MSWI plant. Organohalogen Compounds 2004; 66: Benfenati E, Mariani G, Lodi M, Reitano G, Fanelli R. Is bioexsiccation releasing dioxins? Organohalogen Compounds 2004; 66: Fattore E, Mariani G, Guzzi A, Di Guardo A, Benfenati E, Lodi M, Fanelli R. Air dioxin concentrations in Seveso area. In: Halogenated Environmental Organic Pollutants, 18th. Symp., Stockholm, Sweden, august 17-21, : Benfenati E, Mariani G, Schiavon G, Lodi M, Fanelli R. Diurnal, weekly and seasonal air concentrations of PCDD and PCDF in an industrial area. Fresenius Journal Analytical Chemistry 1994; 348: Benfenati E, Pastorelli R, Castelli M G, Fanelli R, Carminati A, Farneti A, Lodi M. Studies on the tetrachlorodibenzo-pdioxins (TCDD) and tetrachlorodibenzofurans (TCDF) emitted from an urban incinerator. Chemosphere 1986; 15: Roberta Pastorelli, Head of Protein and Gene Biomarkers Unit since 2004, Researcher , Research fellow at the Mario Negri Institute. Doctoral Degree in Biological Sciences (University of Milan, 1982), Postgraduate degree in Pharmacological Research, Mario Negri Institute (1986), Postdoctoral fellow at the Massachusetts Institute of Technology, Cambridge, MA ( and 1991). Research areas: Toxicoproteomic investigation of global protein expression profiles and their modulation evoked by environmental compounds in different biological compartments. Critical targets and pathways in toxicology. Pharmacogenetics: effects of genetic polymorphisms in the human population on the individual susceptibility to environmental xenobiotic and carcinogen effects. Selected publications: 1. Pastorelli R, Saletta F, Campagna R, Carpi D, Dell?osta C, Schiarea S, Vineis P, Airoldi L, Matullo G. Proteome characterization of a human urothelial cell line resistant to the bladder carcinogen 4-aminobiphenyl. Protome Science Moretti M, Dell'omo M, Villarini M, Pastorelli R, Muzi G, Airoldi L, Pasquini R. Primary DNA damage and genetic polymorphisms for CYP1A1, EPHX and GSTM1 in workers at a graphite electrode manufacturing plant. BMC Public Health : Pastorelli R, Carpi D, Campagna R, Airoldi L, Pohjanvirta R, Viluksela M, Hakansson H, Boutros P C, Moffat I D, Okey A B, Fanelli R. Differential expression profiling of the hepatic proteome in a rat model of dioxin resistance: correlation with genomic and transcriptomic analyses. Mol Cell Proteomics 2006; 5: Pastorelli R, Carpi D, Airoldi L, Chiabrando C, Bagnati R, Fanelli R, Moverare S, Ohlsson C.Proteome analysis for the identification of in vivo estrogen-regulated proteins in bone. Proteomics 2005; 5: Airoldi L, Magagnotti C, Pastorelli R, Fanelli R. Enzyme polymorphisms influencing the metabolism of heterocyclic aromatic amines. J Chromatogr B Analyt Technol Biomed Life Sci 2004; 802: Vineis P,V eglia F, Anttila S, Benhamou S, Clapper M L, Dolzan V, Ryberg D, Hirvonen A, Kremers P, Le Marchand L, Pastorelli R, Rannug A, Romkes M, Schoket B, Strange R C, Garte S, Taioli E. CYP1A1, GSTM1 and GSTT1 polymorphisms and lung cancer: a pooled analysis of gene-gene interactions. Biomarkers 2004; 9:

58 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The Department works to investigate environmental factors and their effects on human health. The main research lines focus on the survey of environmental contaminants, the assessment of human exposure with related health risks, and toxicity mechanisms of pollutants. The assessment of environmental contamination is carried out not only for well-known and widespread compounds, like dioxins and PCBs, but also for new classes of "unconventional" pollutants, e.g., endocrine disruptors, potentially toxic "natural" compounds, and drugs entering the environment after human or veterinary use. The identification for the first time of illicit drugs in urban waste and river waters, led to a new original tool for the evidence-based monitoring of community drug abuse. For all these survey activities sophisticated analytical methods based on advanced mass spectrometric techniques are developed. The Department is active in the assessment of human exposure to toxic compounds in the atmosphere and the diet, which is the main source of priority pollutants (PCBs, dioxins and other endocrine disruptors). Assessment of the risk associated to contamination in real-life scenarios has recently gained much importance. In order to respond to the growing demand for information, the Department is more and more involved in toxicological and ecotoxicological risk analysis, based on studies in field and predictive models of toxicity. Molecular epidemiology studies are used to identify genetic and/or environmental factors posing risks to human health. By this approach, we search for new useful biological markers" to identify susceptible subjects, in view of finding appropriate preventive strategies. The Department has implemented an advanced technological proteomic platform, in order to identify proteins differentially expressed in biological compartments in various experimental and clinical conditions. This approach is particularly relevant in toxicology, since it can contribute to find new biomarkers of toxicity or pathology, and to identify molecular targets and toxic effect mechanisms of pollutants and drugs. FINDINGS/MAIN RESULTS The lack of retinoic acid (knock-out mice for the retinal dehydrogenase 1 gene, RALDH1) modifies the proteome profile of the bone, through changes in the expression of proteins involved in chondrogenesis and osteoclastogenesis. Resistance to the bladder carcinogen 4-aminobiphenyl in human urothelial cells is mediated by deregulation of apoptosis and membrane trafficking proteins. Bone protein profile in a murine model of osteoporosis. Identification of novel protein targets responsive to the effects of estrogens in bone. TCDD's effect on the liver proteome profile of exposed rats. Determination of a subset of rat hepatic proteins indicative of differences in dioxin susceptibility. The presence of 4-aminobiphenyl-hemoglobin adducts may help identify nonsmokers at high risk of cancers related to environmental tobacco smoke exposure. Reference values of allele and genotype frequency of several metabolic genes in 15,000 control subjects. CYP1A1 polymorphism affects lung tumor risk. Identification of CYP2C9 genetic polymorphism as a determinant of severe adverse reactions to phenytoin. On-line in silico models to predict ecotoxicity of pesticides for regulatory purposes. A new model for identification of endocrine disruptors using molecular docking. A method aimed at characterizing environmental odors to identify odor sources in complex environments. Moderate-to-high fish consumption can result in exceeding the daily intake safety levels of PCBs and dioxin-like compounds established by the European Commission. The same food type may contain significantly different concentrations of PCBs and dioxin-like compounds, depending on the geographic origin (this may help lower the risk for the consumers by understanding and controlling the causes of the differences). The environmental levels of several drugs exceed the safety limits established for them. Environmental pollution from pharmaceuticals is a general phenomenon that can be described by 56

59 controllable variables (environmental load and mass balance). Illicit drug residues and their metabolites were found in urban waste and river waters. Environmental levels can be used as a new tool to estimate illicit drugs consumption in the population. The distribution of dietary intake values of dioxins, dioxin-like PCBs and non dioxin-like PCBs was characterized for the general Italian population. The higher intake of PCBs due to consumption of farmed fish vs. wild fish is mainly due to the higher fat content in farmed fish. Development of novel mass spectrometric methods for the selective measurement of therapeutic and illicit drugs in environmental samples. NATIONAL COLLABORATIONS ARPA Veneto ASL di Brescia CESTEC, Regione Lombardia, Milano CNR IRSA CSPO-Firenze CSRA-Asti Fondazione 'S. Maugeri' Fondazione ISI, Torino INRAN-Istituto Nazionale di Ricerca sugli Alimenti e la Nutrizione Istituto Clinico Humanitas, Milano Istituto Superiore di Sanità I.Z.S.L.T - Istituto Zooprofilattico Sperimentale del Lazio e Toscana Ministero dell'ambiente Politecnico di Milano Politecnico di Torino Provincia di Vercelli Provincia Pordenone Università degli Studi di Cagliari Università degli Studi di Genova Università degli Studi di Napoli "Federico II" Università degli Studi di Palermo Università degli Studi di Pavia Università degli Studi di Perugia Università degli Studi di Roma "La Sapienza" Università degli Studi di Torino Università dell Insubria, Varese 57

60 INTERNATIONAL COLLABORATIONS BASF Agricultural Centre, Limburgerhorf, Germany Central Science Laboratory, York, UK Danish Institute of Agricultural Sciences, Research Centre Foulum, Tjele, Denmark Department of Analytical and Pharmaceutical Chemistry, The Royal Danish School of Pharmacy, Denmark Department of Anatomy and Cell Biology, University of Oulu, Oulu, Finland Department of Analytical and Pharmaceutical Chemistry, The Royal Danish School of Pharmacy, Denmark Department of Computer Science and Engineering, University of Galati, Romania Department of Electrical and Computer Engineering, University of Patras, Greece Department of Environmental Health, National Public Health Institute, Kuopio, Finland Department of Epidemiology & Public Health, Imperial College, London, United Kingdom Department of Inland Fisheries, Institute of Freshwater Ecology and Inland Fisheries, Berlin, Germany Department of Molecular Biology, University of Bergen, Bergen, Norway Department of Organic Chemistry, Universidad de Cadiz, Spain Division of Endocrinology, Department of Internal Medicine, Sahlgrenska University Hospital, Goteborg, Sweden Environmental Chemistry, IIQAB-CSIC, Barcelona, Spain Environmental Hygiene and Chemistry Department, Institute of Environmental Medicine and Hospital Epidemiology, University of Freiburg, Germany Environmental Hygiene and Chemistry Department, Institute of Environmental Medicine and Hospital Epidemiology, University of Freiburg, Germany Faculté de Médicine et de Pharmacie, Université de Mons-Hainaut, Mons, Belgium Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands Forschungzentrum Jülich Gmbh, Jülich, Germany Gruppo Collaborativo sulla Suscettibilità Genetica ai Cancerogeni Ambientali (GSEC), Milano, Italy Institute of Environmental Medicine. Karolinska Institute, Stockholm, Sweden Institute of Pharmaceutical Chemistry, University of Pécs, Pecs, Hungary Institute of Phytomedicine, Biological Control, Horticulture and Nematology, Wien, Austria Institute of Soil Science and Plant Cultivation, Pulawy, Poland Interuniversitaeres Forchunginstitut fuer Agrarbiotechnologie, Tulln, Austria Istituto di Chimica di São Carlos, Università di São Paulo, Brazil KnowledgeMiner Software, Berlin, Germany In Vitro Testing Industrial Platform, Tres Cantos (Madrid), Spain Laboratory of Chemometrics & Bioinformatics, University of Orléans, Orléans, France Lithuanian Institute of Agricultrure, Vilnius, Lithuania Liverpool John Moores University, Liverpool, United Kingdom National Institute of Chemistry, Kemijski Institut Ljubljana, Ljubljana, Slovenia Natural Resources Research Institute, University of Minnesota, Duluth, MN National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands Pesticide Safety Directorate, York, UK Plant Protection Institute, Hungarian Academy of Sciences, Budapest, Hungary School of Biomedical Sciences, University of Ulster, Coleraine, Northern Ireland Syngenta Crop Protection AG, Basel, Switzerland UFZ Leipzig, Germany University of Tartu, Tartu, Estonia 58

61 EDITORIAL BOARD MEMBERSHIP Journal of Environmental Science and Health, Part B (Emilio Benfenati), Journal of Environmental Science and Health, Part C (Emilio Benfenati), International Journal of Computational Intelligence (Emilio Benfenati), International Journal of Information Technology (Emilio Benfenati), International Journal of Signal Processing (Emilio Benfenati), Chemistry Central Journal (Emilio Benfenati), Current Computer Aided Drug Design (Emilio Benfenati), Advances in Chemoinformatics and Computational Methods (Emilio Benfenati).. PEER REVIEW ACTIVITIES Addiction, Analytical and Bioanalytical Chemistry, Chemometrics and Intelligent Laboratory Systems, CHEMOLAB, Chemosphere, Chemical Biology & Drug Design, Environmental Modelling & Software, Environmental Science & Technology, Journal of Chemical Information and Modeling, International Journal of Molecular Science, Journal of Hazardous Materials, Molecular Diversity, Proteomics, Waste Management. NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP CCPF Commissione Consultiva Prodotti Fitosanitari (Ministero della Salute, Ministero dell'ambiente) IGQ - Commissione Ambiente e Sicurezza IGQ - ETS Commission EVENT ORGANIZATION NOSE2008. INTERNATIONAL CONFERENCE ON ENVIRONMENTAL ODOUR MONITORING and CONTROL. Rome, Italy 6-8 July

62 PARTICIPATION IN EVENTS IN WHICH THE DEPARTMENT WAS INVOLVED SETAC Europe, 17th annual meeting, May 2007, Porto, Portugal 55th American Society for Mass Spectrometry Conference, June 3-7, 2007, Indianapolis, IN, USA The International Society of Pharmacovigilance. 7 th Annual Meeting October 2007, Bournemouth, UK. ASMS sponsored meeting. The Art of Open Air Ionization on Surfaces. November Chemical Heritage Foundation. Philadelphia, PA, USA Proteomix, XI Meeting, November 29-30, 2007, Torino, Italy. Workshop of the ATHON EC project, Valencia, Spain, November 29- December 30, GRANTS AND CONTRACTS ACEGAS S.p.A, Trieste ASL Mantova CSRA Associazione Italiana Ricerca sul Cancro Comune di Lomello Comune di Mazzano e Rezzato Consorzio Quadrifoglio S.p.A. ECODECO, Pavia European Commission (MEBFOOD; SAFEFOODNET; DEMETRA; EUFRAM; HERBICBIOREM; BONETOX, ATHON, CASCADE, HAIR, CAESAR, RAINBOW, CHEMOMENTUM) Ferrero, Alba (Cuneo) FIAT Auto S.p.A. Fondazione CARIPLO, Milano Fondazione Italo Monzino, Milano I.Z.S.L.T - Istituto Zooprofilattico Sperimentale del Lazio e Toscana Lucart, Cartiera Lucchese S.p.A. Porcari, Lucca Ministero dell'istruzione, dell'università e della Ricerca, Italia Ministero della Salute, Italia Ministero dell'ambiente, Italia Provincia di Pordenone Provincia di Vercelli SO.GE.NU.S. S.p.A Telethon TM.E. S.p.A 60

63 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Pastorelli R, Saletta F, Campagna R, Carpi D, Dell?osta C, Schiarea S, Vineis P, Airoldi L, Matullo G. Proteome characterization of a human urothelial cell line resistant to the bladder carcinogen 4-aminobiphenyl. Proteome Science 2007 Moretti M, Dell'omo M, Villarini M, Pastorelli R, Muzi G, Airoldi L, Pasquini R. Primary DNA damage and genetic polymorphisms for CYP1A1, EPHX and GSTM1 in workers at a graphite electrode manufacturing plant. BMC Public Health : 270 Vineis P, Veglia F, Garte S, Malaveille C, Matullo G, Dunning A, Peluso M, Airoldi L, Overvad K, Raaschou- Nielsen O, Clavel-Chapelon F, Linseisen JP, Kaaks R, Boeing H, Trichopoulou A, Palli D, Crosignani P, Tumino R, Panico S, Bueno-De-Mesquita HB, Peeters PH, Lund E, Gonzalez CA, Martinez C, Dorronsoro M, Barricarte A, Navarro C, Quiros JR, Berglund G, Jarvholm B, Day NE, Key TJ, Saracci R, Riboli E, Autrup H. Genetic susceptibility according to three metabolic pathways in cancers of the lung and bladder and in myeloid leukemias in nonsmokers. Ann Oncol Jul;18: Vineis P, Hoek G, Krzyzanowski M, Vigna-Taglianti F, Veglia F, Airoldi L, Overvad K, Raaschou-Nielsen O, Clavel-Chapelon F, Linseisen J, Boeing H, Trichopoulou A, Palli D, Krogh V, Tumino R, Panico S, Bueno-De- Mesquita HB, Peeters PH, Lund E E, Agudo A, Martinez C, Dorronsoro M, Barricarte A, Cirera L, Quiros JR, Berglund G, Manjer J, Forsberg B, Day NE, Key TJ, Kaaks R, Saracci R, Riboli E. Lung cancers attributable to environmental tobacco smoke and air pollution in non-smokers in different European countries: a prospective study. Environ Health ; 6:7. Manuguerra M, Matullo G, Veglia F, Autrup H, Dunning AM, Garte S, Gormally E, Malaveille C, Guarrera S, Polidoro S, Saletta F, Peluso M, Airoldi L, Overvad K, Raaschou-Nielsen O, Clavel-Chapelon F, Linseisen J, Boeing H, Trichopoulos D, Kalandidi A, Palli D, Krogh V, Tumino R, Panico S, Bueno-De-Mesquita HB, Peeters PH, Lund E, Pera G, Martinez C, Amiano P, Barricarte A, Tormo MJ, Quiros JR, Berglund G, Janzon L, Jarvholm B, Day NE, Allen NE, Saracci R, Kaaks R, Ferrari P, Riboli E, Vineis P. Multi-factor dimensionality reduction applied to a large prospective investigation on gene-gene and gene-environment interactions. Carcinogenesis : Toropov A A, Benfenati E SMILES in QSPR/QSAR modeling: Results and perspectives Current Drug Discovery Technologies : Benigni R, Netzeva T, Benfenati E, Bossa C, Franke R, Helma C, Hulzebos E, Marchant C, Richard A, Woo Y-T, Yang C The expanding role of predictive toxicology: An update on the (Q)SAR models for mutagens and carcinogens J Environ Sci Heal C : Mandich A, Bottero S, Benfenati E, Cevasco A, Erratico C, Maggioni S, Massari A, Pedemonte F, Viganò L In vivo exposure of carp to graded concentrations of bisphenol A Gen Comp Endocr : Benfenati E The specificity of the QSAR models for regulatory purposes: the example of the DEMETRA project SAR QSAR Environ Res : Maran U, Sild S, Mazzatorta P, Casalegno M, Benfenati E, Romberg M Grid computing for the estimation of toxicity: Acute toxicity on fathead minnow (Pimephales promelas) Lect Notes Comput Sci : Urbatzka R, van Cauwenberge A, Maggioni S, Viganò L, Mandich A, Benfenati E, Lutz I, Kloas W Androgenic and antiandrogenic activities in water and sediment samples from the river Lambro, Italy, detected by yeast androgen screen and chemical analyses Chemosphere : Fratev F, Lo Piparo E, Benfenati E, Mihaylova E Toxicity study of allelochemical-like pesticides by combination of 3DQSAR, docking, local binding energy (LBE) and GRID approaches SAR QSAR Environ Res :

64 Toropov A A, Benfenati E Optimisation of correlation weights of SMILES invariants for modelling oral quail toxicity Eur J Med Chem : Kahn I, Maran U, Benfenati E, Netzeva T I, Schultz T W, Cronin M T D Comparative quantitative structure-activity-activity relationships for toxicity to Tetrahymena pyriformis and Pimephales promelas Altern Lab Anim : Toropov A A, Benfenati E SMILES as an alternative to the graph in QSAR modelling of bee toxicity Comput Biol Chem : Benfenati E, Azimonti G, Auteri D, Lodi M Environmental and ecological toxicology: Computational risk assessment in: Ekins S (Ed.), Computational Toxicology: Risk Assessment for Pharmaceutical and Environmental Chemicals, John Wiley & Sons Inc., Hoboken NJ, USA (2007), Benfenati E, Craciun M, Neagu C D The use of the DEMETRA models in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007), Benfenati E The quality criteria of the DEMETRA models for regulatory purposes: Specificity, general lessons and future perspectives in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007), Amaury N, Benfenati E, Boriani E, Casalegno M, Chana A, Chaudhry Q, Chretien J R, Cotterill J, Lemke F, Piclin N, Pintore M, Porcelli C, Price N, Roncaglioni A, Toropov A A Results of DEMETRA models in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007), Benfenati E, Chretien J R, Gini G, Piclin N, Pintore M, Roncaglioni A Validation of the models in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007), Amaury N, Benfenati E, Bumbaru S, Chana A, Craciun M, Chretien J R, Gini G, Guo G, Lemke F, Minzu V, Mueller J A, Neagu C D, Pintore M, Stroia S A, Trundle P Hybrid systems in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007), Benfenati E, Casalegno M, Cotterill J, Price N, Spreafico M, Toropov A A Characterization of chemical structures in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007), Benfenati E, Boriani E, Craciun M, Malazizi L, Neagu C D, Roncaglioni A Databases for pesticide ecotoxicity in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007), Benfenati E, Clook M, Friday S, Hart A QSARs for regulatory purposes: the case for pesticide authorization in: Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007),

65 Lo Piparo E, Mazzatorta P, Benfenati E Computational methods to study properties of allelochemicals and modelling of molecular interactions in allelopathy in: Roshchina V V, Narwal S S (Eds.), Plant Cell Diagnostics - Images, Biophysical and Biochemical Processes in Allelopathy, Science Publishers, Enfield NH, USA (2007), Pomati F, Cotsapas C, Castiglioni S, Zuccato E, Calamari D. Gene expression profiles in zebrafish (danio rerio) liver cells exposed to a mixture of pharmaceuticals at environmentally relevant concentrations. Chemosphere 2007, 70: Castiglioni S, Zuccato E, Chiabrando C, Fanelli R, Bagnati R. Detecting illicit drugs and metabolites in wastewater using high performance liquid chromatography-tandem mass spectrometry. Spctroscopy Europe 2007, 19 (4): Bagnati R, Bianchi G, Marangon E, Zuccato E, Fanelli R, Davoli E. Direct analysis of isopropylthioxanthone (ITX) in food by HPLC/MS/MS. Rapid Commun Mass Spectrom 2007, 21: Berrie C P, Iurisci C, Piccolo E, Bagnati R, Corda D. Analysis of phosphoinositides and their aqueous metabolites. Methods Enzymol 2007; 434 : Carubelli G, Fanelli R, Mariani G, Nichetti S, Crosa G, Calamari D, Fattore E. PCB contamination in farmed and wild sea bass (Dicentrarchus labrax L.) from a coastal wetland area in central Italy. Chemosphere 2007 ; 68 : LAY PRESS SELECTION PUBLISHED IN 2007 Zuccato E, Castiglioni S, Bagnati R, Fanelli R. I farmaci: inquinanti ambientali ubiquitari. Quaderni acp (5): Zuccato E, Castiglioni S, Fanelli R, Bagnati R. Inquinamento da farmaci: le evidenze (parte I). Ricerca & Pratica 2007; 23 (134): Zuccato E, Castiglioni S, Fanelli R, Bagnati R. Inquinamento da farmaci: la regolamentazione e gli interventi (parte II). Ricerca & Pratica 2007; 23 (135): Fattore E, Davoli E. La qualità dell'ambiente e le problematiche tossicologiche relative alle emissioni di microinquinanti organ ici In: Recupero di energia e materia da rifiuti solidi: i processi, le tecnologie, le esperienze, le norme. AMRA, Napoli, 2007; E. Davoli, G. Bianchi, A. Colombo, M. Lodi, R. Marras, G. Rotella, V. Senese, E. Benfenati. Studi ambientali sui livelli di microinquinanti per e post insediamento di impianti di termovalorizzazione. Nuova Gea- Quaderni Ambiente 2007 ; 2 : OTHER PRODUCTS PUBLISHED IN 2007 Benfenati E (Ed.), Quantitative Structure-Activity Relationships (QSAR) for Pesticide Regulatory Purposes, Elsevier Science Ltd, Amsterdam, The Netherlands (2007),

66 RESEARCH ACTIVITIES Laboratory of Molecular Toxicology Toxicoproteomics Studies are ongoing on the characterization of changes in the proteome profile induced by environmental toxic compounds, with the aim of obtaining protein biomarkers with the ability to differentiate two or more biological states. Proteome changes in tissues and target organs of animals, and cells treated with endocrine disruptors, estrogens, or environmental carcinogens, are related to functional changes during toxicological processes. Qualitative and quantitative proteome changes resulting from the exposure to environmental toxic compounds or in pathological conditions are monitored also in humans and focused on plasma and urine. Proteome analysis includes protein separation by two-dimensional gel electrophoresis and protein identification by mass spectrometry (MALDI-TOF-MS, LC-ESI-MS/MS) coupled to the use of existing databases. Alternatively, peptides resulting from the digestion of protein mixtures with specific proteases are separated by two-dimensional liquid chromatography. Molecular Epidemiology The laboratory works mainly on the measurement of biological markers used to assess human exposure to environmental toxic compounds. Our studies include DNA- and blood proteinadduct formation by several environmental carcinogens. In addition, we study whether the polymorphism of genes coding for enzymes involved in the activation and detoxification of carcinogens are determinants of adduct formation. Genotypes are detected by restriction fragment length polymorphism analysis, after the amplification by polymerase chain reaction of specific nucleotide sequences of the genes under study. The laboratory participates in an international cooperation study aimed at the collection of reference values on allele and genotype frequency of the most common metabolic enzyme polymorphisms in control populations. Pharmacogenetics The study of the human genome has established that part of the observed individual variability in the reaction to pharmacological therapies is due to genetic traits. The analysis of the genetic polymorphism of drug metabolism and disposition allows optimizing the therapy according to the individual genetic makeup. This provides useful information not only for a correct drug dosage, but also for the prevention of adverse drug reactions. Laboratory of Analytical Biochemistry Identification and characterization of proteins by mass spectrometry Our laboratory is developing different analytical and instrumental techniques for the identification and characterization of proteins and peptides in biological samples. This activity is mainly aimed at 1) characterizing different protein isoforms that are found significantly altered in differential proteomics studies with two dimensional gel electrophoresis, 2) profiling proteins in biological fluids for discovery and identification of biomarkers of physiopathological and toxicological relevance, 3) identifying and characterizing endogenous degradation products of proteins, 4) identifying proteins produced by cells in vitro in response to given stimuli, 5) selectively isolating biologically relevant proteins by immunoaffinity-based micro-techniques for subsequent identification by mass spectrometry, and characterization of post-translational modifications by LC-MS/MS. 64

67 Ongoing projects include the study of exogenous protein degradation in renal tubular cells in relation to antigen presentation mechanisms, the search for a protease responsible of the abnormal cleavage of von Willebrand Factor in patients with thrombotic thrombocytopenic purpura, and the characterization of the secretome of cancer cell lines in vitro to identify factors affecting immune cells. Method development in proteomics The laboratory works on the optimization of various analytical methodologies for proteomics, i.e. various complementary techniques of protein identification by mass spectrometry (MALDI- TOF-MS, LC-ESI-MS/MS), isolation and purification of proteins, and characterization of their post-translational modifications. Laboratory of Environmental Chemistry and Toxicology Development and use of analytical methods to evaluate contamination in water bodies, soil, biota, human samples in exposed population Analytical methods are developed to study environmental pollutants in water ecosystems, landfills, contaminated sites. Qualitative and quantitative analyses of organic pollutants are done by mass spectrometry (GC-MS, LC-MS, LC-MS/MS). Typical analyses include PCDD/F, PCB, PAH, polybrominated diphenylethers, pesticides, endocrine disruptor chemicals, and industrial pollutants. Studies on environmental, toxicological and ecotoxicological properties of chemicals Research is carried out on pollutant properties, searching literature data, comparing and evaluating different sources, and mainly developing predictive models to cope with the lack of experimental data. Thus, we develop models starting merely from the chemical structure. The research addresses the different kinds of chemical descriptors and chemical fragments, obtained with different software. Then, we develop models using algorithms such as neural network, fuzzy logic, genetic algorithms, classifiers, multivariate analysis, etc. Different methods are compared and integrated within a structured ensemble. Standardized methods for pesticides were developed and validated according to OECD guidelines. Risk assessment of pollutants Studies are aimed at assessing the risk of pollutants for human population and environment. For this we model transport and diffusion of pollutants, to obtain a predicted concentration on given space and time scales. Such an activity is integrated with those above described on chemical analyses and toxicity prediction, to achieve a continuous transfer of data and research. Research on pollutants emitted in the atmosphere (Unit of Industrial and Environmental Hygiene) Studies address different aspects of atmospheric pollution. Research deals with: sampling areas around the pollution source, chemical analyses, transport modeling depending on meteorological conditions and orography, risk assessment for population and environment. Qualitative and quantitative analyses are done by gas chromatography-mass spectrometry using high resolution for PCDDs/PCDFs, and negative ion-chemical ionization for PCBs. 65

68 Laboratory of Mass Spectrometry Particulate air pollution Epidemiological studies consistently show an association between an increasing number of pathologies, both acute and chronic, and particulate air pollution. This has been shown not only in respiratory, but in cardiovascular diseases as well. Airborne particulate sampling and analysis strategies are developed to characterize both adsorbed compounds and exposition in different activities. Method development in environmental sciences Methods, analytical methodologies, instrumentation and software for data acquisition and reduction, are developed for environmental studies. High-sensitivity instrumentation, mainly based on mass spectrometry, is developed for trace and ultra-trace analysis. Also, transportable instrumentation is developed for field studies or continuous monitoring. Characterization of environmental odor annoyance Characterization of odors poses several analytical problems because they result from a complex mixture of compounds (odorants) stimulating receptors in the nasal cavity. Most odorants are volatile organic compounds (VOC) generated by bacterial degradation of organic matter. They are often present at trace levels, while numerous sources can contribute to the total odor. Using sampling techniques specifically developed for olfactometry, solid phase microextraction and GC/MS analysis, we can detect traces (low ppb to high ppt) of a wide polarity/volatility range of airborne VOC odorant compounds. With a chemometric approach, we can characterize the sources of emissions, assess odor control methods, and identify emissions that contribute to odors in ambient air. Laboratory of Food Toxicology Chemical contaminants in food We are studying human exposure to dietary PCBs and dioxins. PCBs were measured in food items in different European countries, showing differences in PCBs exposure of European consumers. Further studies were aimed at measuring PCBs and dioxins in food from an Italian area at high risk of contamination. Ongoing studies are focused on other emerging food contaminants. Therapeutic and illicit drugs in the environment Pharmaceuticals are a class of emerging environmental pollutants. We have organized a campaign to detect the presence of pharmaceuticals and their metabolites in Italian rivers and sewage treatment plants, with the aim of better characterizing the contamination and assessing related risks. Human and environmental risks are evaluated by studying the toxic effects of pharmaceuticals at environmental levels, on cultures of human and zebra fish cells. Further ongoing studies are aimed at investigating a possible relationship between antibiotic occurrence and resistance in environmental bacteria. The possible presence of illicit drugs in water samples from sewage treatment plants and rivers was investigated, starting with cocaine and its metabolites. Their levels, used to estimate drug abuse in the local population, revealed that cocaine consumption greatly exceeds official estimates. 66

69 Farming methods and emerging pollutants A project has been outlined to set up new methods for the assessment of GMO safety in human nutrition. This project is based on the use of new techniques to detect possible changes induced in living organisms by GMO-derived foods. Regulatory activities On behalf of the Ministry of Health, we carried on the evaluation of the dossiers required for pesticide registration within the European Union. Unit of Environmental Pollutants Risk Assessment Exposure to environmental pollutants Research activities include both quantitative measurement of contaminants in environmental samples, and assessment of exposure. Specific projects are the following: Risk assessment on health effects due to exposure to pollutants from emissions of incinerators and landfills Measurments of persistent organic pollutants such as dioxins and furans polybrominated, and perfuorooctanoic acid and perfluorooctanoic acid (PFOA) and perfluorooctane sulphonate (PFOS) in farmed and wild fish coming from Mediterranean Sea. These data will be used to assess dietary exposure of the general Italian population through fish consumption. Exposure assessment methodology development New methods for exposure assessment are developed, employing probabilistic approaches and more refined statistical models, starting from real cases of contamination. A current study deals with the exposure of the Seveso population to dioxin from contaminated soil. Evaluation of toxicological data Toxicological data resulting from in vivo sub-chronic studies in rats exposed to individual dioxin-like and non dioxin-like PCBs are evaluated in detail, in order to investigate the doseresponse relationship and the applicability of the benchmark dose approach. Unit of Analytical Instrumentation Development and application of analytical methods for compounds of biological and environmental interest Methods are developed mainly using solid phase extraction (SPE) followed by liquid chromatography - mass spectrometry (LC-ESI-MS/MS) or gas chromatography - mass spectrometry (GC-MS). Substances of interest include proteins, peptides, hormones, pharmaceuticals, drugs of abuse, pesticides, and other environmental contaminants (PCBs, hydroxy-pcbs, perfluorinated compounds). 67

70 DEPARTMENT OF NEUROSCIENCE STAFF Head Gianluigi FORLONI, Biol.Sci.D. Laboratory of Biology of Neurodegenerative Disorders Head Cell Death and Neuroprotection Unit Head Gianluigi FORLONI, Biol.Sci.D. Tiziana Borsello, Biol.Sci.D. Laboratory of Drug Metabolism Head Silvio CACCIA, Farm.D. Laboratory of Experimental Neurology Head Annamaria VEZZANI, Biol.Sci.D. Laboratory of Experimental Psychopharmacology Head Luigi CERVO, Ph.D. Laboratory of Geriatric Neuropsychiatry Head Ugo LUCCA, MSc Epidemiology and Social Psychiatry Unit Head Barbara D AVANZO, Philos.D. Geriatric Epidemiology Unit Head Geriatric Pharmacology Unit Head Mauro TETTAMANTI, Biol.Sci.D. Emma RIVA, M.D. Laboratory of Inflammation and Nervous System Diseases Head Maria Grazia DE SIMONI, Biol.Sci.D. Laboratory of Molecular Neurobiology Head Caterina BENDOTTI, Farm.D. 68

71 Laboratory of Neurochemistry abd Behavior Head Roberto William INVERNIZZI, Biol. Sci D Pharmacology of Cognitive Behavior Unit Head Mirjana CARLI, Ph.D. Laboratory of Neurological Disorders Head Ettore BEGHI, M.D. Laboratory of Quality Assessment of Geriatric Services Head Alessandro NOBILI, M.D. 69

72 CURRICULA VITAE Gianluigi Forloni, obtained the Degree of Biological Science at the University of Milan in After two years of post doc at the Department of Neuroscience and Psychiatry at Johns Hopkins University in Baltimore, USA, he came back to the Mario Negri Institute and between 1992 and 1996 he was the head of the Neurobiology of Alzheimer's disease Unit; since 1996 he is the Head of the Biology of Neurodegenerative Diseases Lab and since 2002 the Head of the Neuroscience Department. His scientific interest is focused on the biological and genetic bases of aging-related disorders in particular Alzheimer s disease, Prion-related encephalopathies and Parkinson s disease. He has been member of several European committees for the examination of projects in the neuroscience field. He is now member of the coordination group and thematic leader on drugs development of European Network of Excellence Neuroprion. He is President of the Italian Association on Brain Aging Research (AIRIC) and member of the European Academy of Sciences. He is the author of more than 150 peer-reviewed scientific articles and about 30 reviews or book chapters. Selected publications Forloni G. Angeretti N., Chiesa R., Monzani E., Salmona M., Bugiani O.,Tagliavini F. Neurotoxicity of a prion protein fragment. Nature 362: (1993) Forloni, G., Tagliavini, F.,Bugiani, O. and Salmona, M. Amyloid in Alzheimer s disease and prion-related encephalopathies: Studies with synthetic peptides. Progr. Neurobiol. 49: (1996) Forloni, G., Bertani, I. Calella, AM., Thaler, F.Invernizzi. R. Alpha-synuclein and Parkinson's disease selective neurodegeneration effect of alpha synuclein fragment on dopaminergic neurons in vitro. Ann. Neurol. 47: (2000) Forloni G. Iussich, S. Awan T. Colombo L. Angeretti, N. Girola, L. Bertani, I. Poli, G. Caramelli, M. Bruzzone, MG.Farina, L. Limido, L. Rossi, G. Giaccone G. Ironside, JW. Bugiani, O.Salmona M. and Tagliavini, F. Tetracyclines affect prion infectivity Proc. Natl. Acad. Sci. New York 99: (2002) Pesaresi M, Lovati C, Bertora P, Mailland E, Galimberti D, Scarpini E, Quadri P, Forloni G, Mariani C. Plasma levels of beta-amyloid (1-42) in Alzheimer's disease and mild cognitive impairment. Neurobiol Aging., 27:904-5 (2006) Fioriti, L. Angeretti, N.. Colombo, L., De Luigi A., Manzoni, C., Colombo A., Morbin, M., Tagliavini, F., Salmona, M. Chiesa, R. Forloni, G. Neurotoxic and gliotrophic activity of a synthetic peptide homologous to Gerstmann-Sträussler- Scheinker disease amyloid protein. J. Neurosci. 27: (2007) Ettore Beghi (EB) graduated in Medicine in 1972 and received his specialty in neurology in 1976 at the University of Milan. He trained in epidemiology with a fellowship at the Department of statistics and Epidemiology of the Mayo Clinic in Rochester, MN (USA). He is Head of the Laboratory of Neurological Disorders at the Mario Negri Institute, Director of the Neurophysiology/Epilepsy Unit and Professor of Neuroepidemiology at the University of Milano-Bicocca, Monza. He is member of the editorial board of the journals Epilepsia, Neuroepidemiology, Inpharma, Drugs in R & D, Clinical Drug Investigation, Neurological Sciences and is a referee of several national and international medical journals. The main areas of interest and research include studies on the descriptive, analytic, and experimental epidemiology in the field of epilepsy, peripheral neuropathies, headache, and amyotrophic lateral sclerosis. Selected publications Leone, MA. Solari, A.,Beghi, E. for the FIRST Group. Treatment of the first tonic-clonic seizure does not affect longterm remission of epilepsy. Neurology 2006; 67: Millul, A., E. Beghi, G. Logroscino, A. Micheli, E. Vitelli, A. Zardi, for the Registro Lombardo SLA (SLALOM). Survival of patients with amyotrophic lateral sclerosis in a population-based registry. Neuroepidemiology 2005; 25: Tonini, C., E. Beghi, A.T. Berg, G. Bogliun, L. Giordano, R.W. Newton, A. Tetto, E. Vitelli, D. Vitezic, S. Wiebe. Predictors of epilepsy surgery outcome: a meta-analysis. Epilepsy Res 2004; 62: Van den Broek, M., and Beghi E., for the RESt-1 Group. Morbidity in patients with epilepsy: type and complications. A European Cohort Study. Epilepsia 2004; 45: Van den Broek, M. and Beghi E. for the RESt-1 Group. Accidents in patients with epilepsy: type and complications. A European Cohort Study. Epilepsia 2004; 45: Musico, M., E. Beghi, A. Solari, F. Viani for the First Seizure Trial Group. Treatment of the first tonic-clonic seizure does not improve the prognosis of epilepsy. Neurology 1997; 49:

73 Caterina Bendotti, got her degree in Pharmacy at the University of Milano in 1984; In she was post doc at the Genetic developmental Lab, Dept. of Physiology of the Johns Hopkins University, Baltimore, USA. In she was research fellow in the laboratory of Neuropharmacology and in the 1992, she became head of the Molecular Neurobiology Unit in Institute, since 1998 she is head of laboratory. The major research interest is the study of pathogenetic mechanisms of familial Amyotrophic Lateral Sclerosis.. Since 2002 she is a member of the editorial board of Journal of Neurochemistry. In has been Member of Scientific Committees of the International Symposia on ALS held in Milano, Novembre,2003. In has been member of the Italian Ministry of Health Committees for the diagnosis, cure, care and assistance of patients with ALS. Since 2005 is member of the Board of Directors of the Italian Society of Neuroscience. Since 2006 is member of the Research Advisory Panel of the MND Association, UK. Scientific reviewer of 11 international scientific journals. In 2007 she has co-organised the first international meeting on Mutant SOD1 and familial ALS:from the molecule to man held in Milano(13-16 September). She is author and co-author of 110 articles 100 of which with peer-review. Rapporteur of many communications in national and international meetings. Selected publications Sau D, De Biasi S, Vitellaro-Zuccarello L, Riso P, Guarnieri S, Porrini M, Simeoni S, Crippa V, Onesto E, Palazzolo I, Rusmini P, Bolzoni E, Bendotti C, Poletti A. Mutation of SOD1 in ALS: a gain of a loss of function. Hum Mol Genet. 16(13): , Massignan T, Casoni F, Basso M, Stefanazzi P, Biasini E, Tortarolo M, Salmona M, Gianazza E, Bendotti C, Bonetto V. Proteomic analysis of spinal cord of presymptomatic amyotrophic lateral sclerosis G93A SOD1 mouse.biochem Biophys Res Commun. 353(3):719-25, 2007 Peviani M, Cheroni C, Troglio F, Quarto M, Pelicci G, Bendotti C. Lack of changes in the PI3K/AKT survival pathway in the spinal cord motor neurons of a mouse model of familial amyotrophic lateral sclerosis.mol Cell Neurosci. 34: , 2007 Carri MT, Grignaschi G, Bendotti C. Targets in ALS: designing multidrug therapies. Trends Pharmacol Sci. 27(5):267-73, 2006 Cheroni C., Peviani M., Cascio P., Debiasi S., Monti C. and Bendotti C. Accumulation of human SOD1 and ubiquitinated deposits in the spinal cord of SOD1G93A mice during motor neuron disease progression correlates with a decrease of proteasome. Neurobiol. Disease. 18(3): , 2005 Bendotti C and MT Carri. Lessons from models of SOD1-linked familial ALS. Trends Mol Med. 10(8): , 2004 Bendotti C., Tortarolo M., Suchak S.K., Calvaresi N., Carvelli L., Bastone A., Rizzi M., Rattray M. and Mennini T. Transgenic SOD1 G93A mice develop reduced GLT-1 in spinal cord without alterations in cerebrospinal fluid glutamate levels. J. Neurochem.,79, , 2001 Migheli A., Atzori C., Piva R., Tortarolo M., Girelli M., Schiffer D. and Bendotti C. Lack of apoptosis in mice with ALS. Nature Medicine: 5, , Silvio Caccia. Laurea in Pharmacy at the University of Milan. Diploma in Industrial Chemistry at the L. Cobianchi Technical Institute (Verbania, NO) and Diploma in Biochemical Research at the Istituto di Ricerche Farmacologiche Mario Negri (Milan). Research fellow, Laboratory of General Pharmacology at the Mario Negri Institute, ; Permanent Researcher, Laboratory of Neuropharmacology, 1975; Head of Pharmacokinetic Unit, 1983; Head of Drug Metabolism Laboratory, 1988 to date, doing research in the field of pharmacology and toxicology with particular focus on pharmacokinetic aspects, both at the pre-clinical and clinical level. He has been member of the scientific assessment teams (acting as expert) for the evaluation of marketing authorisation applications submitted to the European and Italian Agencies.He is author and co-author of more than 200 articles, including reviews, monographs and book chapters. Selected publications Caccia S. N-dealkylation of arylpiperazine derivatives: disposition and metabolism of the 1-aryl-piperazines formed. Curr Drug Metab 2007 ; 8 : Caccia S. Main active components of St. John's Wort (Hypericum Perforatum) extracts: current analytical procedures for pharmacokinetics and concentration-response studies. Curr Pharm Anal 2006; 2: Caccia S. Antidepressant-like components of Hypericum perforatum extracts: An overview of their pharmacokinetics and metabolism. Curr Drug Metab 2005; 6: Caccia S. Metabolism of the newest antidepressants: Comparisons with related predecessors IDrugs 2004; 7: Caccia S. Biotransformation of post-clozapine antipsychotics. Pharmacological implications. Clin Pharmacokinet 2000; 38: 39. Caccia S. Metabolism of the newer antidepressants. An overview of the pharmacological and pharmacokinetic implications. Clin Pharmacokinet 1998; 34:

74 Luigi Cervo got the PhD degree from the Open University, Milton Keynes, United Kingdom in 2005 in recognition of a program of research undertaken at the Mario Negri Institute for Pharmacological Research. From 2006 he is the head of the Laboratory of Experimental Psychopharmacolgy. Since 1997 he has been the chief of the Behavioural Pharmacology Unit. From 1978 to 2001 he has been a research fellow in the laboratory of Neuropharmacology and in 1981 he was awarded the degree in Biochemical Research from the Mario Negri Institute. Between 1981 and 1983 he spent two years as a research fellow in the Department of Psychiatry at the Chicago University, Illinois, U.S.A. His main research interest concerns Neuropsychopharmacology and the mechanism of action of psychotropic drugs. In particular the role of receptors subtypes for serotonin, dopamine, noradrenaline and glutamate in drug dependence and drug craving, depression and anxiety. Mechanisms underlying acute as well as chronic pain are also of interest. He is author of numerous peer-reviewed articles in international scientific journals, author of communications in international meetings, and ad-hoc reviewer for several international scientific journals. Selected publications Cervo L, Carnovali, F, Stark JA, Mennini T. Cocaine-seeking behavior in response to drug-associated stimuli in rats: involvement of D 3 and D 2 dopamine receptors. Neuropsychopharmacology 2003; 28: Cervo L, Cocco A, Carnovali F. Effects on cocaine and food self-administration of (+)-HA-966, a partial agonist at the glycine/nmda modulatory. Psychopharmacology (Berl) 2004; 173: Grignaschi G, Burbassi S, Zennaro E, Bendotti C, Cervo L.A single high dose of cocaine induces behavioural sensitization and modifies mrna encoding GluR1 and GAP-43 in rats. Eur J Neurosci. 2004, 20: Cervo L, Mennini T, Rozio M, Ekalle-Soppo CB, Canetta A, Burbassi S, Guiso G, Pirona L, Riva A, Morazzoni P, Caccia S, Gobbi M. Potential antidepressant properties of IDN 5491 (hyperforin-trimethoxybenzoate), a semisynthetic ester of hyperforin. Eur Neuropsychopharmacol. 2005, 15: Cervo L, Canetta A, Calcagno E, Burbassi S, Sacchetti G, Caccia S, Fracasso C, Albani D, Forloni G, Invernizzi RW. Genotype-dependent activity of tryptophan hydroxylase-2 determines the response to citalopram in a mouse model of depression. J Neurosci Sep 7;25(36): Cervo L, Burbassi S, Colovic M, Caccia S. Selective antagonist at D3 receptors, but not non-selective partial agonists, influences the expression of cocaine-induced conditioned place preference in free-feeding rats. Pharmacol Biochem Behav. 2005, 82: Cervo L, Cocco A, Petrella C, Heidbreder CA. Selective antagonism at dopamine D3 receptors attenuates cocaineseeking behaviour in the rat. Int J Neuropsychopharmacol. 2007, 10: Calcagno E, Canetta A, Guzzetti S, Cervo L, Invernizzi RW. Strain differences in basal and post-citalopram extracellular 5-HT in the mouse medial prefrontal cortex and dorsal hippocampus: relation with tryptophan hydroxylase-2 activity. J Neurochem. 2007, 103: Maria Grazia De Simoni got the Doctoral Degree in Biological Sciences in 1977 at the University of Milano, Italy. 1981: Research Specialist in Pharmacology (PhD), Mario Negri Institute, Milan, Italy : European Community fellowship for "Advanced Professional Training", INSERM U 171, Universitè Claude Bernard, Lyon, France; 1984 Department of Histology, Karolinska Institute, Stockholm. Working experience: : Chief of the Neurochemistry Unit, Mario Negri Institute, Milano; 1998-present: Chief of the Laboratory of Inflammation and Nervous System Diseases, Mario Negri Institute. Scientific interests: pathogenesis of cerebral ischemia/reperfusion and traumatic brain injury; inflammatory response and apoptotic mechanisms as targets of therapeutic strategies; animal models and clinical studies. She is member of the board of Master in Tecnologie Avanzate Applicate alle Patologie Neurodegenerative", University of Milan and member of the board of Associazione Italiana per la Ricerca sull Invecchiamento Cerebrale (AIRIC). Selected pubblications De Simoni MG, Storini C, Barba M, Catapano L, Arabia AM, Rossi E, Bergamaschini L. Neuroprotection by complement (C1)-inhibitor in mouse transient brain ischemia. J Cereb Blood Flow Metab, 23: , De Simoni M G, Rossi E, Storini C, Pizzimenti S, Echart C, Bergamaschini L. The powerful neuroprotective action of C1-inhibitor on brain ischemia-reperfusion injury does not require C1q. Am J Pathol., 164: , Bergamaschini L, Rossi E, Storini C, Pizzimenti S, Distaso M, Perego C, De Luigi A, Vergani C and De Simoni MG. Peripheral treatment with enoxaparin, a low-molecular weight heparin, reduces plaques and β-amyloid accumulation in a mouse model of Alzheimer s disease. J. Neurosci. 24: , 2004 Troglio F, Echart C, Gobbi A, Pawson T, Pelicci PG, De Simoni MG & Pelicci G. The neuron-specific Rai (Shc C) adaptor regulates the PI3K-Akt pathway in vivo and protects against cerebral ischemia. Proc Natl Acad Sci U S A 101(43): , Storini C, Bergamaschini L, Gesuete R, Rossi E, Maiocchi D, De Simoni MG. Selective inhibition of plasma kallikrein protects brain from reperfusion injury. JPET 318: ,

75 Capone C, Fabrizi C, Piovesan P, Principato MC, Marzorati C, Ghirardi O, Fumagalli L, Carminati P and De Simoni MG. 2-Aminotetraline derivative protects from ischemia/reperfusion brain injury with a broad therapeutic window, Neuropsychopharmacology, 32: , 2007 Capone C, Frigerio S, Fumagalli S, Gelati M, Principato M C, Storini C, Montinaro M, Kraftsik R, De Curtis M, Parati E, De Simoni MG. Neurosphere - derived cells exert a neuroprotective action by changing the ischemic microenvironment. PLoS ONE 2 e373, Roberto W. Invernizzi started his career in the laboratory of Neuropharmacology of the Istituto di Ricerche Farmacologiche Mario Negri in 1976, where, at present, he heads the Laboratory of Neurochemistry and Behavior. In 1986 he got his degree in Biological Sciences at the Università Statale di Milano and in 1996 he was nominated head of the Intracerebral Microdialysis Unit. Of particular interest to Invernizzi s research team is the study of the neurochemical mechanisms and neuronal circuitries involved in the pathology of the main psychiatric diseases, such as depression and schizophrenia and in the mechanism of action of psychotropic drugs. Since 1987 he applied the intracerebral microdialysis technique to study the in vivo release of monoamines. Using this technique, Invernizzi s team first contributed to clarifying the role of serotonergic and adrenergic autoreceptors in the effect of antidepressant drugs suggesting new hypotheses on their mechanism of action. Currently, Invernizzi s team is involved in two main collaborative projects aimed at clarifying the neurochemical mechanisms involved in the resistance to antidepressant drugs and the role of glutamatergic and serotonergic mechanisms in attentional processes. Reviewer for various international journals in the field of pharmacology and neurochemistry. Author and co-author of more than 60 peer-reviewed articles. Member of the Italian Society of Neuroscience and the Italian Society of Pharmacology. Selected publications Baviera M, Invernizzi RW, Carli M. Haloperidol and clozapine have dissociable effects in a model of attentional performance deficits induced by blockade of NMDA receptors in the mpfc. Psychopharmacology (Berl) 2007 published online. Calcagno E, Canetta A, Guzzetti S, Cervo L, Invernizzi RW. Strain differences in basal and post-citalopram extracellular 5-HT in the mouse medial prefrontal cortex and dorsal hippocampus: relation with tryptophan with tryptophan hydroxylase-2 activity. J Neurochem 2007; 103 : Invernizzi RW, Pierucci M, Calcagno E, Di Giovanni G, Di Matteo V, Benigno A, Esposito E. Selective activation of 5HT2C receptors stimulates GABA-ergic function in the rat substantia nigra pars reticulata: a combined in vivo electrophysiological and neurochemical study. Neuroscience : Carli M, Baviera M, Invernizzi RW, Balducci C Dissociable contribution of 5-HT1A and 5-HT2A receptors in the medial prefrontal cortex to different aspects of executive control such as impulsivity and compulsive perseveration in rats Neuropsychopharmacology 2006; 31: Calcagno E, Carli M, Invernizzi RW The 5-HT1A receptor agonist 8-OH-DPAT prevents prefrontocortical glutamate and serotonin release in response to blockade of cortical NMDA receptors J Neurochem 2006; 96: Cervo L, Canetta A, Calcagno E, Burbassi S, Sacchetti G, Caccia S, Fracasso C, Albani D, Forloni G, Invernizzi RW Genotype-dependent activity of tryptophan hydroxylase-2 determines the response to citalopram in a mouse model of depression J Neurosci 2005; 25: Greco B, Invernizzi RW, Carli M Phencyclidine-induced impairment in attention and response control depends on the background genotype of mice: reversal by the mglu2/3 receptor agonist, LY Psychopharmacology (Berl) 2005; 179: Ugo Lucca got his Master of Science, University of Aberdeen - UK, At the Mario Negri Institute he was investigator from , head of the "Clinical Evaluation of Antidementia Drugs Unit" ( ) and, since 1996, head of the "Laboratory of Geriatric Neuropsychiatry". The main areas of interests include epidemiology and clinic features of dementia; natural history of dementia; neuropsychiatric disorders of the elderly; instruments for the screening diagnosis and clinical course assessment of dementia; clinical evaluation of anti dementia treatments and CNS active drugs (phase I, II, III, IV and observational studies). Selected publications Spagnoli A, Lucca U, Menasce G, Bandera L, Cizza G, Forloni G, Tettamanti M, et al. Long-term acetyl-l-carnitine treatment in Alzheimer's disease. Neurology 1991; 41: Lucca U, Comelli M, Tettamanti M, Tiraboschi P, Spagnoli A. Rate of progression and prognostic factors in Alzheimer s disease: a prospective study. J Am Geriats Society 1993; 41: Lucca U, Tettamanti M, Forloni G, Spagnoli A. Nonsteroidal anti-inflammatory drug use in Alzheimer s disease. Biological Psychiatry 1994; 36:

76 Imbimbo BP, Martelli P, Troetel WM, Lucchelli F, Lucca U, Thal LJ, and the Eptastigmine Study Group. Efficacy and safety of eptastigmine for the treatment of patients with Alzheimer s disease. Neurology 1999; 52: Quadri P, Fragiacomo C, Pezzati R, Zanda E, Forloni G, Tettamanti M, Lucca U. Homocysteine, folate, and vitamin B- 12 in mild cognitive impairment, Alzheimer s disease and Vascular Dementia. Am J Clinical Nutr 2004; 80: Lucca U, Tettamanti M, Quadri P. Homocysteine lowering and cognitive performance. New England Journal of Medicine 2006; 355: Alessandro Nobili got his degree in Medicine (Milan, 1990). Master in Biotechonological Research, Regione Lombardia, Milan International School of Pharmacology, 31 Course on: Drug Epidemiology and Post-marketing Surveillance, Erice, September Course on: Methods in Epidemiological Research, Milan, October Course: Long Term Clinical Trials, Cogne January Main areas of interest Methodology of Randomized Clinical Trials; Pharmacoepidemiology and postmarketing surveillance research; Drug utilization studies; Quality assessment of geriatric services; Qualitative studies on caregiver role in the care of patients with dementia; Methodological evaluation of the Special Care Unit for Alzheimer Disease patients; Methodology of drug information. Employment and research experience Chief of the Unit of Quality Assessment of Geriatric Services Chief of the Drug Information Services for the Elderly, Laboratory of Geriatric Neuropsychiatry, Istituto di Ricerche Farmacologiche Mario Negri, Milan. Editorial Board of the MICROMEDEX Inc., Englewood, Colorado USA. National Expert accredited by Italian Ministry of Health for The Italian (AIFA) and European Agency for the Evaluation of Medicinal Products (EMEA). Head of the Laboratory of the Quality Assessment of Geriatric Services at the Mario Negri Institute since Selected publications Nobili A, Tettamanti M, Frattura L, et al. Drug use in the elderly in Italy. Ann Pharmacother 1997; 31: Nobili A, Gebru F, Rossetti A, et al. Doctorline a private toll-free telephone medical information service. Ann Pharmacother 1998; 32: Nobili A, Riva E, Tettamanti M, et al. The effect of a structured intervention on caregivers of patients with dementia and problem behavior: a randomized controlled pilot study. Alzheimer Dis Assoc Disord 2004; 18: Lucca U, Nobili A, Riva E, Tettamanti M. Low level of B vitamins and the risk of cognitive and functional decline in the very-old: results from the Monzino 80-Plus Study. Neurobiol Aging 2004; 25: 31. Lucca U, Nobili A, Riva E, Tettamanti M Cholinesterase inhibitor use and age in the general population Arch Neurol 2006; 63: Tettamanti M, Garri' M T, Nobili A, Riva E, Lucca U Low folate and the risk of cognitive and functional deficits in the very old: The Monzino 80-plus study J Am Coll Nutr 2006; 25:

77 Annamaria Vezzani got her Degree in Biological Science at the University of Milan in 1978 and she specialized in Neuropharmacology at the Mario Negri Institute in She spent her post-doctoral period in Baltimore at the University of Maryland in working on the mechanisms of epileptogenesis in experimental models of epilepsy. She spent additional post-doctoral periods at the University of Stockholm and at the Karolinska Institute between 1985 and She was on sabbatical at the Albert Einstein College of Medicine in 2002 in the laboratory of Developmental Epilepsy. She is involved in studies on the biochemical and molecular mechanisms involved in the etiopathogenesis of seizures disorders using experimental models of epilepsy. The present research is focused on the functional role of neuroactive peptides and inflammatory mediators in the modulation of neuronal excitability and seizure-related neurodegeneration. Focus of the research is also on the mechanisms of pharmacoresistance. Since 1997 she is the Head of the Laboratory of Experimental Neurology at the Mario Negri Institute. She is member of the Editorial Board of Epilepsy Currents and Neuroscience and Associate Editor for Exp Models of Epilepsia. She is appointed of the Chair of the Commission on Neurobiology of International League Against Epilepsy which is promoting initiatives for improving translational research in epilepsy. Selected publications Balosso S, Ravizza T, Perego C, Peschon J, Campbell I, De Simoni MG, Vezzani A. TNF-alpha inhibits kainic acidinduced seizures in mice via p75 receptors (2005) Ann Neurol, 57, 804 Dube C., Vezzani A., Behrens M., Bartfai T., Baram TZ. (2005) Interleukin-1beta contributes to the generation of experimental febrile seizures. Ann Neurol, 57,152. Richichi C, E-J. D. Lin, D. Stefanin, D. Colella, T. Ravizza,G. Grignaschi, G. Sperk, M. J. During and A. Vezzani Anticonvulsant and antiepileptogenic effects mediated by adeno-associated virus vector neuropeptide Y expression in the rat hippocampus (2004) J Neurosci, 24,3051 Rizzi M, Caccia S, Guiso G, Richichi C, Gorter JA, Aronica E, Aliprandi M, Bagnati R, Fanelli R, D'Incalci M, Samanin R, Vezzani A. Limbic seizures induce P-glycoprotein in rodent brain: functional implications for pharmacoresistance (2002) J Neurosci, 22, 5833 Vezzani A., Moneta D., Conti M., Richichi C., Ravizza T., De Luigi A., De Simoni M.G., Sperk, Andell-Jonsson S., Lundkvist J., Iverfeldt K. and Bartfai T. (2000) "Powerful anticonvulsant action of IL-1 receptor antagonist upon intracerebral injection and astrocytic overexpression in mice" Proc. Natl. Acad. Sci. USA, 97, Vezzani A., Moneta D., Conti M., Richichi C., Ravizza T., De Luigi A., De Simoni M.G., Sperk, Andell-Jonsson S., Lundkvist J., Iverfeldt K. and Bartfai T. (2000) "Powerful anticonvulsant action of IL-1 receptor antagonist upon intracerebral injection and astrocytic overexpression in mice" Proc. Natl. Acad. Sci. USA, 97, Tiziana Borsello got her Degree in Biological Science at the University of Torino in 1990 and she then obtained a PhD in Neuroscience at the University of Turin Medical School. She won 1 year fellow of the European Science Foundation scholarship for work at the Netherlands Research Institute of Amsterdam. From 1997 to 1999 she was a Researcher at the Institute of Neurobiology, CNR, Rome Italy. In the period she was Premier Assistant, Département de Biologie Cellulaire et de Morphologie, Université de Lausanne, Switzerland, and then became Maitre Assistant and group leader in the same institute. In 2004 joined the Biol. Neurodeg. Disorders Lab at the "Mario Negri Institute. In 2005 won the Prize of the Pfizer Foundation, Neuroscience and Diseases Nervous System. Since 2006 she is the Head of the Unit: Neuronal Death and Neuroprotection. Her main scientific interest is understanding the role of signalling pathways in neuronal death after different stress-stimuli and the neuroprotection. In particular, the present research is focused on the study of the mechanisms of excitotocic stress, ischemia, Traumatic Brain Injury and the cell death pathways in neurodegenerative diseases as Alzheimer, with the challenge to define the neuronal death pathways to design more specific methods of neuroprotection. Selected pubblications Repici M, Centeno C, Tomasi S, Forloni G, Bonny C, Vercelli A, Borsello T.Activation After Cerebral Ischemia And Effect Of D-Jnki1 On C-Jun And Caspase-3 Activation. Neuroscience. 2007, 150: 40-9 Borsello T., Centeno C., Riederer IM, Haeflinger JA and Riedere BM. Phosphorylation-dependent dimerization and subcellular localization of islet-brain 1/c-Jun N-terminal kinase-interacting protein 1. J Neurosci Res. 2007, 85: Borsello T Ed Neuroprotection: Methods In Molecular Biology Published By Humana Press, Usa Humana Press, USA, Methods in Molecular Biology, June 2007 Colombo A, Repici M, Pesaresi M, Santambrogio S, Forloni G, Borsello T. The Tat-Jnk Inhibitor Peptide Interferes With Beta Amyloid Protein Stability Cell Death Differ. 2007, 14: Borsello T and Forloni G. JNK signalling: a possible target to prevent neurodegeneration. Current Pharmaceutical Design 2007, 13, Centeno C., Repici M., Chatton J. Y., Riederer B. M., Bonny C., Nicod P., Price M., Clarke P. G., Papa S., Franzoso G. and Borsello T. Role of the JNK pathway in NMDA-mediated excitotoxicity of cortical neurons. Cell Death Differ, 2007, 14:

78 Borsello T. and Bonny C.Use of cell-permeable peptides to prevent neuronal degeneration. Trend in Mol. Med. 2004, 10: Borsello T, Clarke PG, Hirt L, Vercelli A, Repici M, Schorderet DF, Bogousslavsky J, Bonny C. A peptide inhibitor of c-jun N-terminal kinase protects against excitotoxicity and cerebral ischemia. Nature Med. 2003, 9: Mirjana Carli started his scientific career in the laboratory of Neuropharmacology of the Istituto di Ricerche Farmacologiche Mario Negri Milan in 1977, where, at present, she is head of the Pharmacology of Cognitive Behaviour Unit. She spent a few years in the laboratory of Cognitive Neuroscience, Dept. of Experimental Psychology, University of Cambridge (UK) directed by Prof. Trevor W. Robbins. Here she took interest in the role of brain monoamines in attention, and for this purpose developed several behavioral tests for rats. In 1986 she returned to the laboratory of Neuropharmacology of the Istituto di Ricerche Farmacologiche Mario Negri. Here she devoted her efforts to the study of the role played by neuronal mechanisms in cognitive processes such as memory, attention and executive functions. Her work has improved the knowledge of the role played by some serotonin receptors in cognitive processes. Selected publications Carli M, Baviera M, Invernizzi R, Balducci C Dissociable contribution of 5-HT1A and 5-HT2A receptors in the medial prefrontal cortex to different aspects of executive control such as impulsivity and compulsive perseveration in rat Neuropsychopharmacology 2006; 31: Greco B, Carli M Reduced attention and increased impulsivity in mice lacking NPY Y2 receptors: Relation to anxiolyticlike phenotype Behav Brain Res 2006; 169: Carli M, Baviera M, Invernizzi R, Balducci C The serotonin 5-HT2A receptors antagonist MI00907 prevents impairment in attentional performance by NMDA receptor blockade in the rat prefrontal cortex Neuropsychopharmacology 2004; 29: Balducci C, Nurra M, Pietropoli A, Samanin R, Carli M Reversal of visual attention dysfunction after AMPA lesions of the nucleus basalis magnocellularis (NBM) by the cholinesterase inhibitor donepezil and by a 5-HT(1A) receptor antagonist WAY Psychopharmacology (Berl) 2003; 167: Carli M, Balducci C, Samanin R. Stimulation of 5-HT1A receptors in the dorsal raphe ameliorates the impairment of spatial learning caused by intrahippocampal 7-chloro-kynurenic acid in naive and pretrained rats Psychopharmacology (Berl) 2000; 158: Carli M, Samanin R The 5-HT1A receptor agonist 8-OH-DPAT reduces rats' accuracy of attentional performance and enhances impulsive responding in a five-choice serial reaction time task: Role of presynaptic 5-HT1A receptors Psychopharmacology (Berl) 2000; 149: Barbara D Avanzo obtained her master in Philosophy in 1989 at the University of Milan. Her main field of interest is epidemiologic research in mental health. She was involved in the analysis of the implementation of the psychiatric reform in Italy and quality evaluation of services and their recent modifications with specific attention to the role of psychiatric residential facilities in the community service networks; evaluation of effectiveness of the most common psychosocial interventions; suicide trend monitoring and study of suicide prevention programs and initiatives. More recently, she is working on issues like recovery-oriented services, consumers empowerment, and methods of participation of consumers to evaluation of services, and acknowledgment of the value of the consumers point of view about psychiatric treatments and services. She worked as researcher in the Laboratory of General Epidemiology between 1991 and 1996, and she is Chief of the Unit of Epidemiology and Social Psychiatry since Member of the Scientific National Board of WAPR Italy and of the World Head Office of the WAPR. Selected publications Barbato A, D'Avanzo B. Marital therapy for depression. Cochrane Database Systematic Reviews 2006; Issue 2. Parabiaghi A, Barbato A, D'Avanzo B, Erlicher A, Lora A. Assessing reliable and clinically significant change on Health of the Nation Outcome Scales: method for displaying longitudinal data. Aust N Z J Psychiatry 2005; 39: Barbato A, D'Avanzo B. Involuntary placement in Italy. Br J Psychiatry 2005; 186: Guaiana G, Andretta M, Corbari L, Mirandola M, Sorio A, D'Avanzo B, Barbui C. Antidepressant drug consumption and public health indicators in Italy, J Clinical Psychiatry 2005; 66: D'Avanzo B, Battino R N, Gallus S, Barbato A. Factors predicting discharge of patients from community residential facilities: A longitudinal study from Italy. Aust N Z J Psychiatry 2004; 38: D'Avanzo B, Barbato A, Barbui C, Battino N, Civenti G, Frattura L. Discharges of patients from public psychiatric hospitals in Italy between 1994 and Int J Social Psychiatry 2003; 49:

79 Emma Riva, Medical Doctor degree in 1984 University of Milan, PhD in 1990 in Cardiovascular Pathophysiology at the University of London (UK) Training: Research Assistant, Department of Pharmacology, Medical School, University of Ottawa, Canada; Internship in Internal Medicine, Ospedale Luigi Sacco, Milan; Cardiac Fellow, St Thomas' Hospital, London, UK. Field of interest: Prevalence and effects of anemia on cognitive, functional and clinical variables in the elderly; Problem behaviors in dementia; Burden for care-givers of Alzheimer Disease patients; End of life care. Present and past roles in Institute Head of the Geriatric Pharmacology Unit, Istituto "Mario Negri", Milan; Scientific Director of the hospice Via di Natale Franco Gallinin, Aviano, Italy; Consultant Istituto Geriatrico Pio Albergo Trivulzio, Milan: Project member of PREDICT (Policy Review and Evaluation of Dementia and Institutional Care Trends): a Transnational Comparison. Selected publications Tettamanti M, Garrì MT, Nobili A, Riva E, Lucca U. Low folate and risk of cognitive and functional deficits in the very old: The Monzino 80-plus study. J Am Coll Nutr 2006;25: Lucca U, Nobili A, Riva E, Tettamanti M. Cholinesterase inhibitor use and age in the general population. Arch Neurol 2006;63: Nobili A, Riva E, Tettamanti M, Lucca U, Liscio MR, Petrucci B, Salvini Porro G. The effect of a structured intervention on caregivers of patients with dementia. Results of a Randomized Controlled Study. Alzheimer Dis and Associated Disorders 2004;18:75-82 Il malato terminale oncologico. Esperienze dall hospice. Ed. Emma Riva. Il Pensiero Scientifico, 2001 Riva E, Tettamanti M, Gallini C. Il ruolo del medico di medicina generale nella gestione dei malati terminali oncologici. Indagine svolta tra i medici di medicina generale in Friuli Venezia Giulia. Ricerca & Pratica 2001 Riva E, Nobili A, Trecate F. Per un impiego "ragionato" dei neurolettici, per la gestione dei disturbi del comportamento in corso di Malattia di Alzheimer. Rec Prog Med 1998;89: Mauro Tettamanti got his Biology Degree at the Università degli Studi di Milano in 1986, and the specialisation in Epidemiology and Medical Statistics in 1993, at the Università degli Studi di Pavia. Teaching experience Introduction course to statistics, Master in Ergonomy, Politecnico di Milano, years Areas of interest: Planning, conduction and analysis of clinical trials and epidemiologic researches in the geriatric field: Phase I, II, III and observational studies on the efficacy of drugs on neurologic disorders, with special emphasis on dementia; Effects of multi-disciplinary interventions on geriatric/dementia patients; Epidemiology and risk factors of dementia; Care of patients with terminal illness; Association of anemia with prevalence of diseases and cognitive problems Scholarship between 1989 and 1998, Senior Researcher since 1999 and Head of the Unit of Geriatric Epidemiology at the Mario Negri Institute since Selected publications Spagnoli A, Lucca U, Menasce G, Bandera L, Cizza G, Forloni G, Tettamanti M, et al. Long-term acetyl-l-carnitine treatment in Alzheimer's disease. Neurology 1991; 41: Lucca U, Comelli M, Tettamanti M, Tiraboschi P, Spagnoli A. Rate of progression and prognostic factors in Alzheimer's disease: A prospective study. J Am Geriatr Soc 1993; 41:45-49 Quadri P, Fragiacomo C, Pezzati R, Zanda E, Forloni G, Tettamanti M, Lucca U. Homocysteine, folate, and vitamin B- 12 in mild cognitive impairment, Alzheimer disease, and vascular dementia. Am J Clin Nutr 2004; 80: Lucca U, Nobili A, Riva E, Tettamanti M. Cholinesterase inhibitor use and age in the general population. Arch Neurol 2006; 63: Lucca U, Tettamanti M, Quadri P. Homocysteine lowering and cognitive performance. N Engl J Med 2006; 355:1390 Tettamanti M, Garri' M T, Nobili A, Riva E, Lucca U. Low folate and the risk of cognitive and functional deficits in the very old: The Monzino 80-plus study. J Am Coll Nutr 2006; 25:

80 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The Department of Neuroscience is formed by ten Laboratories; the activities of research are devoted to the study of neurological and psychiatric diseases, evaluated by the biological point of view, clinical and epidemiological aspects and the quality of care. Together with these activities, in the Department other more general expertise are present. Pharmacokinetics studies, drug information service and preparation of protocols for clinical trial and epidemiological studies are activities in charge of the Neuroscience Department. Traditionally part of the Department was devoted to the creation of experimental models for the pharmacological, neurochemical and pathogenetic studies in Alzheimer or prion's diseases, epilepsy, depression and cognitive impairment. More recently, consolidated expertise were created in the pathogenesis of amyotrophic lateral sclerosis (ALS), cerebral stroke and drug abuse. Some of these disorders, like epilepsy, ALS and Alzheimer's disease are investigated from the clinical and epidemiological points of view for the evaluation of drug and care efficacy. The activities of the Department are aimed to an integration of the different expertise to develop multidisciplinary approaches. The purpose is to address at different levels, knowledge, therapy and clinical practice to the numerous questions, largely unresolved, proposed by the disorders of nervous system. FINDINGS/MAIN RESULTS 2007 In a cellular model it has been shown that the peptide of D-JNK-1-TAT inhibiting the JNK phosphorylation activity mediated by the enzyme JNK, exerts a control on β amyloid production, these data indicate possible therapeutic perspectives in Alzheimer s disease The transgenic mice overexpressing the mutated form of human protein precursor of amyloid (APP) have a cognitive deficit associated to glutametergic dysfunction independent from the β amyloid deposits In cellular model the presence of α synuclein mutations associated to Parkinson s disease, does not influence the neuroprotective activity of the protein, this indicates that these mutations affected the aggregation state more then physiological activity of α synuclein. In the transgenic mice overexpressing a mutated form of human prion protein associated to CJD generated in the Institute exhibit some behavioral features reminiscent of the clinical condition in humans carrying the same mutation. In an epidemiological study we found that the values of some plasmatic parameters (IGF, IL-6) showed an age-dependent trend until 90 years old, for the older population the correlation with age disappeared. In a prospective population-based study in the very old (Monzino 80-plus Study), depressive symptoms were common prior to dementia. However, the risk of developing dementia associated with depression, if present, seemed modest. In the same prospective population-based study in the very old (Monzino 80-plus Study), walking one hour or more a day was associated with an almost 40% decreased risk of developing dementia. 78

81 In a rural population of elderly aged 75 years and older, dividing the subjects in three groups on the basis of disability (no disability at all, instrumental disability and basic activity disability) gave groups with similar size. People with major disability were older, had a higher number of diagnoses, received more specialist visits (but a similar number of primary care physician visits and hospitalizations) and needed more paid assistance (double than those with minor disability and ten times more than those without disability). In a prospective ambulatory population of cognitively normal or mildly cognitively impaired elderly, individuals with baseline elevated homocysteine concentrations have an almost 3-fold increased risk of developing dementia in the following 3-4 years. More than 1 out of 10 elderly persons (65-84 years) were anemic and most of the cases had a mild grade anemia. Hemoglobin concentrations decreased and prevalence of (mild) anemia increased with increasing age. After controlling for many potential confounders, mild anemia was found to be associated with poorer health conditions and with increased risk of clinically relevant outcome such as hospitalization and mortality. Patients with dementia resident in Alzheimer s special care units (ASCU) had a lower rate of hospitalisation and use of physical restraints than those in traditional nursing homes. In ASCU 60% of patients with dementia were taking at least one antipsychotic, 49% typical and 51% atypical. More than 50% of patients exposed to antipsychotics at baseline, were still taking the drug after 18 months of follow-up. The use of antipsychotic agents was strongly related to the presence of agitation, irritability, delusions, anxiety, night-time behaviour and aberrant motor behaviour. From January to December 2003, in the Lecco Local Health Authority, 15.5% of elderly patients were exposed to potential severe drug-drug interactions (60.8% were women). The risk of drug interaction increased with age and the number of prescription and chronic therapies. Marked differences in the quality of acute care offered across various acute wards in Italy. Seriuos inadequancies were reported in physical and process indicators in the private wards. Whereas efficacy on crisis was evaluated as rather good, satisfaction of patients with the care and assistance received during the admission was modest. The Natural Networks patients, followed according to a naturalistic design, showed significant improvements in quality of life, needs satisfaction, and social functioning. In amyotrophic lateral sclorosis loss of ambulation, percutaneous gastrostomy and non-invasive mechanical ventilation are outcome measures to be used in epidemiological surveys and therapeutic trials. The long-term prognosis of epilepsy is the same in patients treated at the first seizure and those treated at the recurrence. These findings suggest that treatment should be started at the first seizure only on a case-by-case basis. The use of a third drug in children refractory to two anticonvulsants does not affect the chance of seizure remission, suggesting that drug resistance in epilepsy can be identified at the time of failure of two drugs. Focal dystonia in the adult is a rare age-related clinical condition. Blepharospasm is the commonest disease variety. 79

82 The proteomic analysis of the blood cells of ALS patients has evidenced variations of some proteins that are also found alterated in the spinal cord of SOD1G93A mice at the presymptomatic stage. Therefore, these proteins can be considered potential candidates for the identification of the pathogenetic mechanisms of ALS and as useful biomarkers for the early diagnosis of the disease We evidenced a specific proteasomal dysfunction in the motor neurons of SOD1G93A mice at the symptomatic stages which may contribute to the accumulation of intracellular protein aggregates. In addition the proteomic characterization of the insoluble proteins from the spinal cord of SOD1G93A mice has identified a series of oxidated proteins suggesting a possible link between oxidative stress and aggregation pathways in ALS pathogenesis. We have demonstrated that specific neuropeptide Y and somatostatin receptor subtypes mediate the anticonvulsant activity of these endogenous peptides. Thus, this knowledge may be exploited to develop new anticonvulsant drugs with a novel mechanism of action We have demonstrated the modulatory role of some pro- and anti-inflammatory cytokines in seizures using experimental models of epilepsy in rodents, thus describing a new etiopathological mechanism which may be relevant for human epilepsy We have demonstrated that membrane-bound drug transport proteins are functionally activated by seizures and have a significant role in decreasing the brain concentrations of antiepileptic drugs in experimental models. Pharmacological intervention to block the activity of these proteins may contribute to reverse multidrug resistance in epilepsy Complement inhibitor (C1-INH) has powerful neuroprotective actions in brain ischemia/reperfusion injury Microglia can explain protective actions in the ischemic environment Neural stem cell reduce ischemia/reperfusion injury by changing the ischemic environment. Agonists at serotonin 2C receptors as well non-selective opioid receptor antagonists selectively modulate, in laboratory rodents, the drug seeking behaviour induced by the environmental stimuli predictive of cocaine availability. DBA/2J and BALB/c mice are not responder to serotonin selective uptake blockers in an animal model predictive of the antidepressant activity and may represent an animal model of resistance to SSRI Agonists at serotonin 2C receptors as well non-selective opioid receptor antagonists selectively modulate, in laboratory rodents, the drug seeking behaviour induced by the environmental stimuli predictive of cocaine availability Genetic differences in serotonin synthesis contribute to the efficacy of SSRIs in mice Co-treatment with tryptophan restores the antidepressant-like effect in mice non-responder to the SSRI alone 80

83 Basal and SSRI-induced serotonin release is strongly suppressed in mice carrying the mutation of TPH-2, the limiting step enzyme in brain serotonin synthesis The blockade of NMDA receptors of the rat prefrontal cortex induces an increase of glutamate release and is deleterious for prefrontal cortex-dependent cognitive functions NATIONAL COLLABORATIONS Associazione Familiari Insonnia Familiare Fatale malattie da prioni, Treviso Associzione Italiana GIST A.I.G. Associazione per la Ricerca Neurogenetica, Lamezia Terme (CS) e ASL 6, Regione Calabria Agenzia di Sanità Pubblica del Lazio, Regione Lazio Assessorato alla Salute, Comune di Milano Azienda Ospedaliera Ospedali Riuniti di Bergamo Azienda Sanitaria Locale di Bergamo CEND, Centro Eccellenza per le Malattie Neurodegenerative, Università di Milano Centro Fatebenefratelli San Giovanni di Dio, Cernusco sul Naviglio Centro di Neurofarmacologia, Dipartimento di Scienze Farmacologiche, Università di Milano Centro Studi in Psichiatra, ASL 2, Torino Centro Parkinson-Istituti Clinici di Perfezionamento Clinica IRCSS S. Maria Nascente, Milano Clinica Neurologica III Università di Milano, Azienda Ospedaliera S. Paolo, Milano Clinica Psichiatrica, Università Milano Bicocca Consorzio Ricerche Luigi Amaducci, CRIC, Arcugnano (Vc) Consorzio MIA, Milano DIBIT, San Raffaele Scientific Insitute, Milano. Dipartimento di Chimica Biologica, Università di Padova Dipartimento di Chimica, Università egli Studi di Firenze Dipartimento Endicronologia, Università di Milano Dipartimento Farmaco Chimico Tecnologico, Università di Siena Dipartimento di Farmacologia Medica, Università di Milano Dipartimento di Fisiologia Umana, Facoltà di Medicina, Università di Milano Dipartimento di Medicina e Sanità Pubblica, Sezione di Psichiatria e Psicologia Clinica, Università di Verona Dip. di Morfofisiologia, Scuola di medicina Veterinaria, Università di Torino, Grugliasco (TO). Dip. Neurologia, IRCCS Fondazione Maugeri, Pavia Dipartimento Neurologia, Ospedale Molinette, Torino Dipartimento di Neurologia Università di Milano, Ospedale Luigi Sacco. Dipartimento di Neuroscienze, Università di Parma, Parma Dipartimento di Salute Mentale di Niguarda, Milano Dipartimento di Salute Mentale ASL 3 Genovese, Genova Dipartimento di Salute Mentale ASL 4, Torino Dipartimento di Salute Mentale, Azienda Ospedaliera Carlo Poma di Mantova, Mantova Dipartimento di Salute Mentale San Carlo, Milano Dip. di Scienze Biomolecolari e Biotecnologie, Università di Milano Dipartimento Scienze Neurologiche, Università di Genova, Genova Dipartimento Scienze Neurologiche, Ospedale Maggiore Policlinico di Milano Direzione Generale Famiglia e Solidarietà Sociale, Regione Lombardia, Milano Direzione Generale Sanità, Regione Lombardia, Milano 81

84 Direzione Regionale Sanità e Servizi Sociali, Regione Umbria Divisione Neurologica, Università di Bologna Evidentia Medica, Grottaferrata, Roma Federazione Alzheimer Italia, Milano Franco Calori Cell Factory, Centro Trasfusionale e di Immunologia dei Trapianti, IRCCS Ospedale Maggiore, Milano Fondazione Clelio Angelino Fondo Edo Tempia Hospice Via di Natale Franco Gallini, Aviano (PN) IRCSS "Casa Sollievo della Sofferenza", San Giovanni Rotondo IRCCS Istituto Auxologico Italiano, Milano Istituto di Ricovero e Cura a Carattere Scientifico IRCCS (I.N.R.C.A.), Ancona IRCSS "San Raffaele", Milano Istituto Europeo di Oncologia, IRCCS, Milano Istituto di Farmacologia e Farmacognosia, Università di Urbino Istituto di Farmacologia, Università di Milano Istituto G. Ronzoni, Milano Istituto Nazionale Neurologico Carlo Besta, Milano Istituto Scientifico Humanitas Istituto di Scienze e Tecnologie della Cognizione, CNR, Roma Istituto "Stella Maris", IRCCS, Calambrone (PI) Istituto Superiore di Sanità, Roma Istituto Zooprofilattico Piemonte Liguria Val D'Aosta,Torino Laboratorio di Immunopatologia Renale, Ospedale San Carlo, Milano Laboratorio di Neuroscienze, Centro Dino Ferrari, Università di Milano Lega Italiana per la Lotta contro i Tumori Ospedale del Bambin Gesu, Roma Ospedale Regionale Ca Fondello, Treviso Ospedale "Molinette", Torino Polo Oncologico, ASL 12, Biella Provincia Lombardo-Veneta Ordine Ospedaliero San Giovanni di Dio, Fatebenefratelli di Cernusco sul Naviglio Scuola di Specializzazione in Psicoterapia IRIS-Insegnamento e Ricerca Individuo e Sistemi, Milano Scuola di Terapia Cognitiva Studi Cognitivi, Milano Società Italiana Medicina Interna, Roma Unione Nazionale delle Associazioni per la Salute Mentale (UNASAM), Milano Unità di Geriatria, Ospedale Maggiore IRCCS, Università di Milano Unità Operativa di Psichiatria, Azienda Ospedaliera Luigi Sacco di Milano, Milano Unità Operativa di Psichiatria, Azienda Ospedaliera San Gerardo di Monza, Monza Unità Operativa di Psichiatria di Garbagnate, Azienda Ospedaliere Salvini di Garbagnate, Garbagnate Milanese Unità Operativa di Psichiatria, Fondazione IRCCS Ospedale Maggiore Policlinico, Mangiagalli e Regina Elena di Milano, Milano Università degli Studi di Foggia Università Cattolica del Sacro Cuore di Roma Università del Piemonte Orientale, Novara Università di Milano, IRCCS Ospedale Maggiore, Milano Università Milano-Bicocca, Monza Università La Sapienza, Roma U.O. Neurologia, Clinica S. Maria, IRCCS, Castellanza (VA). 82

85 INTERNATIONAL COLLABORATIONS Albert Eistein College of Medicine, Bronx, NY, USA Atomic Energy Commission, Service de Neurovirologie, Fontenay-aux-Roses, France Beaumont Hospital, Dublin, Ireland Cambridge Centre for Brain Repair, University of Cambridge, UK Centre for Neuroscience Research and Division of Biomolecular Sciences, GKT School, King s College, London, UK Chorley & South Ribble General Hospital, Chorley, Columbia Univ, Haverstraw, NY, USA Department of Anatomy and Physiology, Laval University, Quebec Department of Cell Biology, Washington University, St Louis, USA Department of Chemistry,The Australian National University, Canberra City, Australia Department of Experimental Psychology, University of Cambridge, UK Department of Pathology and Infectious Diseases Royal Veterinary College, Herts, UK Department of Psychiatry, Medical Center University of Mississippi, Jackson, USA Directorate General for the Health and Consumer Protection, European Commission, Luxembourg Division of Medical Genetics, CHUV Lausanne, Switzerland European Union of Family Associations of People with Mental Illness (EUFAMI) Geriatric Division and Department of Metabolic Diseases, Ospedali Regionali of Lugano and Mendrisio, Switzerland HSPH Harvard University, Boston, USA IBCM, University of Lausanne, Lausanne, Switzerland Institut de Génétique Humaine du CNRS, Montpellier, France Jefferson Med Coll, Philadelphia, USA Karolinska Institutet, Stockholm, Sweden King s College Hospital, London, UK Max-Delbrück-Center for Molecular Medicine, Berlin, Germany National Insitute on Aging, NIH, Baltimore, USA Neuroprion, Network of Excellence, WP VI, EC Neurological Department of the University of Tirana, Albania Ninewells Hospital and Medical School, Dundee, Scotland UK Northern Illinois University, DeKalb, IL, USA Novartis Pharma, Basel, Switzerland Robarts Research Institute, London, Ontario, Canada Royal Manchester Children's Hospital, Manchester, UK Royal Preston Hospital, Preston, UK Sergievsky Center, Columbia University, New York, NY, USA Servizio di Geriatria, Ospedale della Beata Vergine, Mendrisio, Switzerland The Scripps Research Institute, Jupiter, Florida, USA University of Alberta, Canada Univ of California at Irvine, Irvine, CA, USA University of Cardiff, United Kingdom Univ of Colorado, Denver, USA University Hospital, London, ON, Canada Univ of Innsbruck, Innsbruck, Austria Univ of Maryland, Baltimore, USA 83

86 University of Maastricht, the Netherlands University of Rijeka Medical School, Rijeka, Croatia University of Szeged, Ungary Université Victor Segalen, Bordeaux, France Virtanen Institute for Molecular Sciences, University of Kuopio, Finland Vrije Universiteit Medical Center, Amsterdam, The Netherlands Walton Hospital, Liverpool, UK WAPR (World Association for Psychosocial Rehabilitation) Weill Cornell Medical College, New York, USA World Mental Health, Department of Mental Health and Substance Abuse, Geneva, Switzerland EDITORIAL BOARD MEMBERSHIP Biochemical Journal (Chiesa, Forloni) Brain Aging (Forloni) Clinical Drug Investigation (Beghi) Clinical Neurology and Neurosurgery (Beghi) Cochrane Collaboration, Epilessia (Beghi) Dialogo sui Farmaci (Nobili) Drugs in the R&D (Beghi) Epidemiologia e Prevenzione (Lucca) Epilepsia (Beghi, Vezzani Assistant editor) Epilepsy Current (Vezzani) Epilepsy Research (Vezzani) Inpharma (Beghi) International Journal of Mental Health (Barbato) Journal of Neurochemistry (Bendotti) Neurological Sciences (Beghi) Neuroepidemiology (Beghi) Neuroscience (Vezzani) Open Aging Journal (Forloni) Open Geriatric Medicine Journal (Forloni) Psichiatria di Comunità (Barbato) Quality of Life Research (Barbato) Ricerca & Pratica (Nobili) Stroke (De Simoni, Associate editor) 84

87 PEER REVIEW ACTIVITIES Acta Neurologica Scandinavica Acta Psychiatrica Scandinava Alzheimer Disease and Associated Disorders American Journal of Clinical Nutrition American Journal of Human Genetics American Journal of Pathology American Journal of Physiology Annals of Neurology Annals of Pharmacotherapy Behavioural Brain Research Biochimica et Biophysica Acta Biochemical Journal Biochemistry BioMed Central Neurology Biological Psychiatry Brain Research Brain Research Bulletin Brain Research Review Clinical Drug Investigation Clinical Neurology and Neurosurgery Clinical Pharmacokinetics Clin Pharm Therapy CNS Drugs Dialogo sui farmaci Drugs Epidemiologia e Psichiatria Sociale Epilepsia Epilepsy & Behavior European Journal of Immunology European Journal of Neuroscience European Journal of Pharmacology European Journal of Public Health Experimental Neurology European Neuropsychopharmacology Expert Opinion on Pharmacotherapy FASEB Journal FEBS letters Fundamental Clinical Psychopharmacology Future Drugs Giornale di Neuropsichiatria dell Età Evolutiva Glia International Journal of Neuropsychopharmacology Journal of the American Board of Family Practice Journal of Biological Chemistry Journal of Cell. Biology Journal of Cerebral Blood Flow and Metabolism Journal of Chemical Neuroanatomy Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Science 85

88 Journal of Headache and Pain Journal of Histochemistry and Cytochemistry Journal of Immunology Journal of Neurochemistry Journal of Neuroimmunology Journal of Neurology, Neurosurgery and Psychiatry Journal of Neuroscience Journal of Pharmacy and Pharmacology Journal of Psychopharmacology Journal of Psychosomatic Research Journal of Structural Biology Life Sciences Lancet Lancet Neurology Molecular Brain Research Molecular and Cellular Neuroscience Molecular Therapy Neuroepidemiology Neurology Neurological Sciences Nerobiology of Aging Neurobiology of Diseases Neuropharmacology Neuropsychopharmacology Neuroscience Neuroscience Letters N.S. Archives Pharmacology Parkinsonism & Related Disorders Pharmacological Research Pharmacoepidemiology and Drug Safety Pharmacology Biochemistry & Behavior PlosOne Proc Natl Acad Sci, USA Psychopharmacology Synapse Trends Molecular Medicine 86

89 NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Advisory Board of the Italian League Against Epilepsy Board of "Master in Advanced Technologies for the Study of Neurodegenerative Diseases", University of Milan Commission on Health Care Policy of the International League Against Epilepsy (ILAE) Commission of Italian Minister of Health for the study of the problems associated to diagnosis, therapy and assistance ALS patients Coordination Group of NoE Neuroprion EC Coordination Group of the European project Quelles professionnalités en santé mentale. Perspectives croisées, usagers, élus professionnels. Council of Italian Association on Brain Aging Research (AIRIC) Expert for European Agency for the Evaluation of Medicines (EMEA) Expert reviewer for the Medical Research Council (MRC), UK Expert of the Minister of Health to EMEA Italian Association of Neuroepidemiology (Past President) Italian Association on Brain Aging Research (AIRIC, President) Italian Society of Neuroscience (Council) International Committee on Epilepsy and the Law International Organizing Committee e coordinator of the Secretariat al Global Forum for Community Mental Health, Department of Mental Health of WHO International Subcommittee of the American Academy of Neurology Medical Research Council Strategic Grant Application Mental Health Working Party of DG-SANCO, Directorate General Public Health and Consumer Protection of the European Union, Brussels, Belgium National expert accredited by AIFA (Italian Medicines Agency) for the European Agency for the Evaluation of Medicinal Products (EMEA) Neurobiology Commission of the International League against Epilepsy Neuroepidemiology Section of the American Academy of Neurology (past Chair) Research Advisory Panel, MND Association, UK Scientific Advisory Board of Sheffield Institute Foundation for MND 87

90 EVENT ORGANIZATION Epidemiology and research methods course. Medicina basata sull evidenza in neuropsichiatria dello sviluppo: la diagnosi come strumento di codifica per una banca-dati. Istituto Scientifico Stella Maris, Calabrone (Pisa) June 7-9, th Annual Meeting of American Academy of Neurology Breakfast Seminar How to manage a patient with a first epileptic seizure: An evidence-based approach April 28 May 5, 2007, Boston, MA, USA. International meeting on Mutant SOD1 and familial ALS: from the molecule to man. Milan, September 13-16, 2007 Course for physicians: The traps of literature: looking for, selecting and reading an article. May 23 rd, Villa Camozzi, Ranica (BG) Course for physicians: What one knows when a new drug is approved. September, 27 th ASL Bergamo. Information week: Who is afraid of generic (bioequivalent) drugs? November 17 th -23 rd ASL Bergamo. 5 a Giornata di studio sulla malattia di Alzheimer I disturbi del comportamento nelle demenze. La gestione dei disturbi del comportamento nelle demenze, March 24, 2007, Ateneo Veneto, Venezia 3 Corso di formazione e aggiornamento per operatori socio-sanitari: La malattia di Alzheimer e le altre demenz e (10 days) October 1 st November 5 th, 2007, IRE Venezia, Venezia Rehabilitation, empowerment, recovery November 30 December 1 st, 2007, Istituto di Ricerche Farmacologiche Mario Negri, Milano 4th Conference on Epileptogenesis, Pisa May th International Epilepsy Congress, Singapore, July 8-12,

91 GRANTS AND CONTRACTS Abbott GmbH & Co. KG Amgen, Milano ASL 2 Piemonte. Assessorato alla Salute, Comune di Milano Association pour la recherché sur la SLA, France Bristol-Myers Squibb Boehringer Ingelheim CURE Epilepsy Dipartimento di Salute Mentale, Azienda Ospedaliera Niguarda Ca Granda, Milano Dana Foundation Evidentia Medica, Grottaferrata (Roma) Fondazione Cariplo, Milano Fondazione Mariani, Milano Fondazione Italo Monzino, Milano FP6, European Union Glaxo-SmithKline, Italy Hospice "Via di Natale Franco Gallini", Aviano (PN) Human Frontiers Scientific Programme IMPHA II, DG-SANCO, Public Health and Consumers' Protection (Directorate General Istituto Comprensivo Statale "G.D. Romagnosi", Carate Brianza (MI) I.R.I.S Istituto Superiore di Sanità Janssen-Cilag H. Lundbeck A/S, Denmark Ministero della Ricerca Scientifica Ministero della Salute MND Association, UK Newron Ospedale Casa Sollievo di San Giovanni Rotondo Pharming Regione Lombardia, Assessorato alla Famiglia e Solidarietà Sociale e Assessorato alla Sanità, Milano Rimoldi e Bergamini Sanofi-Aventis SELECTA MEDICA, Pavia Servier Laboratories, Parigi Sigma-Tau Telethon Unione Nazionale Associazioni per la Salute Mentale UNASAM Vertex 89

92 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Albani D., Batelli, S., Pesaresi, M. Prato, F., Polito L. Forloni G. Pantieri R. A novel PSENEN mutation in a MCI patient with positive family history for Alzheimer s Disease. Alzheimer & Dementia 3: (2007) Albani, D., Artuso V. Roiter, I. Batelli, S., Prato, F. Pesaresi, M. Galimberti, D. Scarpini, E., Bruni, A., Franceschi, M. Piras MR., Confaloni, A. and Forloni G. Presenilin-1 mutation E318G represents a risk factor for familial Alzheimer s disease in the Italian population and affects A(1-40) production. Neurobiol Aging, 28: (2007) Barbato A, Agnetti G, D'Avanzo B, Frova M, Guerrini A, Tettamanti M. Outcome of a community-based rehabilitation program for people with mental illness who are considered difficult to treat. J Rehabil Res Devel 44: (2007) Bartfai T, Sanchez-Alavez M, Andell-Jonsson S, Schultzberg M, Vezzani A, Danielsson E, Conti B. Interleukin-1 system in CNS stress: seizures, fever, and neurotrauma. Ann N Y Acad Sci. 1113:173-7 (2007) Baviera M, Invernizzi RW, Carli M. Haloperidol and clozapine have dissociable effects in a model of attentional performance deficits induced by blockade of NMDA receptors in the mpfc. Psychopharmacology (Berl) published online. (2007) Beghi. E. Epilepsy. Current Opinion in Neurology 20: (2007) Beghi, E., A. Millul, A. Micheli, E Vitelli, G. Logroscino, and SLALOM Group. Incidence of ALS in Lombardy, Italy. Neurology 68: (2007) Beghi, E., T. Mennini, C.Bendotti, P. Bigini, G. Logroscino, A.Chiò, O. Hardiman, D. Mitchell, Swingler, B.J. Traynor, A. Al Chalaby. The heterogeneity of amyotrophic lateral sclerosis: a possible explanation of treatment failure. Curr Med Chem 14: (2007) Begley, CE., G.A. Baker, E. Beghi, J. Butler, D. Chisholm, J.T. Langfitt, P. Levy, C. Pachlatko, S. Wiebe, K.L. Donaldson on behalf of the ILAE Commission on Healthcare Policy. Cross-country measures for monitoring epilepsy care. Epilepsia 48: (2007) Biasini E, Medrano AZ, Thellung S, Chiesa R, Harris DA. Multiple biochemical similarities between infectious and non-infectious aggregates of a prion protein carrying an octapeptide insertion. J Neurochem [Epub ahead of print] Bigini P, Repici M, Cantarella G, Fumagalli E, Barbera S, Cagnotto A, De Luigi A, Tonelli R, Bernardini R, Borsello T, Mennini T. Recombinant human TNF-binding protein-1 (rhtbp-1) treatment delays both symptoms progression and motor neuron loss in the wobbler mouse. Neurobiol Dis Nov 12; [Epub ahead of print] Borsello, T. Forloni, G. JNK signaling: a target to prevent neurodegeneration. Current Pharmaceutical Design. 13: (2007) Borsello T, Centeno C, Riederer IM, Haefliger JA, Riederer BM. Phosphorylation-dependent dimerization and subcellular localization of islet-brain 1/c-Jun N-terminal kinaseinteracting protein 1. J Neurosci Res. 85: (2007) Burattini C, Burbassi S, Aicardi G, Cervo L.Effects of naltrexone on cocaine- and sucrose-seeking behaviour in response to associated stimuli in rats. Int J Neuropsychopharmacol. Epub 2007 Mar 5 Burbassi S, Cervo L. Stimulation of serotonin(2c) receptors influences cocaine-seeking behavior in response to drug-associated stimuli in rats. Psychopharmacology (Berl). Epub 2007 Sep 27 Caccia, S. N-dealkylation of 1-arylpiperazine derivatives: disposition and metabolism of the formed 1-arylpiperazines. Curr. Drug Met., 8: (2007) Calcagno E, Canetta A, Guzzetti S, Cervo L, Invernizzi RW. Strain differences in basal and post-citalopram 90

93 extracellular 5-HT in the mouse medial prefrontal cortex and dorsal hippocampus: relation with tryptophan hydroxylase-2 activity. J Neurochem. 103: (2007) Capone C, Fabrizi C, Piovesan P, Principato M C, Marzorati P, Ghirardi O, Fumagalli L, Carminati P, De Simoni M G. 2-Aminotetraline derivative protects from ischemia/reperfusion brain injury with a broad therapeutic window. Neuropsychopharmacology 2007 ; 32 : Capone C, Frigerio S, Fumagalli S, Gelati Maurizio, Principato M C, Storini C, Montinaro M, Kraftsik R, De Curtis M, Parati E, De Simoni M G. Neurosphere-derived cells exert a neuroprotective action by changing the ischemic microenvironment. PLoS One 2007; 2(4):e373 Cavestro, C., A. Rosatello, G.M. Micca, M. Ravotto, M.P. Marino, G. Asteggiano. E. Beghi. Insulin metabolism s altered in migraineurs: a new pathogenic mechanism for migraine? Headache 47: (2007) Colombo, A., Repici M., Pesaresi M. Santambrogio S. Forloni G. and Borsello T. The Amyloid Precursor Protein processing is regulated by JNK phosporylation in the cytoplasmic domain: D-JNKI1 inhibits its proteolytic cleavage, Cell Death & Diff. 14: (2007) Cornaggia, CM., Beghi, M. Beghi E. for the REST-1 Group. Psychiatric events in epilepsy. Seizure 16: (2007) De Girolamo G, Barbato A, Bracco R, Gaddini A, Miglio R, Morosini P, Norcio B, Picardi A, Rossi E, Rucci P, Santone G, Dell Acqua G. The Characteristics and Activities of Acute Psychiatric Inpatient Facilities: National Survey in Italy. British J. Psych 191: (2007) Del Bo, R., Ghezzi, S., Scarlato, M., Albani, D. Galimberti, D., Lucca, U., Tettamanti, M., Scarpini, E., Forloni, G., Bresolin, N, Comi, GP. Role of VEGF gene variability in longevity: A lesson from the Italian population Neurobiol Aging, 2007 June 15 [Epub ahead of print] Fenoglio C, Galimberti D, Piccio L, Scalabrini D, Panina P, Buonsanti C, Venturelli E, Lovati C, Forloni G, Mariani C, Bresolin N, Scarpini E. Absence of TREM2 polymorphisms in patients with Alzheimer's disease and Frontotemporal Lobar Degeneration. Neurosci Lett. 411:133-7 (2007) Fioriti, L. Angeretti, N.. Colombo, L., De Luigi A., Manzoni, C., Colombo A., Morbin, M., Tagliavini, F., Salmona, M. Chiesa, R. Forloni, G. Neurotoxic and gliotrophic activity of a synthetic peptide homologous to Gerstmann- Sträussler-Scheinker disease amyloid protein. J. Neurosci. 27: (2007) Fossati, R., G. Apolone, E. Negri, A. Compagnoni, C. La Vecchia, S. Mangano, L. Clivio, S. Garattini, for the General Practice Tabacco Cessation Investigators Group. A double-blind, placebo-controlled, randomized triail of Bupropion for smoking cessation in primary care. Arch Intern Medicine 167: (2007). Furlan R, Bergami A, Brambilla E, Butti E, De Simoni M G, Campagnoli M, Marconi P, Comi G, Martino G. HSV- 1-mediated IL-1 receptor antagonist gene therapy ameliorates MOG35-55 induced experimental autoimmune encephalomyelitis in C57BL/6 mice. Gene Ther 14 : (2007) Gallucci M., Amici, GP. Ongaro F. Gajo G.B., De Angeli, S. Forloni G., Albani, D. Prato F. Polito, L. Zanardo A and Regini C. Associations of the plasma interleukin 6 (IL-6) levels with disability and mortality in the elderly in the treviso longeva (trelong) study. Arch Geriatr. Geront. 44 Suppl: (2007) Gallucci M., Ongaro F. Bresolin F. Bernardi, U. Salvato, C. Minello A., Amici, GP., Barasciutti, E., Mazzucco, S., Gajo G.B, De Angeli, S. Forloni G., Albani, L. Zanardo A and Regini C. The treviso longeva (trelong) study: a biomedical, demographic, economic and social investigation on people 70 years and over in a typical town of northeast of Italy. Arch Geriat Geront 44 Suppl: (2007) Grignaschi G, Zennaro E, Tortarolo M, Calvaresi N, Bendotti C. Erythropoietin does not preserve motor neurons in a mouse model of familial ALS. Amyotroph Lateral Scler. 8:31-5, (2007). Invernizzi R Role of TPH-2 in brain function: news from behavioral and pharmacologic studies J Neurosci Res :

94 Invernizzi R, Pierucci M, Calcagno E, Di Giovanni G, Di Matteo V, Benigno A, Esposito E Selective activation of 5HT2C receptors stimulates GABA-ergic function in the rat substantia nigra pars reticulata: a combined in vivo electrophysiological and neurochemical study Neuroscience 144: (2007) Li A, Christensen HM, Stewart LR, Roth KA, Chiesa R, Harris DA. Neonatal lethality in transgenic mice expressing prion protein with a deletion of residues EMBO J. 26: (2007) Lo Coco D, Veglianese P, Allievi E, Bendotti C. Distribution and cellular localization of high mobility group box protein 1 (HMGB1) in the spinal cord of a transgenic mouse model of ALS. Neurosci Lett. 412(1):73-7 (2007) Logroscino, G.,E. Beghi, O. Hardiman, A. Chiò, J.D. Mitchell, R.J. Swingler, B. Traynor, for EURALS. Effects of referral bias on assessing survival in ALS. Neurology 69: 939 (2007) Ludolph AC, Bendotti C, Blaugrund E, Hengerer B, Loffler JP, Martin J, Meininger V, Meyer T, Moussaoui S, Robberecht W, Scott S, Silani V, Van Den Berg LH; ENMC Group for the Establishment of Guidelines for the Conduct of Preclinical and Proof of Concept Studies in ALS/MND Models. Guidelines for the preclinical in vivo evaluation of pharmacological active drugs for ALS/MND: report on the 142nd ENMC international workshop. Amyotroph Lateral Scler. 8: (2007) Massignan T, Casoni F, Basso M, Stefanazzi P, Biasini E, Tortarolo M, Salmona M, Gianazza E, Bendotti C, Bonetto V. Proteomic analysis of spinal cord of presymptomatic amyotrophic lateral sclerosis G93A SOD1 mouse. Biochem Biophys Res Commun. 353: (2007) Noe' F, Nissinen J, Pitkänen A, Gobbi M, Sperk G, During M, Vezzani A. Gene therapy in epilepsy: the focus on NPY. Peptides. 28: (2007) Parabiaghi A, Lasalvia A, Bonetto C, Cristofalo D, Salvi G, Ruggeri M. Predictors of caregiving burden in relatives of people with schizophrenia. A 3-year follow-up in a community mental health service. Acta Psych Scand Suppl 437: (2007) Pesaresi M, Batelli S, Prato F, Polito L, Lovati C, Scarpini E, Quadri P, Mariani C, Albani D, Forloni G. The urokinase-type plasminogen activator polymorphism PLAU_1 is a risk factor for APOE-epsilon4 non-carriers in the Italian Alzheimer's disease population and does not affect the plasma Abeta(1-42) level. Neurobiol Dis. 25: (2007) Peviani M, Cheroni C, Troglio F, Quarto M, Pelicci G, Bendotti C. Lack of changes in the PI3K/AKT survival pathway in the spinal cord motor neurons of a mouse model of familial amyotrophic lateral sclerosis. Mol Cell Neurosci. 34: (2007) Renoldi G, Calcagno E, Borsini F, Invernizzi R Stimulation of group I mglu receptors in the ventrotegmental area enhances extracellular dopamine in the rat medial prefrontal cortex J Neurochem 100 : (2007) Repici M, Centeno C, Tomasi S, Forloni G, Bonny C, Vercelli A, Borsello T. Time-course of c-jun N-terminal kinase activation after cerebral ischemia and effect of D-JNKI1 on c-jun and caspase-3 activation. Neuroscience.150: 40-9 (2007) Ruggeri M, Salvi G, Bonetto C, Lasalvia A, Allevi L, Parabiaghi A, Bertani M, Tansella M. Outcome of patients dropping out from community-based mental health care: a 6-year multiwave follow-up study. Acta Psych Scand Suppl 437:42-52 (2007). Savica, R., E. Beghi, G. Mmazzaglia, F. Innocenti, O. Brignoli, C. Cricelli, A.P. Caputi, R. Musolino, E. Spina, G. Trifirò. Prescribing patterns of antiepileptic drugs in Italy: a natiowide population-based study in the years Eur J Neurology 14: (2007) Sau D, De Biasi S, Vitellaro-Zuccarello L, Riso P, Guarnieri S, Porrini M, Simeoni S, Crippa V, Onesto E, Palazzolo I, Rusmini P, Bolzoni E, Bendotti C, Poletti A. Mutation of SOD1 in ALS: a gain of a loss of function. Hum Mol Genet. 16: (2007) Vezzani A. On Demand Up-regulation of Therapeutic Genes in the Brain: Fiction or Reality? Epilepsy Curr. ;7: (2007) Vezzani A, Baram TZ. New roles for interleukin-1 Beta in the mechanisms of epilepsy. 92

95 Epilepsy Curr.7:45-50 (2007) Zoccolella, S., E. Beghi, G. Palagano, A. Fraddosio, V. Guerra, V. Samarelli, V. Lepore, I.L. Simone, P. Lamberti, L.Serlenga and G. Logroscino for the SLAP registry. Riluzole and amyotrophic lateral sclerosis survival: a population-based study in southern Italy. Eur J Neurol. 14: (2007) Zoccolella, S., E. Beghi, G. Palagano, A. Fraddosio, V. Guerra, V. Lepore, I.L. Simone, P. Lamberti, L. Serlenga, G. Logroscino. ALS multidisciplinary clinic and survival: results from a population-based study in Southern Italy. J Neurol 254: (2007).. LAY PRESS SELECTION PUBLISHED IN 2007 Albani D, Polito L, Vittori A, Forloni G Fattori di rischio genetici per le demenze e screening per la loro indentificazione pre-clinica Ricerca & Pratica 2007 ; n.137 : Barbato A, D Avanzo B, Ferrannini L, Parabiaghi A, Vaggi M. Un occasione per la ricerca clinica in Italia. Lo studio GISAS su aripipr, olanz e aloper nel trattamento dei disturbi schizofrenici, Psichiatria di Comunità, in press. Cerzani M, Pasina L, Clavenna A, Nobili A, Garattini L Revisione critica degli studi italiani di farmacoeconomia sull'uso dei farmaci antinfiammatori non steroidei in medicina generale Quaderni Farmacoeconomia 2007 ; 3 : D Avanzo B. L empowerment come orizzonte di lavoro con il disagio psichico. Animazione Sociale 2007; 10: Dell Acqua G, Norcio B, de Girolamo G, Barbato A, Bracco R, Gaddini A, Miglio R, Morosini P, Picardi A, Rossi E, Rucci P, Santone G. Caratteristiche e attività delle strutture di ricovero per pazienti psichiatrici acuti: i risultati dell indagine nazionale Progres-Acuti, Giornale Italiano di Psicopatologia 2007; 13: Clavenna A, Bonati M, Campi R, Labate L, Nobili A, Pasina L, Tettamanti M, Monesi L, Marzona I, Roncaglioni M C, Bortolotti A, Fortino I, Locateli G W, Giuliani G La prescrizione di farmaci per i bambini e gli anziani nella ASL di Lecco Ricerca & Pratica 2007 ; n.135 : La Vecchia C, Beghi E Epidemiologia delle demenze La demenza in Italia. Aggiornamento e casi clinici interattivi UTET, Torino, 2007; Nobili A, Garattini S La Caffeina Aggiornamento Medico 2007 ; 31 : Nobili A, Pasina L, Garattini S Il paziente allergico Aggiornamento Medico 2007 ; 31 : Nobili A, Pasina L, Garattini S. Il paziente con emicrania. Aggiornamento Medico 2007; 31: Nobili A, Pasina L, Garattini S Il paziente con dolore cronico: gli analgesici oppioidi Aggiornamento Medico 2007 ; 31 : Nobili A, Pasina L, Garattini S. I fluorochinoloni. Aggiornamento Medico 2007; 31: Nobili A, Piana I, Balossi L, Tettamanti M, Trevisan S, Lucca U, Tarantola M Il malato di demenza in RSA: risultati di una valutazione comparativa tra i pazienti degenti nei Nuclei Alzheimer e in RSA senza Nuclei Alzheimer In-formazione Alzheimer. Alla ricerca di nuove connessioni nella rete lombarda dei servizi alle demenze Franco Angeli, Milano, 2007; Sterzi R, Toso V, Guidetti D, Provinciali L, Consoli D, Leone M A, Beghi E Indagine epidemiologica italiana su "La neurologia nell'emergenza-urgenza": il Progetto NEU Neurol Sci 2007 ; 28 : S1-S36 93

96 RESEARCH ACTIVITIES Laboratory of Biology of Neurodegenerative Disorders Alzheimer's disease: genetic studies and clinical investigations In collaboration with different neurological centers and the laboratory of Geriatric Neuropsychiatry it has been created a bank of blood samples for DNA of patients with Alzheimer s disease (AD), in familial (FAD) or sporadic form (SAD), and patients with vascular dementia (VD). In all subjects the diagnosis of dementia is performed according to the international guidelines. Since 2005 we started also the collection of blood samples from subjects with fronto-temporal dementia. The genetic studies are aimed to the identification of causal factors in FAD and risk factors in SAD. Mutations on genes encoding proteins involved in the physiopathology of AD were investigated. The pathogenic role of these mutations is under investigation using fibroblasts obtained from skin biopsy. Furthermore, we continued the screening of FAD samples for the genes encoding for presenilin 1 and 2 (PS-1 and PS-2) and APP, missense mutations in these three genes were associated with AD. Based on the evidence obtained in experimental studies, in collaboration with the Geriatric Neuropsychiatry lab and the Inflammation and CNS diseases lab, we elaborated a protocol for a clinical trial to investigate the effects low molecular weight heparin on the CSF and plasma β amyloid levels in Alzheimer patients Alzheimer's disease: preclinical studies The formation of β amyloid (Aβ) deposits in brain parenchyma and on the wall of cerebral blood vessels is an early event in AD and there are now numerous genetic, biochemical and neuropathological studies pointing to a causal role of Aβ in the pathogenesis of AD. Thus, prevention the formation of Aβ aggregates or their elimination once formed is a potential therapeutic approach to the disease. This aim is strongly persecuted with different strategies including the regulation of enzymes responsible of the synthesis and degradation of Aβ and the enzymes influencing the metabolism of amyloid precursor protein (APP). In the lab, we developed the idea to interfere directly with the Aβ deposits formation using anti-amyloidogenic drugs. The experimental studies have shown the potential therapeutic activity of these drugs in AD, and now they will be tested in a clinical setting. In collaboration with the Department of Biochemistry and Molecular Pharmacology, we tested new molecules that can bind the amyloid aggregates to identify either anti-dementia drugs or potential diagnostic markers. Furthermore, using in vitro and in vivo models, we are evaluating new approaches to reduce the beta amyloid production either by affecting directly on beta secretase or through the modulation of JNK pathway, Furthermore, the modulation of this pathway has been considered for the potential neuroprotective activity in AD and other neurodegenerative disorders. Genetics of aging In collaboration with Geriatric Neuropsychiatry Lab for the Monzino 80-plus study and with dr. Maurizio Gallucci from the ARGel Association in Treviso for Trelong study we collected a large number of blood samples from subjects over seventy. In these samples we are performing a genetic analysis to identify genetic profiles associate to the longevity and /or to the agingassociated pathologies with specific attention to the dementias. The aim is to cross the genotype/phenotype profile with pathologies and environmental aspects including style of life, diet and economical conditions to identify risks and protective factors. Initially the subjects were genotypized for ApoE, whom allele E4 is a well-known risk factor for Alzheimer s disease and several other disorders and sirt-1 a gene codified for protein member of a enzymatic family 94

97 of sirtuins associated to the longevity in several experimental models. The results are interesting but before drawing any conclusion we need to consider the numerous other parameters collected in our database. Prion's disease: in vitro studies Prion s diseases (TSE) are neurodegenerative disorders of sporadic inherited origin but also transmissible, they have different clinic and neuropathological features but all are characterized by the cerebral accumulation of an altered form of prion protein (PrPsc). TSE are rare diseases but the transmission from bovine (BSE) to humans induced a public health alarm in UK and successively in all Europe. PrPsc is involved in the pathogenesis of the disease and it is also an essential component of the infective agent. In the lab numerous projects were developed to understand the association between the presence of PrPsc and the neurodegenerative process. The biological effects of peptides homologous to large fragment of PrP were investigated. In particular PrP , synthesized in the Protein Chemistry and Biochemistry lab, homologous to the fragment found in the cerebral deposits in TSE patients, is neurotoxic and spontaneously structured in beta sheet conformation. The pathogenetic role of PrP was investigated not only through the external application of peptides but also by the evaluation of intracellular mechanisms potentially involved in the formation of PrP aggregates. Prion s diseases: studies in vivo The lab has the facilities to study the experimental scrapie, mice and hamsters are inoculated with infected brain homogenate. The hamsters after intracerebral inoculation develop the disease in days and died within a month. The histopathological analysis of the brain show the presence of PrPsc deposits, neuronal damage, diffuse astrogliosis and the typical spongiosis at cortical and thalamic level. The anti-amyloidogenic activity of tetracyclines has been investigated also in this experimental contest. After the ex-vivo approach, where the homogenate was treated before the inoculation, the curative effect of tetracyclines was tested in collaboration with the lab of Chemistry and Biochemistry of Proteins by treatment the experimental scrapie in hamsters with intramuscular doxycycline, the treatment prolonged the survival of the animals. Other drugs are now under investigation to verify the curative effects in TSE. Other animal models of TSE are available in the lab, transgenic mouse developed by dr. Chiesa. These mice express the mutated forms of PrP associated to the familial TSE. The homozygotes exhibit at different level of severity the symptoms reminiscent of the pathologies associated with the mutations. From the neuropathological point of view some lines exhibit an accumulation of PrP and clear neurodegeneration of the cerebellar granular cells, in the other brain regions no evident alterations were found. In the brain tissue of the mice was found a PrP with some of the characteristic of PrPsc, however the brain material is not infected. The pathogenesis of TSE is investigated in these models through different approaches including the proteomic approach and the electronic microscopy. Parkinson s Disease: genetic studies Parkinson s disease (PD) is the second more diffuse neurodegenerative disorder with an unknown pathogenesis, however for PD several therapies are available and, although at the symptomatic level, their efficacies is well-established. In the etiological studies on PD the genetic component has been traditionally considered with scarce interest whereas the environmental causes were carefully evaluated. This orientation was based on the evidence that the exposure to several toxins can mimic the PD pathology. However the genetic studies in the last few years have completely changed the perspective with the identification of mutations on two genes, encoding for alpha-synuclein and parkin, associated to the juvenile forms of the disease. A mutation on alpha synuclein gene is an event extremely rare, only three mutations 95

98 identified until now, the parkin mutations are numerous ether in puntiform or in deletion form. The mutations on alpha-synuclein gene are dominant while the parkin mutations are associated with PD in recessive form. We collected, in collaboration with several neurological centers, blood samples from PD subjects and the screening of the samples involved genes like alphasynuclein, parkin, DJ-1 and other factors potentially involved in PD. Parkinson s disease: studies in vitro The identification of the mutations associated to Parkinson s disease (PD) gave a substantial contribute to understand the disease and allowed the develop of cellular models to investigate the pathogenesis of the disease. In past we showed the potential neurotoxic activity of alphasinuclein using the synthetic peptide homologous to the fibrillogenic fragment (NAC) of the protein. Successively with help of dr. Negro at the Department of Biochemistry at the University of Padova we prepared cdna vectors including the sequence of wild type and mutated alpha-synuclein Their transfection to the PC12 cells induced in specific conditions a cellular damage. More recently alpha-synuclein was associated to a TAT sequence capable to transport inside the cells the protein. With this method the intracellular concentration of alphasinuclein was better controlled. In a micromolar range alpha-synuclein was toxic, but in nanomolar range, it exerted neuroprotective effect against oxidative stress induced by hydrogen peroxide. This double effect dose-dependent was confirmed in an inducible model. More recently again in collaboration with Dr. Negro, we obtained the recombinant form of DJ-1 associated with TAT (TAT-DJ-1). This protein is similar to alpha-synuclein, mutations of its sequence has been associated to PD. TAT-DJ-1 silencing by small interference RNA (sirnai) were used to study the interaction between DJ-1 and alpha synuclein.. Laboratory of Neurological Disorders Epidemiological studies on amyotrophic lateral sclerosis (ALS) Included are studies on the incidence, risk factors and mortality of ALS. The data are obtained from a regional registry of the disease activated in 1998 and including all patients with newly diagnosed ALS identified in eight provinces of Lombardy. Using similar study protocols, the same data are collected in two additional regional registries (from Piemonte and Puglia) included in a network with the Lombard registry. Information obtained from patients enrolled in the Lombard registry and from cases examined by members of the Italian ALS Study Group has been used to assess the validity and reliability of diagnostic criteria for ALS and selected disability scales. Based on the data recorded, the annual incidence of ALS is comparable to that obtained in other Western countries where ALS registries have been activated, and is among the highest ever published (1.9 per 100,000). Mortality of ALS has been found to be comparable to that of studies from similar populations studied with the same protocol. The study on the validation of the current diagnostic criteria for ALS (the El Escorial criteria) showed that to be considered valid and reliable, the criteria should be used after proper training of the investigators. In October 2004, the Laboratory of Neurological Disorders has started a European collaborative group for the ALS registries (EURALS) with the intent to create a common database (completed in the year 2005) with the participation of the existing regional and national disease registries. Three major scientific activities are in course: 1. A comparative study of the clinical characteristics of patients with ALS enrolled in the registries as compared to those seen in secondary and tertiary health care facilities; 2. A meta-analysis of the incidence of ALS, performed by pooling data from the cohorts of patients enrolled in the populationbased registries; 3. A case-control study on trauma and risk of ALS; 4. A case-control study on ALS, physical exercise and sport. Study # 3 is ongoing only in Italy. In 2007 a scientific report 96

99 was completed (now in press) on the progression of ALS (marked by loss of ambulation, percutaneous gastrostomy and non-invasive assisted ventilation). Innovative therapeutic strategies in patients with epilepsy A cohort of patients with a first unprovoked seizure, randomised since 1988 by several Italian centers to immediate treatment or to treatment only at the time of a seizure relapse, was followed to verify the impact of the two therapeutic strategies on the long-term prognosis of epilepsy, measured by the chance of achieving 5-year remission. To provide a pragmatic definition of drug resistance in childhood epilepsy, children refractory to two antiepileptic drugs (in sequence or in combination) were randomised to the use of a third drug or to the optimization of the existing treatment and followed for up to three years. Therapeutic response was measured by the achievement of a six-month period of remission. The study has been conducted in collaboration with the IRCCS Stella Maris of Calambrone (PI). Epidemiology of neurological disorders in Albania With the collaboration of the Fondazione Mariani and the Neurological Department of the University of Tirana, an epidemiological survey has been started to assess the prevalence and incidence of several neurological conditions (stroke, epilepsy, headache, dementia, peripheral neuropathy, multiple sclerosis) comparing an urban and a rural community (Tirana and Saranda). A study on the validation of the diagnostic criteria is in course. Quality of life in children with neuromuscular disorders With a financial support of the Telethon organization, a study has been completed on the validation of a questionnaire on the quality of life in children and adolescents with different neuromuscular disorders. This is an Italian multicenter study coordinated to the Child Neurology Clinic of Pavia. Cerebrovascular disorders and risk of epilepsy Epilepsy is a frequent complication of stroke. Acute symptomatic seizures (i.e. seizures occurring in the seven days after stroke) can occur in up to two-thirds of cases and epilepsy (i.e. repeated unprovoked seizures) in 2-4%. There are no consistent findings on the risk factors for acute symptomatic seizures, unprovoked seizures and epilepsy in patients with stroke. For these reasons, in 2007 a multicenter national prospective survey has been started to assess the risk of seizures and epilepsy (and the main risk factors) in a cohort of patients with a first ischemic or hemorrhagic stroke followed for a maximum period of 24 months. The study was also implemented to assess the feasibility of a pragmatic therapeutic trial on the prophylaxis of seizures and epilepsy in stroke. Diagnosis and prevalence of dystonia As part of a collaboration with the institute San Raffaele of Milano, a study has been completed on the prevalence of adult focal dystonia in the province of Foggia. Cases are ascertained through different sources (hospital admissions, neurology, ENT, ophthalmology, and ortopedics outpatient visits) in the two major local hospitals. A total of 41 women (59%) and 28 men (41%) with a mean age of 58 years (range 23-87) were identified as having a primary focal or segmental dystonia. The crude prevalence rate was 12.6 per 100,000 (95% CI ) The age-adjusted prevalence rate was 13.5 (95% CI ). Blepharospasm was the commonest variety. The prevalence of the disease increases up to 27.8 at age years and declines thereafter. 97

100 Therapeutic trials in neurological disorders During the year 2007 three therapeutic trials sponsored by the Italian Drug Agency (AIFA) were started or continued. Included are: 1. A randomized double-blind parallel-group placebocontrolled trial on the efficacy and tolerability of L-acetylcarnitine in ALS; 2. A randomized open-label parallel-group trial comparing Erythropoietine to Metyl-prednisolone in patients with acute spinal cord injury; 3. A randomized double-blind parallel-group placebo-controlled trial on the efficacy and safety of valproate in medication-overuse headache. The first trial aims at finding a potentially effective drug in a clinical condition for which there is only one product (Riluzole) with at best modest efficacy on survival. L-acetylcarnitine has been found to improve survival in experimental models of motor neuron disease. The second trial intends to verify the efficacy of erythropoietin, a drug shown to mitigate the effects of traumatic spinal shock and accelerate recovery in experimental animals. The drug chosen for comparison (Methylprednisolone at high doses) has been selected for being the present gold standard in clinical practice. The third trial aims at verifying whether valproate (a drug commonly used for the prophylaxis of migraine) abates symptoms occurring in drug-overuse headache, a common and frequently invalidating variety of chronic idiopathic headache. All three trials are multicenter and nationwide. The laboratory of neurological disorders is the coordinator of the first trial and a partner in the other two trials, where the main tasks include protocol and CRF preparation, statistical analysis, and preparation of the final scientific report.. Laboratory of Drug Metabolism 1-Aryl-piperazine as active metabolites of centrally acting drug Dichloro(substituted) phenylpiperazine has been incorporated in the structure of drugs and pharmacologically active compounds which, in analogy with structurally related compounds, may undergo CYP3A4-mediated N-dealkylation (oxidative N-C cleavage) resulting in the formation of phenylpiperazine derivatives. However, there was no information on how these metabolites contribute to the pharmacological activity of their parent compound(s), which include psychotropic drugs and experimental compounds currently under pre-clinical evaluation. We therefore developed a reliable high-performance liquid chromatography-electrospray ionization mass spectrometric method for the detection of dichlorophenylpiperazine isomers and their parent compound(s) in body fluids and brain tissue. This enabled us to confirm that the metabolism of these phenylpiperazine derivatives includes N-dealkylation of the piperazinyl nitrogen to 2,3-, 2,4- or 2,5-dichlorophenyl-piperazine, although with differences in the rate and extent of the process. As phenylpiperazines concentrate in the brain, causing a variety of serotonin receptor-related pharmacological effects, we also examined their concentrations in brain of rodents given the various phenylpiperazine derivatives, and the metabolite-to-parent drug ratios at steady state. The metabolites had higher brain uptake than the parent compound(s), so the metabolite-to-parent drug ratio was higher in brain than in blood. For the recently introduced antipsychotic aripiprazole the concentrations of the metabolite 2,3-dichlorophenylpiperazine averaged only 0.08 in serum but about 0.4 in whole brain (on a molar basis) at approximately the time of the aripiprazole maximum concentration in the rat. Obviously, these measurements cannot be made in humans but assuming that the compounds concentrate in human brain to the same extent as in the rat, at the site of action 2,3- dichlorophenylpiperazine should account for only a small percentage of aripiprazole. It is therefore conceivable that this metabolite contributes to the central effects of aripiprazole in rats, but it should be of less clinical significance for human patients, although its pharmacological profile is still largely unknown and there may be physiological and pathological factors that raise the metabolite-to-parent drug ratio at the site of action. 98

101 Pharmacological role of the constituents of Hypericum perforatum extracts The chemical composition of extracts of hypericum perforatum L. (St. John s wort) is essentially known but is still not clear which constituent(s) account, wholly or in part, for the antidepressant activity of the extracts, and through what neurochemical mechanism(s). The phloroglucinol hyperforin shares most of the in vitro and in vivo pharmacological properties of the extracts, and is possibly a main antidepressant component, but there is also evidence for other pharmacologically active components. However, identifying the roles of the various derivatives and the mechanism(s) of their activity is complicated by the scarcity of information about their ability to cross the blood-brain barrier and the concentrations reached in brain after administration of the extracts. This is also true for the biflavone biapigenin and particularly its I3,II8 analog amentoflavone which, although present in smaller amounts in extracts, shows a multitude of pharmacological actions in vitro and in vivo in animal models. The lack of pharmacokinetic data in man and animals and questions about the brain uptake of amentoflavone and biapigenin prompted us to examine their brain uptake and concentrations and the relationships with plasma concentrations after pharmacologically effective doses in mice. After doses of Hypericum perforatum extracts the brain concentrations of biapigenin and amentoflavone were below the limit of quantification. The same was true for amentoflavone after a biflavone-enriched extract of Ginkgo biloba. Levels were consistently detected only after intraperitoneal biapigenin or amentoflavone but were low and mostly related to the residual biflavone in the circulation. Poor brain-to-blood permeability is common to other polar components of Hypericum perforatum, resulting in brain concentrations generally too low for any direct interaction with neurotransmitter transporters and receptors which are obviously important for the action of conventional antidepressants. Likewise, the in vitro interactions of biapigenin and amentoflavone with known central mechanisms are apparently not relevant for the in vivo effects of the extracts because they occur at biflavone concentrations far exceeding those found in the brain after pharmacologically effective doses. However, this does not exclude that tissues other than brain may concentrate biapigenin or amentoflavone sufficiently to exert beneficial effects after daily intake of the extracts. Resistence to antidepressant drugs: studies in animal models The selective serotonin reuptake inhibitors are the drugs of choice in the treatment of depression. However, they are not or only partially effective in a fraction of depressed patients. The reasons are substantially unknown, though pharmacogenetic studies have linked the response to serotonin reuptake inhibitors to polymorphisms in various genes coding for serotonin mechanisms, particularly the promoter of the serotonin transporter molecule. These studies are therefore aimed to investigate the neurobiological mechanism(s) of resistance to antidepressant drugs in strains of mice carrying different isoforms of tryptophan hydroxylase-2, the enzyme responsible for the synthesis of brain serotonin. These studies are conducted in collaboration with the laboratory of Experimental Psycopharmacology (L. Cervo) and the laboratory of Neurochemistry and Behaviour (R.W. Invernizzi), who will provide a brief description of recent results with the potent serotonin reuptake inhibitors citalopram and paroxetine. 99

102 Laboratory of Experimental Neurology Role of inflammatory molecules in ictogenesis and epileptogenesis We are studying the role of IL-1beta and TNF-alpha systems in the genesis and propagation of seizures and in the associated neurodegenerative phenomena. We have demonstrated that epileptic activity induces the synthesis of these cytokines and related molecules involved in inflammatory processes. IL-1beta has proconvulsant actions while its naturally occurring antagonist (IL-1Ra) and TNF-alpha have anticonvulsant activities. We are now evaluating the role of these molecules in seizure-associated neuronal damage in mature brain and during postnatal development. We are studying pharmacological approaches to block IL-1betasignaling involved in the proconvulsant effects of this cytokine. Research activity 2: Mechanisms of drug resistance in epilepsy: This study is aimed at investigating whether multidrug resistance in epilepsy is dependent on the activity of membrane bound glycoproteins able the extrude antiepiletic drugs from the brain tissue back into the blood stream. These proteins (in particular those produced by the MDR or MPR genes) appear to have a significant role in determining the resistance to some anticancer agents. Our studies indicate that the MDR gene product, P-glycoprotein or P-gp, is increased in specific brain regions during epileptic activity. The increase in protein determines a decreased brain concentration of phenytoin and transgenic mice lacking this protein accumulate higher concentration of this antiepileptic drug in their brain. Since an increased production of this protein occurs in the brain of human epileptics, it is possible that it contributes to drug refractoriness by impairing the attainment of clinical relevant concentrations of these drugs in the epileptic tissue. We recently found that the inhibition of the activity of P-gp in experimental models of epilepsy using specific blockers of its function, enhances the efficacy of anticonvulsant treatments. We have also demonstrated a role P-gp in brain drug disposition in experimental models of focal cortical dysplasia, a clinical condition associated with drug-refractory seizures. Mechanisms of drug resistance in epilepsy This study is aimed at investigating whether multidrug resistance in epilepsy is dependent on the activity of membrane bound glycoproteins able the extrude antiepiletic drugs from the brain tissue back into the blood stream. These proteins (in particular those produced by the MDR or MPR genes) appear to have a significant role in determining the resistance to some anticancer agents. Our studies indicate that the MDR gene product, P-glycoprotein or P-gp, is increased in specific brain regions during epileptic activity. The increase in protein determines a decreased brain concentration of phenytoin and transgenic mice lacking this protein accumulate higher concentration of this antiepileptic drug in their brain. Since an increased production of this protein occurs in the brain of human epileptics, it is possible that it contributes to drug refractoriness by impairing the attainment of clinical relevant concentrations of these drugs in the epileptic tissue. We recently found that the inhibition of the activity of P-gp in experimental models of epilepsy using specific blockers of its function, enhances the efficacy of anticonvulsant treatments. We have also demonstrated a role P-gp in brain drug disposition in experimental models of focal cortical dysplasia, a clinical condition associated with drug-refractory seizures. New therapeutic approaches of In vivo gene transfer This study concerns the use of adeno-associated viral vectors to introduce genes with therapeutic potential in the brain, thus increasing the synthesis of specific proteins to produce long-lasting anticonvulsant effects. We have demonstrated that adeno-associated viral vectors carrying the human neuropeptide Y gene under the control of a neuronal promoter, significantly increase the brain concentration of this peptide after intrahippocampal injection for a prolonged time (up to 3 months after a single intracerebral injection). The rats overexpressing this peptide are less 100

103 susceptible to limbic seizures and to epileptogenesis. Future development of this study concerns the optimization of the transgene transfer technology to inhibit spontaneously recurring seizures and envisaging a possible clinical application. Laboratory of Geriatric Neuropsychiatry Survey on the health status of old people living in the rural community of Moltrasio Data on the recruited subjects (95% of the target population) were cleaned and analyzed together with the primary care physicians. A full report and a short one dedicated to a larger public were written. The availability of the information to the local administrators enabled them to identify which interventions were most requested/needed and to begin to implement them. The results were also the subject of a conference specifically held for the population which was the target of survey. Population study on the prevalence of dementias in the older-old Parallel to the progressive increase of individuals aged 80 years or older within the elderly population (65+), the number of demented patients of 80 years or older makes up an ever increasing fraction of the total population affected by dementia. As very often happens, the exclusion from studies of subjects in the oldest age classes tends to inevitably underestimate the total number of individuals affected by dementia present in the population. To fill this gap, a door-to-door population study on the prevalence, incidence, risk factors and evolution of dementias and age-associated cognitive deficits has been set up in an elderly population aged 80 years or older living in eight small towns of Varese Province. The study is funded by a grant from the Fondazione Italo Monzino, Milano. Effects of anemia in the elderly A previous large survey in old resident of Biella (65-84 years old) has been conducted in collaboration with the Local Health Authority of Biella (ASL 12) to determine the prevalence of anemia. We have now extended the investigation to the oldest old residents (85 years or older; n= 1.775) in order to estimate the prevalence and impact of mild anemia also in this segment of the elderly population. Evaluating risk profiles in hospitalised elderly subjects In collaboration with the Geriatric Division of the Beata Vergine Hospital, Mendrisio, Switzerland, hospitalized and ambulatory patients are evaluated from a neuropsychological, functional and mobility point of view in order to estimate the impact of these factors on heath-related outcomes and disease progression. Longitudinal follow-up of individuals with mild cognitive impairment (MCI) In collaboration with the Geriatric Unit of the Beata Vergine Hospital, Mendrisio, Switzerland, follow-up study to estimate the rate of conversion to dementia of all Mild Cognitive Impairment or Questionable Dementia (CDR 0.5) patients seen at the Memory Clinic of the Hospital. Quality of care of terminally ill oncological subjects In 1999 we started a collaborative programme with the hospice via di Natale Franco Gallini in Aviano (PN). The aim of the research project was to assess the quality of care given in hospice to terminally ill oncological patients at the end of life. Two studies have been planned, one retrospective and one prospective. Present aim of the study is the 101

104 assessment of the hospice activities after its opening and to provide continuous training to nurses on use of databank. Randomised controlled trial of the Italian Group for the Study of the Second Generation Antipsychotics (GISAS) The study compares three antipsychotics, aloperidole, olanzapine, aripiprazole, according to efficacy and tolerability through a pragmatic experimental design on 800 patients, identified and randomized in 80 centres (10 patients per centre with about inhabitants catchment areas). The study is the first one conducted on such large numbers in Italy on this topic. Tools and procedures were defined and procedures for the Ethical Committees approval started. The study was presented in its scientific background in several occasions, and recruitment of centres was conducted, mainly in Northern Italy. At end of 2007, 38 centres were recruited, some of which are already active, some waiting for the ethical committee s approval, some organizing the setting for the recruitment and randomization phases. Suicides We are working on two sides: one concerns temporal trends analyses in the Italian population, currently with particular attention to suicides in adolescents and gender differences; the second one concerns the definition of a multicenter study on attempted suicides aiming at improving identification, first contact, registration, quantification at the emergency services and referral to adequate services. A network of participating services is in course of definition. At present, the study consists of the administration of a structured questionnaire to the professionals responsible of the emergency services (currently tested in three services), extraction and analysis of information from the clinical records of cases of attempted suicide identified in the emergency services (one service), comparison of data from clinical records and those from the informative system where available, adoption of specific codes to be added into the informative system where available, and a follow-up of cases seen at the emergency services. The ProgresAcuti Project of care in psychiatric acute services Analysis of data from the second phase of the descriptive study of acute psychiatric services from all Italy. In the second phase, a cluster randomization selected a subsample of 64 services. An analysis was conducted on outcomes of patients with short admissions in these services. Innovative Regional Program. The Empowerment Project The Department of Mental Health of the Niguarda Hospital in Milan continues the activity to spread and introduce empowerment values and practices in all the psychiatric community services of Milan in the perspective to encourage real opportunities for users autonomy and group identity. Tools to investigate expectations and motivations of the professionals involved in the project were designed and applied, and other tools were design for self-evaluation of the professionals in their activity of sensibilisation and to evaluate quality usefulness of this activity according to the targeted service professionals. We are conducting an observational prospective evaluation of the project Natural Social Networks (patients are accompanied by friends, acquaintances, neighbors, or colleagues for emotional support or support in various activities); we have started the observational prospective evaluation of patients admitted to housing facilities in Milan. 102

105 The revolving door effect in psychiatric residential facilities: monitoring the problem and identifying strategies The second and third phases of the project are going on: we evaluate follow-up pathways of care and clinical and functional outcomes of patients of the psychiatric residential facilities of the community service networks of several areas (for a total catchment area of 500,000 inhabitants). A total of 97 patients were recruited in the cohort of the discharge patients (discharged patients are followed up since their discharge from a community residential facility, with concern to their pathway of care in the community), and 85 in the cohort of the admitted patients (patients are followed since their admission to psychiatric residential facilities with concern to their care pathways in the facility, and, if discharged in the two years of prospective observation, to their pathway of care in the community), and evaluation tools are administered every six months. The aim is to investigate outcomes of patients during and after residential treatment, follow the care projects and pathways, identify barriers to discharge, and, for those discharged, assess quality of community tenure and subsequent admissions to residential facilities or acute services. Self-accreditation of quality of the housing facilities in the Health Trusts 1 and 2 of Turin Groups composed by users of the housing facilities, family members, professionals working in the same housing facilities and professional working in the community services discuss and the key features of the housing facilities according to their own points of view. From this material, a group of professionals develops a scheme for attribution of scores to the housing facilities. The scores will be periodically attributed to the housing facilities by the multidisciplinary groups. We assist and train the groups in the whole process. Satisfaction of users of mental health services of Milan who receive support from the Social Fund provided by the City of Milan The investigation was commissioned by the City of Milan, which provides the community psychiatric services with a fund to be used for patients considered in need, and aimed either at describing the main features of the patients assisted through the fund, and to estimate their satisfaction with it. Of the 18 community service networks of Milan, 8 provided data for more than 70% of the subjects supported by the social fund, and 4 for less than 50%. We evaluated a total of 633 patients with reference to satisfaction, and 627 of them with reference to their general and social features. UNASAM Project Quality improvement and evaluation in mental health with the active participation of the associations During the first semester a questionnaire investigating family members satisfaction with psychiatric services, professionals and interventions was designed and discussed with family members from four Italian regions in various focus groups. Groups of family members were also trained about basic concepts and procedures in service evaluation and conduction of the investigation. A total of 100 family members administered the questionnaire in 41 community mental health services of four regions, and 1450 questionnaires at end of 2007 had been collected. 103

106 Laboratory of Inflammation and Nervous System Diseases Role of selected aspects of the inflammatory response in ischemia/reperfusion injury Previous studies of ours have indicated that complement and related inflammatory systems such as contact/kinin and fibrinolytic systems may represent novel targets for reducing ischemia/reperfusion injury. We have shown that C1-INH, an endogenous serine-protease inhibitor that acts as a major regulator of both complement and kinin systems, markedly improves neurological deficits and reduces infarct volume in mice with focal transient as well as permanent ischemia induced by middle cerebral artery occlusion. We have further extended this finding defining its effectivness on different strains of mice (displaying different levels of complement expression), the time-window and the dose-response curves. Since C1-INH may act on different substrates, we have evaluated the specific involvement of the different complement pathways and of the other inflammatory systems, ie kinin and coagulation systems. To explore the mechanisms of C1-INH neuroprotection, we have also investigated the expression (protein and mrna) of inflammatory cytokines, adhesion molecules, NO synthase isoforms, apoptosis markers. The results obtained show that: i) C1-INH effectively and markedly reduces brain ischemia/reperfusion injury, inducing a decrease of the 90% of the ischemic lesion; ii) C1-INH actions lead to inhibition of cell recruitment, inflammation and apoptosis; iii) C1-INH neuroprotection is at least partially independent from the complement classical pathway and inhibition of other complement pathways, of other inflammatory systems such as the contact/kinin system, or of thrombotic events associated with ischemia-reperfusion injury may be involved in its powerful protective action (De Simoni et al. 2003; De Simoni et al. 2004; Storini et al. 2005; Storini et al. 2006). Thus C1-INH, which is presently used as replacement therapy in patients with C1-INH deficiency, represent a novel promising strategy for stroke therapy. Studies are presently going on to further define the mechanism of neuroprotection by C1-INH Stem cells as a therapeutic approach in stroke The aim of the project is to verify the conditions for the effectivenss of microsphere-derived stem cells (MSC) in reducing the ischemic injury and to investigate the mechanisms triggered by their infusion in the ischemic brain. MSC, isolated from newborn mice are infused to syngenic mice in which transient ischemia is induced by middle cerebral artery occlusion. At different time points, up to 14 days, several parameters are evaluated: distribution and phenotype of injected cells, neurodegeneration, behavioral deficits, cytokine and trophic factor gene expression, microglia activation. The results obtained show that: i) MSC effectively counteract ischemia / reperfusion injury: they can decrease neuronal loss and reverte functional impairments related to exploratory behaviour and sensory/motor activity; ii) they elicite an early response: 24 h after infusion, chemokines, angiogenic and neurotrophic factor transcripts are activated; these factors are no more activated at longer times or when cells are infused after 7 days; iii) the protective mechanism of MSC in this model of mild ischemia seems to be mainly due to the induction of beneficial factors; iv) activation of microglial cells is one of the aspects of changes in ischemic environment induced by MSC; v) their presence in the brain tissue is enhanced by the ischemic injury. Thus the reciprocal interaction between MSC and ischemic environment is crucial for stem cells protective actions (Capone et al, 2007). Ongoing studies include: 1) definition of the mechanisms of homing of stem cells in the injuried brain; 2) analysis of the protective/toxic role of microglia; 3) investigation of the early events triggered by stem cells in the ischemic brain 104

107 Identification of ischemic tolerance mediators in cerebral ischemia Recent studies indicate that cell death resulting from ischemic injury can be reduced when a sublethal ischemic episode occurs hours or days before a severe ischemic insult. This phenomenon is known as Ischemic PreConditioning (IPC) and the induced neuroprotection is called Ischemic Tolerance (IT). Several models of induction and maintenance of tolerance have been described, but the molecular mechanisms of the IPC-induced neuroprotection are not identified yet and a clear view of the mechanisms responsible for ischemic preconditioning is still lacking. Specific aims of the project are: i) to define the characteristics of IT induced by small transient ischemic attack (TIA) in experimental models of cerebral ischemia; ii) to elucidate the pathways and/or the mediators involved in ischemic tolerance that may represent potential therapeutic targets for stroke; iii) to study of direct effect of relevant protective molecules in in vitro and in vivo ischemia models. Thus ischemic preconditioning provides an opportunity to identify the putative candidate that can confer neuroprotection against stroke. A major goal is to identify the underlying endogenous protective cellular receptor/signaling cascades, with the long-term goal to allow therapeutic augmentation of the endogenous protective mechanisms in cerebral ischemia. Blood-brain barrier and preconditioning Although most attention has focused on the neuronal effects of ischemic preconditiong (IPC, see above) recent studies have shown that IPC reduces also ischemia-induced damage of the cerebrovascular unit. Together with astrocytes, pericytes and microglia, the endothelial cells in the cerebral microvasculature are the main component of the Blood-Brain Barrier (BBB). The integrity of this complex structure is essential for the maintenance of the nervous system microenvironment and its dysfunction is involved the pathophysiology of cerebral ischemia. The major goal of the research is to identify possible endogenous protective cellular receptor/signalling cascades, activated by IPC on BBB. To this purpose we have established a bidimensional BBB model by means of co-cultures of mouse brain endothelial cells and glial cells. In this model we can show the typical BBB features: presence of tight junctions, high transendothelial electrical resistance, typical paracellular and transcellular permeability values. We are specifically addressing the following aspects: 1) changes in BBB characteristics and protein expression following IPC ex vivo; 2) identification of mediators and/or pathways involved in activation and maintenance of ischemic tolerance in the cerebrovascular unit. Traumatic brain injury in mice: pathophysiological mechanisms Traumatic brain damage is the result of a primary injury (irreversible and due to the biomechanical effects of the impact) and of secondary molecular and cellular events that are initiated minutes after the injury and that interact in a complex network leading to cell death or recovery. In patients who survive the initial injury, this secondary injury cascade is now believed to account for the majority of brain damage observed following trauma. In addition the injured brain is vulnerable to systemic and intracranial secondary insults that may occur and exacerbate traumatic brain damage. Since the major part of the damage evolves over days following the impact, there is an opportunity for pharmacological therapeutic intervention. Animal models of TBI have been developed to recapitulate many clinical and pathologic aspects of head injury and have provided the basis for the dramatic increase in the understanding of the pathophysiology of brain damage after trauma. We have recently set up a mouse model of controlled cortical impact (CCI) brain injury, which yields a consistent and predictable degree of injury and recapitulates many aspects of human cerebral contusion, including increased intracranial pressure, changes in cerebrovascular reactivity (regional hypoperfusion), cerebral edema and neuronal damage in the cortex and hippocampus resulting in neurological motor deficits and cognitive dysfunction. Even if to date no specific neuroprotective therapies have been proven to be effective in attenuating the neurological sequelae of TBI patients, a better understanding of the molecular and cellular mechanisms leading to post-traumatic cell death and 105

108 recovery and their relation to functional impairment remains an important goal of experimental TBI research. Current areas of our research include: 1) neuroinflammatory response following TBI by using genetically engineered mice with the aim of investigating the role of selected genes; 2) cellular and molecular mechanisms of post-traumatic brain vulnerability to secondary insults; 3) identification of possible endogenous protective response activated by trauma similar to the ischemic preconditioning; 4) translational research in which we investigate the molecular mechanisms of damage and recovery in our model and in human brain samples obtained from patients during surgical removal of cerebral contusions. Laboratory of Molecular Neurobiology Study on pathogenic mechanisms of Amyotrophic Lateral Sclerosis Role of protein aggregation A pathological feature of ALS is the accumulation of protein aggregates in the perykaria and axons of motor neurons. Our hypothesis is that this may be due to an impairment of the ubiquitin/proteasome system (UPS). To investigate whether motor neurons of SOD1 mutant mice may specifically display an impairment of the proteasome activity we have developed double transgenic mice expressing the SOD1G93A and carrying in all cells a fusion protein functioning as marker for the activity of the proteasome, the UbG76V-GFP. We have recently concluded the project with the following results: in some motor neurons of symptomatic double transgenic mice, showing neuropathological features such as the vacuolisation and accumulation of phosphorylated neurofilaments, there is an increase in the immunoreactivity for GFP indicating a dysfunction of the proteasome. However, the accumulation of GFP in the motor neurons does not occur in the presymptomatic mice indicating that this phenomenon is likely involved in the rapid progression rather than in the pathogenesis of the disease. This is compatible with the massive accumulation of insoluble proteins found in the spinal cord of SOD1G93A at the advanced stage of the disease. The results are reported in the PhD thesis of Cristina Cheroni and in a manuscript submitted for publication. In addition, in collaboration with the Proteomic Unit of the Molecular Biochemistry and Pharmacology department we have completed the proteomic characterization of the insoluble proteins components of the aggregates isolated from the spinal cord of SOD1G93A mice. They are cytoskeletal proteins, mainly intermediate filaments, several mitochondrial proteins, chaperones, proteins of the endoplasmic reticulum, proteins involved in metabolic pathways and signaling. Post-translational modification such as nitration and carbonylation have also been found in some proteins suggesting a possible link between oxidative stress and aggregation. Role of glutamate AMPA receptors in the pathogenesis of ALS To further study the role of glutamate AMPA receptors in the susceptibility of motor neurons we have examined the expression and distribution of other two proteins regulating the trafficking of the AMPA receptor and in particular that of the subunit GluR2, which plays the major role in the homeostasis of intracellular calcium. The proteins are GRIP-1 and alsin whose interaction determine a stabilisation of the GluR2 subunit to the membrane. We observed an increase of both proteins in the motor neurons of presymptomatic SOD1G93A mice. These results, together with the previous one showing an increase of NSF, which is mainly involved in the transport of the GluR2 subunit from the cytosol to the membrane, suggest that this response may be an attempt of the motor neurons to protect them from the excitototoxity by increasing the calcium impermeable subunit of AMPA receptor at the membrane. 106

109 In vitro studies on neuron-glia interaction The interaction neurons-glia plays an important role in the inflammatory and excitotoxic mechanisms leading to motor neuron degeneration. To assess these mechanisms we set up an in vitro paradigm made by co-colture of astrocytes and spinal neurons from SOD1G93A mice embryos and non transgenic mice. We have found that motor neurons degenerate selectively when the mutant SOD1 is expressed only in the motor neurons or only in the astrocytes or in both cell type, suggesting an active role of the astrocytes in this process. Selective inhibitors of p38mapk exhibited a neuroprotective effect on motor neuron when applied to the different coltures confirming the hypothesis of an important involvement of this signalling protein in the pathogenesis of ALS. However, we have not observed significant alteration in the expression and release of TNF alpha, as found in the in vivo studies, suggesting that high levels of this cytokine found in the spinal cord of SOD1G93A mice might derive from microglia and/or other immuno cells. Further studies are in progress to assess the involvement of other cytokines and their interaction with the mechanisms of excitotoxicity. Studies on the axo-nuclear communication in the motor neurons of FALS models. The aim of this unit is to investigate whether an impairment of axonuclear communication may contribute to the degeneration of motor neurons or if it is possible to use this mechanism to vehicle a potential neuroprotective gene therapy from the nerves to the nucleus by a retrograde transport. To this purpose we plan to analyse the expression of some proteins contributing to this process in the spinal cord and in the nerves(sciatic and frenic) of SOD1G93A mice. Initially, we mainly focused our attention to the analysis of importin beta which play a crucial role in assembly dinein and other signaling proteins that must be rapidly transported from the axon to the nucleus after an axon injury. In the motor neurons which show clear signs of degeneration as the accumulation of phosphorylated neurofilaments, the immunostaining for importing beta appeared remarkably increased in the nucleus in respect to the cytoplasm suggesting that this may be a response signal to the axonal sufferance of these motor neurons. Also in the sciatic nerves of presymptomatic SOD1G93A mice, analysed by immunohistochemistry we have found an increase of this protein. Whether this may represent an accumulation or a slower transport of the protein in the axons triggered by the presence mutant SOD1 remains needs to be elucidated. We are now confirming these results in the axoplasm extract from sciatic and frenic nerves of SOD1G93A transgenic rats that allow to have more material to be analized. A colony of these rats that develop a phenotype similar to that of SOD1G93A mice have been recently introduced in our laboratory. Therapeutic interventions in mouse model of ALS Development of gene therapies for motor neuron protection The results obtained so far in our laboratory and in other groups indicate that one of the early response occurring in the motor neurons of SOD1G93A mice is the activation of the MAP kinases cascade while the mechanisms of defence such is that mediated by PI3K/Akt is not activated. Based on these observations we plan to develop a neuroprotective intervention for motor neurons using two strategies: one aimed to inhibit the intracellular activation of MAP kinases and the other aimed to activate the intracellular mechanisms of survival. In the first case we have developed viral vector carrying the shrna against p38mapk. Preliminary tests on neuron and astrocytes coltures have allowed to select sequences of shrna able to silencing almost completely the expression of p38 and to inhibit its activation by cytokines. For the second strategy we developed viral vectors expressing the activated form of Akt, importing for the inhibition of cell death process. At the present we are developing new vectors with specific promoters in attempt to vehicle the Akt selectively to the motor neurons. 107

110 Studies aimed to identify biomarkers for the diagnosis and progression of the disease in ALS patients. The diagnosis of ALS is based mainly on neurophysiological parameters associated to the progression of the disease and requires about one year to be formulated with certainty. Such interval is quite long considering that the prognosis of the disease is two-five years since the onset of symptoms. This has a negative impact on the possibility to apply a prompt therapeutic intervention. In collaboration with the department of Neurology of the Fondazione Salvatore Maugeri, IRCCS, of Pavia, we have started a series of studies aimed to investigate through the proteomic analysis, the protein profile in the PBMC and CSF of SLA patients at different stages of the disease in comparison to health controls. So far we have observed a series of interesting proteins differentially expressed potentially involved in the disease progression and useful as potential biomarkers. We are now validating the changes in some of these proteins in a wider number of ALS patients by western blot. In parallel, we are completing the characterization of nitrated proteins in the PBMC and in the CSFof ALS patients compared to healthy controls. The protein nitration on tyrosine is an oxidative mechanism that alters the function of proteins inducing their inactivation or a gain of toxic functions. Laboratory of Experimental Psychopharmacology Drug Abuse. Neural basis of drug self-administration, drug craving and relapse in the drug abuse assumption Drug craving, defined as the desire to experience the effect(s) of a previously experienced psychoactive substance is a cardinal feature of drug addiction and is clinically significant because of its potential link to relapse. To provide useful indications to the development of novel therapeutic approaches to prevent the use and abuse and the relapse of drug assumption following the outcome of craving, we elaborated experimental models of self-administration and relapse induced by cocaine-associated cues, after a period of abstinence. It was found that agonists at serotonin 2C receptors and non selective antagonists at opioids receptors selectively modulate rats seeking behaviour induced by cocaine-associated cues after a long period of abstinence and in the absence of any further cocaine. Ongoing studies are evaluating whether this modulation is peculiar for cocaine or could be generalized to other abused drugs. The role of several other neurochemical mechanisms potentially involved in the drug-seeking behaviour are also in progress. Resistance to antidepressant drugs: experimental and clinical studies This project arises from a collaboration between the laboratories of Neurochemistry and Behavior (R.W. Invernizzi), Drug Metabolism (Silvio Caccia), Biology of Neurodegenerative Disorders (Gianluigi Forloni) and focus on behavioral and biochemical characterization of an experimental model of resistance to the antidepressant drugs. Using an animal model predictive of the antidepressant activity, the effects of selective serotonin reuptake blockers (SSRI) was evaluated in several mice strains. It was found that DBA/2J and BALB/c do not respond to the antidepressant-like activity of the SSRI. The lack of effect was attributed to genotype-dependent impairment of 5-HT synthesis since DBA/2J and BALB/c carrying a single nucleotide polymorphism (C1473G mice) in the gene for the brainspecific isoform of tryptophan hydroxylase-2, the rate-limiting enzyme in the synthesis of serotonin are characterized by a decreased serotonin synthesis. This hypothesis seems to be supported by the observation that DBA/2J and BALB/c mice had less dialysate 5-HT in the medial prefrontal cortex and dorsal hippocampus than C57BL/6J amice. Moreover, in DBA/2J 108

111 and BALB/c the SSRI raised significantly less extracellular 5-HT when compared to C57BL/6J mice. Studies are ongoing to clarify whether 1) whether DBA/2J and BALB/c are responder to SSRI effects other than the anti-depressant-like 3) whether DBA/2J and BAB/c are responder to other class of antidepressant drugs. Laboratory of Neurochemistry and Behavior Resistance to antidepressant drugs Despite the wide range of antidepressant drugs available for the treatment of mood disorders the delayed onset of the antidepressant effect and the partial or no response in a considerable portion of patients still limits their efficacy. Studies in mice show that the response to antidepressant drugs is strain-dependent, suggesting that genetic differences influence the antidepressant response. Ongoing studies in collaboration with the Laboratories of Experimental Psychopharmacology and Drug Metabolism are aimed at assessing the neurobiological mechanisms involved in the resistance to antidepressant drugs in mice. The gene for the brain-specific isoform of tryptophan hydroxylase-2 (TPH-2), the rate-limiting enzyme in the synthesis of serotonin, is mutated in DBA/2J and BALB/c mice. These mice synthesize less serotonin than C57BL/6 mice (that carry the wild type form of TPH-2) and do not respond to antidepressants inhibiting selectively the reuptake of serotonin (SSRI) in the forced swimming test, a procedure used to screen compounds for antidepressant effects. In addition, mutation of TPH-2 attenuated the effect of SSRI on the extracellular concentrations of the neurotransmitter These results suggest that genetic differences in serotonin synthesis could contribute to the efficacy of SSRI. Ongoing studies are aimed at assessing the molecular mechanisms involved in the antidepressant response and identify pharmacological strategies to restore the response in non-responder to the SSRI alone. Animal model of cognitive deficit of schizophrenia; typical and atypical antipsychotics The cognitive deficit is a core symptom of schizophrenia, which has been linked to functional outcome and is relatively independent of psychotic symptoms. The antipsychotics, either typical or atypical, are able to control positive symptoms such as delirium, hallucinations and paranoia. However, the currently available atypical antipsychotics when compared to conventional antipsychotics show somewhat superior efficacy for the management of cognitive deficits in patients with schizophrenia. The cognitive deficit of schizophrenia was modelled in rats, by using a test of attention such as the 5-choice serial reaction time task (5-CSRTT) and injections of glutamate NMDA receptor antagonists into the medial prefrontal cortex (mpfc). This model makes clear links with psychopathology as dysfunctional glutamate neurotransmission in the mpfc has been implicated in cognitive deficits of schizophrenia and the 5-CSRTT is the rat analogue of the continuous performance test used to assess attention and vigilance in schizophrenic patients. Antipsychotics possess a complex pharmacology across the biogenic amine receptor families as shown by affinity constants derived from radioligand-binding techniques. The ability to antagonise the DA D 2 receptor function is shared by the conventional and by the atypical antipsychotics. However, atypical antipsychotics show a high affinity also for serotonin 5- HT 2A, 5-HT 2C and 5-HT 1A receptors. Our studies compared the effects of conventional and atypical antipsychotics in this model of cognitive deficit of schizophrenia. The results show that antipsychotics may be differentiated by a selective effect of typical antipsychotics on compulsive perseveration, and atypical antipsychotics on impulsivity. Intracerebral microdialysis studies show that attentional deficits induced by NMDA receptor antagonists is 109

112 associated with excessive glutamate in the medial prefrontal cortex of the rat and this effect was prevented by atypical antipsychotics. 110

113 111

114 DEPARTMENT OF CARDIOVASCULAR RESEARCH STAFF Head Maria Grazia FRANZOSI, Biol.Sci.D. Laboratory of Cardiovascular Clinical Pharmacology Head Roberto LATINI, M.D. Bio-imagin Unit Head Cardiovascular Endocrine Unit Head Tissue Culture Unit Head Fabio FIORDALISO, Biol.Sci.D. Serge MASSON, Ph.D. Giovanna BALCONI, Univ.Dipl. Laboratory of Clinical Drug Evaluation Head Maria Grazia FRANZOSI, Biol.Sci.D. Bioinformatics Unit Head Enrico NICOLIS, Comp.Sci.Stud. Laboratory of General Practice Research Head M.Carla RONCAGLIONI, Biol.Sci.D. Laboratory of Medical Statistics Head Simona BARLERA, Dr.Sci.Pol., MSc. Laboratory of Clinical Pharmacology Head Nursing Research Unit Head Gianni TOGNONI, M.D. Paola DI GIULIO, R.N. 112

115 CURRICULA VITAE Maria Grazia Franzosi got her Biological Science degree in 1972 at the University of Milan. Education 1972 Doctoral degree in Biological Sciences, University of Milan, Italy 1978 Postdoctoral degree in Pharmacological Research, Istituto di Ricerche Farmacologiche "Mario Negri di Milano, Italy Main fields of activity Coordination of multicentric randomised clinical trials. Relationship between genetic and environmental risk factors in coronary events. Pharmacogenetics. Pharmacoeconomics. Drug Epidemiology and Post-Marketing Surveillance. Position from 2002 Director of the Department of Cardiovascular Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy from 2004 Member of Steering Committee, Studio GISSI-AF Study, Milano, Italy from 2001 from 1998 Member of Steering Committee, Studio GISSI-HF Study, Milano, Italy Member of Steering Committee of the PROCARDIS Research Programme - A genome-wide strategy to identify susceptibility loci in precocious coronary artery disease - University of Oxford, UK from 1997 Member of Antithrombotic Trialists Collaboration, Oxford, UK from 1996 Member of Steering Committee and National Coordinator for Italy of the Organization to Assess Strategies for Ischemic Syndromes (OASIS-2, OASIS-5, OASIS 6, OASIS-7, CURE, INTER- HEART studies), Canadian Cardiovascular Collaboration, McMaster University, Hamilton, Canada and of ACTIVE and RELY studies, Canadian Cardiovascular Collaboration, McMaster University, Hamilton, Canada Director of European Coordinating Centre and Member of Steering Committee, Collaborative from 1993 from 2002 Organization for RheothRx Evaluation (CORE), McMaster University, Hamilton, Canada Member of Steering Committee, Studio GISSI-Prevenzione, Milano, Italy Member of Fibrinolytic Therapy Trialists s Collaboration, Oxford, UK e del Collaborative Group on Angiotensin Converting Enzyme Inhibitors Trials, National Institutes of Health, Bethesda, Washington, USA Head of the Laboratory of Clinical Drug Evaluation, Istituto di Ricerche Farmacologiche "Mario Negri" Head of the Clinic Drug Evaluation Unit of the Laboratory of Clinical Pharmacology, Istituto di Ricerche Farmacologiche "Mario Negri" from 1984 Member of the Scientific and Organising Secretariat, Gruppo Italiano per lo Studio della Sopravvivenza nell'infarto Miocardico (GISSI-1, GISSI-2, GISSI-3 studies) Milano, Italy Researcher at the Laboratory of Clinical Pharmacology, Istituto di Ricerche Farmacologiche "Mario Negri" and at the Regional Center for Drug Information of the Lombardy Region Selected publications Anand SS, Xie CC, Mehta S, Franzosi MG, Joyner C, Chrolavicius S, Fox KA, Yusuf S, CURE Investigators. Differences in the management and prognosis of women and men who suffer from acute coronary syndromes. J Am Coll Cardiol 2005, 46: Yusuf S, Howken S, Ounpuu S, Bautista L, Franzosi MG, Commerford P, Lang CC, Rumboldt Z, Onen CL, Lisheng L, Tanomsup S, Wangai Jr P, Razak F, Shama AM, Anand SS, on behalf of the INTERHEART Study Investigators. Obesity and the risk of myocardial infarction in participants from 52 countries: a case-control study. Lancet 2005, 336: Levantesi G, Macchia A, Marfisi R M, Franzosi M G, Maggioni A P, Nicolosi G L, Schweiger C, Tavazzi L, Tognoni G, Valagussa F, Marchioli R, on behalf of the GISSI-Prevenzione Investigators. Metabolic syndrome and risk of cardiovascular events after myocardial infarction. J Am Coll Cardiol 2005; 46: Franzosi MG. Should we continue to use BMI as a cardiovascular risk factor? Lancet 2006; 368: Farrall M, Green FR, Peden JF, Olsson PG, Clarke R, Hellenius ML, Rust S, Lagercrantz J, Franzosi MG, Schulte H, Carey A, Olsson G, Assman G, Tognoni G, Collins R, Hamsten A, Watkins H, on behalf of the PROCARDIS Consortium. Genome-wide mapping of susceptibility to coronary artery disease identifies a novel replicated locus on chromosome 17. PLoS Genet Chiodini B, Franzosi M G, Barlera S, Signorini S, Lewis C M, D'Orazio A, Mocarelli P, Nicolis E, Marchioli R, Tognoni G, GISSI, SIBioC-GISSI Prevenzione Group. Apolipoprotein E polymorphisms influence effect of pravastatin on survival after myocardial infarction in a Mediterranean population: the GISSI-Prevenzione study. Eur Heart J 2007 ; 28:

116 Simona Barlera got her degree in Political Science, area Statistics at the Università degli Studi di Milano in Milano in 1992, followed by a master in Medical Statistics at the London School of Hygiene and Tropical Medicine, University of London in Education Degree in Political Science, area Statistics at the Università degli Studi di Milano in Milano Master post-lauream in Medical Statistics at the London School of Hygiene and Tropical Medicine, University of London, London Visiting Scientist in the Department of Statistical Genetics, Wellcome Trust Centre for Human Genetics, University of Oxford (UK). Main fields of activity Methodology of Clinical Trials in the cardiovascular field. Preparation and viewing of research protocols, planning and conduct of statistical analyses and the reporting of findings on scientific journals. Genetic epidemiology: genome-wide strategies (linkage analysis) to identify susceptibility genes in coronary artery disease; case-control studies in order to identify candidate genes involved in the cardiovascular pathology. Position from Oct 2006 Head of the Laboratory of Medical Statistics, Department of Cardiovascular Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy May 99-Sept 06 Head of the Medical Statistics Unit, Department of Cardiovascular Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Visiting Scientist in the Department of Statistical Genetics, Wellcome Trust Centre for Human Genetics, University of Oxford (UK) Researcher in the Unit of Applied Statistics and Information Technology, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Selected publications Chiodini B, Barlera S, Franzosi M G, Labarta V, Introna M, Tognoni G.APO B gene polymorphisms with coronary artery disease: A meta-analysis. Atherosclerosis 2003; 167: Latini R, Masson S, Anand I, Salio M, Hester A, Judd D, Barlera S, Maggioni AP, Tognoni G, Cohn J N, Val-HeFT Investigatore. The comparative prognostic value of plasma neurohormones at baseline in patients with heart failure enrolled in Val-HeFT. Eur Heart J 2004; 25: Roncaglioni M C, Avanzini F, Roccatagliata D, Monesi L, Tamayo-Benitez D, Tombesi M, Caimi V, Longoni P, Lauri D, Barlera S, Tognoni G, Collaborative Group Risk Prevention Study. How general practitioners perceive and grade the cardiovascular risk of their patients. Eur J Cardiovasc Prev Rehabil 2004; 11: Maggioni AP, Latini R, Carson PE, Singh SN, Barlera S, Glazer R, Masson S, Cere` E, Rognoni G, Cohn JN. Valsartan reduces the incidence of atrial fibrillation in patients with heart failure: Results from the Valsartan Heart Failure Trial (Val-HeFT). Am Heart J 2005; 149: 1-10 Masson S, Latini R, Anand IS, Vago T, Angelici L, Barlera S, Missov ED, Clerico A, Tognoni G, Cohn JN, Val-HeFT Investigators. Direct comparison of B-type natriuretic peptide (BNP) and amino-terminal probnp in a large population of patients with chronic and symptomatic heart failure. The Valsartan Heart Failure (Val-HeFT) data. Clin Chem 2006; 52: Barlera S, Specchia C, Farrall M, Chiodini BD, Franzosi MG, Rust S, Green F, Nicolis E, Peden J, Assmann G, Collins R, Hamsten A, Tognoni G, PROCARDIS Consortium. Multiple QTL influence the serum Lp(a) concentration: a genome-wide linkage screen in the PROCARDIS study. Eur J Hum Genet 2007; 15:

117 Roberto Latini got his Medical Doctor degree in 1978 at the University of Milan. Education University of Milan School of Medicine, degree in Medicine Merck Sharp & Dohme International Fellow in Clinical Pharmacology Main fields of activity Mechanisms of cardiac damage following ischemia, with focus on eurohumoral activation. Use of stem cells for cardiac repair. Biohumoral investigations within large scale clinical trials in heart failure and atrial fibrillation. Positions from 1990 Head of the Cardiovascular Clinical Pharmacology Laboratory (Cardiovascular Research Department) Istituto di Ricerche Farmacologiche Mario Negri, Milan, Italy from 2001 Member of the GISSI-HF Steering Committee from 2004 Member of the GISSI-AF Steering Committee from 2005 Member of the CandHeart Steering Committee from 1999 Visiting Professor Dept of Medicine, New York Medical College, Valhalla, NY, USA Cardiology Fellow (Dr. R. E. Kates, Laboratory) Stanford University Medical Center, CA, USA Member of the Sub-Group RMs for Drugs (Community Bureau of Reference, Commission of the European Communities) Fellow at the Laboratory of Clinical Pharmacology of the Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Selected publications Latini R, Maggioni AP, Peri G, Gonzini L, Lucci D, Mocarelli P, Vago L, Pasqualini F, Signorini S, Soldateschi D, Tarli L, Schweiger C, Fresco C, Cecere R, Tognoni G, Mantovani A, LATIN Investigators. Prognostic significance of the long pentraxin PTX3 in acute myocardial infarction. Circulation 2004; 110: Corada M, Chimenti S, Cera M R, Vinci M, Salio M, Fiordaliso F, De Angelis N, Villa A, Bossi M, Staszewsky L, Vecchi A, Parazzoli D, Motoike T, Latini R, Dejana E. Junctional adhesion molecule-a-deficient polymorphonuclear cells show reduced diapedesis in peritonitis and heart ischemia-reperfusion injury. Proc Natl Acad Sci USA 2005; 102: Galli D, Innocenzi A, Staszewsky L, Zanetta L, Sampaolesi M, Bai A, Martinoli E, Carlo E, Balconi G, Fiordaliso F, Chimenti S, Cusella G, Dejana E, Cossu G, Latini R. Mesoangioblasts, vessel-associated multipotent stem cells, repair the infarcted heart by multiple cellular mechanisms. A comparison with bone marrow progenitors, fibroblasts, and endothelial cells. Arterioscler Thromb Vasc Biol 2005; 25: Latini R, Masson S, Anand I S, Missov E, Carlson M, Vago T, Angelici L, Barlera S, Parrinello G, Maggioni A P, Tognoni G, Cohn J N, Val-HeFT Investigators. Prognostic value of very low plasma concentrations of troponin T in patients with stable chronic heart failure. Circulation 2007; 116: Sarto P, Balducci E, Balconi G, Fiordaliso F, Merlo L, Tuzzato G, Pappagallo G L, Frigato N, Zanocco A, Forestieri C, Azzarello G, Mazzucco A, Valenti M T, Alborino F, Noventa D, Vinante O, Pascotto P, Sartore S, Dejana E, Latini R. Effects of exercise training on endothelial progenitor cells in patients with chronic heart failure. J Card Fail 2007; 13:

118 Maria Carla Roncaglioni got her Biological Science degree in 1987 at the University of Milan. Education 1987 Doctoral degree in Biological Sciences, University of Milan, Italy Research Fellow at the Dept. of Biochemistry, Faculty of Medicine, Rijksuniversiteit of Limburg, Maastricht, The Netherland (Prof. C.Hemker); Visiting Scientist at the Cardiovascular Research Unit, Hammersmith Hospital, London, UK (Prof. A. Maseri) Main fields of activity Coordination of multicenter clinical trials and observational studies in different cardiovascular areas (neurological, angiological, cardiological). Coordination of a network of more than 1000 GPs actively involved in epidemiological and experimental studies in the prevention of cardiovascular diseases. Position from 2001 Head of the Laboratory for General Practice Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy from 1989 Senior Researcher in the Clinical Pharmacology Laboratory, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy from 1974 Researcher in the Laboratory for the Study of Haemostasis and Thrombosis, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Selected publications Avanzini F, Palumbo G, Alli C, Roncaglioni MC, Ronchi E, Cristofari M, Capra A, Rossi S, Nosotti L, Costantini C, Pietrofeso R, PPP Primary Prevention Project. Effects of low-dose aspirin on clinic and ambulatory blood pressure in treated hypertensive patients. Am J Hypertens 2000; 13: Tognoni G, Avanzini F, Pangrazzi J, Roncaglioni M C, Bertele V, de Gaetano G, Caimi V, Tombesi M, Colombo Fabio, Barlera S, PPP - Primary Prevention Project. Low-dose aspirin and vitamin E in people at cardiovascular risk: A randomized trial in general practice. Lancet 2001; 357: Sacco M, Pellegrini F, Roncaglioni MC, Avanzini F, Tognoni G, Nicolucci A, PPP - Primary Prevention Project. Primary prevention of cardiovascular events with low-dose aspirin and vitamin E in type 2 diabetic patients. Results of the Primary Prevention Project (PPP) trial. Diabetes Care 2003; 26: Roncaglioni MC, Avanzini F, Roccatagliata D, Monesi L, Tamayo-Benitez D, Tombesi M, Caimi V, Longoni P, Lauri D, Barlera S, Tognoni G, Collaborative Group Risk Prevention Study. How general practitioners perceive and grade the cardiovascular risk of their patients Eur J Cardiovasc Prev Rehabil 2004; 11: Monesi L, Avanzini F, Barlera S, Caimi V, Lauri D, Longoni P, Roccatagliata D, Tombesi M, Tognoni G, Roncaglioni MC. Appropriate use of antiplatelets: is prescription in daily practice influenced by the global cardiovascular risk? Eur J Clin Pharmacol 2005; 61: Berger JS, Roncaglioni MC, Avanzini F, Pangrazzi J, Tognoni G, Brown DL. Aspirin for the primary prevention of cardiovascular events in women and men: A sex-specific meta-analysis of randomized controlled trials. JAMA 2006; 295:

119 Gianni Tognoni got his Medical Doctor degree in 1970, University of Milan. Education 1964 Philosophy Doctor, University of Rome 1970 Medical Doctor degree, University of Milan Main fields of activity Randomised Clinical Trials. Pharmacoepidemiology and pharmacoeconomics. Outcomes research Genetic epidemiology. Technology transfer and drug policy in developing countries. Cardiocerebrovascular disorders; primary and secondary prevention; heart-failure; diabetes; aging; psychiatric epidemiology. Health and human rights. Position from 2004 Member, Commission of Human Experimentation of the Italian Drug Agency (AIFA) Member, Commissione Unica del Farmaco (CUF), Ministry of Health from 2002 Director, Consorzio Mario Negri Sud, S. Maria Imbaro, Chieti Coordinator, Department of Clinical Pharmacology and Epidemiology, Consorzio Mario Negri Sud, S. Maria Imbaro, Chieti Coordinator, Department of Cardiovascular Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano from 1990 Co-Director, Scuola Superiore di Ricerca in Medicina Generale (CSeRMEG) from 1988 Coordinator of training and research programs in community medicine, clinical epidemiology, pharmaco-epidemiology in Bolivia, Argentina, Chile, Brazil, Ecuador, Perù from 1986 Founding member of the International Society of Drug Bulletins (ISDB) Coordinator, Commission of Human Experimentation, Regione Lombardia from 1983 Member of the Editorial Board of the nursing research Journal Rivista dell'infermiere/assistenza Infermieristica e Ricerca from 1977 Consultant to WHO and other UN agencies for drug selection and policy; training in methods of clinical and epidemiological research in developing countries mainly in Latin America (Costa Rica, Nicaragua, Bolivia, Perù) and Africa (Angola, Ethiopia, Burkina Faso) from 1976 Head, Laboratory of Clinical Pharmacology of the Istituto di Ricerche Farmacologiche "Mario Negri", Milano from 1975 Head, Regional Centre for Drug Information (CRIF), Regione Lombardia, Istituto di Ricerche Farmacologiche "Mario Negri", Milano Research Assistant, Laboratory of Clinical Pharmacology, Istituto di Ricerche Farmacologiche "Mario Negri", Milano Selected publications Anand I S, Fisher L D, Chiang Y T, Latini R, Masson S, Maggioni A P, Glazer R D, Tognoni G, Cohn J N, Val-HeFT Investigators. Changes in brain natriuretic peptide and norepinephrine over time and mortality and morbidity in the Valsartan Heart Failure (Val-HeFT). Circulation 2003; 107: Franciosi M, Pellegrini F, De Berardis G, Belfiglio M, Di Nardo B, Greenfield S, Kaplan S H, Rossi M C E, Sacco M, Tognoni G, Valentini M, Nicolucci A, QuED Study Group. Impact of physicians' beliefs and practices on cholesterol levels in patients with type 2 diabetes: A longitudinal assessment. Am Heart J 2005; 149: Maggioni A P, Latini R, Carson P, Singh S N, Barlera S, Glazer R, Masson S, Cere' E, Tognoni G, Cohn J N.Valsartan reduces the incidence of atrial fibrillation in patients with heart failure: Results form the Valsartan Heart Failure Trial (Val-HeFT). Am Heart J 2005; 149: Monesi L, Avanzini F, Barlera S, Caimi V, Lauri D, Longoni P, Roccatagliata D, Tombesi M, Tognoni G, Roncaglioni M C. Appropriate use of antiplatelets: is prescription in daily practice influenced by the global cardiovascular risk? Eur J Clin Pharmacol 2005; 61: Evangelista V, De Berardis G, Totani L, Avanzini F, Giorda CB, Brero L, Levantesi G, Marelli G, Pupillo M, Iacuitti G, Pozzoli G, Di Summa P, Nada E, De Simone G, Dell'Elba G, Amore C, Manarini S, Pecce R, Maione A, Tognoni G, Nicolucci A. Persistent platelet activation in patients with type 2 diabetes treated with low doses of aspirin. J Thromb Haemost 2007; 5: Silletta MG, Marfisi RM, Levantesi G, Boccanelli A, Chieffo C, Franzosi MG, Geraci E, Maggioni AP, Nicolosi GL, Schweiger C, Tavazzi L, Tognoni G, Marchioli R, GISSI-Prevenzione. Coffee consumption and risk of cardiovascular events after acute myocardial infarction. Results from the GISSI (Gruppo Italiano per lo Studio della Sopravvivenza nell'infarto miocardico)-prevenzione trial. Circulation 2007; 116:

120 Giovanna Balconi got her degree at the School for Technicians of Biomedical Institutes of the University of Milan, with a specialisation in Histology in the Pathological Anatomy Laboratory of the same University (1968). Main fields of interest Isolation, culture and characterization of peripheral blood circulating progenitor cells of patients with heart failure. In vitro culture and characterization of stem cells for repair of myocardial infarction in experimental animal models. Positions from July 2005 Head of Tissue Culture Unit, Cardiovascular Clinical Pharmacology Laboratory, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Oct June 2005 Head of Tissue Culture Unit, Vascular Biology Laboratory, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Dec Oct 1995 Head of Tissue Culture Unit, Anticancer Chemotherapy Laboratory, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Oct Nov 1983 Researcher, Anticancer Chemotherapy Laboratory, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Selected publications Balconi G, Spagnuolo R, Dejana E Development of endothelial cell lines from embryonic stem cells. A tool for studying genetically manipulated endothelial cells in vitro Arterioscler Thromb Vasc Biol 2000; 20: Condorelli G, Borello U, De Angelis L, Latronico M, Sirabella D, Coletta M, Galli R, Balconi G, Follenzi A, Frati G, Cusella De Angelis M G, Gioglio L, Amuchastegui C S, Adorini L, Naldini L, Vescovi A, Dejana E, Cossu G Cardiomyocytes induce endothelial cells to trans-differentiate into cardiac muscle: Implications for myocardium regeneration Proc Natl Acad Sci USA 2001; 98: Cattelino A, Liebner S, Gallini R, Zanetti A, Balconi G, Corsi A, Bianco P, Wolburg H, Moore R, Oreda B, Kemler R, Dejana E The conditional inactivation of the beta-catenin gene in endothelial cells causes a defective vascular pattern and increased vascular fragility J Cell Biol 2003; 162: Cusella De Angelis M G, Balconi G, Bernasconi S, Zanetta L, Boratto R, Galli D, Dejana E, Cossu G Skeletal myogenic progenitors in the endothelium of lung and yolk sac Exp Cell Res 2003; 290: Galli D, Innocenzi A, Staszewsky L, Zanetta L, Sampaolesi M, Bai A, Martinoli E, Carlo E, Balconi G, Fiordaliso F, Chimenti S, Cusella G, Dejana E, Cossu G, Latini R Mesoangioblasts, vessel-associated multipotent stem cells, repair the infarcted heart by multiple cellular mechanisms. A comparison with bone marrow progenitors, fibroblasts, and endothelial cells Arterioscler Thromb Vasc Biol 2005; 25: Sarto P, Balducci E, Balconi G, Fiordaliso F, Merlo L, Tuzzato G, Pappagallo G L, Frigato N, Zanocco A, Forestieri C, Azzarello G, Mazzucco A, Valenti M T, Alborino F, Noventa D, Vinante O, Pascotto P, Sartore S, Dejana E, Latini R Effects of exercise training on endothelial progenitor cells in patients with chronic heart failure J Card Fail 2007; 13: Paola Di Giulio got her Nursing Diploma in the Nursing School of Istituto Nazionale dei Tumori in Milano and her Master in Oncology Nursing at Guildford University (UK) in Main fields of activity Coordination of multicentre and observational studies studies in cardiology and palliative care. Coordination of nursing networks. Position from March 2001 Associated professor at the Turin University. from 1997 Responsible of the Nursing Research Unit from 1995 Senior researcher of the Cardiovascular Research Department from 1989 Consultant of the Clinical Phrmacology Laboratory since 1998 Coordinator of the Editorial Board of Assistenza Infermieristica e Ricerca Selected publications Laquintana D, Di Giulio P: Per un ruolo infermieristico nella farmacovigilanza. Assistenza Infermieristica e ricerca 2002; 21: Di Giulio P, Saiani L, Laquintana D, Palese A, Gruppo PARI-ETLD. Studio clinico randomizzato controllato in doppio cieco sull efficacia dei trattamenti delle lesioni da decubito. Assistenza Infermieristica e Ricerca 2004; 23: Toscani F, Brunelli C, Miccinesi G, Costantini M, Gallucci M, Tamburini M, Paci E, Di Giulio P, Peruselli C and the Italian Co-operative Research Group on Palliative Medicine Predicting survival in terminal cancer patients: clinical observation or quality of life evaluations? Palliative Medicine 2005; 19: Toscani F, Di Giulio P, Brunelli C, Miccinesi G. Laquintana D. How How people die in hospital general wards: a descriptive study. J Pain Symptom Manage 2005; 30:

121 Saiani L, Di Giulio P, Gruppo PARI-FV (Percorsi Assistenziali e Ricerca Infermieristica-Farmaco Vigilanza) Epidemiologia dei problemi assistenziali legati a farmaci e presidi in RSA e distretto. Assistenza Infermieristica e Ricerca 2007; 26: Lepore V, Cecchetto G, Di Giulio P, Saiani L, Samarelli V, Saugo M, Romero M, Scurti V, Tognoni G, Valerio M. Età anziana-molto-anziana, e aspettativa di vita? Assistenza Infermieristica e Ricerca 2007; 26: Fabio Fiordaliso got his Biological Science degree in 1995 at the University of Milan. Education 1998 Postdoctoral degree in Pharmacological Research, Istituto di Ricerche Farmacologiche Mario Negri, Milan, Italy 1995 Doctoral degree in Biological Sciences, University of Milan, Italy Main fields of activity Therapeutical potential of stem cell and antioxidant treatments in experimental model of diabetic cardiomyopathy and in primary myocyte cultures exposed to hyperglycemia. Morphological and structrural analysis of cells and tissue by optical, confocal and electron microscopy. Positions from 2007 from 2006 from 2005 from 2005 from 2001 Head of Bio-imaging Unit, Department of Cardiovascular Research, Istituto di Ricerche Farmacologiche Mario Negri, Milan Member of the Heart Failure Association (HFA) of the European Society of Cardiology Member of the Working group on myocardial function (WG 4) of the European Society of Cardiology Member of the steering committee of the Consorzio of Microscopy and Image Analysis (MIA) Senior Research Scientist, Laboratory of Cardiovascular Clinical Pharmacology (Department of Cardiovascular Research), Istituto di Ricerche Farmacologiche Mario Negri, Milan Post-Doctoral Research Fellow at Cardiovascular Research Institute (Department of Medicine), New York Medical College, Valhalla, New York Research Fellow, Laboratory of Cardiovascular Clinical Pharmacology (Department of Cardiovascular Research), Istituto di Ricerche Farmacologiche Mario Negri, Milan Research training, Institute of General Pathology, University of Milan (Italy) Selected publications Fiordaliso F, Li B, Latini R, Sonnenblick EH, Anversa P, Leri A, Kajstura J. Myocyte death in streptozotocin-induced diabetes in rats is angiotensin II dependent. Laboratory Investigation 2000; 80: Fiordaliso F, Leri A, Cesselli D, Limana F, Safai B, Nadal-Ginar B, Anversa P, Kajstura J. Hyperglycemia activates p53 and p53-regulated genes leading to myocyte cell death. Diabetes 2001; 50: Fiordaliso F, Bianchi R, Staszewsky.,Cuccovillo I, Doni M, Laragione T, Salio M, Savino C, Melucci S, Santangelo F, Scanziani E, Masson S, Ghezzi P, Latini R. Antioxidant N-Acetyl-L-Cysteine attenuates hyperglycemia-induced cardiomyocyte death in rats. Journal of Molecular and Cellular Cardiology 2004; 37: Fiordaliso F, Chimenti S, Staszewsky L, Bai A, Carlo E, Cuccovillo I, Doni M, Mengozzi M, Tonelli R, Ghezzi P, Coleman T, Brines M, Cerami A, Latini R. A non-erythropoietic derivative of EPO effectively ameliorates experimental cardiac ischemia with reperfusion. Proceedings National Academy Sciences (PNAS) 2005; 102: Fiordaliso F, Cuccovillo I, Bianchi R, Bai A, Doni M, Salio M, De Angelis N, Ghezzi P, Latini R, Masson S. Cardiovascular oxidative stress is reduced by an ACE inhibitor in a rat model of streptozotocin-induced diabetes. Life Sciences 2006; 79: Fiordaliso F, De Angelis N, Cuccovillo I, Bai A, Salio M, Serra DM, Bianchi R, Razzetti R, Latini R, Masson S. Effect of β-adrenergic and renin-angiotensin system blockade on myocyte apoptosis and oxidative stress in diabetic hypertensive rats. Life Sciences 2007; 81: Serge Masson obtained his doctorate (PhD) in Biochemistry and Cellular Biology in 1990 at the University of Marseilles (France), followed by a postdoctoral stay at the Panum Institute in Copenhagen (Denmark) Education Doctorate fellow, Faculty of Medicine, University of Aix-Marseilles, France 119

122 Post-doctoral Researcher, Panum Institute and Assistant Lecturer, University of Copenhagen, Denmark 1993 Research Scientist, NMR Laboratory, Hospital San Raffaele, Milan, Italy from 1994 Research Scientist, Department of Cardiovascular Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Main fields of activity Physiopathology, diagnostic and prognostic role of the activation of neuroendocrine systems in cardiovascular disease Position from 2002 from 2002 from 2002 Head of the Cardiovascular Endocrine Unit, responsible for Quality Assurance for the Department of Cardiovascular Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Tutor of fellows of the School of Specialists in Pharmacological Research, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Fellows of the American Heart Association (Basic Council) and the Working Group on Myocardial Function of the European Society of Cardiology Selected publications Latini R, Masson S, Anand IS, Salio M, Hester A, Judd D, Barlera S, Maggioni AP, Tognoni G, Cohn NJ, for the Val- HeFT Investigators. The comparative prognostic value of plasma neurohormones at baseline in patients with heart failure enrolled in Val-HeFT. Eur Heart J 2004; 25: Anand IS, Latini R, Florea VG, Kuskowski MA, Masson S, Signorini S, Mocarelli P, Hester A, Glazer R, Cohn JN, for the Val-HeFT Investigators. C-reactive protein in heart failure: prognostic value and the effect of valsartan. Circulation 2005; 112: Masson S, Latini R, Anand I S, Barlera S, Judd D, Salio M, Perticone F, Perini G, Tognoni G, Cohn JN, on behalf of the Val-HeFT Investigators. The prognostic value of Big endothelin-1 in more than 2,300 patients with heart failure enrolled the Valsartan in Heart Failure Trial (Val-HeFT). J Card Fail 2006; 12: Masson S, Latini R, Anand IS, Vago T, Angelici L, Barlera S, Missov ED, Clerico A, Tognoni G, Cohn JN, on behalf of the Val-HeFT Investigators. Direct comparison of B-type natriuretic peptide (BNP) and amino-terminal probnp in a large population of patients with chronic and symptomatic heart failure. The Valsartan Heart Failure (Val-HeFT) data. Clin Chem 2006; 52: Latini R, Masson S, Anand I S, Missov E, Carlson M, Vago T, Angelici L, Barlera S, Parrinello G, Maggioni AP, Tognoni G, Cohn J N, Val-HeFT Investigators. Prognostic value of very low plasma concentrations of troponin T in patients with stable chronic heart failure. Circulation 2007; 116: Staszewsky L, Wong M, Masson S, Barlera S, Carretta E, Maggioni AP, Anand IS, Cohn JN, Tognoni G, Latini R, Valsartan Heart Failure Trial Investigators. Clinical, neurohormonal, and inflammatory markers and overall prognostic role of chronic obstructive pulmonary disease in patients with heart failure: data from the Val-HeFT Heart Failure Trial. J Card Fail 2007; 13: Enrico Bjørn Nicolis has attempted some courses in Computer Science at the University of Milan. Education Research fellow, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Main fields of activity Data management and analysis of randomized clinical trials. Developing of database and tools for studies of population genetics, particularly for linkage analysis. Position from 2001 Head of the Bioinformatics Unit, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy from 1999 Research fellow of the Laboratory of Clinical Drugs Evaluation from 1997 System administrator at the EDP center, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy from 1991 Research fellow at the Medical Informatics and Applied Statistics Unit, Istituto di Ricerche Farmacologiche "Mario Negri", Milano, Italy Selected publications GISSI-Avoidable Delay Study Group. Epidemiology of avoidable delay in the care of patients with acute myocardial infarction in Italy. A GISSI-generated study. Arch Intern Med 1995; 155: Nobili A, Gebru F, Rossetti A, Schettino F, Zahn R W, Nicolis E, Macario G, Celani L, Acik V O, Farina M L, Naldi L. Doctorline: A private toll-free telephone medical information service. Five years of activity: Old problems and new perspectives. Ann Pharmacother 1998; 32:

123 Santoro E, Nicolis E, Franzosi MG.Telecommunication technology for the management of large scale clinical trials: The GISSI experience. Comput Methods Programs Biomed 1999; 60: Santoro E, Nicolis E, Franzosi M G, Tognoni G. Internet for clinical trials: Past, present, and future. Control Clin Trials 1999; 20: Tognoni G, Franzosi MG, Nicolis E, Barlera S, Specchia C, Chiodini B, Crociati L, Ferrario L, PROCARDIS Consortium. A trio family study showing association of the lymphotoxin-alfa N26 (804A) allele with coronary artery disease. Eur J Hum Genet 2004; 12: Barlera S, Specchia C, Farrall M, Chiodini BD, Franzosi MG, Rust S, Green F, Nicolis E, Peden J, Assmann G, Collins R, Hamsten A, Tognoni G, PROCARDIS Consortium. Multiple QTL influence the serum Lp(a) concentration: a genome-wide linkage screen in the PROCARDIS study. Eur J Hum Genet 2007; 15:

124 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The areas of interest of the Department of Cardiovascular Research include the experimental, clinical, genetic, epidemiological aspects of acute myocardial infarction, cardiac failure, cardiac arrhythmias, as well as the clinical and epidemiological investigation of cardiovascular prevention, hypertension and stroke. Following the successful experience of the GISSI-trials (Gruppo Italiano per lo Studio della Sopravvivenza nell'infarto), the activation of large collaborative networks in the setting of the National Health Service hospitals and in general practice has become a key characteristics of the Department, which can now rely on the permanent collaboration of over 300 clinical groups and of several hundred general practitioners. Over the years, firm links have also been established with international leading research groups. The experimental research activity concerns the physiopathology, the pharmacological modulation and the prognostic role of the activation of the renin-angiotensin-aldosterone system, as well as other neurohormonal systems, in myocardial infarction and heart failure, the physiopathology, the pharmacological modulation and prognostic role of the activation of the inflammatory processes in myocardial infarction and heart failure; a more recent research topic is the cell therapy of myocardial infarction. The activity in clinical research includes the clinical assessment of therapeutic strategies with large scale clinical trials in the field of acute coronary syndromes, congestive heart failure and atrial fibrillation. A recently developing area is the genetic epidemiology of myocardial infarction and heart failure. Several studies have been conducted in the area of clinical epidemiology and risk factors assessment of myocardial infarction. The collaboration with a large network of General Practitioners in the area of cardiovascular prevention allowed to test new hypotheses through large scale clinical trials and to evaluate the actual transferability of evidence based interventions in the every day practice through epidemiological or outcome research studies. Pharmacoepidemiological studies through the analysis of a large sample of Local Health Units drug prescriptions were also performed. A research network of nurses has been developed with the main focus on the assessment of healthrelated quality of life of patients and on the epidemiology of nursing interventions and their implications for patients' well being and outcomes. FINDINGS/MAIN RESULTS The short pentraxin C-reactive protein (CRP) and the long pentraxin (PTX3) are markers of cardiovascular risk and of severity of heart disease. However, the pthophysiologic role of pentraxins in these diseases is largely unknown. Using knockout mice for ptx3 we have shown that PTX3, a highly conserved protein from mouse to man, reduces inflammatory response in acute myocardial infarction and consequently ischemia/reperfusion injury. Thus PTX3 is more than a simple marker of cardiovascular risk. The study has been done in collaboration with Istituto Clinico Humanitas, Rozzano, Milano (Prof Alberto Mantovani). We conducted a genetic study in the patients of the GISSI-Prevenzione trial to assess the effect of the APOE gene on mortality and on the response to statin treatment in secondary prevention after myocardial infarction (MI). We analysed 3304 Italian patients with MI randomized to pravastatin or no treatment, with a median follow-up time of 24.3 months. We found that epsilon-4 allele is a determinant of pravastatin response in terms of survival. Though in the entire population investigated we found a beneficial effect of pravastatin in terms of survival, only the epsilon-4 carriers seemed to have gained a significant benefit from this treatment. We suggest that the effect of statins is of particular interest in this fraction of the 122

125 population and that genetic markers can help in identifying patients that benefit more from statin treatment. PROCARDIS data were analyzed with the goal to identify chromosomal regions associated to the variability of quantitative phenotypes. The serum concentration of lipoprotein Lp(a), known to be highly heritable and associated with cardiovascular risk, was analyzed using a genome-wide linkage analysis. A highly significant linkage was confirmed at LPA locus on chromosome 6q27, as well as another region of significant linkage was detected on chromosome 13q These results have been published on the European Journal Human Genetics 2007: 15, and provide a basis for further studies of candidate genes in these regions. NATIONAL COLLABORATIONS Angiogenesis and Tumor Targeting Research Unit & Telethon Institute for Gene Therapy, Ospedale San Raffaele, Milano ANMCO (Associazione Nazionale Medici Cardiologi Ospedalieri) CINECA (Consorzio Interuniversitario per il Calcolo Automatico dell'italia Nord-Orientale) CSeRMEG (Centro Studi e Ricerche in Medicina Generale) Gruppi organizzati di MMG (FIMMG, CoS, Ass.Cu.M.I., AMISI) IEO (Istituto Europeo di Oncologia), Milano IFOM-FIRC, Milano Istituto di Ricerca in Cure palliative Lino Maestroni Laboratorio di Endocrinologia, Ospedale Luigi Sacco, Milano Ematologia, Ospedale Sant Anna, Torino Regione Lombardia SIBioC (Società Italiana di Biochimica Clinica e Biologia Molecolare) SIFO (Società Italiana di Farmacia Ospedaliera) Stem Cell Research Institute, Ospedale San Raffaele, Milano Università degli Studi di Milano, Dipartimento di Medicina Interna Università degli Studi di Torino, Dipartimento di Anatomia, Farmacologia e Medicina Forense Università degli Studi di Torino, Dipartimento di Sanità Pubblica e Microbiologia Università degli Studi di Verona, Dipartimento di Sanità Pubblica Università degli Studi di Verona, Istituto di Anatomia Umana INTERNATIONAL COLLABORATIONS Cecomet (Centro de Epidemiologia comunitaria y Medicina tropical, Esmeraldas) Ecuador Cochrane Collaboration, Oxford, UK Clinical Trial Research Unit, Auckland University, New Zealand CTSU (Clinical Trial Service Unit) /ISIS (International Studies on Infarct Survival), Oxford, UK Division of Genetics and Development, Guy's, King's and St Thomas' School of Medicine, King's College, London, UK ECLA (Estudios Cardiologicos de Latino-America) EVGN (European Vascular Genomics Network) VI Framework Program, Unione Europea JW Goethe University, Department of Cardiology, Frankfurt, Germany Karolinska Institutet, Stockholm, Sweden PHRI (Population Health Research Institute), McMaster University, Hamilton, Ontario, Canada 123

126 SIOP (International Society of Paediatric Oncology) Europe University of Minnesota, Minneapolis, USA University of Oslo, Norway Wellcome Trust Centre for Human Genetics, University of Oxford, UK WONCA (World Organization of Family Doctors), Europe EDITORIAL BOARD MEMBERSHIP Circulation, International Journal of Health Services, Journal of Clinical Epidemiology (Gianni Tognoni) Journal of Cardiac Failure, Journal of Cardiovascular Medicine (Roberto Latini) Assistenza Infermieristica e Ricerca, European Journal of Cancer Care, European Journal of Oncology Nursing, International Nursing Perspectives (Paola Di Giulio) PEER REVIEW ACTIVITIES American Heart Journal, Atherosclerosis Thrombosis and Vascular Biology, Cardiovascular Research, Circulation, Circulation Research, Clinical Pharmacology and Therapeutics, European Heart Journal, European Journal of Cardiovascular Nursing, International Journal of Cardiology, International Journal of Obesity, Italian Journal of Cardiology, Journal of Cardiac Failure, Journal of Internal Medicine, Journal of Cardiovascular Medicine, Life Sciences, The Lancet, PharmacoEconomics. NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Ethical Committee of Lombardy Region Ethical Committee of the Provincia of Verona Agenzia Italiana del Farmaco, Direzione Generale Farmaci e Dispositivi Medici EVENT ORGANIZATION Investigator's Meeting - Stato di avanzamento dello studio RE-LY 02/03/07, Boheringer Ingelheim, Reggello (Firenze) Master di I Livello in Ricerca Clinica dell Università degli Studi di Milano, Facoltà di Medicina e Chirurgia, Dipartimento di Medicina Interna. Lezioni di Metodologia e Statistica: 16/04/07 La ricerca clinica oggi: profit e no-profit. Il protocollo dello studio 17/04/07 Il disegno degli studi clinici 18/04/07 Legislazione sulla sperimentazione clinica e ruolo dei Comitati Etici 26/04/07 Statistica descrittiva 08/05/07 Statistica inferenziale (1) 124

127 10/05/07 Statistica inferenziale (2) 16/05/07 Monitoraggio degli studi clinici 22/05/07 Analisi della sopravvivenza Istituto di Ricerche Farmacologiche Mario Negri, Milano Investigator's Meeting - Stato di avanzamento dello studio Prono/Supino II 01/06/07, Sala Gialla, Milano Convention Centre-MIC, SMART Fiera, Milano Investigator's Meeting Valutazione di differenti strategie anestesiologiche per interventi neurochirurgici sopratentoriali "NeuroMorfeo" 14/11/07, Aula E, Istituto di Ricerche Farmacologiche Mario Negri, Milano PARTICIPATION IN EVENTS IN WHICH THE DEPARTMENT WAS INVOLVED American College of Cardiology. 56th Annual Scientific Session, 24-27/03/07, New Orleans, USA - Microalbuminuria in chronic heart failure: prevalence and determinants in more that 2000 patients enrolled in the GISSI-HF trial - Brain natriuretic peptide and severity of diastolic dysfunction: interaction with LV geometry and function in mild-moderate heart failure. The AREA-IN-CHF Study IUSS Ferrara Università degli studi di Ferrara Master di II Livello, Scuola di Ricerca Clinica ed Epidemiologica, 28/03/07, Università di Ferrara, Italy - Introduzione alla farmacocinetica - Farmacocinetica di fattori endogeni e biomarcatori Annual Meeting of the Norwegian Society of Cardiology. Kardiologisk Varmote 2007, Radisson SAS Plaza Hotel, 3-5/05/07 Oslo, Norway - Changes over time vs single measurements of probnp in risk stratification and monitoring of patients with chronic heart failure. Recent data with use of Roche TnT in relation to HF European Society of Cardiology. Annual Meeting of the Working Group: Myocardial Function Adverse Cardiac Remodelling: mechanism and repair, 4-6/05/07, Kracow, Polonia - Cardiac mesoangioblasts are committed, self renewable progenitors, associated with small vessels of adult mouse ventricle - Modulation of homing and survival of cardiac mesoangioblasts transplanted in the infarcted mouse heart Roche Diagnostics GmbH. procardio Symposium. 5th International Symposium on NTproBNP. NT-proBNP and other biomarkers for the management of cardiac patients, 11-12/05/07, Seville, Spain - The prognosti value of high sensitive troponin T in patients with stable chronic heart failure. Data from the Val-HeFT trial Convegno Regionale ANCE Lombardia. Il cuore nella donna. Un appuntamento annuale. 26/05/07, Hotel Michelangelo, Milano, Italy - Scarsa presenza femminile nella popolazione dei trial rispetto al mondo reale. Rischio di un indagine deformata e possibili correzioni 125

128 Associazione Nazionale Medici Cardiologi Ospedalieri. XXXVIII Congresso Nazionale di Cardiologia, 3-5/06/07, Firenze, Italy - Le evidenze sperimentali - Marcatori biologici circolanti in pazienti con bronco-pneumopatia cronica ostruttiva ed insufficienza cardiaca sintomatica. Dati dello studio GISSI-HF - La microalbuminuria nell insufficienza cardiaca: prevalenza e correlati clinici in più di 2000 pazienti dello studio GISSI-HF - Implementazione di un protocollo di infusione di insulina a gestione infermieristica per il controllo intensivo della glicemia nel diabete con sindrome coronarica acuta - Validazione di un protocollo standardizzato per la transizione della terapia insulinica infusiva a quella sottocutanea nel diabetico con sindrome coronarica acuta Heart Failure Association of the European Society of Cardiology. Heart Failure Congress 2007, 9-12/06/07, Hamburg, Germany - Prevalence and significance of microabuminuria in a large population of patients with chronich heart failure. Data from the Italian GISSI-HF trial - Metabolic syndrome in chronic heart failure: prevalence and relation to clinical features and aetiology - Cardioprotection by erythropoietin and its derivatives emeeting. Cardiac Stem Cells: what we need for clinical translation, 27-28/06/07, Roma, Italy - Cardiac mesoangioblasts are committed, self renewable progenitors, associated with smal vessels of adult mouse ventricle - Modulation of homing and survival of cardiac mesoangioblasts transplanted in the infarcted mouse heart European Society of Cardiology. ESC Congress 2007, 1-5/09/07, Vienna, Austria - Circulating biomarkers in patients with chronic obstructive pulmonary disease and symptomatic heart failure. Data from the GISSI-HF trial European Vascular Biology Organization. 4th European Meeting on Vascular Biology and Medicine, 17-20//09/07, Bristol, United Kingdom - Workshop V, Vascular Progenitors 2 WONCA, CNGE. 13th WONCA 2007 Europe Conference 1er Congrès de la Médicine Générale en France. Re-thinking primary care in the European context. A new challenge for General Pratice, 17-20/10/07, Paris, France. - Feasibility of a large scale randomised trial in General Practice: The Risk & Prevention Study Società Italiana Attività Regolatorie, Società Italiana di Farmacia Ospedaliera, Società di Scienze Farmacologiche Applicate. La sperimentazione clinica dei farmaci e dispoistivi medici. I corso di aggiornamento, 18-19/10/07, Roma, Italy - Gli studi no-profit Azienda Ospedaliera S. Giovanni Addolorata. I primi 30 giorni dopo l infarto, 19-20/10/07, Roma, Italy - Ruolo della inibizione neurormonale (betabloccanti inibizione del sistema renina angiotensina aldosterone) Casa di Cura Polispecialistica Presidio Ospedaliero ASL 22, Regione Veneto. Strategie di prevenzione cardiovascolare, 20/10/07, Peschiera del Garda (VR), Italy - Terapia cellulare cardiaca. A che punto siamo? 126

129 American Heart Association. AHA Scientific Session 2007, 4-7/11/07, Orlando, Florida - Levels of circulating progenitor cells in patients with chronic heart failure predict outcomes Clinical Forum Srl, Next Stop Knowledge, 9-10/11/07, Istituto di Ricerche Farmacologiche Mario Negri, Milano, Italy - Statine in studi di popolazione e nel cardiopatico non ischemico con insufficienza cardiaca: dati disponibili - Sottoprogetti del GISSI-HF: obiettivi e dati basali Regione Toscana Consiglio Regionale Pianeta Galileo - La sperimentazione dei farmaci sull uomo, 17/11/07, Lucca, Italy - La sperimentazione dei farmaci sull uomo European Section of the Aldosterone Council ESAC Meeting st International Meeting of the French Society of Hypertension, 13-14/12/07, Paris, France - Effects of canrenone on BNP, PIIINP and LV remodeling in patients with mild chronic heart failure (AREA-IN-CHF study): final results. GRANTS AND CONTRACTS AIFA (Agenzia Italiana del Farmaco), AstraZeneca, Azienda Ospedaliera San Gerardo Monza, Chiesi Farmaceutici, Boehringer Ingelheim Italia SpA, BRAHAMS AG, Collegio Interprovinciale Milano-Lodi, Fondazione Cariplo, Fondazione Don Gnocchi Milano, Fondazione San Raffaele Milano, Heart Care Foundation, Istituto Auxologico di Milano, Istituto Nazionale Neurologico C. Besta, Kinetic Concepts Inc., Ministero della Salute, Novartis Pharma, Oslo University, Oxford University, Population Health Research Institute-Mc Master University, Pfizer Italia, Roche Diagnostics, Sigma Tau, SPA Società Prodotti Antibiotici S.p.A., Takeda Italia S.p.A. 127

130 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Abdulla J, Barlera S, Latini R, Kjoller-Hansen L, Sogaard P, Christensen E, Kober L, Torp-Pedersen C A systematic review: Effect of angiotensin converting enzyme inhibition on left ventricular volumes and ejection fraction in patients with a myocardial infarction and in patients with left ventricular dysfunction Eur J Heart Fail 2007; 9: Barlera S, Specchia C, Farrall M, Chiodini BD, Franzosi MG, Rust S, Green F, Nicolis E, Peden J, Assmann G, Collins R, Hamsten A, Tognoni G, PROCARDIS Consortium Multiple QTL influence the serum Lp(a) concentration: a genome-wide linkage screen in the PROCARDIS study Eur J Hum Genet 2007; 15: Boccanelli A, Cacciatore G, Mureddu GF, De Simone G, Clemenza F, De Maria R, Di Lenarda A, Gavazzi A, Latini R, Masson S, Porcu M, Vanasia M, Gonzini L, Maggioni AP, AREA IN-CHF Baseline characteristics of patients recruited in the AREA IN-CHF study (Antiremodelling Effect of Aldosterone Receptors Blockade with Canrenone in Mild Chronic Heart Failure) J Cardiovasc Med 2007; 8: Chiodini B, Franzosi MG, Barlera S, Signorini S, Lewis CM, D'Orazio A, Mocarelli P, Nicolis E, Marchioli R, Tognoni G, GISSI, SIBioC-GISSI Prevenzione Group Apolipoprotein E polymorphisms influence effect of pravastatin on survival after myocardial infarction in a Mediterranean population: the GISSI-Prevenzione study Eur Heart J 2007; 28: Cicoira M, Maggioni AP, Latini R, Barlera S, Carretta E, Janosi A, Soler Soler J, Anand I, Cohn J, Val-HeFT Investigators Body mass index, prognosis and mode of death in chronic heart failure: Results from the Valsartan Heart Failure Trial Eur J Heart Fail 2007; 9: Evangelista V, De Berardis G, Totani L, Avanzini F, Giorda CB, Brero L, Levantesi G, Marelli G, Pupillo M, Iacuitti G, Pozzoli G, Di Summa P, Nada E, De Simone G, Dell'Elba G, Amore C, Manarini S, Pecce R, Maione A, Tognoni G, Nicolucci A Persistent platelet activation in patients with type 2 diabetes treated with low doses of aspirin J Thromb Haemost 2007; 5: Fiordaliso F, De Angelis N, Bai A, Cuccovillo I, Salio M, Serra DM, Bianchi R, Razzetti R, Latini R, Masson S Effect of Beta-adrenergic and renin-angiotensin system blockade on myocyte apoptosis and oxidative stress in diabetic hypertensive rats Life Sci 2007; 81: Grasselli G, Gattinoni L, Kavanagh B, Latini R, Laupacis A, Lemaire F, Pesenti A, Suter P, Slutsky A, Tognoni G Feasibility, limits and problems of clinical studies in Intensive Care Unit Minerva Anestesiol 2007; 73: Krum H, Latini R, Maggioni AP, Anand I, Masson S, Carretta E, Ingrilli' F, Pettinati G, Glazer R, Tognoni G, Cohn J Statins and symptomatic chronic systolic heart failure. A post-hoc analysis of 5010 patients enrolled in Val-HeFT Int J Cardiol 2007; 119: Latini R, Masson S, Anand I S, Missov E, Carlson M, Vago T, Angelici L, Barlera S, Parrinello G, Maggioni AP, Tognoni G, Cohn J N, Val-HeFT Investigators Prognostic value of very low plasma concentrations of troponin T in patients with stable chronic heart failure Circulation 2007; 116: Sarto P, Balducci E, Balconi G, Fiordaliso F, Merlo L, Tuzzato G, Pappagallo GL, Frigato N, Zanocco A, Forestieri C, Azzarello G, Mazzucco A, Valenti M T, Alborino F, Noventa D, Vinante O, Pascotto P, Sartore S, Dejana E, Latini R Effects of exercise training on endothelial progenitor cells in patients with chronic heart failure J Card Fail 2007; 13: Saunders CL, Chiodini B, Sham P, Lewis CM, Abkevich V, Adeyemo AA, de Andrade M, Arya R, Berenson GS, Blangero J, Boehnke M, Borecki IB, Chagnon YC, Chen W, Comuzzie AG, Deng HW, Duggirala R, Feitosa MF, 128

131 Froguel P, Hanson RL, Hebebrand J, Huezo-Dias P, Kissebah AH, Li W, Luke A, Martin LJ, Nash M, Ohman M, Palmer LJ, Peltonen L, Perola M, Price RA, Redline S, Srinivasan SR, Stern MP, Stone S, Stringham H, Turner S, Wijmenga C, Collier DA Meta-analysis of genome-wide linkage studies in BMI and obesity Obesity 2007; 15: Silletta MG, Marfisi RM, Levantesi G, Boccanelli A, Chieffo C, Franzosi MG, Geraci E, Maggioni AP, Nicolosi GL, Schweiger C, Tavazzi L, Tognoni G, Marchioli R, GISSI-Prevenzione Coffee consumption and risk of cardiovascular events after acute myocardial infarction. Results from the GISSI (Gruppo Italiano per lo Studio della Sopravvivenza nell'infarto miocardico)-prevenzione trial Circulation 2007; 116: Staszewsky L, Wong M, Masson S, Barlera S, Carretta E, Maggioni AP, Anand IS, Cohn JN, Tognoni G, Latini R, Valsartan Heart Failure Trial Investigators Clinical, neurohormonal, and inflammatory markers and overall prognostic role of chronic obstructive pulmonary disease in patients with heart failure: data from the Val-HeFT Heart Failure Trial J Card Fail 2007; 13: LAY PRESS SELECTION PUBLISHED IN 2007 Avanzini F, Marelli G, Donzelli W, Sorbara L, Palazzo E, Bellato L, Colombo E, De Martini M, per il Gruppo di Studio DDD Protocollo di infusione di insulina a gestione infermieristica per il controllo intensivo della glicemia. G It Diabetol Metab 2007; 27: Clavenna A, Bonati M, Campi R, Labate L, Nobili A, Pasina L, Tettamanti M, Monesi L, Marzona I, Roncaglioni M C, Bortolotti A, Fortino I, Locateli G W, Giuliani G La prescrizione di farmaci per i bambini e gli anziani nella ASL di Lecco Ricerca & Pratica 2007; n.135: Marchioli R, Marfisi R M, Borrelli G, Chieffo C, Franzosi M G, Levantesi G, Maggioni A P, Nicolosi G L, Scarano M, Silletta M G, Schweiger C, Tavazzi L, Tognoni G Efficacy of n-3 polyunsaturated fatty acids according to clinical characteristics of patients with recent myocardial infarction: insights from the GISSI-Prevenzione trial J Cardiovasc Med 2007; 8: s34-s37 Masson S, Latini R An update on N-terminal pro-brain-type natriuretic peptide for risk stratification in chronic heart failure Eur Cardiov Dis 2007; 1: Masson S, Latini R, Tacconi M T, Bernasconi R Incorporation and washout of n-3 polyunsaturated fatty acids after diet supplementation in clinical studies J Cardiovasc Med 2007; 8 suppl 1: s4-s10 129

132 RESEARCH ACTIVITIES Laboratory of Cardiovascular Clinical Pharmacology The effects of mesoangioblasts and of different progenitor cells on injury after experimental myocardial infarction in the mouse Many studies have demonstrated that autologous and homologous cells of various origins can repair myocardium damaged due to an acute ischemic insult. Mesangioblasts are potentially interesting when compared with bone marrow precursors because (a) they are easily expanded and (b) obtainable by a biopsy of skeletal muscle in man. Mesoangioblasts isolated from human heart biopsies migrate and home in the heart of immunodeficient mice after coronary ligation, and they survive for at least 4 weeks. Studies are ongoing to establish their protective and regenerative potential, as a basis for possible clinical studies. The same model of coronary ligationin immunodeficient mice is being used for testing in vivo the angiogenic potential of other progenitor cells such as mononuclear cells Tie-2+ (collaboration with Michele De Palma, HSR, Milano), CD34+ (collaboration with Franca Fagioli,Osp S. Anna, Torino), CD133+ (collaboration with Giulio Pompilio, Centro Cardiologico Monzino, Milano). This research is partly done within the sixth EU program, European Vascular Genomics Network (EVGN, Characterization of the cardiac phenotype of PTX3 in knock-out mice PTX3 has been shown to have a non redundant role in reproduction and in the resistance to fungal infection (Aspergillus fumigatus). However, its role in the cardiovascular apparatus has not been well defined, though it is an independent prognostic marker after acute myocardial infarction in man. The role of PTX3 in myocardial infarction appears to be protective, in fact PTX3-null mice had larger infarct size than mice wild type. The observation can be explained by the decrease in inflammatory response induced by PTX3 through complement cascade. Therapeutic potential of stem cell therapy in animal models of diabetic cardiomyopathy Diabetes is a major risk factor for the development of various cardiovascular complications. Morbidity and mortality associated with diabetes are essentially related to vascular lesions. The accumulation of extracellular matrix in the diabetic aorta is involved in increases in wall stiffness and eventually in the development of cardiac dysfunction. Neural Stem Cells (NSC) are multipotent cells which have the ability to differentiate also to endothelial progenitors cells (EPCs). Decreased levels of EPCs have been reported in both type 1 and type 2 diabetic patients. The large decrease in elastin/collagen ratio, index of aortic elasticity, and altered angiogenesis found in diabetic rats in our study could reflect the progressive deterioration of vascular structure and function that may lead to turbulent blood flow and eventually hampering the hemodynamic function. We observed, for the first time, that treatment with mouse NSC attenuated the remodeling of the arterial extracellular matrix and restored the correct angiogenic response, and eventually attenuated the mild hypertensive state of diabetic rats suggesting a potential role of these cells in reducing the vascular dysfunction in diabetes. A new study has been already planned to evaluate the mobilization and homing of endogenous progenitor cells (EPCs) and exogenous (mesoangioblast) progenitor cells injected into the left ventricle chamber in a genetic model of obesity and type II diabetes, and by this way their contribution to the regeneration of myocardium and blood vessels after an experimental myocardial infarction. 130

133 Pulmonary injury by hydrochloric acid in the mouse: a model of Aspiration pneumonitis to test protective interventions Aspiration pneumonitis (AP) occurs when the acid content of the stomach makes his way through the larynx in the lower respiratory tract. Patients with consciousness disturbance are at risk for this event. Specifically, it has been shown that Pulmonary Aspiration can complicate between % general anesthesia procedures. The main injurious mechanism of AP is the chemical insult due to the low ph of gastric secretions, which causes a chemical burn of the airway tree and of the alveolar structures. The course of AP can be extremely variable, ranging from the silent aspiration characterized by a modest desaturation to the dramatic sequelae of Acute Lung Injury (ALI) and Acute Respiratory Distress Syndrome (ARDS), requiring prolonged mechanical ventilation and potentially leading to death. The purpose of this research is to establish and characterize under different aspects a murine model of regional AP, allowing a two week survival of the animals and the evaluation of the long term evolution of the injury by multiple tests, computed tomography, biochemical assays, histological evaluation, lung function and gas-exchange. Conditional transgenic model of cardiac HGF expression to promote neovascularization and to recruit stem cells in the myocardial infarction Growth factors such as hepatocyte growth factor (HGF) through angiogenic and anti-apoptotic effects may promote cardiac repair after myocardial infarction and in heart failure. The cardiacspecific α-myosin Heavy Chain (MHC-α) transactivator was used to direct expression of HGF to cardiomyocytes: by this way the effects of HGF can be tested under cardiac ischemia and reperfusion, without the need for administration of exogenous HGF. The Heart-HGF transgenic model is being used to verify the ability of HGF in vivo to promote neovascularisation, to protect cardiomyocytes from apoptosis, to recruit and activate endogenous or transplanted stem cells and to sustain their cellular replication and differentiation into cardiomyocytes after ischemic damage and reperfusion. Roles of macrophages in cardiac ischemia/reperfusion injury and in cardiac repair Macrophages either resident or from blood-borne monocytes play several key roles in the response of the heart to ischemic injury. They may be useful in particular during cardiac repair, when collagen deposition occurs and neonagiogenesis, stimulated by growth factors produced by macrophages. The aim of the project is to assess the relevance of macrophages in myocardial repair and scar formation after myocardial infarction, in the attempt to dissect the role of inflammatory cells in myocardial injury vs repair. The effects of coronary ligation (with/without reperfusion) will be tested in mice in which machrophages can be ablated by administration of Difteria toxin: transgenic mice expressing human difteria toxin receptor (CD11b-DTR) will respond to the toxin. Since machrophages peak in mouse heart on day 1 after MI and return to baseline by day 7, the ablation should last no more than 7 days coronary ligation and reperfusion. The heart will be evaluated histologically for infarct size, myocyte loss, immunostain for machropages and PMN,capillary density (angiogenic response to ischemia). Cardiac function will be evaluated by echocardiography at week 6-8. Other experiments might be envisaged to assess the role of machrophages in modulating homing and/or retention of stem cells administered to mice with infarction. GISSI-HF: biohumoral sub-study and microalbuminuria The clinical trial GISSI-HF (follow up has been completed by 2007) is designed to assess whether two treatments (a statin and n-3 polyunsaturated fatty acids or PUFA) can improve the prognosis of patients with heart failure of any etiology, with preserved or compromised left 131

134 ventricular ejection fraction. A biohumoral substudy aims at exploring the pathophysiology of heart failure and mechanisms of action of the treatments in study. In 1200 patients enrolled in the substudy, blood samples are collected at enrolment and after three months to measure plasma PUFA, and markers of ventricular myocyte stress (brain natriuretic peptides, BNP and N-terminal probnp), myocardial damage (cardiac troponin T, measured with the standard assay or with a newly developed high sensitive method) and inflammation (C-reactive protein, CRP, pentraxin-3, PTX3). Baseline fraction of n-3 PUFA averages 3.4 and increases by about 30% over 3 months. Baseline BNP concentration in 1223 patients is 141 pg/ml (median) and correlates with the severity of heart failure. The number of circulating endothelial progenitor has been measured in a smaller group of 68 patients, in collaboration with the laboratory of E. Dejana (IFOM). Number of endothelial progenitor cells in patients with heart failure is 30-50% lower than in healthy volunteers was inversely correlated to morbidity/mortality in a population of patients from 5 Italian centers and a Cardiology center in Frankfurt (collaboration with Andreas Zeiher). Microalbuminuria (defined as the ratio between urinary concentrations of albumine and creatinine) is being measured in more than 2000 patients enrolled in the GISSI- HF trial as an indicator of renal endothelial dysfunction. Microalbuminuria (albumine/creatinine = mg/g) is present in 19% of the patients and is associated to inflammation, endothelial dysfunction and neurohormonal activation. Novel and more stable circulating biomarkers, have been successively assayed. They belong to the family of natriuretic peptides (mid-regional proatrial natriuretici peptide, MR-proANP), endothelin (C-terminal pro-endothelin-1, CT-proET-1), vasopressin (C-terminal pro-vasopressin, CT-proAVP) and adrenomedullin (mid-regional proadrenomedullin, MR-proADM). Other markers related to infection and innate immunity are currently measured. Anti-remodeling effect of aldosterone receptors blockade with canrenone in mild chronic heart failure: the AREA IN-CHF The RALES study has shown that spironolactone, an aldosterone receptor antagonist, reduces the risk of morbidity and mortality both from progressive heart failure (HF) and sudden death in patients with moderate to severe HF. Aldosterone antagonists may be effective because they oppose the effects of aldosterone on sodium retention, loss of magnesium and potassium, sympathetic activation, vascular compliance and cardiac fibrosis. In AREA IN-CHF, a multicentre randomized, placebo-controlled clinical trial, the effect of canrenone, a metabolite of spironolactone, is assessed in patients with mild chronic HF (n= 500) in terms of echocardiographic left ventricular remodeling and a combined end-point of mortality and morbidity. The design of the study includes blood collection in all patients at randomization and after 6 months to determine the plasma concentration of aldosterone, brain natriuretic peptide and a circulating marker of collagen turnover (propeptide N-terminal of collagen III). These biohumoral factors are assayed centrally at the Department of Cardiovascular Research and should give insight on the effect of canrenone on these markers and on their relation with study outcome. There is no significant difference between canrenone and placebo for the primary endpoint, changes in end-diastolic left ventricular volume. On the other hand, canrenone improves systolic function and reduces left ventricular mass compared to placebo. BNP concentration, but not PIIINP, is drastically reduced in patients randomized to canrenone. PTX-3, a novel long pentraxin is an marker of severity of disease and of outcome in cardiovascular diseases, independent of C-reactive protein PTX-3 is a novel long pentraxin whose expression is induced by cytokines in endothelial and mononuclear cells, mostly in striated muscle and heart, while C-reactive protein (CRP) is mainly synthesized in the liver. PTX3 was shown to peak in plasma around 7 h after onset of symptoms of MI and to be an independent predictor of 3-month mortality. PTX3 has been assayed with a more accurate method in 1200 patients with symptomatic heart failure (GISSI- 132

135 HF), and will be assayed in 350 patients with atrial fibrillation (GISSI-AF) and in 1400 patients with heart failure (CandHeart) to explore its role in other cardiovascular diseases. First results indicate that PTX3 is associated with different clinical characteristics in patients with heart failure, including advanced age, ventricular dysfunction, functional class (NYHA class), and comorbidities such as atrial fibrillation and diabetes. Echocardiographic and biohumoral substudy GISSI-AF trial The GISSI Atrial Fibrillation trial (GISSI-AF) tests the efficacy of Valsartan, an angiotensin II AT1-receptor blocker, in the prevention of atrial fibrillation recurrence in 1400 patients. A substudy of the GISSI-AF is currently ongoing to evaluate the potential role of biohumoral factors and cardiac structural remodeling in the reoccurrence and severity of atrial fibrillation. In approx. 400 patients three serial echocardiographic exams (at randomization, 6 months and 1 year) and contemporaneous blood collection are performed. Left ventricular and atrial dimensions will be determined by echocardiography, whereas plasma levels of BNP, NTproBNP, troponin T (high sensitive method), endothelin-1 and inflammatory markers (CRP, PTX3) will be measured. Besides giving clues on the pathophysiology of atrial fibrillation, the most common arrhythmia in elderly, this substudy aims at providing mechanical insights of the potential benefits of the study drug. The study was completed on January 31, Central readings of echocardiograms and assay of circulating biomarkers are ongoing. CandHeart: effects of candesartan on BNP and left ventricular function in patients with symptomatic heart failure Candesartan, an antagonist of angiotensin II type 1 receptors, significantly reduces mortality and morbidity in heart failure, as shown by the CHARM trials. The principal objective of the trial is to assess the effects Candesartan on circulating levels of brain natriuretic peptide (BNP) in patients suffering from CHF with depressed or preserved left ventricular (LV) systolic function. The patients (approx. 1400) will be followed for 1 year. Serial biohumoral and echocardiographic determinations will be performed at randomization, and after 3 and 12 months (end of study). Besides BNP, other biohumoral markers such as aldosterone, C-reactive protein, PTX3 and the microalbuminuria will be measured. The first patient was randomized in Dec About 450 patients have been randomized by the end of Trial Prone/Supine 2: effects of prolonged prone position on survival in severe acute respiratory distress syndrome (ARDS) This trials extends the findings of the previous Prone/Supine trial that showed in 304 patients with ARDS that 6-hour pronation in the first 10 days improved blood gas parameters, but did not reduce mortality (around 50% in the first 6 months). This was one of the rare example of a multicenter trial in Intensive Care performed entirely in Italy. The same group of Centers (headed by the Istituto di Anestesia e Rianimazione from the Policlinico di Milano, L Gattinoni with data management by Mario Negri) has launched a new trial that aims at evaluating the effect of a protocol of prolonged pronation (20 hours a day) in patients with severe ARDS, a subgroup that showed a trend towards improved survival on pronation in the previous study. Three hundred and twenty five patients out of the target of 340 have been enrolled in 20 centers; enrollment should be concluded by May 2008 and data analyzed by the statisticians at the Department of Cardiovascular Research. DyDa: left ventricular dysfunction in diabetes. Prevalence and incidence of left ventricular dysfunction in diabetics patients without clinical cardiac disease This is a prospective, multicentric, national and epidemiological trial aimed at evaluating the prevalence of left ventricular dysfunction (systolic or diastolic) in 1000 patients with type 2 133

136 diabetes mellitus but no clinical cardiovascular disease at enrolment. The incidence of left ventricular dysfunction will be monitored during a 2-year follow-up using ECG and echocardiography. The Biomarker Core Laboratory will evaluate the biohumoral profile of these patients at study entry, measuring the circulating levels of brain natriuretic peptide, C-reactive protein, microalbuminuria and glycated hemoglobin. Enrolment started in July 2006 will be concluded in March In a subgroup of patients from the study, levels of circulating progenitor cells and their angiogenic potential in vitro will be measured to assess whether they are associated to risk of developing cardiovascular disorders in diabetics. Albumin Italian Outcome Sepsis Study. The ALBIOS Study (AIFA) ALBIOS is a multicenter, controlled, randomized clinical trial that compares the efficacy of human albumin and a crystalloid solution for volume replacement in patients with severe sepsis or septic shock. The primary endpoint is survival at 28 and 90 days after enrolment. Secondary endpoints include the number of organ dysfunctions, severity of organ dysfunction (SOFA scale), and lengths of stay in (intensive care unit) ICU and in hospital. More than 150 ICU in Italy are expected to participate to this large study, coordinated by the Ospedale Maggiore Policlinico in Milan and the Consorzio Mario Negri Sud. In a substudy coordinated by the Laboratory of Cardiovascular Clinical Pharmacology from the Mario Negri Institute in Milan, a centralized blood bank will be collected to provide insights on the potential effects of albumin on markers of inflammation, infection, cardiac function and coagulation in about 700 patients. The first patient should be enrolled in the first semester of Clinical, biochemical and instrumental predictors of outcome in rehabilitation after cardiac surgery (MIUR) Due to the short length of hospital recovery in patients with cardiac disease, the period of rehabilitation becomes crucial and is indicated for almost all the cardiomyopathies, including myocardial revascularization and surgery of cardiac valves. The objective of this study is to evaluate the prognostic role of circulating biomarkers in 250 patients enrolled in 5 Centers for rehabilitation after cardiac surgery. A plasma bank will be collected under the responsibility of the Laboratory of Cardiovascular Clinical Pharmacology from the Mario Negri Institute to assay potential markers of interest. The project is coordinated by the Fondazione Don Gnocchi in Milan. The first patient should be enrolled in the first semester of PUFAxStatin: a clinical study on the interaction between statins and n-3 polyunsaturated acids (PUFA) on the lipid and inflammatory profile of diabetic patients at cardiovascular risk There are potential beneficial pharmacological interactions between statins and n-3 PUFA on lipid and inflammatory metabolism. The present study assesses the effects of these two drugs, alone or in combination on circulating biomarkers of inflammation and on the lipid profile of diabetic patients at high cardiovascular risk (hypertension, ischemic heart disease, smoking). A total of 123 patients have been enrolled. In collaboration with the Laboratory of Neuroimmunology, the effect of statin and n-3 PUFA on the ex-vivo production of proinflammatory cytokines by stimulated whole blood has been evaluated. First results indicate that the serum concentrations of LDL-cholesterol and triglycerides are significantly reduced in patients receiving the statin, alone or in association with n-3 PUFA. Markers of ventricular function (N-terminal probrain natriuretic peptide, NT-proBNP) and inflammation (C-reactive protein) are significantly improved in patients enrolled in the group treated with statin. There are no overt differences among the experimental groups for the other markers of inflammation and endothelial dysfunction. The study is conducted in collaboration with the Laboratory of Clinical Drug Evaluation. 134

137 Evaluation of different anesthesiological strategies for supratentorial neurosurgery. The NeuroMorfeo Study (AIFA) The aim of the study is to evaluate whether an anesthesia with volatile anesthetics is equivalent to endovenous anesthetics for elective supratentorial surgery. This is a multicenter, randomized, controlled and opened study, based on a design of equivalence for comparison of different anesthesiological strategies. Patients to be included (n= 393) will be selected in 14 neurosurgical centers in Italy for elective intracranial surgery with supratentorial lesions without signs of endocranial hypertension (range of age years). The biohumoral response to surgical stress will be measured as an indicator of homeostasis and neurovegetative status. The urinary excretion of catecholamines and cortisol and the plasma concentration of cortisol will be measured in a central laboratory. The study is coordinated by the San Gerardo Hospital in Monza and by the Department of Cardiovascular Research from the Mario Negri Institute. The first patient has been enrolled in December Laboratory of Clinical Drug Evaluation PROCARDIS: A genome-wide strategy to identify susceptibility loci in precocious coronary artery disease The PROCARDIS research programme, a genome-wide strategy to identify susceptibility loci in precocious coronary artery disease (CAD) supported by the 5th Framework Programme of the EC, was initiated as a collaboration between the universities of Oxford and Münster, the Karolinska Institute, the Mario Negri Institute with the support of the GISSI group, Oxagen, a biotechnology company, and AstraZeneca. The objectives of the first stage of this programme were to collect a minimum of 2000 affected sibling pairs (ASPs) and families with precocious CAD and to apply genome-wide linkage mapping techniques, by typing anonymous highly informative markers spaced throughout the genome, to identify chromosomal regions which are linked to the susceptibility to early-onset CAD. The study design is based on family ascertainment, to allow non-parametric linkage analyses (in ASPs). The PROCARDIS collected 2,036 CAD families from four European countries, in order to maximise the power of detecting genes that confer modest risks. A genome-wide linkage scan identified three promising regions for intensive study; one of the linked regions (Chromosome 17) was confined to families with multiple cases of myocardial infarction and was replicated in a second independent series of families. In addition the linkage scan confirmed a previously identified locus on Chromosome 2. These results demonstrate that novel CAD susceptibility genes are tractable to positional cloning which promises to lead to the identification of new molecular insights into this condition, and hopefully, new treatments. Additionally, the program has collected nuclear families (e.g. parent-offspring trios) for transmission-based tests of association for fine mapping by linkage disequilibrium analysis and testing of positional candidate genes. Extensive clinical and biochemical intermediate phenotype data have also been collected by bringing study subjects in to dedicated clinics; these data will allow mapping of known and novel quantitative risk factors which will be important in prioritising positional candidate genes at mapped chromosomal loci. The serum concentration of lipoprotein Lp(a), known to be highly heritable and associated with cardiovascular risk, was analyzed using a genome-wide linkage analysis. A highly significant linkage was confirmed at LPA locus on chromosome 6q27, as well as another region of significant linkage was detected on chromosome 13q These results provide a basis for further studies of candidate genes in these regions. Further quantitave phenotypes have been already analyzed (Body Mass Index) or are in progress (Homocisteine). The second stage of PROCARDIS, supported by the EC 6th Framework Programme, is conducting a large genome wide association study, where the patients with myocardial infarction enrolled in the first stage will be compared with control subjects to identify novel 135

138 candidate genes. The results will be replicated in different populations. The collaboration has been extended to include complementary expertise, as proteomics, bioinformatics, functional analysis. GISSI-HF: A large scale clinical trial testing the effects of n-3 PUFA and Rosuvastatin on mortality/morbidity of patients with symptomatic Congestive Heart Failure The GISSI (Gruppo Italiano per lo Studio della Sopravvivenza nell'insufficienza cardiaca) is a collaborative group endorsed by ANMCO (Associazione Nazionale Medici Cardiologi Ospedalieri) and by the Istituto Mario Negri, active from 20 years in the cardiovascular research field. The GISSI-HF is the fifth large scale clinical trial conducted by the Group. While pharmacological treatments specifically targeted to the cardio-circulatory system have been largely investigated, scanty controlled data are available concerning the role of dietary and metabolic approaches in the management/outcome of patients with heart failure. The GISSI-HF is a prospective, multicenter, randomized, double blind, placebo controlled study, with randomized allocation of patients with a clinical diagnosis of heart failure to: Randomization 1 (R1): n-3 PUFA vs corresponding placebo; Randomization 2 (R2): rosuvastatin vs corresponding placebo. The primary objective of the GISI-HF is to demonstrate that, in patients with heart failure treated at the best of recommended treatment, long term administration of n-3 PUFA and/or rosuvastatin is more effective than the corresponding placebo in the reduction of all-cause mortality and all-cause mortality or cardiovascular hospitalizations. The study follow-up will last at least 3 years. The study started in September 2002 with the participation of 364 departments of cardiology. The recruitment phase has been completed with more than 7000 patients randomized. Several substudies focus on possible mechanistic effects of the study treatments: ventricular remodeling (echo); biohumoral; genetic; arrhythmic and autonomic pattern (Holter monitoring); exercise capacity; cognitive function; burden of care. The follow-up was completed and final results are expected to be presented in The project is conducted in collaboration with the Laboratory of Cardiovascular Clinical Pharmacology. GISSI-HF Genetic Substudy The role of genetic factors in causes, evolution, prognosis and treatment of heart failure is largely unexplored, with the exception of heart failure originated by specific cardiomyopathies (such as dilated, hypertrophic, arrhythmogenic right ventricular cardiomyopathies), for which the role of heritable gene mutations is increasingly well understood. Heart failure (HF) is a syndrome with different etiologies, and more than one half is caused by coronary heart disease (CHD). A genetic substudy to GISSI-HF (see "ad hoc" section) offers the opportunity to improve knowledge on the role of genetic factors involved in heart failure, through a collection of blood samples of a large population of patients, involving cases of heart failure of different etiologies, i.e. non-ischaemic and ischaemic heart disease. The objective of the genetic substudy is 1) to assess the relationships between the polymorphysms of various candidate genes and the clinical outcome in patients enrolled in GISSI-HF study; 2) to assess whether these relationships are modified by the experimental treatments. The genetic markers to be studied will initially focus on polymorphisms of genes involved in lipid metabolism and inflammation. However, as new genetic information is being accumulated continually, at present it is not known if other polymorphisms/genes will be important at the time of study completion. The possibility to assay other allelic variants is then foreseen. The GISSI-HF genetic substudy is conducted by nearly 100 Centres that have included 2200 patients. 136

139 GISSI-Prevenzione-Genetic Study Myocardial infarction is a multifactorial disease. While the role of known risk factors on coronary heart disease susceptibility is well defined, the impact of the genetic components and its interaction with environmental factors need investigation. The GISSI-Prevenzione trial investigated the effects of pharmacological treatments with n-3 PUFA and pravastatin on morbidity and mortality after myocardial infarction. During the study more than 8000 samples of a large population of patients affected by this disease has been collected and stored with the collaboration of SIBioC (Società Italiana di Biochimica Clinica e Biologia Molecolare). The GISSI-Prevenzione-Genetic Study investigates the role of genetic factors in ischaemic heart disease. The objectives of the project are 1) to assess the relationships between the polymorphysms of various candidate genes and the clinical outcome in patients enrolled in the large clinical trial GISSI-Prevenzione study; 2) to assess whether these relationships are modified by the pharmacological treatments. According to these objectives, we investigated the relationship between APOE, mortality and the response to treatment in 3304 myocardial infarction survivors randomized to pravastatin or no treatment. We found that epsilon 4 allele is a determinant of pravastatin response in terms of survival. Though in the entire population investigated we found a beneficial effect of pravastatin in terms of survival, only the epsilon 4 carriers seemed to have gained a significant benefit from this treatment. We suggest that the effect of statins is of particular interest in this fraction of the population and that genetic markers can help in identifying patients that benefit more from statin treatment. Association studies in the same population on the adiponectin gene and C-reactive protein gene variants are in progress. GISSI-AF. Clinical trial testing the efficacy of an angiotensin II AT1- receptor blocker in Atrial Fibrillation The GISSI-AF is a randomized, prospective, parallel group, placebo-controlled, multicenter study on the use of valsartan, an angiotensin II AT1-receptor blocker in the prevention of Atrial Fibrillation (AF) recurrence. Atrial fibrillation is the most frequent form of arrhythmia in clinical practice, affecting 6% of people over 65 years old. The traditional therapies are often able to restore the sinus rhythm but are subject to a very high percentage of relapses, above all when the AF has been present for a long time and when there are structural changes at both atrial and ventricular level. The reninangiotensin-aldosterone system (RAAS) plays a key role in the remodeling phenomenon, which makes increasingly irreversible the electrical, mechanical and structural properties of the atrial tissue. Existing experimental and clinical data do not allow any definite conclusion regarding the efficacy of an angiotensin II AT1-receptor blocker in the prevention of AF. The GISSI group decided to conduct a specific trial of adequate size versus placebo aimed to assess if the addition of valsartan on top of established therapies can reduce the recurrence of atrial fibrillation in patients with a history of recent AF associated with cardiovascular diseases/comorbidities. The study has recruited and treated for 12 months 1400 patients. A biohumoral and echo substudy including 400 patients will be integral part of the main study focusing on how possible prognostic factors can be affected by the study treatment. According to the GISSI philosophy, GISSI-AF is a pragmatic trial, with broad selection criteria to mimic real clinical practice as much as possible. The final results are expected to be presented in The project is conducted in collaboration with the Laboratory of Cardiovascular Clinical Pharmacology. The Population Health Research Institute (PHRI), McMaster University, Hamilton, Ontario, is the coordinating center of a multinational network of cardiology clinics that collaborate to the 137

140 conduction of multicenter large scale clinical trials (nearly 40 Countries and more than 600 cardiology clinics). The Laboratory of Clinical Drug Evaluation is responsible for the scientific coordination in Italy of some of these trials (ACTIVE, RE-LY, CURRENT). ACTIVE study (Atrial Fibrillation Clopidogrel Trial with Irbesartan for prevention of Vascular Events) The aims of the study were to evaluate whether clopidogrel plus acetylsalicylic acid (ASA) is superior to ASA alone (A study) and non-inferior to standard oral anticoagulant therapy (W study) in preventing vascular events in patients with atrial fibrillation and to evaluate whether blood pressure lowering with irbesartan is superior to placebo in preventing vascular events in patients with atrial fibrillation (I study). The primary efficacy outcome is the first occurrence of stroke, myocardial infarction, vascular death over the duration of follow-up, (a minimum of 2 and maximum of 4 years approximately). A sample size of patients was planned, 6500 in the W study (testing the efficacy of warfarin vs ASA + clopidogrel) and 7500 in the A study (testing the efficacy of ASA + clopidogrel vs ASA alone). The W study was stopped early by the Data and Safety Monitoring Board in September 2005 after an interim analysis showing a significant difference in favor of warfarin over the combination of ASA + clopidogrel. The details of the ACTIVE W have been published (Lancet 2006; 367: ). The A and the I studies are continuing. RE-LY study (Randomized Evaluation of Long term anticoagulant therapy) Non-valvular atrial fibrillation is implicated in nearly 15% of strokes. Dose-adjusted warfarin decreases the risk of stroke by 62%. However, in practice, the risk of bleeding, the variability of anticoagulation intensity and the need of frequent monitoring and dose adjustments limits treatment with warfarin, leaving patients outside the therapeutic range almost half the time. Underuse of warfarin in patients with atrial fibrillation at high risk of bleeding calls for safer, more reliable alternatives. The direct thrombin inhibitor dabigatran could offer fixed oral dosing without need for coagulation monitoring, rapid onset and offset of action, stable pharmacokinetics with little potential for drug interactions, and no known food interactions. For these reasons an international multicentre, randomized, active controlled, parallel group, noninferiority, clinical trial (RE-LY study) was designed to evaluate the efficacy and safety of dabigatran etexilate compared with open label adjusted warfarin for the prevention of stroke and systemic embolism in patients with non-valvular atrial fibrillation. The recruitment of patients was completed and the long-term follow-up is ongoing. CURRENT OASIS-7 study OASIS 7 is a randomized, multinational, 2X2 factorial design, parallel-group, double-blind study, comparing a high loading dose regimen of clopidogrel versus standard dose and high dose regimen of aspirin versus standard dose, in patients with acute coronary syndrome managed with an early invasive strategy. The study involves approximately 14,000 patients in 800 clinical centers worldwide. The primary objective of the study is to determine whether a high dose regimen of clopidogrel is superior to a standard dose of clopidogrel in preventing CV death, myocardial infarction or stroke and to determine if high dose of aspirin is as safe as low dose in terms of TIMI major bleeding rate. Secondary objective is to evaluate the safety of the clopidogrel high dose regimen compared to the standard dose regimen in terms of TIMI major bleeding. INTER-HEART Study INTER-HEART is an epidemiological study sponsored by the World Health Organization and the World Heart Federation aimed to determine the association between risk factors and acute myocardial infarction within populations defined by ethnicity and/or geographic region, and to 138

141 assess the relative importance of risk factors across these populations. The project is coordinated by the PHRI, McMaster University, Hamilton, Canada. The Clinical Drug Evaluation Laboratory serves as National Coordination Office for Italy. INTER-HEART was conducted in 52 countries in Asia, Europe, Middle East, Africa, Australia, and North and South America, utilizing a standardized protocol. The study evaluated the importance of conventional and emerging risk factors within each geographic region, and whether their impact varies by region in a population of nearly individuals (cases with acute myocardial infarction and matched controls). The study collected data on demographic factors (country of origin, first language), socioeconomic status (education, occupation, income), lifestyle (tobacco use, physical activity, dietary patterns), and personal and family history of cardiovascular disease and risk factors. The INTER-HEART has documented that nine simple modifiable risk factors can account for over 90% of the population attributable risk for heart disease globally. More importantly, these risk factors appear to have similar predictability in all regions of the world, as well as in all ethnic groups. A further analysis published in the 2005 redefined obesity: traditional definitions have been based on Body Mass Index (BMI), but this paper very clearly showed that, irrespective of BMI, the waist-to-hip ratio (WTHR) is a far better predictor of myocardial infarction risk across diverse populations. The observations carry important implications for people and populations hitherto considered to be low risk on the basis of their BMIs. In a further analysis, the INTERHEART study established that all forms of tobacco exposure, such as smoking, chewing tobacco or inhaling second hand smoke, increase the risk of heart attack. Compared to people who had never smoked, smokers had a three-fold increased risk of a heart attack. An ad hoc assessment of the gender influence, comparing risk factors for acute myocardial infarction (MI) between women and men globally, the INTER-HEART showed that women experience their first acute MI on average 9 years later than men. Nine modifiable risk factors are significantly associated with acute MI in both men and women and explain greater than 90% of the PAR. The difference in age of first MI is largely explained by the higher risk factor levels at younger ages in men compared to women. The approach to prevention of MI in men and women can be based on similar principles in all regions of the world. Laboratory of General Practice Research Risk and Prevention Study (R&P) R&P is a study on the optimization of cardiovascular prevention of subjects at high risk performed at national level by General Practitioners. Study objective and design - Controlled clinical trial, double-blind and randomised, of the efficacy of a n-3 PUFA treatment in reducing the incidence of cardiovascular events, both fatal and non-fatal, in a population defined as at high risk by participating GPs. - Practicability and overall yield of the preventive interventions adopted (outcome study) The epidemiological and care history of this population shall form the object of a specific evaluation according to a plan of formal predefined analyses. Study population Inclusion criteria Among the subjects deemed by GPs to be at high cardiovascular risk, patients are selected if presenting: - multiple risk factors (e.g. hypertension, hypercholesterolemia, diabetes, smoking, family history of myocardial infarction, obesity, sex and old age) - previous cardio-cerebrovascular events or clinical manifestations of atherosclerotic disease (stroke, TIA, peripheral arteriopathy, previous arterial revascularisation procedures, angina 139

142 pectoris). Exclusion criteria - serious co morbidity with an unfavorable prognosis over the short term (e.g. cancer); - expected non-compliance over a long period of time; contraindications (known allergies to n- 3PUFA) - indications (previous MI) for treatment with n-3 PUFA. Efficacy measures The primary objective is to evaluate if a long-term administration of n-3 PUFA is more effective than placebo in reducing: overall mortality and major cardiovascular events (non fatal myocardial infarction and stroke) overall major cardiovascular events (cardiovascular mortality, non-fatal strokes and myocardial infarction); coronary-related mortality and sudden death. Randomisation is central, stratified by GP. The experimental treatment consists of one capsule containing 1g of n-3 PUFA, or the corresponding placebo, to be taken daily. The duration of follow-up is 5 years. In order to document with sufficient statistical reliability that the experimental treatment with n- 3 PUFA reduces the incidence of events [-15% total death, non fatal myocardial infarction and stroke; -20% cardiovascular death and non fatal myocardial infarction and stroke; -30% cardiovascular death; -40% sudden death] a total of 12,000 patients is required. Up-date of the study: From February 2004 to March ,521 patients have been enrolled by a network of 860 GPs. The Local Health Authorities involved are 57 and in each one investigator s meeting has been organized. The characteristics of the population so far enrolled are the following: mean age 65 years, males 62%, hypertension 79%, hypercholesterolaemia 62%, diabetes 56%, smokers 16%, obesity 35%, family history of premature myocardial infarction 20%. Twenty five% of patients have a clinical manifestation of atherosclerotic diseases, 50% have diabetes in association with another risk factor and 23% have multiple risk factors. The attributability of cardiovascular events in the GP s perception A cardiovascular event is the sudden manifestation - more or less expected - of clinical history, life-style habits, social environment, patient and medical attitudes and health strategies adopted. To carefully examine the reasons why a cardiovascular event occurred is part of a good care in the context of general practice. To render explicit this process for all the representative cases that occurred during a defined period of care could transform a usual attitude of good clinical practice into an epidemiology of the attributability. The aim of this study is to evaluate the epidemiology of the attributability of cardiovascular events through GPs perception. Clinical, socio-economic, psychological, risk factors and lifestyle variables will be investigated. The study started in October 2007 and a network 167 GPs are actively involved. Pharmaco-epidemiology studies on administrative databases: the Drugdrug interactions" Project This is a study aimed to develop and implement a database on drug-drug interaction to be put into a prescription computerised system of the Regione Lombardia (Italy) as a support for the general practitioners (GPs) to assess the risk of potential drug interactions. The study was also set up to assess the prevalence of drug-drug interaction in a sample of prescriptions of a cohort of GPs. A model for the epidemiological analyses of the drug prescriptions database of Lecco Local Health Authority has been developed. The analyses have been performed on the cohort of subjects > 65 years ( subjects, mean age of 75.2 years) on a total of prescription records. Eighty-six % of the subjects received al least one prescription per year, overall men were more treated than woman and the class of drugs most frequently prescribed was the 140

143 cardiovascular one (63.8% of the population). ). The prevalence of potentially severe drug-drug interactions was 16 %, and increased at rising of the patient s age and of the number or chronic drug prescribed. Epidemiological and clinical profile of diabetic patients in Lombardy Region using administrative databases. The study is part of an ongoing pharmacoepidemiological project in collaboration with the Health Department of the Lombardy Region. Its main objective is the definition of a model to assess and control the use of health resources of diabetic patients by means of integrated administrative database. Specific aims of the study are: To identify the diabetic population by specific drug consumption (ATC group = A10) To assess the co morbidities by co-administered cardiovascular and non-cardiovascular drugs To describe the diagnostic procedures by the assessment of laboratory, instrumental and specialist examinations To estimate the rate and causes of hospitalization by the analyses of hospital admission data The stratification of global cardiovascular risk in hypertensive patients of the district of Borbon Ecuador The Laboratory is involved in a collaborative project with the Cecomet (Centro de Epidemiologia comunitaria y Medicina tropical) in Esmeralda, Ecuador, on the prevalence and treatment of hypertension in the district of Borbon, a rural zone of Ecuador in the northern part of the country. In this area, 36% of the adult population is affected by hypertension and more than half of hypertensive patients present blood pressure levels > 160/110 mmhg. From 2001, in the District is ongoing an intensive follow-up of the hypertensive population with the following aims: to evaluate the global cardiovascular risk of the population, to better control blood pressure levels increasing the number of subjects treated with hypertensive therapy (in particular those at high cardiovascular risk) and monitoring of the clinical complications. Preliminary data show that: Patients treated with hypertensive therapy are increased from 39% to 59% Antihypertensive drugs are mainly prescribed to subjects with high blood pressure levels (80% of those with systolic blood pressure >180mmHg are actually under treatment) or at high cardiovascular risk (82%) Blood pressure control is improved (patients with systolic blood pressure levels > 180mmHg decreased from 33% to 24% and those with levels < increased from 26% to 34%) The fraction of patients at high or very high cardiovascular risk is decreased from 40% to 33% However, the compliance to antihypertensive treatment is still unsatisfactory since only half of the subjects are compliant with the prescribed therapy. Laboratory of Clinical Pharmacology Quality of Life, Depression and Cognitive problems in heart failure patients (QDF-GISSI-HF) The project is a sub-project of the GISSI-HF study. The aim of the study is to describe the evolution of depression, cognitive problems and the quality of life in a planned sample of 1500 heart failure patients; assess the use of common instruments that measure QDF variables and compare the assessment of the instrument (Geriatric Depression scale, Mini Mental State 141

144 Examination, Kansas City Cardiomiopathy Questionnaire) with the clinical perception of the nurses; describe if assessed or perceived patients' problems (low quality of life, high depression or compromised cognitive function) lead to any caring intervention. The baseline clinical characteristics of the 1564 patients included in the QDF study are closely comparable with those of the GISSI-HF population. The study instruments could be validly administered to the greatest majority of patients (KCQQ 97.2%, GDS 94.9%, MMSE 80.6% of patients >70 years). The nurses network nested in a major clinical trial, has produced one of the largest prospective cohort of HF patients who are comprehensively assessed and prospectively monitored, to allow an integrated evaluation of the relevance and implications of QDF measurements also on the clinical outcomes of this population. Epidemiology of nursing problems and drug-surveillance in nursing homes and home care The aim of the study is to describe problems nurses that require nursing decisions, and nurses encounter in the care of patients admitted to nursing home and home care, with specific attention to drug-related problems. In fact an adverse drug reaction is likely to be the cause of most unexpected problems that arise in elderly patients. The project has been completed and more than 350 nurses have been involved in 95 Nursing Homes and Districts. In the 6 observation days overall 1500 patients were observed. Overall 2224 problems were identified and 25% attributed to drugs and devices. Knowledge that patients have on their drugs at discharge from hospital The project is in its initial phase and involves several hospitals in the northen regions of Italy. Data on a first group of 163 patients from 6 hospitals were collected interviewing patients at discharge on their old and new therapies: 78% of patients is not informed on drugs/behaviours to avoid and 70% don t know side effects. No major differences were observed between old and new therapies. The data collection is still ongoing in other centres. 142

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146 DEPARTMENT OF MOLECULAR BIOCHEMISTRY AND PHARMACOLOGY STAFF Head Mario SALMONA, Sci.Prep.Alim.D., Ph.D. Laboratory of Biochemistry and Protein Chemistry Head Mario SALMONA, Sci.Prep.Alim.D.,Ph.D. Medical Biochemistry Unit Head Valentina BONETTO, Chem.Pharm.D. Synaptic Transmission Unit Head Marco GOBBI, Pharm.D. Laboratory of Molecular Biology Head Enrico GARATTINI, M.D. Pharmacogenomics Unit Head Maddalena FRATELLI, Biol.Sci.D. Gene Structure and Regulation Unit Head Mineko TERAO, Bioch.D., Ph.D. Laboratory of Receptor Pharmacology Head Tiziana MENNINI, Pharm.D. Laboratory of Neuroimmunology Head Pietro GHEZZI, Ph.D. Pharmacology of Septic Shock Unit Head Pia VILLA, Pharm.D. Metabolic Neuropathies Unit Head Roberto BIANCHI, Biol.Sci.D. Laboratory of Molecular Pathology Head Lavinia CANTONI, Biol.Sci.D. Laboratory of Systems Biology Head Gianfranco BAZZONI, M.D. 144

147 CURRICULA VITAE Mario Salmona obtained his doctorate degree in Biochemistry and Food Technology at the University of Milan in His background is in biochemistry, biophysics and pharmacology. His scientific interests relate to problems of human and animal diseases originating from the aberrant folding of proteins. In this context, a major portion of his studies was devoted to the etiopathogenesis and therapy of prion diseases. He has published over 200 articles on peer reviewed scientific journals Ph.D in Pharmacology, Mario Negri Institute 1975 Visiting Fellow in the Department of Biology of the Weizmann Institute of Science, Rehovot, Israel Head, Laboratory of Enzyme Research, Mario Negri Institute 1995 to date Dean of the School of Advanced Pharmacology, Mario Negri Institute 1997 to date Head, Department of Biochemistry and Molecular Pharmacology, Mario Negri Institute 2003 to date Member of the American Society of Biochemistry and Molecular Biology Referee for international scientific journals Selected publications Gerstmann-Straussler-Scheinker disease amyloid protein polymerizes according to the "dock-and-lock" model. Gobbi M, Colombo L, Morbin M, Mazzoleni G, Accardo E, Vanoni M, Del Favero E, Cantù L, Kirschner D A, Ceci P, Ubezio P, Manzoni C, Forloni G, Tagliavini F, Salmona M J Biol Chem. 2006; 281: Tetracycline and its analogues as inhibitors of amyloid fibrils: searching for a geometrical pharmacophore by theoretical investigation of their conformational behaviour in aqueous solution U. Cosentino, M.R.Varì, A.A. G. Saracino, D. Pitea, G. Moro, M. Salmona Journal of Molecular Modelling, 2005, 11: Role of Plasminogen in Propagation of Scrapie Salmona M., Capobianco R., Colombo L., De Luigi A., Rossi C., Mangieri M., Giaccone G., Quaglio E., Chiesa R., Donati M.B., Tagliavini F. and Forloni G. Journal of Virology 2005, 79: The role of platelet activating factor in prion and amyloid-beta neurotoxicity Bate C, Salmona M, Williams A Neuroreport 2004; 15: Squalestatin cures prion-infected neurons and protects against prion neurotoxicity. Bate C, Salmona M, Diomede L, Williams A. J Biol Chem. 2004; 279: Channels formed with a synthetic mutant prion protein PrP(82-146) homologous to a 7 kda fragment in diseased brain of GSS patients Bahadi R., Farrelly P.V., Kenna B.L., Kourie J.I., Tagliavini F., Forloni G., Salmona M. Am.J.Physiol. (Cell Physiology) 2003; 285: C862-C872 Gianfranco Bazzoni got his Medicine and Surgery degree in 1988 (at the University of Milan) and the specialisation in Pharmacological Research in 1992 (at the Mario Negri Institute, Milan). His area of expertise is cell biology, with focus on the processes of cell adhesion and migration Research Fellow, Mario Negri Institute Post-doctoral Fellow, Dana Farber Cancer Institute and Harvard Medical School, Boston, MA Research Scientist, Mario Negri Institute 2003 Head, Unit of Cell Adhesion, Mario Negri Institute 2004 to date Head, Laboratory of Systems Biology, Mario Negri Institute 2004 Regular Member of The American Physiological Society, Bethesda, MD Referee for international scientific journals Selected publications Paris L, Tonutti L, Vannini C, Buzzoni G. Structural organization of the tight junction. Biochim Biophys Acta 2007; (Epub ahead of print) Huang H, Cruz F, Bazzoni G. Junctional Adhesion Molecule-A Regulates Cell Migration and Resistance to Shear Stress. J Cell Physiol 2006; 209: Bazzoni G. Endothelial tight junctions: permeable barriers of the vessel wall. Thromb Haemost 2006; 95(1): Martinez-Estrada OM, Manzi L, Tonetti P, Dejana E, Bazzoni G. Opposite effects of Tumor Necrosis Factor and soluble fibronectin on Junctional Adhesion Molecule-A in endothelial cells. Am J Physiol (Lung Cell Mol Physiol) 2005; 288: L1081-L1088. Bazzoni G, Tonetti P, Manzi L, Cera MR, Balconi G, Dejana E. Expression of Junction Adhesion Molecule-A prevents spontaneous and random motility. J Cell Sci 2005; 118: Bazzoni G, Dejana E. Endothelial cell-to-cell junctions: molecular organization and role in vascular homeostasis. Physiol Rev 2004; 84(3):

148 Lavinia Cantoni obtained her degree in Biological Sciences in 1973 at the University of Milan. Then she specialized in pharmacological research at the Mario Negri Institute ( ). Research areas 1) biochemical-molecular mechanisms activated by oxidative stress 2) drug metabolism 3) porphyrias Post-doctoral Fellow, Medical Research Council, Toxicology Unit, Carshalton, UK (Winner of a Welcome Trust Research Fellowship) Research Scientist, Mario Negri Institute for short periods Visiting Scientist, Toxicology Unit, Carshalton, UK, and Cornell Medical Center, New York, USA Head, Unit of Heme and Hemoprotein Metabolism, Mario Negri Institute to date, Head, Laboratory of Molecular Pathology at the Mario Negri Institute. Member of the National Roll of Biologists and of the Italian Toxicology Society. Referee for international scientific journals, tutor for university degree theses and teacher in the Pharmacological Research Specialisation Course for graduates held at the Mario Negri Institute. Selected publications Rizzardini M., Chiesa R., Angeretti N., Lucca E., Salmona M., Forloni G., Cantoni L. Prion protein fragment differentially induces heme oxygenase-1 mrna in cultured neurons and astroglial cells. J.Neurochem. 68: , 1997 Rizzardini M., Zappone M., Villa P, Gnocchi P., Sironi M., Diomede L., Meazza C., Monshouwer M., Cantoni L. Kupffer cell depletion partially prevents hepatic heme oxygenase 1 messenger RNA accumulation in systemic inflammation in mice: role of interleukin 1 beta. Hepatology 27: , 1998 Cantoni L.,Valaperta R.,.Ponsoda X., Castell, J.V., Barelli D., Rizzardini M., Mangolini A., Hauri L., Villa P. Induction of hepatic heme oxygenase-1 by diclofenac in rodents: role of oxidative stress and cytochrome P-450 activity. J. Hepatology, 38: , 2003 Babetto E., Mangolini A., Rizzardini M., Lupi M., Conforti L., Poletti A., Rusmini P., Cantoni L. Tetracycline-regulated gene expression in the NSC-34-tTA cell line for investigation of motor neuron diseases. Mol. Brain Res. 140: 63-72, 2005 Rizzardini M., Lupi M., Mangolini A., Babetto E., Ubezio P., Cantoni L. Neurodegeneration induced by complex I inhibition in a cellular model of familial amyothrophic lateral sclerosis. Brain Res. Bull., 69: , 2006 Raimondi A., Mangolini A., Rizzardini M., Tartari S., Massari S., Bendotti C., Francolini M., Borghese N., Cantoni L., Pietrini G. Cell culture models to investigate the selective vulnerability of motoneuronal mitochondria to familial ALSlinked G93ASOD1. Eur. J. Neurosci. 24: , 2006 Enrico Garattini obtained his degree in Medicine and Surgery with full marks (110/110) in 1982 at the University of Milan. His scientific interests relate to problems of Cellular Biology and Molecular Biology Research Fellow of the National Research Council, Mario Negri Institute Postdoctoral Researcher at the Roche Institute of Molecular Biology, Department of Neurosciences Nutley, New Jersey, US Senior Researcher Regione Lombardia and Head of the Molecular Biology Unit, Mario Negri Institute 1997 to date Head, Laboratory of Molecular Biology, Mario Negri Institute From 2005 Dean, Advanced School of Pharmacology (Philosophy Doctor), Mario Negri Institute Member of the Editorial Board of the European Journal of Cancer and of Current Cancer Therapy Reviews Member of the American Society of Biochemistry and Molecular Biology (ASBMB) Selected publications Gianni M, Parrella E, Raska I Jr, Gaillard E, Nigro EA, Gaudon C, Garattini E, Rochette-Egly C. P38MAPK-dependent phosphorylation and degradation of SRC-3/AIB1 and RARalpha-mediated transcription. EMBO J Feb 22;25(4): Parrella E, Gianni M, Fratelli M, Barzago MM, Raska I Jr, Diomede L, Kurosaki M, Pisano C, Carminati P, Merlini L, Dallavalle S, Tavecchio M, Rochette-Egly C, Terao M, Garattini E. Antitumor activity of the retinoid-related molecules (E)-3-(4'-hydroxy-3'-adamantylbiphenyl-4-yl)acrylic acid (ST1926) and 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphthalene carboxylic acid (CD437) in F9 teratocarcinoma: Role of retinoic acid receptor gamma and retinoid-independent pathways. Mol Pharmacol Sep;70(3): Garattini E, Parrella E, Diomede L, Gianni M, Kalac Y, Merlini L, Simoni D, Zanier R, Ferrara F F, Chiarucci I, Carminati P, Terao M, Pisano C. ST1926, a novel and orally active retinoid-related molecule inducing apoptosis in myeloid leukemia cells: Modulation of intracellular calcium homeostasis. Blood 2004; 103: Vila R, Kurosaki M, Barzago M M, Kolek M, Bastone A, Colombo L, Salmona M, Terao M, Garattini E. Regulation and biochemistry of mouse molybdo-flavoenzymes. The DBA/2 mouse is selectively deficient in the expression of aldehyde 146

149 oxidase homologues 1 and 2 and represents a unique source for the purification and characterization of aldehyde oxidase. J Biol Chem 2004; 279: Kurosaki M, Terao M, Barzago M M, Bastone A, Bernardinello D, Salmona M, Garattini E. The aldehyde oxidase gene cluster in mice and rats: Aldehyde oxidase homologue 3, a novel member of the molybdo-flavoenzyme family with selective expression in the olfactory mucosa. J Biol Chem 2004; 279: Pisano C, Kollar P, Gianni M, Kalac Y, Giordano V, Ferrara F F, Tancredi R, Devoto A, Rinaldi A, Rambaldi A, Penco S, Marzi M, Moretti G, Vesci L, Tinti O, Carminati P, Terao M, Garattini E. Bis-indols a novel class of molecules enhancing the cytodifferentianting properties of retinoids in myeloid leukemia cells. Blood 2002; 100: Pietro Ghezzi Research Areas: Cytokines and inflammation; redox regulation : Researcher, Mario Negri Institute 1991 to date: Head, Laboratory of Neuroimmunology, Mario Negri Institute : Research Associate at Stanford University School of Medicine, Department of Genetics 2000 to date: Member, Kenneth Warren Laboratory, Ossining, NY (USA) Referee for international scientific journals Selected publications Leist M, Ghezzi P, et al.. Derivatives of erythropoietin that are tissue protective but not erythropoietic. Science Jul 9;305(5681): Fratelli M, Goodwin LO, Orom UA, Lombardi S, Tonelli R, Mengozzi M, Ghezzi Gene expression profiling reveals a signaling role of glutathione in redox regulation.proc Natl Acad Sci U S A Sep 27;102(39): Villa P, Bigini P, Mennini T, Agnello D, Laragione T, Cagnotto A, Viviani B, Marinovich M, Cerami A, Coleman TR, Brines M, Ghezzi P. Erythropoietin selectively attenuates cytokine production and inflammation in cerebral ischemia by targeting neuronal apoptosis. J Exp Med Sep 15;198(6): Siren AL, Fratelli M, Brines M, Goemans C, Casagrande S, Lewczuk P, Keenan S, Gleiter C, Pasquali C, Capobianco A, Mennini T, Heumann R, Cerami A, Ehrenreich H, Ghezzi P. Erythropoietin prevents neuronal apoptosis after cerebral ischemia and metabolic stress. Proc Natl Acad Sci U S A Mar 27;98(7): Laragione T, Bonetto V, Casoni F, Massignan T, Bianchi G, Gianazza E, Ghezzi P. Redox regulation of surface protein thiols: identification of integrin alpha-4 as a molecular target by using redox proteomics. Proc Natl Acad Sci U S A Dec 9;100(25): Tiziana Mennini got her degree in Pharmacy at the University of Milano (1975). In the same year she obtained a fellowship from the European Molecular Biology Organization, to learn sub-cellular fractionation techniques and synaptosomes utilization in neurochemistry, at the laboratory of Prof. VP Whittaker ( Stockholm, Sweden). In 1882 she spent a further period in Prof. Whittaker s laboratories (Max-Plank-Institut fur Biophysikalische Chemie, Abteilung Neurochemie Am Fassberg, Gottingen, Germany). She spent all her scientific career at the Mario Negri Institute: Research Assistant in the Laboratory of Drug Metabolism Chief of the Unit of Neurochemical Transmission, 1988 to date Chief of the Laboratory of Receptor Pharmacology Speaker, chairman and organizer at many congresses and courses, author of more than 200 articles published in international journals in the area of receptor pharmacology and neuropharmacology. Selected publications Beghi E, Bendotti C, Mennini T Merits of a new drug trial for ALS? Science 308: Gobbi M, Mennini T Is St John's wort a 'Prozac-like' herbal antidepressant? Trends Pharmacol Sci 22: The Italian ALSSG Ceftriaxone in amyotrophic lateral sclerosis. Eur J Neurol 3: Mennini T, Mocaer E, Garattini S Tianeptine, a selective enhancer of serotonin uptake in rat brain. Naunyn- Schmiedebergs Arch Pharmacol 336: Mennini T, Garattini S Benzodiazepines receptor binding in vivo: pharmacokinetic and pharmacological significance. Advances Biochemical Psychopharmacol 38: Mennini T, Bernasconi S, Manara L, Samanin R, Serra G The effect of intracerebral 6-hydroxy dopamine on 3Hreserpine binding to different brain regions of the rat. Pharmacol Res Commun 9:

150 Roberto Bianchi got his degree in Biological Sciences at University of Milan, Italy in Since 1975 he served as student teacher and supervisor at Mario Negri Institute and from 1989 to 1997 at Houston University. His main interests are diabetic complications (peripheral and autonomic neuropathies) and neurodegenerative disorders (Multiple Sclerosis, drugs induced neuropathies) Technician in the Laboratory of Biochemical Pharmacology, Mario Negri Institute Visiting Scientist in the Center for Neurosciences Behavioral Research, The Weizmann Institute of Science, Rehovot, Israel Research Assistant in the Laboratory of Biochemical Pharmacology, Mario Negri Institute Research Fellow in the Dept. Biochemical Biophysical Sciences, University of Houston, US Research Fellow in the Dept. Medicine, Case Western Reserve University, Cleveland, US Since 1996 Head, Unit of Metabolic Neuropathies, Mario Negri Institute Selected publications Bianchi R., Berti-Mattera L.N., Fiori M.G., Eichberg J.:Correction of altered metabolic activities in sciatic nerves of streptozotocin-induced diabetic rats. Effects of ganglioside treatment. Diabetes 39: (1990). Scarpini E., Bianchi R., Moggio M., Sciacco M., Fiori M.G., Scarlato G.: Decrease of nerve Na+,K+-ATPase activity in the pathogenesis of diabetic neuropathy. J. Neurol. Sci. 120: (1993). Conti G., Scarpini E., Baron P.L., Livraghi S., Tiriticco M., Bianchi R, Vedeler C., Scarlato G.: Macrophage infiltration and death in the nerve during the early phases of experimental diabetic neuropathy: a process concomitant with endoneurial induction of IL-1 and p75ntr. J. Nuerol. Sci. 195: (2002) Bianchi R., Buyukakilli B., Brines M., Savino C., Cavaletti G., Oggioni N., Lauria G., Borgna M., Lombardi R., Cimen B., Comelekoglu U., Kanik A., Tataroglu C., Cerami A., Ghezzi P. Erythropoietin both protects from and reverses experimental diabetic neuropathy. Proc Natl Acad Sci USA 101: (2004) Leist M., Ghezzi P., Grasso G, Bianchi R., Villa P., Fratelli M., Savino C., Bianchi M., Nielsen J., Gerwien J., Kallunki P., Larsen A.K., Helboe L., Christensen S., Pedersen L.O., Nielsen M., Troup L., Sager T., Sfacteria A., Erbayktar S, Erbayktar Z., Gokmen N., Yilmaz O., Cerami-Hand C., Xie, Q-W., Coleman T., Cerami A., Brines M. Erythropoietinderived tissue-protective cytokines that do not bind to the classical erythropoietin receptor. Science, 305(5681) , Savino C., Pedotti R., Baggi F., Furlan R., Ubiali F., Gallo B, Nava S., Bigini P., Barbera S., Fumagalli E., Mennini T., Vezzani A., Rizzi M., Coleman T., Cerami A.,Brines M., Ghezzi P., Bianchi R.Delayed administration of erythropoietin and its non-erythropoietic derivatives ameliorates chronic murine autoimmune encephalomyelitis. Journal of Neuroimmunology, 2005, E-pub December 6, 2005 Valentina Bonetto has got the degree in Pharmaceutical Chemistry and Technology at the University of Padua, Italy in She has got the Ph.D. in Medical Biochemistry and Biophysics at Karolinska Institutet, Stockholm, Sweden. Her principal lines of research are: 1) Study of the pathogenetic mechanisms at the basis of amyotrophic lateral sclerosis (ALS); 2) Identification of biomarkers of ALS; 3) Role of the oxidative modification in neurological disorders. These issues are investigated by different experimental approaches, including proteomics and mass spectrometry. Since 2000 she is at the Mario Negri Institute in the Laboratory of Biochemistry and Protein Chemistry, from 2002 is also Assistant Telethon Scientist at Dulbecco Telethon Institute. Since October 2007 she is the Head of the Unit of Medical Biochemistry inside the Laboratory of Biochemistry and Protein Chemistry. She is author of 26 publications from 1994 to 2007, in peer-reviewed journals. Among them she is first author in 11 and last author in 4. She is also author of 2 reviews. She is reviewer for scientific journals in the field of Proteomics and Neuroscience. Selected publications Massignan, T., Casoni, F., Basso, M., Stefanazzi, P., Biasini, E., Tortarolo, M., Salmona, M., Gianazza, E., Bendotti, C., Bonetto V. (2007) Proteomic analysis of spinal cord of presymptomatic amyotrophic lateral sclerosis G93A SOD1 mouse. Biochem. Biophys. Res. Commun., 353: Basso, M., Massignan, T., Samengo, G., Cheroni, C., De Biasi, S., Salmona, M., Bendotti, C., Bonetto, V. (2006) Insoluble mutant SOD1 is partly oligoubiquitinated in amyotrophic lateral sclerosis mice. J. Biol. Chem., 281: Ghezzi, P., Casagrande, S., Massignan, T., Basso, M., Bellacchio, E., Mollica, L., Biasini, E., Tonelli, R., Eberini, I., Gianazza, E., Dai, W.W., Fratelli, M., Salmona, M., Sherry, B., Bonetto. V. (2006) Redox regulation of cyclophilin A by glutathionylation. Proteomics, 6: Casoni, F., Basso, M., Massignan, T., Gianazza, E., Cheroni, C., Salmona, M., Bendotti, C., Bonetto, V. (2005) Protein nitration in a mouse model of familial amyotrophic lateral sclerosis: Possible multifunctional role in the pathogenesis. J. Biol. Chem., 280: Laragione, T., Bonetto, V., Casoni, F., Massignan, T., Bianchi, G., Gianazza, E., Ghezzi, P. (2003) Redox regulation of surface protein thiols: identification of integrin alpha-4 as a molecular target by using redox proteomics. Proc. Natl. Acad. Sci. U S A 100:

151 Casagrande, S.*, Bonetto, V.*, Fratelli, M., Gianazza, E., Eberini, I., Massignan, T., Salmona, M., Chang, G., Holmgren, A., Ghezzi, P. (2002) Glutathionylation of human thioredoxin: a possible crosstalk between the glutathione and thioredoxin systems. Proc. Natl. Acad. Sci. U S A 99, *These authors contributed equally to the study. Maddalena Fratelli got her degree in Biological Sciences at the University of Pisa and at the Scuola Normale Superiore di Pisa in Then the specialization in Pharmacological Research at the Mario Negri Institute in Her main fields of interest are: 1. High throughput genomic systems for the study of drug action and pharmacoresistance. 2. Redox regulation of protein function and gene expression: glutathionylation and gene expression profiling of glutathione dependent responses to oxidant challenge Postdoctoral Research Fellow in the Medical Research Council, Neurobiology Unit, Cambridge, UK. Since 1995, Head, Unit of Mediators of inflammation, Laboratory of Neuroimmunology, Mario Negri Institute Since 2005, Head, Unit of Pharmacogenomics, Laboratory of Molecular Biology, Mario Negri Institute Selected publications Fratelli M, Goodwin LO, Orom UA, Lombardi S, Tonelli R, Mengozzi M, Ghezzi P. Gene expression profiling reveals a signaling role of glutathione in redox regulation. Proc Natl Acad Sci U S A. 2005;102: Brines M, Grasso G, Fiordaliso F, Sfacteria A, Ghezzi P, Fratelli M, Latini R, Xie QW, Smart J, Su-Rick CJ, Pobre E, Diaz D, Gomez D, Hand C, Coleman T, Cerami A. Erythropoietin mediates tissue protection through an erythropoietin and common beta-subunit heteroreceptor. Proc Natl Acad Sci U S A. 2004; 101: Leist M, Ghezzi P, Grasso G, Bianchi R, Villa P, Fratelli M, Savino C, Bianchi M, Nielsen J, Gerwien J, Kallunki P, Larsen AK, Helboe L, Christensen S, Pedersen LO, Nielsen M, Torup L, Sager T, Sfacteria A, Erbayraktar S, Erbayraktar Z, Gokmen N, Yilmaz O, Cerami-Hand C, Xie QW, Coleman T, Cerami A, Brines M. Derivatives of erythropoietin that are tissue protective but not erythropoietic. Science. 2004; 305: Fratelli M, Minto M, Crespi A, Erba E, Vandenabeele P, Del Soldato P, Ghezzi P. Inhibition of nuclear factor-kappab by a nitro-derivative of flurbiprofen: a possible mechanism for antiinflammatory and antiproliferative effect. Antioxid Redox Signal. 2003; 5: Fratelli M, Demol H, Puype M, Casagrande S, Eberini I, Salmona M, Bonetto V, Mengozzi M, Duffieux F, Miclet E, Bachi A, Vandekerckhove J, Gianazza E, Ghezzi P. Identification by redox proteomics of glutathionylated proteins in oxidatively stressed human T lymphocytes. Proc Natl Acad Sci U S A. 2002; 99: Siren AL, Fratelli M, Brines M, Goemans C, Casagrande S, Lewczuk P, Keenan S, Gleiter C, Pasquali C, Capobianco A, Mennini T, Heumann R, Cerami A, Ehrenreich H, Ghezzi P. Erythropoietin prevents neuronal apoptosis after cerebral ischemia and metabolic stress. Proc Natl Acad Sci U S A. 2001; 98: Marco Gobbi got his degree in Pharmacy at the University of Milan, Italy, in Currently, his main fields of interest are: 1) the study of presynaptic mechanisms, such as neurotransmitter release/reuptake with a particular focus on plasma membrane transporters; and 2) the study of protein misfolding and aggregation and in particular the characterization of the prion protein amyloid formation, investigated by different approaches including surface plasmone resonance. Since 1981, Researcher in the Laboratory of Neuropharmacology and, from 1988, in the Laboratory of Receptor Pharmacology, Mario Negri Institute Starting from 1989 Chief, Unit of Synaptic Transmission, Mario Negri Institute In Jan 2006, his group joined the Laboratory of Biochemistry and Protein Chemistry, Mario Negri Institute Co-author in more than 70 scientific publications on peer-reviewed international journals. First or last author in more than 40 of them. Reviewer for international scientific journals operating in the Neuroscience/Neuropharmacology fields. Selected publications Gerstmann-Straussler-Scheinker disease amyloid protein polymerizes according to the "dock-and-lock" model. Gobbi M, Colombo L, Morbin M, Mazzoleni G, Accardo E, Vanoni M, Del Favero E, Cantù L, Kirschner D A, Ceci P, Ubezio P, Manzoni C, Forloni G, Tagliavini F, Salmona M J Biol Chem. 2006; 281: Funicello M, Conti P, De Amici M, De Micheli C, Mennini T, Gobbi M Dissociation of [3H]L-glutamate uptake from L-glutamate-induced [3H]D-aspartate release by 3-hydroxy-4,5,6,6a-tetrahydro-3aH-pyrrolo[3,4-d]isoxazole-4- carboxylic acid and 3-hydroxy-4,5,6,6a-tetrahydro-3aH-pyrrolo[3,4-d]isoxazole-6-carboxylic acid, two conformationally constrained aspartate and glutamate analogs. Mol Pharmacol 66: Gobbi M, Moia M, Pirona L, Ceglia I, Reyes-Parada M, Scorza C, Mennini T p-methylthioamphetamine and 1- (m-chlorophenyl)piperazine, two non-neurotoxic 5-HT releasers in vivo, differ from neurotoxic amphetamine derivatives in their mode of action at 5-HT nerve endings in vitro. J Neurochem 82: Gobbi M, DallaValle F, Ciapparelli C, Diomede L, Morazzoni P, Verotta L, Caccia S, Cervo L, Mennini T Hypericum perforatum L. extract does not inhibit 5-HT transporter in rat brain cortex. Naunyn Schmiedebergs Arch Pharmacol 360:

152 Gobbi M, Gariboldi M, Piwko C, Hoyer D, Sperk G, Vezzani A Distinct changes in peptide YY binding to, and mrna levels of, Y1 and Y2 receptors in the rat hippocampus associated with kindling epileptogenesis. J Neurochem 70: Crespi D, Mennini T, Gobbi M Carrier-dependent and Ca(2+)-dependent 5-HT and dopamine release induced by (+)-amphetamine, 3,4-methylendioxymethamphetamine, p-chloroamphetamine and (+)-fenfluramine. Br J Pharmacol 121: Mineko Terao obtained her doctorate degree in Pharmaceutical Science from the Kobe Women s College of Pharmacy, Japan in Her scientific interests relate to problems of Cellular Biology and Molecular Biology Ph.D in Molecular Biology, Kyoto University, Japan Research Fellow, Department of Medical Chemistry, Kyoto University Faculty of Medicine, Japan Postdoctoral Associate of the Institute for Cancer Research, Philadelphia, US From 1987 Visiting Scientist of Mario Negri Institute From 1998 Head of the Unit of Gene Structure and Regulation, Mario Negri Institute Selected publications Terao M, Kurosaki M, Barzago MM, Varasano E, Boldetti A, Bastone A, Fratelli M, Garattini E. Avian and canine aldehyde oxidases. Novel insights into the biology and evolution of molybdo-flavoenzymes. J Biol Chem Jul 14;281(28): Garattini E, Parrella E, Diomede L, Gianni M, Kalac Y, Merlini L, Simoni D, Zanier R, Ferrara F F, Chiarucci I, Carminati P, Terao M, Pisano C. ST1926, a novel and orally active retinoid-related molecule inducing apoptosis in myeloid leukemia cells: Modulation of intracellular calcium homeostasis. Blood 2004; 103: Vila R, Kurosaki M, Barzago M M, Kolek M, Bastone A, Colombo L, Salmona M, Terao M, Garattini E. Regulation and biochemistry of mouse molybdo-flavoenzymes. The DBA/2 mouse is selectively deficient in the expression of aldehyde oxidase homologues 1 and 2 and represents a unique source for the purification and characterization of aldehyde oxidase. J Biol Chem 2004; 279: Kurosaki M, Terao M, Barzago M M, Bastone A, Bernardinello D, Salmona M, Garattini E. The aldehyde oxidase gene cluster in mice and rats: Aldehyde oxidase homologue 3, a novel member of the molybdo-flavoenzyme family with selective expression in the olfactory mucosa. J Biol Chem 2004; 279: Parrella E, Gianni M, Cecconi V, Nigro E, Barzago MM, Rambaldi A, Rochette-Egly C, Terao M and Garattini E. Phosphodiesterase 4 inhibition by piclamilast potentiates the cyto-differentiating action of retinoids in myeloid leukemia cells. J Biol Chem 2004; 279: Garattini E, Gianni M and Terao M. Retinoid related molecules an emerging class of apoptotic agents with promising clinical potential in oncology: pharmacological activity and mechanisms of action. Curr Pharm Design 2004, 10: Pia Emilia Villa got her degree in Pharmaceutical Chemistry and Technology at the University of Pavia in 1975 and the specialisation in Pharmacological Research at the Mario Negri Institute, Milan in Her scientific interests are the physiopathologic factors of sepsis and their pharmacological modulation, the cellular and molecular mechanisms of neurodegeneration and neuroprotection in experimental cerebral ischemia : Research fellow, laboratory of Perfusion of Isolated Organs and Toxicology, Mario Negri Institute : Visiting fellow, laboratory of Cultured Hepatocytes, Toxicology Unit, MRC, Carshalton, England 1983: Visiting fellow, Unité de Recherche Hépatologique, Rennes, France : Research Scientist, laboratory of Neuroimmunology, Mario Negri Institute 1995 to date: Head, Unit of Pharmacology of Septic Shock, Mario Negri Institute 1982 Regular member of the Italian Society of Toxicology 1992 Regular member of Celltox 1996 Regular member of the International Cytokine Society. Selected publications Villa P, Shaklee CL, Meazza C, Agnello D, Ghezzi P, Senaldi G. Granulocyte colony-stimulating factor and antibiotics in the prophylaxis of a murine model of polymicrobial peritonitis and sepsis. J Infect Dis. 1998; 178: Villa P, Saccani A, Sica A, Ghezzi P. Glutathione protects mice from lethal sepsis by limiting inflammation and potentiating host defense. J Infect Dis. 2002; 185: Erbayraktar S, Grasso G, Sfacteria A, Xie QW, Coleman T, Kreilgaard M, Torup L, Sager T, Erbayraktar Z, Gokmen N, Yilmaz O, Ghezzi P, Villa P, Fratelli M, Casagrande S, Leist M, Helboe L, Gerwein J, Christensen S, Geist MA, Pedersen LO, Cerami-Hand C, Wuerth JP, Cerami A, Brines M. Asialoerythropoietin is a nonerythropoietic cytokine with broad neuroprotective activity in vivo. Proc Natl Acad Sci U S A. 2003;100 (11):

153 Villa P, Bigini P, Mennini T, Agnello D, Laragione T, Cagnotto A, Viviani B, Marinovich M, Cerami A, Coleman TR, Brines M, Ghezzi P. Erythropoietin selectively attenuates cytokine production and inflammation in cerebral ischemia by targeting neuronal apoptosis. J Exp Med. 2003;198 (6): Leist M, Ghezzi P, Grasso G, Bianchi R, Villa P, Fratelli M, Savino C, Bianchi M, Nielsen J, Gerwien J, Kallunki P, Larsen AK, Helboe L, Christensen S, Pedersen LO, Nielsen M, Torup L, Sager T, Sfacteria A, Erbayraktar S, Erbayraktar Z, Gokmen N, Yilmaz O, Cerami-Hand C, Xie QW, Coleman T, Cerami A, Brines M. Derivatives of erythropoietin that are tissue protective but not erythropoietic. Science. 2004;305: Garau A, Bertini R, Colotta F, Casilli F, Bigini P, Cagnotto A, Mennini T, Ghezzi P, Villa P. Neuroprotection with the CXCL8 inhibitor repertaxin in transient brain ischemia. Cytokine. 2005;30:

154 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The Department comprises six laboratories. Research is heterogeneous in terms of scientific interests and aims, but it is unified by the structural and functional study of specific, pharmacologically important gene products, using a common body of techniques. Classical biochemistry and molecular biology methods are used to define proteins that might be targets for the pharmacological activity of drugs. Potential direct interactions between drugs and proteins are studied at the molecular level by a variety of approaches ranging from animal studies to computer simulation. FINDINGS/MAIN RESULTS Identification of tetracylines as potential anti-prion drugs. Synthesis and physicochemical and biological characterization of peptides deduced from the primary sequence of prion protein. Identification of a relationship between cholesterol synthesis and production of prion protein. Protein identifications by mass spectrometry and data base searching using a combination of techniques. Characterization of the Pentraxin 3 role in the organization of the cumulus oophorus extracellular matrix and in female fertility. Characterization of the role of Junctional Adhesion Molecule-A (JAM-A) in the control of cell motility. Characterization of the effect of inflammatory cytokines on JAM-A function. Development of a constitutive and of an inducible motor neuron cellular model to unravel the toxicity of mutant G93A superoxide dismutase 1 responsible for some forms of familial amyotrophic lateral sclerosis. Conditions of oxidative stress or the presence of compounds impairing the electron transport chain are a risk factor to motor neurons of individuals carrying mutant forms of superoxide dismutase 1. Mitochondrial damage due to mutant G93A superoxide dismutase 1 occurs selectively in motor neurons. Mitochondrial damage by mutant G93A superoxide dismutase 1 in motor neurons is modulated by the level of expression of the mutant protein. Identification and characterization of a novel class of retinoids endowed with strong and selective apoptogenic acivity on the neoplastic cell. Pre-clinical development of these agents for the treatment of acute leukemia. Identification and characterization of novel retinoid-based pharmacological combinations for the treatment of acute myelogenous leukemia. Molecular cloning and characterization of the cdnas and genes of four novel members of the mammalian molybdo-flavoprotein family. Definition of a novel gene cluster on human chromosome 2 and mouse chromosome 1. Development of knok-out animals for molybdo-flavoproteins: AOX1, AOH1, AOH2, AOH3. Identification of erythropoietin as a neuroprotective agent and of new molecules with neuroprotective activity. Identification of the pharmacological action of erythropoietin against peripheral neuropathy of diabetes. Identification of erythropoietin derivatives that retain its neuroprotective actions but have lost its hemopoietic ones. Discovery of proteins that are regulated by the redox state through the formation of reversible 152

155 disulfide bonds with glutathione (protein glutathionylation). Identification of exofacial proteins undergoing thiol redox regulation. The use of antioxidant molecules in models of sepsis and inflammation diminishes the inflammatory response while potentiates the innate immune response. Identification of the gene expression profile regulated by thiols. The treatment with a non haematopoietic derivate of Erythropoietin (CEPO) reduces motor neuron loss and clinical progression in a mouse model of ALS related to alterations in vesicle trafficking, the wobbler mouse. Treatment with a soluble TNF receptor in the wobbler mouse, reduces motor neuron degeneration and the phosphorylation of the two main stress kinases (p38 e JNK) activated by TNF receptors. Riluzole treatment reduces motor neuron loss and clinical progression of wobbler mouse by increasing the endogenous BDNF expression. Oxidative stress, glial activation and inflammation occur in the retinopathy as well as in cerebral and spinal cord dysfunction in the mnd mouse, a model of progressive epilepsy with mental retardation related to mutation in the CLN8 gene. These findings provide further evidence for the implication of TNF death receptor signaling in the pathology of Neuronal Ceroid Lipofuscinosis The affinity of pergolide for human cloned 5-HT 2A and 5-HT 2B receptors is similar and higher that that for the human cloned D 2L receptor. These findings, together with the fact that it acts as agonist at both h5-ht 2A and h5-ht 2B receptors, could explain its potential toxicity mediated by activation of cardio-pulmonary 5-HT 2B receptor. New conformationally constrained aspartate and glutamate analogues dissociate glutamate uptake inhibition and reverse transport-mediated release. Dimethyl sulfoxide, a solvent commonly utilized to dissolve hydrophobic compound for in vitro experiments, interferes with the 5-HT6 agonists activity when the scintillation proximity assay is used for evaluating 35S-GTP-γ-S binding; but does not interfere with the europium labelled GTP binding determined by time-resolved fluorescence. NATIONAL COLLABORATIONS Advanced Biology Center, Genoa Dip. Anatomia, Farmacologia, Medicina Legale, University of Turin Dip. Biotecnologie, Università degli Studi, Milan Dip. Chimica Biochimica e Biotecnologie per la Medicina, Università degli Studi, Milan Dip. Chimica Farmaceutica e Tossicologica, Università degli Studi, Milan Dip. Farmaco-Chimico, Università degli Studi, Messina Dip. Farmaco-Chimico-Tecnologico, University of Siena Dip. Farmacologia Medica, Università degli Studi, Milan Dip. Scienze Biochimiche, University of Florence Dip. Scienze Farmaceutiche, University of Catania Dip. Scienze Farmaceutiche, University of Genoa Dip. Scienze Farmacologiche, Università degli Studi, Milan Dip. Scienze Fisiologiche e Farmacologiche,University of Pavia Dip. Scienze Molecolari, University of Milan Dip. Studi pre-clinici, University of Milan Facoltà di Biologia, Università degli Studi, Milan Facoltà di Chimica, Università degli Studi, Milan Facoltà di Chimica, University of Ferrara 153

156 GlaxoSmithkline, Verona Harlan, Milan IRCCS Fondazione "Istituto C. Mondino", Laboratorio di Neurobiologia Sperimentale, Pavia Istituto di Biologia Molecolare Buzzati Traverso, Naples Istituto di Biomedicina e Immunologia Molecolare CNR, Palermo Istituto di Endocrinologia, Centro di Eccellenza per le Malattie Neurodegenerative, Università degli Studi, Milan Istituto di Clinica Neurologica, Ospedale Maggiore Policlinico, Milan Istituto Clinico Humanitas, Milan Istituto di Neuroscienze C.N.R., Pisa Istituto Nazionale dei Tumori, Milan Istituto Nazionale dei Tumori, Naples Istituto Nazionale Neurologico "C. Besta", Milan Istituto Oncologico Europeo, Milan Istituto Regina Elena, Rome Newron Pharmaceuticals, Milan Ospedale Maggiore Policlinico, Milan Ospedale Pediatrico Bambino Gesu', Rome Ospedale Pediatrico "Gaslini", Genoa Ospedale S. Gerardo, Monza, Milan Sigma-Tau, Pomezia, Rome Zambon, Milan INTERNATIONAL COLLABORATIONS Babraham Institute, Cambridge, UK Boston College, Boston, MA, USA Case Western Research University, Cleveland, OH, USA Dept. of Neurology, Keio University, Tokyo, Japan Dept. de Quimica-Fisica de Macromoleculas Biologicas, CSIC, Madrid, Spain Faculdad de Ciencias Medicas, Universidad de Santiago de Chile, Chile Dept. of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem, Israel Flanders Interuniversity Institute for Biotechnology (VIB) University of Gent, Belgium FMP, Berlin, Germany Giessen Polyclinic University, Giessen, Germany Houston University, TX, USA IBSN CNRS, Marseille, France Indiana University, Indianapolis, IN, USA Institut de Genetique et Biologie Moleculaire et Cellulaire, Strasbourg, France Institut Pasteur, Paris, France John Innes Centre, Norwich, UK Kenneth S. Warren Institute, Ossining, NY, USA Max-Planck-Institut für experimentelle Medizin, Göttingen, Germany National Institute of Health, Bethesda, MD, USA Nippon University, Tokyo, Japan North Shore University Hospital, Manhasset, NY, USA Pepscan System BV, Lelystad, Holland Polichem S.A., Lugano, Switzerland Stanford University School of Medicine, Stanford, CA, USA 154

157 Technical University Braunschweig, Germany Trinity College, Dublin, Ireland Universidad de La Laguna, Tenerife, Spain Universidad Nova, Lisbon, Portugal Universitat des Saarlandes, Hamburg, Germany Universitat Freiburg, Germany Université Paris, France Université Victor Segalen Bordeaux 2, Bordeaux, France University of Birmingham, UK University of Cardiff, UK University of Colorado, School of Medicine, Denver, CO, USA University of Glasgow, UK University of Gottingen, Germany University of Muenster, Germany University of Southampton, UK University of Sussex, UK University of Vienna, Austria Waring-Webb Institute, University of Colorado, Denver CO, USA Weizmann Institut, Rehovot, Israel Westfaelische Wilhelms-Universitaet Muenster, Germany EDITORIAL BOARD MEMBERSHIP Neurobiology of Lipids (L. Diomede) Neuroimmunomodulation (P. Ghezzi) Newsletters of the International Cytokine Society (P. Ghezzi) European Journal of Cancer (E. Garattini) PEER REVIEW ACTIVITIES American Journal Physiology, Biochemical Journal, Biochemical Pharmacology, Biochimica Biophysica Acta, Brain Research, Cancer Research, Cell Death and Differentiation, Cell Research, Circulation, Drug Investigation, European Journal of Cancer, European Journal of Immunology, European Journal of Neuroscience, International Journal of Cancer, Journal of Cell Biology, Journal of Hepatology, Journal of Immunology, Journal of Investigative Dermatology, Journal of Lipid Mediators, Journal of Neurochemistry, Journal of Translational Medicine, Neuroscience Letters, Pharmacological Research, Physiological Genomics, Proceedings of the National Academy of Sciences, Life Sciences. 155

158 PARTICIPATION IN EVENTS IN WHICH THE DEPARTMENT WAS INVOLVED Symposium: Keystone Symposia on Molecular and Cellular Biology - Molecular Mechanisms of Neurodegeneration, Synaptic dysfunction in the cerebellum of transgenic mice expressing a prion protein with an insertional mutation, January, Taos, New Mexico Meeting : III Meeting on the Molecular Mechanisms of Neurodegeneration, Use of steroidogenic acute regulatory protein (STAR) in study of mitochondrial ATP-dependent proteases potentially involved in familial amyotrophic lateral sclerosis ; Increased formation of glutathione mixed disulfides in a motor neuron-like cell model for familial amyotrophic lateral sclerosis ; Adaptive response to stable expression of human mutant G93A superoxide dismutase 1 in motor neuron-like cells involves alterations of the glutathione pool, In vitro Mechanisms of Neuroinflammation in a Co-culture system of Astrocytes and Microglia from Neonatal Hamsters, PrP amyloid: the role of oligomers in triggering neuronal degeneration, Early alterations of glutamate exocytosis in the cerebellum of transgenic mice expressing a PrP insertional mutation, May, Milan, Italy Meeting: Joint Annual Meeting ISMRM ESNR, Neural stem cells tracking by MRI in wobbler mouse, a model of motoneuron disease, May, Berlin, Germany Congress: 5 Congresso della Società Italiana di Immunologia, Immunologia Clinica e Allergologia, The long pentraxin PTX3 recognizes Neisseria meningitidis, binds selected outer membrane proteins and amplify the induced inflammatory response, 6-9 June, Trieste, Italy Meeting: 5th Joint Meeting Medical Chemistry, Arylpiperazinylalkylthio benzothiazole or benzoxazole derivatives as selective 5-HT1A serotonin receptor ligands, (-)-HIP-A and (+)- HIP-B: investigation of the pharmacological profile and potential therapeutic applications, June, Portoroz, Slovenia Meeting: Mutant SOD1 and familial ALS: from the molecole to man, Altered redox environment in a motor neuron-like cell model for SOD1-related forms of amyotrophic lateral sclerosis, Glutathione and protein-glutathione mixed disulfides compartmentalization in intact motor neuron-like cells, September, Milan, Italy Congress: XVIII Convegno Nazionale della Divisione di Chimica Farmaceutica della Società Chimica Italiana, Nuovi Arilpiperazinilalchilpirrolopirimidindioni come ligandi potenti e selettivi per i recettori alfa1-adrenergici, September, Chieti, Italy Conference: Prion 2007, Evaluation of neuroinflammation mechanisms in Co-cultures of neurons, astrocytes and microglia from newborn hamsters, Use of heparin derivatives for therapeutic intervention of transmissible spongiform encephalopathies, Role of oligomers in the neurotoxicity of Gerstmann-Straussler-Scheinker disease amyloid protein, September, Edinburgh, Scotland Congress: Congresso Nazionale della Società italiana di Neuroscienze, Mitochondrial toxicity in cell models for familial amyotrophic lateral sclerosis, September, Verona, Italy 156

159 Symposium: 18th International Symposium on ALS/MND, Proteomic analysis of nitrated proteins from peripheral blood mononuclear cells of patients with sporadic ALS, Characterization of protein aggregates in the G93A SOD 1 mouse reveals a possible link between oxidative stress and aggregation pathogenetic pathways, 1-3 December, Toronto, Canada GRANTS AND CONTRACTS Agenzia Italiana del Farmaco, Rome, Italy Biotecnologies BT - Perugia, Italy Dompè, L' Aquila, Italy European Union, Bruxelles, Belgium Istituto Auxologico Italiano, Milan, Italy Istituto Nazionale Neurologico "C. Besta", Milan, Italy Italian Association for Cancer Research (AIRC), Milan, Italy Italian Ministry of University and Research (MIUR), Rome, Italy Kenneth S. Warren Institute, NY, USA Lundbeck A/S, Copenhagen, Denmark Cariplo Foundation, Milan, Italy Don Gnocchi Foundation, Milan, Italy Mariani Foundation, Milan, Italy Monzino Foundation, Milan, Italy Ministry of Health, Rome, Italy National Research Council (CNR), Palermo, Italy North Shore University Hospital, NY, USA Perfetti-Van Melle, Lainate (Mi), Italy Sigma Tau, Pomezia (Rome), Italy Telethon, Milan, Italy University of Florence, Italy University of Milan-Bicocca, Italy University of Siena, Italy Weizmann-Pasteur-Negri Foundation, Paris, France Zambon Group, Bresso (Mi), Italy 157

160 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Beghi E, Mennini T, Bendotti C, Bigini P, Logroscino G, Chiò A, Hardiman O, Mitchell D, Swingler R, Traynor BJ, Al-Chalabi A. The heterogeneity of amyotrophic lateral sclerosis: a possible explanation of treatment failure. Curr Med Chem : Bernhagen J, Krohn R, Lue H, Gregory JL, Zernecke A, Koenen RR, Dewor M, Georgiev I, Schober A, Leng L, Kooistra T, Fingerle-Rowson G, Ghezzi P, Kleemann R, McColl SR, Bucala R, Hickey MJ, Weber C. MIF is a noncognate ligand of CXC chemokine receptors in inflammatory and atherogenic cell recruitment. Nat Med : Bianchi R, Gilardini A, Rodriguez-Menendez V, Oggioni N, Canta A, Colombo T, De Michele G, Martone S, Sfacteria A, Piedemonte G, Grasso G, Beccaglia P, Ghezzi P, D'Incalci M, Lauria G, Cavaletti G Cisplatin-induced peripheral neuropathy: neuroprotection by erythropoietin without affecting tumour growth Eur J Cancer : Bigini P, Atzori C, Fumagalli E, Cagnotto A, Barbera S, Migheli A, Mennini T Lack of caspase-dependent apoptosis in spinal motor neurons of the wobbler mouse Neurosci Lett : Bolchi C, Pallavicini M, Rusconi C, Diomede L, Ferri N, Corsini A, Fumagalli L, Pedretti A, Vistoli G, Valoti E Peptidomimetic inhibitors of farnesyltransferase with high in vitro activity and significant cellular potency Bioorg Med Chem Lett : Fiordaliso F, De Angelis N, Bai A, Cuccovillo I, Salio M, Serra D M, Bianchi R, Razzetti R, Latini R, Masson S Effect of Beta-adrenergic and renin-angiotensin system blockade on myocyte apoptosis and oxidative stress in diabetic hypertensive rats Life Sci : Fioriti L, Angeretti N, Colombo L, De Luigi A, Colombo Alessio, Manzoni Claudia, Morbin M, Tagliavini F, Salmona M, Chiesa R, Forloni G Neurotoxic and gliotrophic activity of a synthetic peptide homologous to Gerstmann-Straussler-Scheinker disease amyloid protein J Neurosci : Garattini E, Gianni M, Terao M Retinoids as differentiating agents in oncology: A network of interactions with intracellular pathways as the basis for rational therapeutic combinations Current Pharmaceutical Design : Garattini E., Giannì M., Terao M. Cytodifferentiation by retinoids, a novel therapeutic option in oncology: rational combinations with other therapeutic agents. Vitam Horm : Ghezzi P, Di Simplicio P. Glutathionylation pathways in drug response. Curr Opin Pharmacol : Review Ghezzi P, Mengozzi M Activities of erythropoietin on tumors: An immunological perspective Eur J Immunol : Leonelli E, Bianchi R, Cavaletti G, Caruso D, Crippa D, Garcia-Segura L M, Lauria G, Magnaghi V, Roglio I, Melcangi R C Progesterone and its derivatives are neuroprotective agents in experimental diabetic neuropathy: a multimodal analysis Neuroscience : Luo X Y, Sharma D, Inouye H, Lee D, Avila R L, Salmona M, Kirschner D A 158

161 Cytoplasmic domain of human myelin protein zero likely folded as Beta structure in compact myelin Biophys J : Luo X, Inouye H, Gross AA, Hidalgo MM, Sharma D, Lee D, Avila RL, Salmona M, Kirschner DA. Cytoplasmic domain of zebrafish myelin protein zero: adhesive role depends on beta-conformation. Biophys J. 2007, 93: Massignan T, Casoni F, Basso M, Stefanazzi P, Biasini E, Tortarolo M, Salmona M, Gianazza E, Bendotti C, Bonetto V Proteomic analysis of spinal cord of presymptomatic amyotrophic lateral sclerosis G93A SOD1 mouse Biochem Biophys Res Commun : Mereghetti I, Cagnotto A, Mennini T Dimethyl sulfoxide: An antagonist in scintillation proximity assay [35S]-GTPgammaS binding to rat 5-HT6 receptor cloned in HEK-293 cells? J Neurosci Methods : Modica M, Salerno L, Pittala' V, Romeo G, Siracusa M, Mereghetti I, Cagnotto A, Mennini T Synthesis and binding properties of new endothelin receptor ligands Lett Drug Des Discov : Noe F, Nissinen J, Pitkanen A, Gobbi M, Sperk G, During M, Vezzani A Gene therapy in epilepsy: the focus on NPY Peptides : Prezzavento O, Campisi A, Ronsisvalle S, Li Volti G, Marrazzo A, Bramanti V, Cannavo' G, Vanella L, Cagnotto A, Mennini T, Ientile R, Ronsisvalle G Novel sigma receptor ligands: Synthesis and biological profile J Med Chem : Roglio I, Bianchi R, Giatti S, Cavaletti G, Caruso D, Scurati S, Crippa D, Garcia-Segura L M, Camozzi F, Lauria G, Melcangi R C Testosterone derivatives are neuroprotective agents in experimental diabetic neuropathy Cell Mol Life Sci : Salerno L, Guerrera F, Modica M, Romeo G, Pittalà V, Siracusa MA, Mereghetti I, Cagnotto A, Mennini T. Synthesis of 1,2,4-triazole derivatives: binding properties on endothelin receptors. Med Chem : Scarchilli L, Camaioni A, Bottazzi B, Negri V, Doni A, Deban L, Bastone A, Salvatori G, Mantovani A, Siracusa G, Salustri A PTX3 Interacts with inter-alpha- trypsin inhibitor. Implications for hyaluronan organization and cumulus oophorus expansion J Biol Chem : Setola V, Terao M, Locatelli D, Bassanini S, Garattini E, Battaglia G Axonal-SMN (a-smn), a protein isoform of the survival motor neuron gene, is specifically involved in axonogenesis Proc Natl Acad Sci USA : Villa P, Triulzi S, Cavalieri B, Di Bitondo R, Bertini R, Barbera S, Bigini P, Mennini T, Gelosa P, Tremoli E, Sironi L, Ghezzi P The interleukin-8 (IL-8/CXCL8) receptor inhibitor reaparixin improves neurological deficits and reduces long-term inflammation in permanent and transient cerebral ischemia in rats Mol Med :

162 RESEARCH ACTIVITIES Laboratory of Receptor Pharmacology Studies on the role of dysfunction of the endoplasmic reticulum (ER) or the Golgi apparatus (GA) in neurodegenerative disease The secretory pathway starts at the endoplasmic reticulum (ER) where proteins are synthesized and folded and chaperone mediated quality control prevents misfolded proteins to reach their destination and interfere with normal metabolism. Protein transport by mean of vesicles continues through the Golgi Apparatus (GA), that is most abundant in neurons, and finishes in the plasma membrane, secretory vesicles or lysosomes. The endocytic pathway enables internalized macromolecules to be delivered via endosomes to lysosomes where they are enzymatically digested. In mammalian cells, the Golgi-associated retrograde protein (GARP) complex is involved in retrograde transport of endosomes to the trans GA network. Defective intracellular membrane trafficking is common to several neurodegenerative diseases. Among the neuronal population, motor neurons, due the their high energy requirement and long axons are, together with retinal cells, the most sensitive ones. The Laboratory of Receptor Pharmacology utilizes two mouse models of neurodegeneration related to cellular transport disruption, carrying mutation in proteins resident in the ER (the mnd mouse) or in the GARP complex (the wobbler mouse). The neuronal ceroid lipofuscinosis (NCLs) are a group of autosomal recessive neurodegenerative diseases and a significant cause of childhood progressive intellectual and neurological deterioration, for which no curative or preventive treatment are available. Among them, the progressive epilepsy with mental retardation is the newest form with mutation in the CLN8 gene encoding a novel ER transmembrane protein with undefined function. An orthologue of CNL8 is mutated in the motor neuron degeneration mouse (mnd) which shows early retinopathy and delayed motor neuron dysfunction without degeneration. How CLN8 mutation leads to NCL defect is unknown. Our laboratory is studying mnd mice since many years: we have already reported decreased spinal cord GLT-1 glial glutamate transporter and increased plasma glutamate concentration already at presymptomatic stage in mnd mice, with increased GluR2 and lowered GluR3 AMPA receptor subunits in the lumbar spinal cord. The AMPA receptor antagonists ZK (non-competitive), like NBQX (competitive), ameliorate motor behavior in mnd mice. We also found that TNF and TNFR1 is increased in the spinal cord of mnd mice already at presymptomatic stage, when intensive astrocytes and microglial proliferation occurs. The rate of oxygen consumption (QO2), and mitochondrial functions were decreased in mnd spinal cord. The level of lipid peroxide derivatives reacting with thiobarbituric acid (TBARS) were increased in mnd spinal cord and retina. L-carnitine treatment delayed the onset of motor behaviour impairment, increased the mitochondrial enzyme activities and was effective in enhancing QO 2 and decreasing TBARS levels. At present we are testing the susceptibility to convulsions in mnd mice at pre-symptomatic stage, in order to find a further link to the human pathology. In these studies new non-invasive approaches, like Optical Imaging, MRI, MicroCT and Confocal Angiography are applied, in order to allow a better translation of the results to the human disease. Amyotrophic Lateral Sclerosis (ALS) is a progressive neurodegenerative disorder involving primarily motor neurons in the spinal cord, brainstem and cerebral motor cortex and leading to denervation, muscular atrophy and paralysis. The disease is sporadic in approximately 90% of 160

163 cases, and the mechanism(s) responsible for the selective motor neuron degeneration are far from being elucidated. A missense mutation in Vps54 has been described in the wobbler mouse, which share many pathological features with ALS patients. In mammalian cells Vps54 forms heterotrimeric complexes with Vps52 and Vps53 to form the GARP complex, involved in retrograde transport of endosomes to the trans GA network. Thus we use the wobbler mouse as a reliable tool to understand the interplay between endosomal dynamics and the selective loss of motor neurons. A series of experiments are in progress in cultured neural cells obtained from wobbler and healthy homozygous mice to investigate the possible effect of Vps54 mutation on intracellular trafficking, mitochondrial activity, and lysosome accumulation. We have already reported that wobbler mice are sensitive to riluzole treatment, without marked changes in AMPA/NMDA receptor subunits expression in motor neurons of early symptomatic mice. In addition we found increased levels of TNF and TNFR1 in the cervical spinal cord and a significant effect of chronic treatment with a soluble h-tnf binding protein, resulting in slower clinical symptoms progression, reduced motor neuron loss and selective inhibition of the two main stress-kinases (p38 and JNK) associated to TNF receptors activation. Finally we investigated two different approaches of cell therapy. Undifferentiated adult neural stem cells (in collaboration with Dr. Parati, Istituto Besta) produced a weak and transient protective effect in clinical progression but significantly reduced motor neuron loss occurring in the wobbler mouse. Transplantation of mononucleate cells from human cord blood (in collaboration with Dott. Lazzari, Policlinico), although did not replace degenerating motor neurons, produced a marked neuroprotective effect by slowing the clinical progression and reducing motor neuron loss, biceps atrophy and neuroinflammation (reactive gliosis). Neuropharmacology of the glutammatergic and serotonergic systems Glutamate is the major excitatory amino acid in the central nervous system; the extracellular glutamate concentration has to be maintained at physiological level by active uptake mediated by specific transporters (Excitatory Amino Acid Transporters, EAATs) located on the plasma membranes of neurons and glia. Alterations in this process might lead to a relevant increase in extracellular glutamate concentrations, that is highly toxic for neurons in the central nervous system. A research project is in progress in order to better characterize the involvement and the role of the neuronal compartment in the general process of glutamate homeostasis. The functional properties are evaluated using biochemical assays and the quantitative characteristics are evaluated by western blot for specific neuronal and glial proteins and flow cytometry analysis (in collaboration with Dr. Bernasconi, Department of Oncology, IRFMN). These evaluations are done on purified preparations from mouse spinal cord. The same characteristics are evaluated on two different animal models of motor neuron degeneration, the wobbler mouse and the transgenic SOD1G93A mouse (in collaboration with Dott. Bendotti, Department of Neuroscience, IRFMN), in order to understand the possible involvement in neurodegenerative diseases. Results obtained to date indicate that there is an active neuronal component in the glutamate uptake. The comparison between the two animal models of neurodegeneration suggest that this component has a different importance in the motor neuron death processes. In the wobbler mouse the excitotoxicity related to alterations of glutamate uptake is likely to be not involved, while in the transgenic SOD1G93A mouse we found a significant reduction of glutamate uptake in the neuronal fraction. This alteration probably contribute to the exacerbation of processes leading to motor neuronal death in this animal model. A research project is in progress in collaboration with professor Bonanno (University of Genova) to characterize mechanisms of glutamate release from synaptosomes obtained from spinal cord of wobbler mouse, in order to evaluate if the motor neuronal death might depend on altered processes of neurotransmitter release in the nerve endings. 161

164 We recently concluded a project focused on the study of the interaction between glutamate transporters and riluzole. Riluzole is a drug with a complex mechanism of action, and is the only drug approved for the treatment of amyotrophic lateral sclerosis (ALS). We used as experimental model cell cultures that stably express the main three glutamate transporters (GLT1, GLAST and EAAC1) and we demonstrated for the first time that riluzole acts on the glutamate uptake mediated by the three EAAT subtypes, significantly increasing the efficiency of the process. This mechanism might be relevant in pathological conditions characterized by glutamate concentrations over the physiological threshold, as occurs in ALS. AMPA receptor-mediated excitotoxicity is one of the main events involved in motor neuron degeneration in ALS pathogenesis. We established a new model of primary cocultures of purified motor neurons over a glial layer. This in vitro model allows to obtain healthier motor neurons maintained in more physiological conditions and was used to demonstrate that AMPA receptor agonists can induce both apoptotic or non-apoptotic death pathways depending on their concentrations. We studied the interaction between excitotoxicity and other potentially neurotoxic factors, such as the inflammatory mediators, and we demonstrated that the proinflammatory chemokine IL-8 induces motor neuron death through the CXCR2 receptor. Preliminary studies on the effect of TNF-α, whose levels were found increased in the spinal cords of animal models of motor neuron degeneration, indicated the pivotal role of glial cells in mediating neurotoxicity of this cytokine. At present, we are studying the main intracellular biochemical pathways involved in neurodegenerative mechanisms (such as calcium influx) and testing new potential treatments to selectively interfere with each of them. We have also started a study on the neurotoxic effect of serum from professional soccer players aimed at identifying potential risk factors present in the blood which could support the high incidence of ALS in this group of athletes. Studies on primary spinal cord or hippocampal neuronal cultures from wild type or mnd mice are in progress in order to investigate their sensitivity to AMPA receptor agonists and antagonists, and to evaluate the role of astrocytes obtained from mnd mice in affecting motor neuron viability. Studies on astrocytes derived from neural stem cells of wobbler or control mice are in progress in order to evaluate whether typical biochemical alterations of wobbler mice are already evident in precursor-derived cells : we study the role of glutamate transporters and possible effects of this kind of cells on healthy motor neurons. Finally, studies on the role of Vsp54 mutation on intracellular trafficking are in progress in primary cultures of astrocytes obtained from the spinal cord of adult wobbler or control mice. The Laboratory of Receptor Pharmacology is maintaining, since many years, collaborations with medicinal chemistry laboratories to characterize the affinity and selectivity of newly synthesized compounds on neurotransmitter receptors using in vitro binding methods. The results are utilized for molecular modelling (QSAR) studies and/or for further pharmacological evaluations. Particularly, a collaboration is ongoing with Prof. S. Grasso (University of Messina) and Prof. C. De Micheli (University of Milan), for the development of new non-competitive AMPA receptor antagonists. An useful application of this technique is also the evaluation of the agonist/antagonist activity of compounds acting on G-protein coupled receptors (GPCR), measuring their effects on 35S- GTP-γ-S binding. New non-radioactive methods based on time-resolved fluorescence are under characterization as functional assays to monitor GPCR activity on cell membranes. Recently we have verified the effect of dimethyl sulfoxide, a solvent commonly utilized to dissolve hydrophobic compound for in vitro experiments, on different in vitro assays utilizing HEK 293 cells expressing the 5-HT6 serotonin receptors. Our results indicate that dimethyl sulfoxide interferes with the agonist activity on 5-HT6 receptor when the scintillation proximity assay 35S-GTP-γ-S binding is used. On the contrary, it does not interferes with europium labelled GTP binding determined by time-resolved fluorescence. 162

165 In addition, the laboratory perform autoradiography binding studies in order to evaluate ex vivo drug-receptor occupancy. Finally, we have ongoing collaborative studies with Dr. Gobbi (Lab. biochemistry and protein chemistry, IRFMN), to characterize the role of the allosteric site at the serotonin transporter on the mechanism of action of SSRI (selective serotonin reuptake inhibitors) antidepressant, like escitalopram. Laboratory of Neuroimmunology Redox regulation The study of the so-called oxidative stress has led to the identification of biochemical events that are modified by antioxidant molecules even in the absence of oxidative stress intended as overproduction of toxic oxygen intermediates (free radicals). We use the term redox regulation to define the pattern of cell functions (gene expression, activity of enzymes or transcription factors) that are in some way modified by the redox state of the cell, defined as the ratio between oxidizing and reducing species (usually: the oxidized glutathione / reduced glutathione ratio). Our work focuses on the molecular mechanisms by which small changes in the redox state can affect proteins, with particular attention to the reversible modification of cysteines to form disulfide bonds (with proteins or with small molecular weight thiols such as glutathione). We recently focused our attention on the identification of the redox state of proteins present on the outside of the plama membrane since these often have key functions (e.g.: transporters, receptors) and are the closest target of extracellular oxidants. We also apply proteomics techniques and gene expression profiling using DNA microarrays to identify the pathways susceptible of redox regulation. Neuroprotection and erythropoietin The pathologies of the central or peripheral nervous systems studied in the lab are: cerebral ischemia, experimental autoimmune encephalomyelitis, and diabetic neuropathy). Using animal models and cell culture, we try to clarify the relationships between neuronal death and inflammation, and to intervene with protective agents. Among the latter, we are studying endogenous molecules that have shown an unexpected anti-apoptotic action on neuronal cells, particularly erythropoietin and anti-inflammatory drugs. Laboratory of Molecular Biology Novel retinoids for the treatment of acute myeloid leukemia The synthetic and natural derivatives of retinoic acid (retinoids) have shown promising activity in the treatment of leukemia and solid cancer. Retinoids exert their therapeutic activity through three distinct types of effects: cyto-differentiation, growth inhibition and apoptosis. The three effects can be dissociated, albeit partially, as retinoids endowed primarily with cytodifferentiating or anti-proliferative activities are known. Recently, we identified and characterized a novel class of retinoids with strong and selective apoptotic activity towards the neoplastic cell. These compounds (RRMs, retinoid related molecules), which were originally developed as selective agonists of the gamma-types of the nuclear receptors of retinoic acid (RARy), induce apoptosis in different types of leukemia and solid cancer cells through a largely unknown mechanism. The process of apoptosis set in motion by RRMs is different from that of other known chemotherapeutic agents and does not require the activation of the nuclear retinoic acid receptors. RRMs are active not only in vitro but also in vivo on a number of pre-clinical models of acute myeloid leukemia. Currently, some of these innovative molecules are in an 163

166 advanced phase of pre-clinical development. Novel retinoic-acid-based pharmacological combinations for the treatment of acute leukemia The clinical use of retinoic acid for the treatment of acute promyelocytic leukemia (APL) is based on the ability of this compound to induce the maturation of the leukemic blast along the normal myelocytic/granulocytic pathway. At present, the clinical use of retinoic acid for the treatment of patients suffering from APL is the sole example of differentiation therapy. Differentiation therapy is worth pursuing as it is theoretically associated with a lower level of toxicity relative to what observed following treatment with the classical cyto-toxic agents. However, the clinical use of retinoic acid is still burdened by a number of problems including natural and induced resistance as well as systemic and local toxicity. One of the possible ways to increase the therapeutic index of retinoic acid is based on the identification of compounds or drugs that potentiate the pharmacological activity of the retinoid. We have recently demonstrated that a series of agents, such as G-CSF, interferons, camp analogs, phosphodiesterase IV inhibitors and a number of other novel compounds sensitize the leukemic blast to the pharmacological activity of retinoic acid. In the long run, it is our objective to develop novel combinations and bring some of the above mentioned retinoic-acid-based combinations to the clinic. In addition we intend to use some of the combinations as pharmacological tools to dissect the intricacies of the cyto-differentiation process set in motion by retinoic acid in the leukemic blast. The family of molybdo-flavoproteins Molybdo-flavoenzymes are proteins of medical and industrial interest. They are the sole enzymes that require molybdenum, in the form of the molybdenum cofactor, for their catalytic activity. The laboratory has a long-standing and specific interest in the biochemistry and biology of mammalian molybdo-flavoproteins. In the past, the laboratory contributed to the elucidation and characterization of the primary structure of the two mammalian molybdoflavoproteins, aldehyde oxidase (AOX1) and xanthine oxidoreductase (XOR). In the last few years, the group identified and cloned the cdnas and the genes coding for three novel mouse molybdo-enzymes (AOH1, AOH2 and AOH3) belonging to the sub-family of molybdoflavoproteins. The long-term goal of our studies is to define the structure, the substrate specificity, the mechanisms of catalysis as well as the physo-pathological function of the three new proteins. We will also continue the biochemical and functional studies on mammalian AOX1 and XOR. To achieve our aims, we have recently developed cell lines over-expressing XOR in a tetracycline inducible fashion. In addition, we have generated knock-out mice for the genes encoding AOH2 and AOH3. Laboratory of Biochemistry and Protein Chemistry Chimico-physical and molecular-biochemical studies on the prion protein and on the peptides deduced from its aminoacid sequence Prion protein fibril formation is associated with neuronal cytotoxicity and astrogliosis observed in prion diseases. The formation of fibrils is the consequence of a conformational switch between the structure of the native and the pathological proteins and it is considered of pivotal importance for the appearance and progression of prion protein diseases. Identifying the molecular determinants responsible for the conformational transition is likely to give insight into the pathogenetic process leading to prion diseases. A reductionist appraoch to the problem calls for the generation of simple experimental models in which the dynamics of the conformational transition can be studied in detail. In our laboratory we developed synthetic peptides that mimic the fibrillogenic properties of pathological prion protein. With the use of 164

167 various types of biochemical and biophysical techniques we studied the conformation of these peptides through the evaluation of their secondary structure, the resistance to protease digestion as well as the aggregating and amyloidogenic properties. Our approach gave detailed informations on the conformational plasticity of various types of prion protein fragments. To understand the correlation between the chemico-physical properties of prion protein-derived peptides and their biological effects we used appropriate cellular models. These experiments, performed in collaboration with the Laboratory of the Biology of Neurodegenerative Disorders, gave informations on the sub-cellular distribution of the peptides to identify the intracellular biological targets. Development of a therapeutical strategy against prion-related diseases Currently, no therapeutic options for the cure of prion-related diseases are available and the identification of new molecules for the prevention and treatment of the infettivity is of great interest. Althought some compounds gave positive results in prion cellular models, in human they exhert low efficacy for toxicity and for their inability to pass the blood brain barrier. A first approach for the development of anti-prion candidates involved molecules capable of interfering with the fibrils formation, directly interacting with the β-sheet conformation of PrPSc causing its disaggregation. In collaboration with the Laboratory of the Biology of the Neurodegenerative Disorders we aimed to identify compounds endowed with anti-fibrillogenic activity and test their efficacy in different in vitro and in vivo pre-clinical models of prion diseases. Our studied indicated that tetracyclines are new good candidates as anti-prion drugs since they act as anti-fibrillogenic compunds increasing the prion sensitivity to protease K digestion. Moreover, they inhibited neuronal cell death and astrogliosis caused by prion peptides and prolonged the survival of prion-infected animals. Another therapeutic approach is based on the development of molecules that inhibited the PrP formation by interacting with the lipid metabolism and destabilizing specific cellular membrane domains. In collaboration with the Department of Infective Pathology and Diseases of the Royal Veterinary School (Hawkshead, UK) we reported that statins, inhibiting cholesterol synthesis, reduced the in vitro prion production. On the other hand, compounds such as polyunsaturated fatty acids, known for their reducing effects on cholesterol levels, increased the PrP production. Our future studies are aimed at understanding the relationship between cholesterol cellular membrane distribution, lipid domain stability and the conversion of cellular prion protein in its pathologic form. Oxidative stress and protein aggregation in amyotrophic lateral sclerosis: a proteomic approach The molecular mechanisms at the basis of neurodegenerative diseases including the geneticallylinked ones, such as amyotrophic lateral sclerosis (ALS), are still unknown. However, there is evidence that oxidative stress and protein aggregation play central roles in the pathogenesis of such diseases. The Unit of Medical Biochemistry, conducted proteome analysis of an animal model of familial ALS. In collaboration with the laboratory of Molecular Neurobiology, we focused the attention on the analysis of protein expression changes and protein modifications, such as tyrosine nitration and ubiquitination, in a transgenic mouse, which over-express human mutated (G93A) superoxide dismutase (SOD1). We analyzed, by proteomic tools, spinal cord of presymptomatic G93A SOD1 mice, we identified nitrated proteins and quantified the level of nitration for each protein in comparison with healthy controls. We have revealed that there is a substantial increase of the nitration level in at least five proteins: actin, alpha and gamma enolase, ATP synthase and a chaperone protein, HSC71. The alteration of the function of these proteins may have important consequences on the cellular metabolism and catabolism, and therefore may be at the basis of the molecular mechanisms leading to neurodegeneration. In addition, by mass spectrometry, we have identified the specific nitrated tyrosines for a number 165

168 of proteins. We have observed that al least enolase and glyceraldehyde 3-phosphate dehydrogenase are nitrated at the same tyrosine site known to be phosphorylated. This is an important finding, which is worthwhile further studying. In fact, it may indicate a possible involvement of nitration in signaling pathways and phosphorylation cascades. Regarding the aggregation studies, we have isolated detergent insoluble-protein fractions from spinal cord of G93A SOD1 mice and we have completed the comprehensive characterization of all the proteins contained. With this study we have identified the protein constituents of aggregates, still unknown, and therefore have contributed to the comprehension of the role of protein inclusions in ALS pathogenesis. Laboratory of Molecular Pathology Novel in vitro models for investigating motor neuron pathologies Presence of mutant forms of specific proteins plays a key role in many neurodegenerative diseases. Experimental models in vivo and in vitro are sorely needed to study the effects of these toxic proteins. Recently it was developed a new methodology (ptet-on/ptet-off) to control gene expression through the level of tetracyclines. We have applied the ptet-off system to a motor neuron-like cell line (NSC-34) establishing NSC-34 tta cell lines for tetracyclineregulated gene expression. These lines are suitable to study the pathogenetic mechanisms of motor neuron diseases after transient/stable transfection with genes of interest for these pathologies. We further used this approach to establish NSC-34 tta cell lines that express in a doxycycline inducible fashion the human G93A mutant Cu/Zn superoxide dismutase. Mutant forms of superoxide dismutase 1 are responsible for some of the familial forms of amyotrophic lateral sclerosis. This model allows to develop novel approaches to study pathogenic mechanisms of amyotrophic lateral sclerosis. Novel intracellular targets in the selective degeneration of motor neurons in amyotrophic lateral sclerosis Amyotrophic lateral sclerosis is a rapidly fatal neurodegenerative disease characterized by loss of motor neurons. The management remains essentially supportive and symptomatic. Understanding the mechanisms underlying the disease is a way to favor more efficient therapeutic strategies. Mitochondrial morphological alterations were observed in the early stages of the disease in the motor nerve terminals of ALS patients and in murine experimental models. For these reasons we addressed our studies to determine biochemical-molecular alterations involved in the mitochondrial damage utilizing different cellular models. We have studied the toxicity of mutant forms of superoxide dismutase 1, responsible for some familial forms of amyotrophic lateral sclerosis. We showed that mutant superoxide dismutase 1 (G93ASOD1) altered the mitochondrial morphology selectively in motor neuronal cells. This damage was modulated by the extent of expression of G93ASOD1. Furthermore presence of G93ASOD1 increased the susceptibility of motor neurons to inhibitors of the electron transport chain (ETC) and to oxidants. Exposure to drugs or exogenous compounds impairing the ETC could thus be a risk to motor neurons of individuals carrying mutant superoxide dismutase 1. Cytochrome P-450 superfamily Cytochrome(s) P-450 have evolved into a large superfamily that varies enormously in substrate affinity and product formation. This system plays a major role in the metabolism of drugs and other chemicals. The majority of existing drugs depends on the P-450 system for terminating their biological effects or for side effects or adverse reaction. The laboratory of Molecular Pathology has a long-standing interest in the induction/degradation mechanisms of specific cytochrome P-450 families due to drug administration or to disease states. Our recent research 166

169 focused on cytochrome P-450 induction by herbal remedies such as Hypericum perforatum extracts (St. John s Wort), which have an alleged activity in mild to moderate depression but interfere with the effect of several drugs. Activation of enzymes of the heme metabolic pathway (heme oxygenase system, biliverdin reductase) as a protective response to stress The enzymatic system of heme oxygenase (HO) is devoted to cellular degradation of heme containing molecules, like cytochromes and hemoglobin, and to recycling of iron. Products formed by the catalytic activity of HO - carbon monoxide and bile pigments - are important regulating factors in the cell. An increase of HO activity (which is usually sustained by activation of the inducible form HO-1) is now considered a protective mechanism against untoward stimuli particularly when oxidative stress is involved. In the past, the laboratory of Molecular Pathology identified cytokines as inducers of HO activity and as transcriptional activators of the HO-1 gene. We are currently investigating the functional significance of HO-1 activation in neurodegeneration. Laboratory for the Study of Biological Systems Novel regulators of cell motility Cell motility plays a central role in several biological processes, under both normal (e.g. embryonic development) and pathological conditions (e.g. tumor cell dissemination). Thus, it is important to identify the molecular mechanisms that regulate cell motility. In recent years, we have characterized Junctional Adhesion Molecule-A (JAM-A), a membrane molecule that localizes to the intercellular tight junctions and binds PDZ-type intracellular proteins. In the course of these studies, we have discovered that JAM-A expression reduces cell motility. In addition, we have found that JAM-A enhances microtubule stability and focal adhesion formation, which are the adhesive points of contact between cells and extracellular matrix. All these functional changes require amino acid residues that mediate binding to PDZ-type intracellular proteins. These findings have highlighted a novel mechanism of motility inhibition that requires the interaction between a membrane protein and PDZ-type intracellular proteins. Effect of inflammatory cytokines on Junctional Adhesion Molecule-A (JAM-A) In the course of inflammatory responses, JAM-A contributes to the leakage of plasma proteins and the transmigration of circulating leukocytes. Although it has been reported that the inflammatory cytokine Tumor Necrosis Factor (TNF) causes the disassembly of JAM-A from the intercellular junctions, the mechanism has not been elucidated fully. Recently, we found that TNF enhances the solubility of JAM-A in non-ionic detergents and increases the amount of detergent-soluble JAM-A at the cell surface. In addition, we found that, upon cell treatment with TNF, higher levels of JAM-A become detectable at the cell surface (by FACS analysis). As these higher levels of JAM-A derive from the intercellular junctions (and not from intracellular stores), we propose that TNF causes not only the disassembly of JAM-A from the junctions and its subsequent redistribution to the cell surface, but also its dispersal in such a way that JAM-A becomes more easily accessible to the antibodies used for FACS analysis. These findings are important to highlight potential mechanisms of permeability regulation during inflammation that might be modulated by inflammatory interventions. 167

170 DEPARTMENT OF EPIDEMIOLOGY STAFF Head Carlo LA VECCHIA, M.D. LABORATORY OF GENERAL EPIDEMIOLOGY Head Carlo LA VECCHIA, M.D. Cancer Epidemiology Unit Head Cristina BOSETTI, Mat.Sci.D. Lifestyle Habits and Prevention Unit Head Liliane CHATENOUD, Biol.Sci.D. Epidemiology for Clinical Research Unit Head Silvano GALLUS, Comp.Sci.D. LABORATORY OF EPIDEMIOLOGICAL METHODS Head Eva NEGRI, Mat.Sci.D. LABORATORY OF EPIDEMIOLOGY AND CHRONIC DISEASES Head Alessandra TAVANI, Biol.Sci.D. LABORATORY OF MEDICAL INFORMATICS Head Eugenio SANTORO, Comp.Sci.D. 168

171 CURRICULA VITAE Carlo La Vecchia holds a Doctor of Medicine from the University of Milan, a Master of Science in Clinical Medicine (epidemiology) from Oxford University and a Diploma from the Post-Graduate School of Pharmacological Research at the Mario Negri Institute for Pharmacological Research in Milan. Work experiences: he is Head of the Department of Epidemiology at the Mario Negri Institute for Pharmacological Research in Milan, Italy. He is also Associate Professor of Epidemiology at the University of Milan, Adjunct Professor of Epidemiology, University of Lausanne, Switzerland and Adjunct Professor of Medicine, School of Medicine, Vanderbilt University, Nashville, TN. Dr. La Vecchia is an Honorary Senior Lecturer in Oral Medicine at the Eastman Dental Institute at the University College of London, a temporary advisor at the International Agency for Research on Cancer IARC/WHO in Lyon and at the WHO in Geneva, and a registered journalist in Milan. He was Adjunct Associate Professor of Epidemiology at the Harvard School of Public Health between 1996 and He is Associate Editor to: European J. of Cancer Prevention and presently serves on the editorial boards of the: American J. of Epidemiology, Asian Pacific J. of Cancer Prevention, Cancer Causes and Control, Current Cancer Therapy Reviews, Digestive and Liver Disease, Economia Politica del Farmaco, European J. of Cancer, European J. of Cancer Prevention, European J. of Clinical Nutrition, European J. of Nutrition, Scope Oncology & Hematology, Int. J. Cancer, J. of Nephrology, Nutrition and Cancer, Oncology, Oral Oncology, Revisiones en Ginecología y Obstetricia, Revista Española de Nutrición Comunitaria, Revue d Epidémiologie et de Santé Publique, Sozial und Praeventivmedizin, The Lancet (Italian edition), Tumori, Alimentazione e Prevenzione. In 1993, he received the Glaxo Prize for medical publication. He has authored or co-authored over 1,360 publications in peer reviewed journals, with over 35,000 quotations. Eva Negri got a degree in Mathematics in 1985 at the University of Milan, School of Mathematics. Awards: EEC scholarship for postgraduate training in Epidemiology (1988). Areas of interest: Design, conduction and analysis of epidemiologic studies on chronic diseases (e.g. cancer and myocardial infarction) and injuries, analysis of mortality of cohorts of workers, analysis of temporal trends and geographic distribution of mortality from cancer, cardiovascular disease, injuries and other selected conditions, analysis of national health surveys, application of linear modeling techniques to the analysis of epidemiological data, collaborative re-analyses and meta-analyses of epidemiological studies. Work experiences: Since 2007: Laboratory Chief, Unit of Epidemiologic Methods, Department of Epidemiology; : Unit Chief, Unit of Epidemiologic Methods, Laboratory Epidemiology; since : Researcher at the Laboratory of Epidemiology; : Collaborator of the Laboratory of Epidemiology. Selected publications Negri E, La Vecchia C, Pelucchi C, Tavani A The risk of acute myocardial infarction after stopping drinking Prev Med 2005; 40: Negri E, Pelucchi C, Talamini R, Montella M, Gallus S, Bosetti C, Franceschi S, La Vecchia C Family history of cancer and the risk of prostate cancer and benign prostatic hyperplasia Int J Cancer 2005; 114: Negri E, Little D, Boiocchi M, La Vecchia C, Franceschi S. B-cell non-hodgkin s lymphoma and hepatitis C virus infection: A systematic review Int J Cancer 2004; 111: 1-8 Negri E, Ron E, Franceschi S, La Vecchia C, Preston-Martin S, Kolonel L, et al. Risk factors for medullary thyroid carcinoma: A pooled analysis Cancer Causes Control 2002; 13: Levi F, La Vecchia C, Boyle P, Lucchini F, Negri E Western and eastern European trends in testicular cancer mortality Lancet 2001; 357:

172 Alessandra Tavani degree in Biological Sciences, University of Milan, Italy (July 1977); Pharmacological Research Specialist, Mario Negri Institute for Pharmacological Research, Milan, Italy (July 1979). Work experiences: : Researcher at the laboratory of Drug Metabolism, Mario Negri Institute for Pharmacological Research. 1981: Researcher at the Unit for Research on Addictive Drugs (director prof. H.W. Kosterlitz), University of Aberdeen, Scotland, U.K : Head of the Unit of Opioid Neuropharmacology, Mario Negri Institute for Pharmacological Research. 1990: Researcher at the Unit of Clinical Perinatal Pharmacology, Mario Negri Institute for Pharmacological Research. From : Head of the Unit of Epidemiology of Chronic Diseases of the Laboratory of Epidemiology, Mario Negri Institute for Pharmacological Research : Head of the Laboratory of Epidemiology of Chronic Diseases of the Department of Epidemiology, Mario Negri Institute for Pharmacological Research. Awards: "Rafaelsen Scholar Award" from the Collegium Internationale Neuro-Psychopharmacologicum (CINP), 16th Meeting, Munich (F.R.G.), Areas of interest: Epidemiology of cancer and coronary heart disease. Selected publications Tavani A, Zucchetto A, Dal Maso L, Montella M, Ramazzotti V, Talamini R, Franceschi S, La Vecchia C. Lifetime physical activity and the risk of renal cell cancer. Int J Cancer 2007 ; 120 : Dal Maso L, Zucchetto A, Tavani A, Montella M, Ramazzotti V, Talamini R, Canzonieri V, Garbeglio A, Negri E, Tonini A, La Vecchia C, Franceschi S. Renal cell cancer and body size at different ages: An Italian multicentre casecontrol study. Am J Epidemiol 2007 ; 166 : Tavani A, Pelucchi C, Parpinel M T, Negri E, Franceschi S, Levi F, La Vecchia C. n-3 Polyunsaturated fatty acid intake and cancer risk in Italy and Switzerland. Int J Cancer 2003; 105: Herrero R, Castellsague X, Pawlita M, Lissowska J, Kee F, Balaram P, Rajkumar T, Sridhar H, Rose B, Pintos J, Fernandez L, Idris A, Sanchez M J, Nieto A, Talamini R, Tavani A, et al. Human papillomavirus and oral cancer: The International Agency for Research on Cancer Multicenter Study. J Natl Cancer Inst 2003; 95: Tavani A, Pelucchi C, Negri E, Bertuzzi M, La Vecchia C. n-3 polyunsaturated fatty acids, fish, and nonfatal acute myocardial infarction. Circulation 2001; 104: Eugenio Santoro got his degree in Computer Science in 1990 at the Milan University. He started to work at the Mario Negri Institute in 1985 as a research fellow. He was Head of the Applied Statistics and Informatics Unit and of the Applied Statistics and Informatics laboratory, which was part of the Department of Cardiovascular Research. Since 2001 he is Head of the Laboratory of Medical Informatics that is currently part of the Department of Epidemiology. His main areas of interest have been biostatistics and clinical informatics with the development of software for data management and data analyses of large scale clinical trials in cardiology, such as the GISSI studies (Gruppo Italiano per lo Studio della Sopravvivenza nell Infarto miocardico). His main current area of interest is the Internet, and more recently the web 2.0, and their application in the medical field, in clinical research, and in medical education through the development of health related websites. He is author or co-author of more than 140 scientific papers published in peer reviewed journals, and of more than 70 scientific abstracts submitted to the main international meetings in the cardiology and in the computer science fields. He is also author of two books (available in Italian) about the use of the Internet in medicine ( Guida alla medicina in rete and Internet in medicina. Guida all uso e applicazioni pratiche, both published by the Pensiero Scientifico Editore, Rome) and of one section about Internet and medicine, included in one of the most important italian medical encyclopedia ( Enciclopedia Medica Italiana, UTET 2007). He also collaborates to the publication of the Italian National Bioethics Committee s guidelines about ethics, health, and the new information technologies. He is a member of the Society for Clinical Trials and of the Society for Internet in Medicine. Selected publications Santoro E. Podcast, wiki e blog: il web 2.0 al servizio della formazione e dell aggiornamento del medico. Recenti Prog Med 2007;98: Santoro E, Rossi Valentina, Pandolfini C, Bonati M. DEC-NET: The development of the European register of clinical trials on medicines for children. Clin Trials 2006; 3: Clivio L, Tinazzi A, Mangano S, Santoro E. The contribution of information technology: Towards a better clinical data management. Drug Dev Res 2006; 67: Santoro E. Internet and information on breast cancer: an overview. Breast 2003; 12:

173 Santoro E, Nicolis E, Franzosi M G, Tognoni G. Internet for clinical trials: Past, present, and future. Control Clin Trials 1999; 20: Franzosi M G, Santoro E, Zuanetti G, Latini R, Maggioni A P, Tognoni G, GISSI. Indications for ACE inhibitors in the early treatment of acute myocardial infarction. Systematic overview of individual data from patients in randomized trial. Circulation 1998; 97: Franzosi M G, Santoro E, De Vita C, Geraci E, Lotto A, Maggioni A P, Mauri F, Rovelli F, Santoro L, Tavazzi L, Tognoni G, GISSI. Ten-year follow-up of the first megatrial testing thrombolytic therapy in patients with acute myocardial infarction. Results of Gruppo Italiano per lo Studio della Sopravvivenza nell Infarto-1 study. Circulation 1998; 98: Franzosi M G, Latini R, Maggioni A P, Barlera S, Negri E, Nicolis E, Santoro E, Santoro L, Tognoni G, Garattini S, GISSI-3. GISSI-3: Effects of lisinopril and transdermal glyceryl trinitrate singly and together on six-week mortality and ventricular function after acute myocardial infarction. Lancet 1994; 343: Maggioni A P, Maseri A, Fresco C, Franzosi M G, Mauri F, Santoro E, Tognoni G, GISSI-2. Age-related increase in mortality among patients with first myocardial infarctions treated with thrombolysis. N Engl J Med 1993; 11: Cristina Bosetti got her degree in Mathematics in 1994 at the University of Milan, School of Mathematics, and the Post-Graduate Diploma in Pharmacological Research in 1999 at the Mario Negri Institute for Pharmacological Research in Milan. Areas of interest: Epidemiology of cancer, cardiovascular diseases and other chronic conditions. In particular case-control studies on cancers of the upper respiratory and digestive sites, thyroid, breast, hormone-related cancers, and on ischemic heart disease. Analysis of risk related to diet, alcohol, tobacco, reproductive and hormonal factors, occupational and environmental exposure to toxic substances, through the application of generalized linear models. She is author/coauthor of more than 130 publications on these issues on peer-reviewed scientific journals. Work experiences: Since Sept. 2005: Unit Head, Unit of Cancer Epidemiology, Department of Epidemiology; : Researcher at the Laboratory of Epidemiology; : Researcher at the Laboratory of Mother and Child Health. Selected publications: Bosetti C, Gallus S, Peto R, Negri E, Talamini R, Tavani A, Franceschi S, La Vecchia C.Tobacco Smoking, Smoking Cessation, and Cumulative Risk of Upper Aerodigestive Tract Cancers.Am J Epidemiol Dec 4; [Epub ahead of print] Bosetti C, Malvezzi M, Chatenoud L, Negri E, Levi F, La Vecchia C. Trends in cancer mortality in the Americas, Ann Oncol 2005; 16: Bosetti C, Spertini L, Parpinel M T, Gnagnarella P, Lagiou P, Negri E, et al. Flavonoids and breast cancer risk in Italy. Cancer Epidemiol Biomarkers Prev 2005; 14: Bosetti C, Micelotta S, Dal Maso L, Talamini R, Montella M, Negri E, et al. Food groups and risk of prostate cancer in Italy. Int J Cancer 2004; 110: Bosetti C, Negri E, Kolonel L, Ron E, Franceschi S, Preston-Martin S, et al. A pooled analysis of case-control studies of thyroid cancer. VII. Cruciferous and other vegetables (International). Cancer Causes Control 2002; 13: Smith J S, Herrero R, Bosetti C, Munoz N, Bosch F X, Eluf-Neto J, et al. IARC Multicentric Cervical Cancer Study Group Herpes simplex virus-2 as a human papillomavirus cofactor in the etiology of invasive cervical cancer. J Natl Cancer Inst 2002; 94: Liliane Chatenoud, Doctor in Science Biology, University of Milan (1987); Postgraduate Doctor in Health Statistics University of Milan (1995). Areas of interest: Epidemiological studies on obstetric diseases. Dermato-epidemiology. Cancer epidemiology (case-control studies on cancers of the breast, female genital tract). Analysis of temporal trends and geographical distribution of perinatal, infant mortality, cancer and other selected conditions (over 100 publications on these topics, ). Work experiences: Since Sept. 2005: Unit Head, Lifestyle and Prevention, Department of Epidemiology : Researcher at the Laboratory of Epidemiology; : Staff Statistician Bracco S.p.A., Milan. Selected publications Chatenoud L, Mosconi P, Malvezzi M, Colombo P, La Vecchia C, Apolone G. Impact of a major thermoelectric plant on self-perceived health status. Prev Med. 2005;41: Naldi L, Chatenoud L, Linder D, Belloni Fortina A, Peserico A, Virgili AR, et al. Cigarette smoking, body mass index, and stressful life events as risk factors for psoriasis: results from an Italian case-control study. J Invest Dermatol. 2005;125:

174 Bosetti C, Malvezzi M, Chatenoud L, Negri E, Levi F, La Vecchia C. Trends in cancer mortality in the Americas, Ann Oncol 2005; 16: Chatenoud L, Tavani A, La Vecchia C, Jacobs D R, Negri E, Levi F, Franceschi S. Whole grain food intake and cancer risk. Int J Cancer 1998; 77: Chatenoud L, Parazzini F, Di Cintio E, Zanconato G, Benzi G, Bortolus R, La Vecchia C. Paternal and maternal smoking habits before conception and during the first trimester: Relation to spontaneous abortion. Ann Epidemiol 1998; 8: Silvano Gallus was born in Milan on the 20th of November 1970, and got his degree in Computer Science in 1999 at the Milan University. Areas of interest: Design, data managing, and statistical analyses of case-control studies on the associations between several risk factors (including in particular tobacco smoking, alcohol drinking and Mediterranean diet) and risk of cancer, coronary heart disease and several other conditions. Analyses of occupational cohort studies. Monitoring of prevalence and trends of smoking habit and obesity in Italy. Author/coauthor of over 130 publication in peer-reviewed journals since Work experiences: Chief of the Unit of Epidemiology for Clinical Research of the Department of Epidemiology (since 2006); computer analyst, graphic designer, and statistical and epidemiological consultant, Milan and Bergamo (since 2002); researcher at the Laboratory of Epidemiology (since 1997); creator, designer and webmaster of the website of one of the major Italian public hospital, Milano ( ). Selected publications Gallus S, Foschi R, Negri E, Talamini R, Franceschi S, Montella M, Ramazzotti V, Tavani A, Dal Maso L, La Vecchia C. Dietary zinc and prostate cancer risk: a case-control study from Italy. Eur Urol. 2007;52: Gallus S, Naldi L, Carli P, La Vecchia C; Italian Group for Epidemiologic Research in Dermatology (GISED). Nevus count on specific anatomic sites as a predictor of total body count: a survey of 3,406 children from Italy. Am J Epidemiol. 2007;166: Gallus S, Scotti L, Negri E, Talamini R, Franceschi S, Montella M, Giacosa A, Dal Maso L, La Vecchia C. Artificial sweeteners and cancer risk in a network of case-control studies. Ann Oncol. 2007;18:40-4. Gallus S, Schiaffino A, La Vecchia C, Townsend J, Fernandez E. Price and cigarette consumption in Europe. Tob Control. 2006;15: Gallus S, Zuccaro P, Colombo P, Apolone G, Pacifici R, Garattini S, La Vecchia C. Effects of new smoking regulations in Italy. Ann Oncol. 2006;17: Clifford GM, Gallus S, Herrero R, Muñoz N, Snijders PJ, Vaccarella S, Anh PT, Ferreccio C, Hieu NT, Matos E, Molano M, Rajkumar R, Ronco G, de Sanjosé S, Shin HR, Sukvirach S, Thomas JO, Tunsakul S, Meijer CJ, Franceschi S; IARC HPV Prevalence Surveys Study Group. Worldwide distribution of human papillomavirus types in cytologically normal women in the International Agency for Research on Cancer HPV prevalence surveys: a pooled analysis.lancet. 2005;366:

175 INTRODUCTION TO THE DEPARTMENT S ACTIVITIES The Department of Epidemiology is involved in the epidemiology of several common cancers (including cancers of the breast, female genital tract, respiratory and digestive sites, prostate and urinary organs, lymphoid malignancies, melanoma, etc.) and of cardiovascular diseases, both through a descriptive and an analytical approach. Among the activities of descriptive epidemiology are the analysis of temporal trends and geographical distribution of mortality from cancer, cardiovascular diseases, and other selected conditions, in Italy and Europe; the analysis of trends in tobacco consumption in the Italian population, and the corresponding effects on incidence and mortality from lung and other tobacco-related neoplasms. The analytic epidemiology activities include the conduction and analysis of case-control studies, aimed at identifying and better quantifying the association between genetic factors (family history), selected lifestyle habits (diet, tobacco, alcohol, etc.), use of exogenous hormones and exposure to various substances and the development of various forms of cancers and cardiovascular diseases. In particular, the Department works on the analysis of dietary correlates of cancer and cardiovascular disease risk; quantification of health effects of tobacco smoking, alcohol consumption and implications for prevention; epidemiological studies on the risk related to oral contraceptive and hormone replacement therapy use; evaluation of the impact of screening in the early diagnosis and prevention of cancer. Other activities include: the conduction of quantitative reviews and meta-analysis of published data; the re-analysis of original data from epidemiological studies of cancers of the thyroid, breast, ovary, and cervix; epidemiological and clinical studies in dermatology in collaboration with the Gruppo Italiano per gli Studi Epidemiologici in Dermatologia (GISED); the analysis of historical cohort studies of occupational exposures to aromatic amines, asbestos, herbicides and other known carcinogens; monitoring and prevention of injuries. Other Department s activities are related to the development of medical websites, the study of the quality of medical information available on the Internet, and the training and research on issues related to medical informatics and those concerning the use in the medical field of the Internet and web 2.0 applications. FINDINGS/MAIN RESULTS In the Italian population, men who stopped smoking before age 50 years avoided more than half of the excess risk of upper aerodigestive tract cancer as men who did not, and men who stopped smoking before age 30 years avoided more than 90% of the risk. Flavonoids, particularly flavanones, are inversely related to the risk of cancers of the upper aero-digestive tract, including oral cavity and pharynx, oesophagus and larynx. A family history of oral, pharyngeal and laryngeal cancer is a strong determinant of oral and pharyngeal cancer risk, independent from tobacco and alcohol use. Among never drinkers, cigarette smoking is associated with an increased risk of head and neck cancer, and among never users of tobacco, alcohol consumption is associated with an increased risk of head and neck cancer only when alcohol is consumed at high frequency. Compared to never smokers, exclusive pipe smokers have a 9-fold increased risk of all upper digestive tract cancers. Pipe smoking and heavy alcohol drinking appear to interact at least on a multiplicative model. 173

176 History of thyroid diseases and goiter does not appear to be a relevant cause of gastric cancer in Italy. There is no relevant role of fried foods consumption on colorectal cancer risk, while there is a possible favorable effect of (fried) olive oil on colon but not on rectal cancer risk. Coffee has a favourable effect on hepatocellular carcinoma. There is an inverse relation between a diet rich in linoleic acid and beta-carotene intake and hepatocellular carcinoma risk. The lifetime number of ovulatory cycles is directly associated with ovarian cancer risk, while pregnancy and oral contraceptive use have a stronger protective effect on ovarian cancer than other anovulatory factors. A starch-rich dietary pattern is an unfavorable indicator of risk for both breast and ovarian cancers, while the animal products and the vitamins and fiber patterns might be associated with a reduced risk of breast and ovarian cancers, respectively. Wine and other types of alcoholic beverages do not influence ovarian cancer risk in the Italian population. High energy and cholesterol intake might increase endometrial cancer risk in Italy. Prostate cancer risk is increased for subjects at the highest quintile of zinc intake, as compared to the lowest one. The direct association is stronger for advanced than for early stage cancers. The major risk factors for prostate cancer observed in middle-aged men (i.e., <60 years) are family history of prostate cancer, high level of education, and low physical activity. A diet rich in refined cereals and poor in vegetables may have an unfavorable role on renal cell cancer. Vitamin E, vitamin C and vegetable fibers have a significant inverse association, that may reflect a real favorable effect, or be an indicator of a beneficial role of a diet rich in vegetable on renal cell cancer risk. Obese subjects (i.e., body mass index 30) have an increased risk of renal cell cancer. Risks among obese persons are apparently higher in never smokers, persons with the clear-cell histologic type, and persons with a Fuhrman nuclear grade of G3-G4. About 10% of cases of renal cell cancer in Italy could be avoided by increasing physical activity. Patients with a history of treated hypertension reported an excess risk of renal cell cancer. There is an inverse association between alcohol drinking and renal cell cancer. Risks continued to decrease even above 100 g/day of alcohol intake, with no apparent leveling in risk. There is no strong association between diabetes and non-hodgkin lymphoma. The small number of cases with diabetes analyzed left open the possibility of a moderate direct relation. 174

177 There are no significant associations between intakes of folate nor other micronutrients involved in the one-carbon metabolism and non-hodgkin lymphoma risk. However, a significant inverse association is observed for all the nutrients examined among abstainers and former drinkers, supporting the possibility of an antagonist effect of alcohol on the one-carbon metabolism in non-hodgkin lymphoma etiology. Non-Hodgkin lymphoma risk is elevated for individuals who reported first-degree relatives with lymphomas and leukemia. The pattern of non-hodgkin lymphoma heritability appear to be uniform across non-hodgkin lymphoma subtypes, but risk patterns differ by specific hematopoietic malignancies and the sex of the relative, revealing critical clues to disease etiology. There is no evidence to support the hypothesis that obesity is a determinant of all types of non- Hodgkin lymphoma combined, while the association observed between severe obesity and diffuse large B-cell lymphoma may warrant further investigation. No direct association emerged between use of saccharin, aspartame and other sweeteners and the risk of various common neoplasms. High dietary levels of glycemic index and load increase the risk of thyroid cancer. Subjects with at least a first-degree relative with cancer of the bladder, kidney or prostate have a 6-fold increased risk of bladder cancer, a 2-fold increased risk of renal cell cancer and a 2-fold increased risks of prostate cancer, respectively. The reliability of the information on family history of cancer in first-degree relatives in our hospital-based case-control study of digestive tract cancers is satisfactory. No substantial reduction in acute myocardial infarction risk resulted from the decrease of cigarette tar yield. Parity and irregular menstrual cycle increased acute myocardial infarction risk, mainly in smokers. Lung cancer mortality in European women is levelling off in Germany, Italy and Poland over the last years, while it is still steadily increasing in France and Spain. Mortality from cirrhosis shows favorable trends in most countries of the world, but is still steadily upward in the UK and central and eastern European countries, due to the persistent increase in the prevalence of alcohol consumption. Mortality from testicular cancer in (young) men remains exceedingly high in most Latin American countries. Body mass index affects early clinical response to systemic treatment for psoriasis irrespectively of the administered drug. Examining the upper limbs only, particularly the lateral arms, is a practical and suitable tool for predicting total nevus count in children. The BPCO.CARE website ( a web based medical index that collect, classify, evaluate, and describe the most useful medical information on the web related to the chronic 175

178 obstructive pulmonary disease) has been developed, including a collection of the web 2.0 tools and applications available in the medical field. NATIONAL COLLABORATIONS Associazione Nazionale dei Medici Cardiologi Ospedalieri (ANMCO) Centro di Riferimento Oncologico, Servizio di Epidemiologia, Aviano (PN) Fondazione LuVI Fondazione SmithKline, Milano Gruppo Italiano per lo Studio della Sopravivenza nell Infarto miocardico (GISSI) Gruppo Italiano Studi Epidemiologici in Dermatologia GISED, Bergamo International Centre for Pesticides and Health Risk Prevention, Milano Istituto Auxologico Italiano, Divisione Malattie Metaboliche III, IRCCS, Piancavallo (VB) Istituto Auxologico Italiano, Laboratorio Sperimentale di Ricerche Endocrinologiche (LSRE), IRCCS, Milano Istituto di Medicina del Lavoro, CTO, Torino Istituto Europeo di Oncologia, Divisione di Epidemiologia e Biostatistica, Milano Istituto Europeo di Oncologia, Divisione di Chirurgia Cervico Facciale, Milano Istituto Nazionale di Ricerca per gli Alimenti e la Nutrizione (INRAN), Roma Istituto Nazionale Neurologico "Carlo Besta", Milano Istituto Nazionale per lo Studio e la Cura dei Tumori, Oncologia Sperimentale, Unità di Eredità Poligenica, Milano Istituto Tumori Fondazione Pascale, Servizio di Epidemiologia, Napoli Novartis Vaccines SpA, Siena Ospedale Niguarda Ca Granda, Dipartimento Trapianti di Fegato, Milano Ospedale Niguarda Ca Granda, Istituto di Fisiologia Clinica CNR, Sezione di Milano, Milano Policlinico di Monza, Unità Operativa di Endoscopia I, Monza (MI) Prima Clinica Ostetrico Ginecologica, Mangiagalli, Milano Società Italiana Attività Regolatorie Università degli Studi di Milano - Bicocca, Dipartimento di Statistica, Milano Università degli Studi di Milano, Milano Università degli Studi di Milano-Bicocca, I Clinica Otorinolaringoiatria, DNTB, Monza Università di Milano, Clinica Pediatrica De Marchi, Milano Università di Milano, Istituto di Statistica Medica e Biometria G.A. Maccacaro, Milano Università di Milano, Prima Clinica Ostetrico Ginecologica, Milano Università di Verona, Clinica Ostetrico Ginecologica, Verona INTERNATIONAL COLLABORATIONS Catalan Institute of Oncology, Institut d Investigaciò Biomédica de Bellvitge (IDIBELL), Cancer Prevention and Control Unit, L Hospitalet de Llobregat, Spain Center of Oncology, Dept. of Epidemiology and Cancer Prevention, Varsavia, Poland Centre for Research in Environmental Epidemiology (CREAL) and Municipal Institute of Medical Research (IMIM), Barcellona, Spain Harvard School of Public Health, Department of Epidemiology, Boston, USA Hôpital Necker - Enfants Malades, Centre of the Association Claude Bernard on Auto-immunes diseases, Parigi, France 176

179 International Agency for Research on Cancer, Lione, France International Epidemiology Institute (IEI), Rockville, USA International Life Science Institute (ILSI), Bruxelles, Belgium National Cancer Institute, Environmental Studies Section, Bethesda, USA National Cancer Institute, Radiation Epidemiology Branch, Bethesda, USA National School of Public Health, WHO, Atene, Greece Registre Vaudois des Tumeurs, Institut Universitaire de Médecine Sociale et Préventive, Losanna, Switzerland Senologic International Society Society for Internet in Medicine Universitat Pompeu Fabra, Department of Experimental and Health Sciences, Barcellona, Spain University of Athens Medical School, Department of Hygiene and Epidemiology, Atene, Greece University of Las Palmas de Gran Canaria, Department of Clinical Sciences, Las Palmas de Gran Canaria, Spain Vanderbilt University, Department of Medicine, School of Medicine, Nashville, USA EDITORIAL BOARD MEMBERSHIP Advances in Therapy (Eva Negri) Alimentazione e Prevenzione (Carlo La Vecchia) Asian Pacific Journal of Cancer Prevention (Carlo La Vecchia) Digestive and Liver Disease (Carlo La Vecchia) European Journal of Cancer Prevention (Carlo La Vecchia, Associate Editor) European Journal of Nutrition (Carlo La Vecchia) Evidence Based Dermatology (Carlo La Vecchia, Liliane Chatenoud) International Journal of Cancer (Carlo La Vecchia) Journal of Nephrology (Carlo La Vecchia) Nutrition and Cancer (Carlo La Vecchia) Portale Partecipasalute.it (Eugenio Santoro) Revisiones en Ginecologìa y Obstetricia (Carlo La Vecchia) Revista Española de Nutriciò Comunitaria (Carlo La Vecchia) Revue d Epidémiologie et de Santé Publique (Carlo La Vecchia) Società Italiana Attività Regolatorie News, SIARNews (Eugenio Santoro) The Lancet, edizione italiana (Carlo La Vecchia) Tumori (Carlo La Vecchia) PEER REVIEW ACTIVITIES Acta Psychiatrica Scandinavica, American Journal of Clinical Nutrition, American Journal of Epidemiology, Annals of Epidemiology, Annals of Oncology, Archives of Internal Medicine, BMC-Public Health, British Journal of Cancer, British Journal of Nutrition, British Medical Journal, Canadian Journal of Physiology and Pharmacology, Cancer, Cancer Causes and Control, Cancer Epidemiology Biomarkers and Prevention, Computer Methods and Programs in Biomedicine, Diabetes/Metabolism Research and Reviews, Digestive Liver Disease, Epidemiologia & Prevenzione, Epidemiology, Epidemiology & Biostatistic, European Heart Journal, European Journal of Cancer, European Journal of Cancer Prevention, European Journal of Clinical Nutrition, European Journal of Epidemiology, European Journal of Public Health, Evidence-Based Healthcare & Public Health, Gynecological Endocrinology, Gut, Hepatology, 177

180 Human Reproduction, International Journal of Cancer, International Journal of Epidemiology, International Journal of Obesity, JAMA, Journal of American College of Nutrition, Journal of Clinical Endocrinology and Metabolism, Journal of Clinical Epidemiology, Journal of Epidemiology and Community Health, Journal of the National Cancer Institute, Maturitas, Nicotine & Tobacco Research, Nutrition and Cancer, Obstetric and Gynecology, Oncology, Preventive Medicine, Radiation Research, Revue d Epidèmiologie et de Santé Publique, The Breast, The Cancer Journal, The Lancet, Tumori. NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Advisory Committee of the Oxford Collaborative group on Aetiological Factors in Cancers of the Female Genital Tract Comitato Scientifico del Gruppo Italiano Studi Epidemiologici in Dermatologia Comitato Scientifico della Società Italiana di Colposcopia e Patologia Cervico Vaginale Consiglio Superiore di Sanità, Gruppo di lavoro sul Metodo Di Bella Data and Safety Monitoring Board of the Phase II therapeutic trial with a humanized nonmitogenic CD3 (ChAgly CD3) monoclonal antibody in recently diagnosed type I diabetic patients IARC/WHO, Lyon, and WHO, Geneva Istituti Clinici di Perfezionamento, Milano Progetto Menopausa Italia, Associazione Ostetrici Ginecologi Italiani Scientific Review Committee of UND/WHO/World Bank Human Reproduction Program Società Italiana della Riproduzione Ministero della Salute, Sottocomitato fumo. Ministero della Salute, Commissione Oncologica Nazionale. 178

181 PARTICIPATION IN EVENTS IN WHICH THE LABORATORY WAS INVOLVED Workshop on Environment & Health: Air quality research needs and opportunities in the EU seventh framework programme of research (FP7). CONCAWE. European studies on ambient particulate matter and health, with a focus on long-term exposure. Brussels, Belgium. 15 January th Annual Inhance Meeting. IARC. Family history of oral cancer. Lyon, France January 2007 Press Lunch AIRC. Le sostanze che fanno bene: ricerca sui flavonoidi e vitamina C per difendersi dal cancro. Milan, Italy. 25 January 2007 IARC Monographs on the Evaluation of Carcinogenic Risks to Humans. Volume 96: Alcoholic beverage consumption, acetaldehyde and urethane. Lyon, France February 2007 Hi-wate Kick-Off Meeting. Barcelona, Spain February 2007 HEM Closing the Gap in central Europe. Meeting of country coordinators. Warsaw, Poland. 5-6 March 2007 Incontro Conferenza stampa. Presentazione dell analisi dei risultati a 5 anni di tre studi, condotti da Mayo Clinic, Lee Moffit Center e INT/IEO, eseguita dagli epidemiologi dello Sloan Kettering, sull utilità della diagnosi precoce con TAC spirale come strumento di individuazione precoce del tumore al polmone, pubblicata su The Journal of American Medical Association e commentata su Nature. Milan, Italy. 7 March 2007 IARC Fellowships Committee Meeting Lyon, France March 2007 IV Convegno Nazionale. La tutela della salute nelle attività sportive e la lotta contro il doping. Un indagine sulla conoscenza, attitudine e prevalenza del doping nell opinione degli atleti italiani. Rome. 3 April 2007 ESMO International Symposium. ESMO. The smoking epidemic and lung cancer. Geneva, Switzerland. 30 Marzo-1 April Porto Cancer Meeting. XVI Edition. Cancer etiology: Brindging worlds. Cancer mortality trends in the European Union: priorities for cancer control. Porto, Portugal April Giornata mondiale senza tabacco. IX Convegno nazionale tabagismo e servizio sanitario nazionale. Politiche di controllo del fumo: quali azioni da intraprendere. Rome. 31 May 2007 The future of medical sciences. Infectious diseases: new frontiers and migration pathologies. Milan. 8 June National Congress SIB Italian Biometric Society. New perspective on parametric confidence intervals for the cost-effectiveness ratio. Pisa, Italy June 2007 Forum dei centri per la disassuefazione dal fumo. Epidemiologia della disassuefazione e stratificazione delle popolazioni che accedono all ambulatorio specialistico. Rome. 5 July Insediamento della Commissione Oncologica Nazionale. Rome. 12 July st Meeting of Collaborators. Collaborative Group on epidemiological studies of endometrial cancer. St Hilda s College, Oxford, UK July

182 56 International congress ISI - International Statistical Institute. New perspective on confidence intervals for the ratio of means of Normal Variables, jointly distributed as a Bivariate Normal. Lisbon, Portugal August ERA-net-2007-rtd evaluation. Evaluation. Brussels, Belgium. 3-4 September Journèes Internationales Huiles Essentielles & Extraits. L Huile d olive: Santè, bien-être et prevention du cancer. Digne Les Bains, France September IV National Congress SISMEC Italian Society of Medical Statistics and Clinical Epidemiology. Bioequivalence assessment from crossover data: a new approach for the construction of confidence intervals for the ratio of two formulation means. Epidemiologia. Monreale September I determinanti della salute: una valutazione quantitativa. Sassari, Italy. 5-6 October Conferenza. Osteonecrosi della mandibola (ONJ) in corso di terapia con bisfosfonati. Milan. 8 October Congresso Nazionale FIMMG-METIS. Caffè e cancro, con riferimento ad altre patologie dell apparato digerente. Villasimius October Conferenza Lega Italiana per la Lotta contro i Tumori. L alimentazione nella prevenzione dei tumori. Castelfranco Veneto. 11 October Le giornate nazionali di nutrizione pratica Dieta mediterranea e cancro. Elementi di prevenzione. Milan.18 October Closing the Gap final conference. The evolution of health following enlargement HEM. Cardiovascular disease. Brussels, Belgium. 23 October PSONET European registry of psoriasis. International safety review board. Rome. 25 October XXXI Congresso Annuale AIE. Fattori di rischio per le cadute nell anziano: scopo per una revisione sistematica, Interventi di prevenzione delle cadute negli anziani con particolare attenzione agli aspetti relativi alla partecipazione dei soggetti. Ostuni October Giornata Nazionale per la Ricerca sul Cancro organizzata Associazione Italiana Ricerca sul Cancro. I danni al nostro DNA provocati dall ambiente. 10 November th Scietific Meeting of the International Epidemiology Association. EMR: epidemiology translating science to health care. Diet and cancer risk in mediterranean countries Riyadh, Saudi Arabia November Vino e salute. Convegno scientifico divulgativo. L aumento della longevità associato al corretto consumo di vino Perché occorre evitare l abuso di bevande alcoliche. Grinzane Cavour. 24 November 2007 XXXXIX Congresso nazionale AIPO. Malattie respiratorie: emergenza sociale. Epidemiology of lung cancer. Florence. 4-7 December Convegno regionale AIOM Lombardia. Riduzione della mortalità per cancro in Europa: illusione o realtà?. Milan. 14 December XII World Congress on Internet in Medicine. GCPBASE: a web-based tool for remote data capture in a clinical trial. Lipsia October

183 XXII Workshop GIDIF, RBM (Gruppo Italiano Documentalisti dell Industria Farmaceutica e degli Istituti di Ricerca Biomedica), Palazzo delle Stelline. Tecnologia RSS e aggregatori di notizie: un nuovo modo di distribuire l informazione. Milan. 4 December 2007 Master Universitario di II livello in Informatica medica, Università di Udine, anno accademico Ruolo di docenza nel modulo Internet e sanità; strumenti e applicazioni al servizio del medico e del cittadino. Udine. 24 May Corso, Internet e l aggiornamento professionale in ambito biomedico promosso da ULSS n.6 Vicenza. Siti e applicazioni Internet in ambito medico. Vicenza. 6 June 2007 Corso, Medicina basata sulle prove di efficacia promosso dall Azienda Unità Sanitaria Locale n.1 di Sassari. Siti e applicazioni Internet in ambito medico: tipologia di informazione e modalità di reperimento. Sassari. 7 May Corso, Medicina basata sulle prove di efficacia promosso dall Azienda Unità Sanitaria Locale n.1 di Sassari. Siti e applicazioni Internet in ambito medico: tipologia di informazione e modalità di reperimento. Sassari. 21 September AIFA Associazione Italiana per la Ricerca sul Cancro AstraZeneca Italia Comune di Milano Lega Italiana Lotta contro i Tumori European Commission (FP6) GISED Ministero della Salute Regione Lombardia GRANTS AND CONTRACTS 181

184 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 La Vecchia C. Hormone replacement therapy in menopause and breast, colorectal and lung cancer: an update. In: Treatment of the Postmenopausal Woman: Basic and Clinical Aspects, ed 3, Lobo R.A. ed. Academic Press, 2007, pp (2007). Gallus S, La Vecchia C. Is there a link between diet and esophageal cancer? Nature Clin. Practice Gastroenterol. & Hepatol., 4: 2-3 (2007). Dal Maso L, La Vecchia C, Augustin LSA, Mantzoros CS, Kendall CWC, Franceschi S. Relationship between a wide range of alcohol consumptions, components of the insulin-like growth factor system and adiponectin. Eur. J. Clin. Nutr., 61: (2007). Galeone C, Talamini R, Levi F, Pelucchi C, Negri E, Giacosa A, Montella M, Franceschi S, La Vecchia C. Fried foods, olive oil and colorectal cancer. Ann. Oncol., 18: (2007). Randi R, Pelucchi C, Gallus S, Parpinel M, Dal Maso L, Talamini R, Augustin LSA, Giacosa A, Montella M, Franceschi S, La Vecchia C. Lipid, protein and carbohydrate intake in relation to body mass index: an Italian study. Publ. Health Nutr., 10: (2007). La Vecchia C., Bosetti C. Diet and cancer risk in Mediterranean countries. Hungarian Med. J., 1: (2007) Colosio C, Fustinoni S, Corsini E, Bosetti C, Birindelli S, Boers D, Campo L, La Vecchia C, Liesivuori J, Pennanen S, Pennanen S, Vergieva T, van Amelsvoort L.G.P.M., Steerenberg P, Swaen GMH, Zaikov C, van Loveren H. Changes in serum markers indicative of health effects in vineyard workers following exposure to the fungicide mancozeb: an Italian study. Biomarkers, 12: (2007). Bravi F, Bosetti C, Negri E, Lagiou P, La Vecchia C. Family history of cancer provided by hospital controls is satisfactorily reliable. J. Clin. Epidemiol., 60: (2007). Chiaffarino F, Parazzini F, Bosetti C, Franceschi S., Talamini R, Canzonieri V, Montella M, Ramazzotti C, La Vecchia C. Risk factors for ovarian cancer histotypes. Eur. J. Cancer., 43: (2007). Scotti L, Tavani A, Bosetti C, Dal Maso L, Talamini R, Montella M, Franceschi S, La Vecchia C. Diabetes and risk of non-hodgkin lymphoma: a case-control study. Tumori, 93: 1-3 (2007). Galeone C, Negri E, Pelucchi C, La Vecchia C, Bosetti C, Hu J. Dietary intake of fruit and vegetable and lung cancer risk: a case-control study in Harbin, northeast China. Ann. Oncol., 18: (2007). Bosetti C, Boffetta P, La Vecchia C. Occupational exposures to polycyclic aromatic hydrocarbons, and respiratory and urinary tract cancers: a quantitative review to Ann. Oncol., 18: (2007). Pelucchi C, Galeone C, Talamini R, Bosetti C, Montella M, Negri E, Franceschi S, La Vecchia C. Lifetime ovulatory cycles and ovarian cancer risk in 2 Italian case-control studies. Am..J Obstet. Gynecol., 196: 83.e1-83.e7 (2007). La Vecchia C, Tavani A. Coffee and cancer risk: an update. 182

185 Eur. J. Cancer Prev., 16: (2007). La Vecchia C., Negri E., International Collaboration of Epidemiological Studies of Cervical Cancer. Cervical cancer and hormonal contraceptives: collaborative reanalysis of individual data on women with cervical cancer and women without cervical cancer from 24 epidemiological studies. Lancet, 370: (2007). Montella M, Polesel J, La Vecchia C, Dal Maso L, Crispo A, Crovatto M, Casarin P, Izzo G, Tommasi LG, Talamini R, Franceschi S. Coffee and tea consumption and risk of hepatocellular carcinoma in Italy. Int. J. Cancer, 120: (2007). Gallus S, Scotti L, Negri E, Talamini R, Franceschi S, Montella M, Giacosa A, Dal Maso L, La Vecchia C. Artificial sweeteners and cancer risk in a network of case-control studies. Ann. Oncol., 18: (2007). Bertuccio P, Tavani A, Gallus S, Negri E, La Vecchia C. Menstrual and reproductive factors and risk of non-fatal acute myocardial infarction in Italy. Eur. J. Obst. Gynecol. Repr. Biol., 134: (2007). Pira E, Pelucchi C, Piolatto PG, Negri E, Discalzi G, La Vecchia C. Role of first and subsequent asbestos exposures on mesothelioma and lung cancer. Br. J. Cancer, 97: (2007). Polesel J, Dal Maso L, La Vecchia C, Montella M, Spina M, Crispo A, Talamini R, Franceschi S. Dietary folate, alcohol consumption, and risk of non-hodgkin Lymphoma. Nutr. Cancer, 57: (2007). Gallus S., Foschi R, Negri E, Talamini R, Franceschi S, Montella M, Ramazzotti V, Tavani A, Dal Maso L, La Vecchia C. Dietary zinc and prostate cancer risk: a case-control study from Italy. Eur. Urol., 52: (2007). Tavani A, Zucchetto A, Dal Maso L, Montella M, Ramazzotti V, Talamini R, Franceschi S, La Vecchia C Lifetime physical activity and the risk of renal cell cancer. Int. J. Cancer, 120: (2007) Pelucchi C, Galeone C, Dal Maso L, Talamini R, Montella M, Ramazzotti V, Negri E, Franceschi S, La Vecchia C. Dietary acrylamide and renal cell cancer. Int. J. Cancer, 120: (2007). Gallus S, Zuccaro P, Colombo P, Apolone G, Pacifici R, Garattini S, Bosetti C, La Vecchia C. Smoking in Italy : Effects of a comprehensive national tobacco regulation. Prev. Med., 45: (2007). Gallus S, Naldi L, Carli P, La Vecchia C, and the Italian Group for Epidemiologic Research in Dermatology (GISED). Nevus count on specific anatomic sites as a predictor of total body count: a survey of 3,406 children from Italy. Am. J. Epidemiol., 166: (2007). Levi F, Randimbison L, Maspoli M, Te VC, La Vecchia C. Second neoplasms after oesophageal cancer. Int. J. Cancer, 121, (2007). Bosetti C. Levi F. Lucchini F, Zatonski WA, Negri E, La Vecchia C. Worldwide mortality from cirrhosis: An update to J. Hepatol., 46: (2007). Fossati R, Apolone G, Negri E, Compagnoni A, La Vecchia C, Mangano S, Clivio L, Garattini S for the General Practice Tobacco Cessation Investigators Group. A double-blind, placebo controlled, randomized trial of bupropion for smoking cessation in primary care. Arch. Intern. Med., 167: (2007). 183

186 Rossi M, Garavello W, Talamini R, La Vecchia C, Franceschi S, Lagiou P, Zambon P, Dal Maso L, Bosetti C, Negri E. Flavonoids and risk of squamous cell esophageal cancer. Int. J. Cancer, 120: , (2007). Bertuccio P, Malvezzi M, Chatenoud L, Bosetti C, Negri E, Levi F, La Vecchia C. Testicular cancer mortality in the Americas ( ). Cancer, 109: (2007). Galeone C, Pelucchi C, Talamini R, Negri E, Dal Maso L, Montella M, Ramazzotti V, Franceschi S, La Vecchia C. Onion and garlic intake and the odds of benign prostatic hyperplasia. Urology, 70: (2007). Zucchetto A, Dal Maso L, Tavani A, Montella M, Ramazzotti V, Talamini R, Canzonieri V, Garbeglio A, Negri E, Franceschi S, La Vecchia C. History of treated hypertension and diabetes mellitus and risk of renal cell cancer. Ann. Oncol., 18: (2007). Wang SS, Slager SL, Brennan P, Holly EA, De Sanjose S, Bernstein L, Boffetta P, Cerhan JR, Maynadie M, Spinelli JJ, Chiu BCH, Cocco PL, Mensah F, Zhang Y, Nieters A, Dal Maso L, Bracci PM, Seniori Costantini A, Vineis P, Severson RK, Roman E, Cozen W, Weisenburger D, Davis S, Franceschi S, La Vecchia C, Foretova L, Becker N, Staines A, Vornanen M, Zheng T, Hartge P. Family history of hematopoietic malignancies an risk of non-hodgkin lymphoma (NHL): a pooled analysis of cases and controls from the International Lymphoma Epidemiology Consortium. (InterLymph). Blood, 109: (2007). Bosetti C, Rossi M, McLaughlin JK, Negri E, Talamini R, Lagiou P, Montella M, Ramazzotti V, Franceschi S, La Vecchia C. Flavonoids and the risk of renal cell carcinoma. Cancer Epidemiol. Biomarkers Prev., 16: (2007). Garavello W, Rossi M, McLaughlin JK, Bosetti C, Negri E, Lagiou P, Talamini R, Franceschi S, Parpinel M, Dal Maso L, La Vecchia C. Flavonoids and laryngeal cancer risk in Italy. Ann. Oncol., 18: (2007). Bravi F., Bosetti C., Tavani A., Bagnardi V., Gallus S., Negri E., Franceschi S., La Vecchia C. Coffee drinking and hepatocellular carcinoma risk: a meta-analysis. Hepatology, 46: (2007) La Vecchia C. Alcohol and liver cancer. Eur. J. Cancer Prev., 16: (2007). Levi F, Lucchini F, Negri E, La Vecchia C. Continuing declines in cancer mortality in the European Union. Ann. Oncol., 18: (2007). Dal Maso L, Zucchetto A, Tavani A, Montella M, Ramazzotti V, Talamini R, Canzonieri V, Garbeglio A, Negri E, Tonini A, La Vecchia C, Franceschi S. Renal cell cancer and body size at different ages: an Italian multicenter case-control study. Am. J. Epidemiol., 166: (2007). Hashibe M, Brennan P, Benhamou S, Castellsague X, Chen C, Curado MP, Dal Maso L, Daudt AW, Fabianova E, Wünsch-Filho V, Franceschi S, Hayes RB, Herrero R, Koifman S, La Vecchia C, Lazarus P, Levi F, Mates D, Matos E, Menezes A, Muscat J, Eluf-Neto J, Olshan AF, Rudnai P, Schwartz SM, Smith E, Sturgis EM, Szeszenia- Dabrowska N, Talamini R, Wei Q, Winn DM, Zaridze D, Witold Zatonski W, Zhang ZF, Berthiller J, Boffetta P. Alcohol drinking in never users of tobacco, cigarette smoking in never drinkers, and the risk of head and neck cancer: pooled analysis in the International Head and Neck Cancer Epidemiology Consortium (Inhance). JNCI, 99: (2007) Gallus S., Foschi R., Talamini R., Altieri A., Negri E., Franceschi S., Montella M., Dal Maso L., Ramazzotti V., La Vecchia C. 184

187 Risk factors for prostate cancer in men aged less than 60 years. A case-control study from Italy. Urology, 70: (2007). Levi F, Bosetti C, Fernandez E, Hill C, Lucchini F, Negri E, La Vecchia. Trends in lung cancer in young European women: the rising epidemic in France and Spain. Int. J. Cancer, 121: (2007). Naldi L, Chatenoud L, Bertuccio P, Zinetti C, Di Landro A, Scotti L, La Vecchia C, and the Oncology Cooperative Group of the Italian Group for Epidemiologic Research in Dermatology (GISED). Improving sun protection behavior among children: results of a cluster-randomized trial in Italian elementary schools. The "SoleSi SoleNo-GISED" project. J. Invest. Dermatol., 127: (2007). Randi G, Scotti L, Bosetti C, Talamini R, Negri E, Franceschi S, Levi F, La Vecchia C. Pipe smoking and cancers of the upper digestive tract. Int. J., Cancer, 121: (2007). Rossi M., Garavello W., Talamini R., Negri E., Bosetti C., Dal Maso L., Lagiou P., Tavani A., Polesel J., Barzan L., Ramazzotti V., Franceschi S., La Vecchia C. Flavonoids and the risk of oral and pharyngeal cancer: a case-control study from Italy. Cancer Epidemiol. Biomarkers Prev., 16: (2007). Polesel J., Talamini R., Montella M., Dal Maso L., Crovatto M., Parpinel M., Izzo F., Tommasi LG, Serraino D, La Vecchia C, Franceschi S. Nutrients intake and the risk of hepatocellular carcinoma in Italy. Int. J. Cancer, 43: (2007). La Vecchia C., Fabbri L.M. RE: Prevention of death in COPD. N Engl J Med May 24;356(21): Galeone C, Pelucchi C, Talamini R, Negri E, Montella M, Ramazzotti V, Zucchetto A, Dal Maso L, Franceschi S, La Vecchia C. Fibre intake and renal cell carcinoma: a case-control study from Italy. Int. J. Cancer, 121: (2007). Boffetta P, McLaughlin JK, La Vecchia C, Autier P, Boyle P. Environment in cancer causation and aetiological fraction: limitations and ambiguities. Carcinogenesis, 28:913-5 (2007). Gallus S, Scottti L, Talamini R, Franceschi S, Dal Maso L, Negri E, La Vecchia C. Re: Alcohol consumption and ovarian cancer risk in a population-based case-control study by Peterson et al. Int. J. Cancer, 121: (2007). Gallus S., La Vecchia C. Reply to L.C. Costello, R.B. Franklin s Letter to the Editor re: Gallus S., Foschi R., Negri E., et al. Dietary zinc and prostate cancer risk: a case-control study from Italy. Eur. Urol., 52: (2007) Deandrea S, Bertuccio P, Chatenoud L, Franceschi S, Serraino D, La Vecchia C. Reply to Alcohol consumption and risk of Hodgkin s lymphoma and multiple myeloma: a multicentre case-control study by Gorini et al, Ann. Oncol., 18: (2007). Ferlay J, Randi G, Bosetti C, Levi F, Negri E, Boyle P, La Vecchia C. Declining mortality from bladder cancer in Europe. BJU Int., 101: (2007). Levi F, Te VC, Maspoli M, Randimbison L, Bulliard JL, La Vecchia C Trends in breast cancer incidence among women under the age of forty. Br. J. Cancer, 97: (2007). Corrao G, Zambon A, Nicotra F, Fornari C, La Vecchia C, Mezzanzanica M, Nappi RE, Merlino L, Cesana G. 185

188 Persistence with oral and transdermal hormone replacement therapy and hospitalisation for cardiovascular outcomes. Maturitas, 57: (2007). Lucenteforte E, Bosetti C, Talamini R, Montella M, Zucchetto A, Pelucchi C, Franceschi S, Negri E, Levi F, La Vecchia C. Diabetes and endometrial cancer: effect modification by body weight, physical activity and hypertension. Br. J. Cancer, 97: (2007). Bravi F, Bosetti C, La Vecchia C. Re: Coffee and hepatocellular carcinoma: cause or confounding? Hepatology, 28: 46:2047 (2007). Randi G, Pelucchi C, Negri E, Talamini R,Galeone C, Franceschi S, La Vecchia C. Family history of urogenital cancers in patients with bladder, renal cell and prostate cancers. Int. J. Cancer, 121: (2007). Levi F, La Vecchia C. Epidemiologie et tendances du cancer en Suisse. Bull. Cancer, 94: (2007). Boccardi D, Menni S, Ferraroni M, Stival G, Bernardo L, La Vecchia C, Decarli A. Birthmarks and transient skin lesions in newborns and their relationship to maternal factors: a preliminary report from Northern Italy. Dermatology, 215: (2007). Santoro E, Clivio L, Mangano S. GCPBASE: a web-based tool for remote data capture in a clinical trial. Technology and Health Care, 15:355 (2007). SELEZIONE PUBBLICAZIONI DIVULGATIVE APPARSE NELL ANNO 2007 La Vecchia C. Tumori. L Eco-difesa. Salviamoci dall autoinquinamento da fumo e alcol. Repubblica, no. 555: 9-10 (2007). La Vecchia C, Beghi E. Epidemiologia delle demenze. In: La demenza in Italia. Aggiornamento e casi clinici interattivi. UTET, Torino, 2007; La Vecchia C. Mediterranean diet and cancer. Element of prevention. Rivista Nutrizione Pratica, no. 1: (2007). La Vecchia C. European studies on ambient particulate matter and health, with a focus on long-term exposure. In: Workshop on environment & health: air quality research needs and opportunities in the EU seventh framework. Programme of research (FP7). Proc. Symp. Brussels, Jan , Concawe, Brussels; 2007: Bertuccio P, Foschi R La salute del bambino nel primo anno di vita in Italia: che cosa è cambiato? NEWS & OPINIONS in Ginecologia : 4-9. Santoro E. Podcast, wiki e blog: il web 2.0 al servizio della formazione e dell aggiornamento del medico. Recenti Prog Med, 98: (2007). Santoro E. Il sito del British Medical Journal. Ricerca&Pratica, n. 136: (2007). Santoro E. Strumenti di rating al servizio del pubblico: è vero empowerment? Ricerca&Pratica, n. 137: (2007). Santoro E. Il web 2.0: dalla partecipazione alla in-formazione. Ricerca&Pratica, n. 138: (2007). Santoro E. Web 2.0 e medicina. CARE, 6: (2007). 186

189 Santoro E. Open access e ricerca biomedica: un nuovo modo di diffondere i risultati della ricerca. SIAR News, 51: (2007). Santoro E. My NCBI: uno strumento per l aggiornamento periodico dei risultati di una ricerca eseguita sul database Medline. SIAR News, 50:47-49 (2007). Santoro E. Informazione sanitaria: ecco le linee guida. Medici Oggi, n. 6: 8-9 (2007). Santoro E. Revolution Health e il coinvolgimento dei cittadini: uno strumento di empowerment? PartecipaSalute, 2007; Santoro E. I blog come strumento di condivisione di esperienze tra pazienti. PartecipaSalute 2007; Santoro E. Open source e web 2.0: una guida per gli utenti di Partecipasalute. PartecipaSalute 2007; Santoro E. Meglio Google o PubMed? II puntata. Va Pensiero 2007; Santoro E. Internet e medicina. In Enciclopedia Medica Italiana, aggiornamento III, Tomo 1, pp UTET 2007 (L. Vella, ed.). Pistotti V, Santoro E. Navigare sulla rete alla ricerca di informazioni di salute. In La dispensa di PartecipaSalute, pp Istituto di Ricerche Farmacologiche Mario Negri, Milano RESEARCH ACTIVITIES Data analysis and management Collection of epidemiological data has continued as planned, and also data checking, entry and input have been pursued. The overall dataset now includes 1,250 cases of oral and pharyngeal cancers, 700 of esophageal, 1,100 of stomach, 6,500 of colorectal, 600 of primary liver, 120 of gallbladder and bile duct cancers, 600 of pancreatic, 850 of laryngeal, 500 of cutaneous malignant melanoma, 7,000 of breast, 1,000 of cervical and 1,000 of endometrial cancers, 200 of gestational trophoblastic disease, 200 of vulvar, 2,000 of ovarian, 1,300 of prostatic, 700 of bladder, 800 of kidney and renal pelvis, 600 of thyroid cancer, 200 of Hodgkin s disease and 500 of other lymphomas, 150 sarcomas, 300 myelomas and about 18,000 controls. Biological samples have been collected for cancers of the upper digestive and respiratory tract, bladder and colorectal cancer to investigate genetic polymorphisms. Flavonoid intake and upper aerodigestive tract cancers We investigated the effect of selected flavonoids in relation to the risk of upper aerodigestive tract neoplasms, and found inverse associations with each cancer site considered. The study of oral and pharyngeal cancer included 805 cases and 2081 hospital controls. The ORs for the highest versus the lowest quintile of intake were 0.51 for flavanones, 0.62 for flavonols, and 0.56 for total flavonoids. The study of esophageal cancer included 304 cases and 743 hospital controls. An inverse association emerged between flavanone intake and esophageal cancer risk (OR=0.38). This study suggested that flavanone intake may account, with vitamin C, for the protective effect of fruit on esophageal cancer. The study of laryngeal cancer included 460 cases and 1088 hospital controls. Significant inverse relations were found with intake of flavan-3-ols (OR=0.64, for the highest versus the lowest quintile), flavanones (OR=0.60), flavonols 187

190 (OR=0.32) and total flavonoids (OR=0.60), providing support for a beneficial effect of selected flavonoids on laryngeal cancer, too. Family history of cancers and oral and pharyngeal cancers With reference to history of neoplasms in first-degree relatives, the OR of oral and pharyngeal cancer was 3.1 for a family history of oral, pharyngeal and laryngeal cancers combined. Significant increases in risk of oral and pharyngeal cancer were also observed for a family history of melanoma (OR=5.8) and lung cancer (OR=1.4). Our study found that a family history of oral, pharyngeal and laryngeal cancer is a strong determinant of oral and pharyngeal cancer risk, independent from tobacco and alcohol use. Alcohol drinking in never-smokers, cigarette smoking in never-drinkers, and upper aerodigestive tract cancers Our data on head and neck cancers were integrated in the International Head and Neck Cancer Epidemiology Consortium, a pooled analysis of 15 case-control studies that included a total of 10,244 cancer case subjects and 15,227 control subjects. One of the aims of the pooled analysis was to analyze the extent to which head and neck cancers are associated with cigarette smoking among never drinkers and with alcohol drinking among never users of tobacco. We found that, among never drinkers, cigarette smoking is associated with an increased risk of head and neck cancer (OR=2.13), and among never users of tobacco, alcohol consumption is associated with an increased risk of head and neck cancer only when alcohol is consumed at high frequency (OR=2.04 for three or more drinks per day versus never drinking). Pipe smoking and upper aerodigestive tract cancer We analysed the association between exclusive pipe smoking and cancers of the upper digestive tract, using data from a series of case-control studies conducted in Italy and Switzerland between 1984 and After excluding cigarette and cigar smokers, 41 male oral and pharyngeal cancer cases, 52 male oesophageal cancer cases and 1,032 male controls were included in the present analysis. Compared to never smokers, exclusive pipe smokers had an OR of 8.7 of all upper digestive tract cancers. The OR was 12.6 for oral and pharyngeal and 7.2 for oesophageal cancer. Pipe smokers who were also heavy alcohol drinkers had an OR of 38.8 as compared to never smokers and light drinkers. Thus, pipe smoking and heavy alcohol drinking appears to interact at least on a multiplicative model. Cumulative risk of upper aerodigestive tract cancer in relation to smoking cessation We estimated the effect of smoking cessation on the cumulative incidence of these cancers by age 75 years (in the absence of competing causes of death), combining odds ratios for males from a network of Italian hospital-based case-control studies ( ) with incidence data for Italian men. The studies included 961 cases with oral/pharyngeal cancer, 618 cases with esophageal cancer, and 613 cases with laryngeal cancer, plus 3,781 controls. For all upper aerodigestive tract cancers, the cumulative risks by 75 years of age were 6.3% for men who continued to smoke any type of tobacco, 3.1% and 1.2% for men who stopped smoking at around 50 and 30 years of age, respectively, and 0.8% among lifelong nonsmokers. Corresponding figures were 3.3%, 1.4%, 0.5%, and 0.2% for oral/pharyngeal cancer; 1.0%, 0.5%, 0.4%, and 0.2% for esophageal cancer; and 2.1%, 1.1%, 0.2%, and 0.2% for laryngeal cancer. In this Italian population, men who stopped smoking before age 50 years avoided more than half of the excess risk of upper aerodigestive tract cancer as men who did not, and men who stopped smoking before age 30 years avoided more than 90% of the risk. 188

191 Goiter and autoimmune thyroid disease and gastric cancer We analyzed data from a hospital-based case-control study conducted between 1985 and 1997 in Milan, including 769 incident cases of gastric cancer and 2081 control subjects. The OR of gastric cancer was 0.70 for history of goiter, 0.61 for thyroid nodules/adenomas, 1.10 for hyperthyroidism, and 0.71 for other benign thyroid diseases. None of the results was statistically significant. Therefore, goiter and other thyroid diseases do not appear to be a relevant cause of gastric cancer in Italy. Fried foods and colorectal cancer This investigation was based on 1394 cases of colon cancer, 886 cases of rectal cancer and 4765 controls, enrolled in Italy and Switzerland between 1992 and Our results did not indicate a relevant role of fried foods on colorectal cancer risk, while we found a possible favorable effect of (fried) olive oil on colon but not on rectal cancer risk. Selected dietary factors and hepatocellular carcinoma A case-control study of hepatocellular carcinoma was conducted in Italy in , including 185 histologically confirmed cases and 412 control subjects admitted to the same network of hospitals. The present study supported the hypothesis of a favorable effect of coffee (OR=0.4 for high consumption), though not decaffeinated coffee and tea, on the risk of hepatocellular carcinoma. Using the same dataset, we considered the effect of various nutrients on the risk of hepatocellular carcinoma, and found an inverse relation with a diet rich in linoleic acid (OR=0.35, for highest versus lowest tertile) and beta-carotene intake (OR=0.48). Lifetime ovulatory cycles and ovarian cancer We considered data from 2 multicentric Italian studies of ovarian cancer, including a total of 2002 cases and 4914 controls, in relation to the number of lifetime ovulatory cycles and to different anovulatory factors. As compared with the lowest quartile, the ORs of ovarian cancer were 1.60, 1.65, and 1.81 for increasing quartiles of lifetime ovulatory cycles. For 1 year of ovulation avoided, the continuous ORs were 0.91 for parity-related anovulations, 0.90 for abortions, 0.92 for oral contraceptive use, 0.99 for age at menarche, and 0.97 for age at menopause. This study found that pregnancy and oral contraceptive use had a stronger protective effect on ovarian cancer than other anovulatory factors. Dietary patterns and breast and ovarian cancers Dietary patterns were identified on a selected set of nutrients through principal component factor analysis. The starch-rich pattern emerged as an unfavorable indicator of risk for both breast and ovarian cancers, while the animal products and the vitamins and fiber patterns might be associated with a reduced risk of breast and ovarian cancers, respectively. Alcohol consumption and ovarian cancer In a separate analysis, we examined the relation between alcohol consumption and ovarian cancer risk. In a population characterized by regular and frequent wine consumption in women, wine was not materially related to ovarian cancer risk, the continuous estimate for wine being Our investigation also confirmed results of earlier studies that none of the other types of alcoholic beverages was associated to ovarian cancer risk. Macronutrients and cholesterol intake and endometrial cancer Previous studies found some evidence that dietary habits may influence the risk of endometrial cancer independently of body mass. In our investigation, significant direct associations were observed with intake of energy (OR=1.7 for the highest versus the lowest quintile), and 189

192 cholesterol (OR=2.1), while a direct borderline association emerged with saturated fatty acids (OR=1.3). On the other hand, there was no association with proteins, sugars, starch, total fat and other selected fatty acids. Energy and cholesterol intake increased endometrial cancer risk. Dietary zinc and prostate cancer In a multicentric case-control study of prostate cancer, including 1294 cases and 1451 male controls, we investigated the relation with dietary zinc intake. This was computed from a validated and reproducible food frequency questionnaire, through the use of an Italian food composition database. Compared with the lowest quintile of intake, the OR of prostate cancer for subjects at the highest quintile of zinc intake was 1.56, with a significant trend in risk. The direct association was stronger for advanced than for early stage cancers. Prostate cancer in men aged less than 60 years We considered the same dataset of prostate cancer to investigate the major risk factors in men younger than 60 years. This age group comprised 219 cases and 431 controls. The major risk factors observed in middle-aged men were family history of prostate cancer (OR=5.5), high level of education (OR=3.3), and low physical activity (OR=0.5, for active versus inactive subjects). Dietary factors and renal cell carcinoma We evaluated the relation between selected risk factors and renal cell cancer (RCC) in an Italian case-control study including 767 patients younger than 79 years with RCC and 1,534 controls. The results of this study suggested that a diet rich in refined cereals and poor in vegetables may have an unfavorable role on RCC. No relation was found for coffee and tea, eggs, red meat, fish, cheese, fruits, desserts and sugars. After allowing for energy and other major covariates, a significant inverse association was found for vitamin E (OR=0.56, for the highest quintile of intake versus the lowest one), and vitamin C (OR = 0.72), although the trend in risk for vitamin C was of borderline significance. No significant trend of decreasing risk was found for other micronutrients analyzed. We also investigated in depth the topic of dietary fibers and RCC, and found an inverse association with vegetable fibers that may reflect a real favorable effect, or be an indicator of a beneficial role of a diet rich in vegetable on RCC risk. Body size at different ages and renal cell carcinoma An increased risk of RCC has been reported in overweight persons in previous studies. In our investigation, we found an OR of 1.3 among obese persons (i.e., those with a body mass index 30) as compared to normal-weight persons (body mass index <25) and an OR of 1.5 among persons in the highest tertile of waist-to-hip ratio. Direct associations emerged for obese persons at ages 30 years (OR=1.5) and 50 years (OR=1.5). RCC risks among overweight and obese persons were apparently higher in never smokers, persons with the clear-cell histologic type, and persons with a Fuhrman nuclear grade of G3-G4. Physical activity and renal cell carcinoma In the same study of RCC, the multivariate OR for the highest as compared to the lowest level of occupational physical activity were 0.65 at age 12 years, 0.67 at age 15-19, 0.74 at age and 0.71 at age years, with significant inverse trends in risk. No significant association was found for leisure-time physical activity. The inverse association between occupational physical activity and RCC risk, if real, may be related to the effects of insulin-like growth factors, or lipid peroxidation and about 9% of cases of RCC in Italy could be avoided by increasing physical activity. 190

193 Hypertension, diabetes mellitus and the risk of renal cell cancer In the same RCC dataset, patients with a history of treated hypertension reported an excess risk of RCC (OR=1.7), and the pattern was confirmed in different strata of sex, education, smoking habits, body mass, tumor histological type, stage, or grade. Therefore, we estimate that the attributable risk of RCC for treated hypertension in this population is 16%. A slight, nonsignificant increased risk was found also for history of diabetes mellitus (OR=1.3). Alcohol consumption and renal cell carcinoma There is some evidence that alcohol consumption is inversely associated with RCC. Since the issue is still unclear, we investigated the relation by pooling data from two Italian case-control studies, including a total of 1115 incident, histologically confirmed cases and 2582 controls. Compared with non-drinkers, the ORs of RCC were 0.87 for 4 drinks per day, 0.76 for >4 to 8 drinks per day and 0.70 for >8 drinks per day of alcoholic beverages. Thus, this pooled analysis found an inverse association between alcohol drinking and RCC. Risks continued to decrease even above 100 g/day of alcohol intake, with no apparent leveling in risk. Diabetes and non-hodgkin lymphoma During year 2007, we examined the role of selected risk factors for non-hodgkin lymphoma (NHL) using data from two Italian case-control studies conducted in and , including 671 cases with incident, histologicallyconfirmed NHL and 1799 controls. We analysed the combined datasets in relation to diabetes. Epidemiological results on the issue are controversial, and diabetes has been related to the risk of several neoplasms. However, the results of our study allow to exclude a strong association between diabetes and NHL. The small number of cases with diabetes in this investigation left open the possibility of a moderate direct relation. Folate, alcohol and non-hodgkin lymphoma With reference to this issue, we examined our recent case-control study, i.e., the only one with detailed information on dietary habits, including 190 cases of NHL and 484 controls. No significant association emerged between NHL risk and intakes of folate (OR=0.9) and other micronutrients involved in the one-carbon metabolism, i.e., vitamin B2 (OR=0.9), vitamin B6 (OR=0.8), and methionine (OR=0.7). However, a significant inverse association was observed for all the nutrients examined among abstainers and former drinkers. Our findings support the possibility of an antagonist effect of alcohol on the one-carbon metabolism in NHL etiology. However, the lack of an overall effect for one-carbon nutrients and the small sample size suggested caution in interpreting our results. InterLymph Pooled Analysis on non-hodgkin lymphoma Our data on hematopoietic malignancies were included in a pooled analysis of 10,211 NHL cases and 11,905 controls from the International Lymphoma Epidemiology Consortium (InterLymph). During year 2007, this pooled analysis examined family history of hematopoietic malignancies and obesity as potential risk factors for NHL. The pattern of NHL heritability remains poorly understood, though there is accumulating evidence of common genetic variations altering NHL risk. In the InterLymph pooled analysis, NHL risk was elevated for individuals who reported first-degree relatives with NHL (OR=1.5), Hodgkin lymphoma (OR=1.6), and leukemia (OR=1.4). The pattern of NHL heritability appeared to be uniform across NHL subtypes, but risk patterns differed by specific hematopoietic malignancies and the sex of the relative, revealing critical clues to disease etiology. Another investigation from the InterLymph Consortium considered the role of obesity on NHL risk. This concluded that there is no evidence to support the hypothesis that obesity is a determinant of all types of NHL 191

194 combined, while the association observed between severe obesity and diffuse large B-cell lymphoma may warrant further investigation. Artificial sweeteners and cancer risk The role of sweeteners on cancer risk has been widely debated over the last few decades. To provide additional information on saccharin and other sweeteners (mainly aspartame), we considered data from a large network of case-control studies, including upper aero-digestive tract, colon, rectum, breast, ovary, prostate and kidney. The ORs for consumption of saccharin were 0.83 for cancers of the oral cavity and pharynx, 1.58 for oesophageal, 0.95 for colon, 0.93 for rectal, 1.55 for laryngeal, 1.01 for breast, 0.46 for ovarian, 0.91 for prostate and 0.79 for kidney cancer. This work indicated a lack of association between saccharin, aspartame and other sweeteners and the risk of these neoplasms. Glycaemic index and load and thyroid cancer Risk of thyroid cancer has been related to refined cereals and starch foods, but the association has never been studied in terms of glycemic index and load. We analyzed the issue using data from an Italian study of 399 cases of thyroid cancer and 616 control subjects. Our study showed that high dietary levels of glycemic index (OR=1.73) and glycemic load (OR=2.17) are associated with thyroid cancer risk. Family history of cancer and urogenital cancers History of urogenital cancers in first-degree relatives was investigated among a total of 1356 subjects with cancer of the bladder, kidney and prostate, and 1067 controls. ORs of urogenital cancers for subjects with at least a first-degree relative with cancer at the same site were 6.1 for bladder, 2.0 for renal cell and 2.0 for prostate cancer. Reliability of information on family history of cancer The reliability of the information on family history of cancer in first-degree relatives in our hospital-based case-control study of digestive tract cancers was tested by re-interviewing at home 294 controls. A satisfactory agreement between the two interviews was reported for family history of any cancer and of cancers of the digestive tract. We found a systematic tendency to report a history of cancer more frequently in the hospital setting than in the home setting. Thus, data on family history of cancer provided by hospital controls show a good reliability. Tar yield in cigarettes and risk of acute myocardial infarction Controversial information is available with reference to the role of type or tar yield of cigarettes on the risk of cardiovascular disease. We considered the issue in a combined dataset of three case-control studies of acute myocardial infarction (AMI) conducted in Italy between 1983 and 2003, including 1990 subjects with a first episode of non-fatal AMI, and 2521 controls. As compared to never smokers, the multivariate OR was 2.70 for smokers of low tar cigarettes (<10 mg), 3.06 for intermediate (10-19 mg) and 3.14 for high tar yield ( 20 mg). After further allowance for duration of smoking and number of cigarettes per day, as compared to low tar yield cigarettes, the OR was 1.14 for intermediate, and 1.28 for high tar yield. Our study confirms that no substantial reduction in AMI risk resulted from the decrease of cigarette tar yield. Menstrual and reproductive factors and risk of acute myocardial infarction We analyzed this association using data from three Italian case-control studies, including a total of 609 female cases of AMI and 1106 female controls. The risk of AMI was slightly increased in women with irregular menstrual cycle (OR=1.36) and those who had at least one child 192

195 (OR=1.45), but without a linear trend in risk with number of children. The association was much stronger in smokers, with an OR of 5.98 for smokers with an irregular menstrual cycle compared with non smokers with a regular one and an OR of 4.77 in smokers who had children compared with non smokers nulliparae. There was no relation with menopausal status, age at menarche and menopause, age at first and last birth and number of abortions. Record-linkage for cohort analyses With reference to our project to create an Italian cohort study based on record-linkage, during 2007 we prepared the archives that will allow to link data between our network of case-control studies (conducted since 1982) and those from the historical archives of the Local Health Unit (ASL) of Milan, that include several health information such as vital status and date of death of the subjects. Several stages of the project have been undertaken and completed during the year. In particular, we finished the identification of subjects recruited in the case-control studies among the historical archives of the ASL, and a database with univocal information on subjects from the two sources is now available, thus allowing a linkage of the archives; at the same time, we completed data collection from all case-control studies in a unique database (using original data and through the preparation of a codebook that allowed to re-codify studies conducted in various periods and on several diseases) that includes information from interviews to approximately 27,000 subjects. During 2007, we searched in various other historical databases individuals that were not linked in historical archives of the ASL. On 18,257 residents in Lombardy, 15,860 (86.9%) subjects, including 7,680 cases (82.2%) and 8,911 controls (91.8%) were linked. Lung cancer in young European women We analyzed age-standardized trends in lung cancer mortality in young women (20-44) for the 6 major European countries, using joinpoint regression. In the early 1970s the highest lung cancer mortality in young women was in the UK (2.1/100,000). UK rates, however, steadily declined and in they were the lowest of all 6 major EU countries (1.2/100,000). The second lowest rate in was in Italy, whose rates remained around 1.1/100,000 between 1970 and 1994, and increased to 1.4 thereafter. In Germany and Poland, lung cancer rates in young women rose from /100,000 in the early 1970s to in the mid 1990s and levelled off during the last decade. Major rises over recent years were observed in France (from 0.8/100,000 in to 2.2 in ) and in Spain (from 0.8 in the to 1.7 in ). Thus, France showed both the highest rate observed over the last 3 decades and the largest rise over the last 2 decades. Worldwide mortality from cirrhosis Cirrhosis mortality was analyzed for 41 countries worldwide over the period using data from the WHO. In the early 1980s, the highest rates were in Mexico, Chile (around 55/100,000 men and over 14/100,000 women), France, Italy, Portugal, Austria, Hungary and Romania (around 30-35/100,000 men and 10-15/100,000 women). Mortality from cirrhosis has been steadily declining in most countries worldwide since the mid or late 1970s. In southern Europe, rates in the early 2000s were less than halved compared to earlier decades. In contrast, rates have been rising in eastern European countries to reach extremely high values in the mid 1990s, and declined only thereafter. In the UK rates were still steadily rising. Mortality from cirrhosis shows favourable trends in most countries of the world, following the reduction in alcohol consumption and hepatitis B and C virus infection. The steady upward trends observed over more recent calendar periods in the UK and central and eastern European countries are attributed to the persistent increase in the prevalence of alcohol consumption. 193

196 Testicular cancer mortality in the Americas Death rates and trends from testicular cancer were compared over the period in all the American countries that provide data. In the early 1980s the highest testicular cancer mortality rates were observed in Chile (1.7/100,000 at all ages, 3.6/100,000 at years) and Argentina (0.9/100,000 at all ages, 1.7/100,000 at years), as compared with 0.4/100,000 for all ages and 0.6/100,000 at 20 to 44 years in Canada, and 0.3/100,000 for all ages and 0.7/100,000 at 20 to 44 years in the US. In , testicular cancer mortality had fallen to 0.2/100,000 in men aged 20 to 44 years in Canada, and to 0.4/100,000 in the US. Conversely, rates were still 1.6/100,000 in Argentina, 2.2/100,000 in Chile and 1.2/100,000 in Mexico, and were around /100,000 in most other Latin American countries that provide data. Mortality from testicular cancer in (young) men remains exceedingly high in most Latin American countries. Urgent intervention is required to provide treatment (essentially modern integrated platinum-based chemotherapy) for this largely curable neoplasm in young men. Smoking prevalence in Italy On 10 January 2005, Italy became the first large European country adopting a comprehensive smoke-free legislation. To provide information on smoking prevalence in Italy and evaluate the effects of the 2005 regulation, we considered data from three companion surveys on smoking conducted in 2004, 2005 and 2006 in Italy. Each survey included more than 3,039 subjects (1,461 men and 1,578 women) aged 15 or over, representative of the general Italian adult population. Current smokers declined from 26.2% (30.0% of men, 22.5% of women) in 2004, to 25.6% (29.3% of men, 22.2% of women) in 2005, and to 24.3% (28.6% of men, 20.3% of women) in While no significant difference was found comparing smoking prevalence in vs , the drop in smoking prevalence in vs was significant (p<0.05) in the total population, in men and in subjects aged years. Smokers consumed a mean of 15.4 cigarettes per day in 2004, 14.6 in 2005 and 13.9 cigarettes per day in The drop in smoking prevalence and consumption may be due, particularly for younger generations, to the comprehensive smoke-free legislation adopted in Italy. Strategies and best practices for the reduction of injuries Some interventions, considered sufficiently promising for the prevention of falls in elderly people, were identified. A search of the published literature and gray literature has been performed and several relevant papers have been retrieved. A database has been created, containing information extracted from the retrieved publications, to classify interventions according to effectiveness, characteristics of the target population, participation and compliance, resources needed for the implementation. The information obtained was analyzed focusing on to the efficacy of the interventions and to subjects participation. Besides the information collected, the principal investigators of each paper have been contacted and invited to fill in a questionnaire on barriers and facilitators found in the implementation of their interventions. Another questionnaire has been developed to perform a survey on the attitudes of the elderly on interventions for the prevention of falls; this survey will be conducted in Italy, Poland and Greece. Furthermore, another bibliographic search has been performed on factors influencing the risk of falls in elderly people, and about 200 risk factors were identified and classified. All papers related to these risk factors were retrieved. Systematic reviews and meta-analyses will be conducted on published studies related to about thirty risk factors for falls. HI-WATE project on colorectal cancer and drinking water by-products In 2007, the Department has been involved in a project of the 6-FP on the Health Impact of long term exposure to disinfection by-products in drinking water (DBPs) (HI-Wate study). Within this project, our Department will conduct a case-control study of colorectal cancer to investigate 194

197 the association with various DBPs, including in particular trihalomethanes (THMs). The study will be conducted in the greater Milan area and in the Provinces of Pordenone and Udine, and will include approximately 500 incident, histologically confirmed cases enrolled in the major general and teaching hospital of the study area, and 500 frequency-matched controls, admitted to the same hospital as cases for acute, nonneoplastic conditions. Cases and controls will be interviewed using an extensive questionnaire. Detailed information on water use and waterrelated habits for etiologically relevant time periods will be collected, including not only water consumption, but also water related activities, such as showering, bathing and swimming, that may influence exposure assessment. Moreover, in order to assess the subjects exposure to DBPs and THMs in water, current and historical THM levels, water source and year of starting chlorination in the study area will be collected from local companies, authorities and municipalities. The relative and attributable risks of colorectal cancer for exposure to THMs and other DBPs will be estimated. Giving the large number of people exposed to chlorinated drinking-water and its DBPs, the study has potentially important implications in terms of public health. Even modest excess risk in relation to DBP exposure may in fact have a relevant impact on a population level, and may be responsible of a considerable number of colorectal cancer cases. Medical History, Drug Exposure and the Risk of Psoriasis The association of psoriasis with selected medical conditions and a number of drugs used before diagnosis was evaluated in a multicenter case-control study involving outpatient services of 20 general and teaching hospitals, conducted in collaboration with the Gruppo Italiano per gli Studi Epidemiologici in Dermatologia (GISED). Entry criteria for cases were a first diagnosis of psoriasis made by a dermatologist and a history of skin manifestations of no more than 2 years after the reported onset of the disease. Controls were the first eligible dermatological patients observed on randomly selected days in the same centers as cases. A total of 560 cases and 690 controls were recruited. The OR of psoriasis was 0.8 in hypertensive subjects, 1.1 in diabetics and 1.1 in hyperlipidemic subjects. Histamine 2 receptor antagonist exposure was negatively associated with psoriasis, with an OR of 0.3. Our study rules out a strong association of psoriasis with common chronic conditions. The reported associations of psoriasis with relatively common conditions such as diabetes mellitus, hypertension and hyperlipidemia may represent a late effect of well-known risk factors for psoriasis, such as smoking and overweight, or reflect factors related to the long course of psoriasis itself. Impact of body mass index and obesity on clinical response to systemic treatment for Psoriasis. Evidence from the PSOCARE project To assess the role of body mass index (BMI) on early clinical response to systemic treatment for psoriasis, we analyzed data from a nationwide registry and cohort study involving compelling registration of patients receiving for the first time in their life a new systemic treatment for psoriasis at reference centres in Italy. Information was gathered by treating physicians with the aid of a web-based electronic form. Patients with a clinical diagnosis of chronic plaque psoriasis and with 8 and 16 weeks of completed follow-up by March 31 th, 2007 were eligible. A relative reduction of Psoriasis Area Severity Index (PASI) of at least 75% at follow-up compared to baseline was evaluated as clinical endpoint (PASI-75). A total of 2,368 patients were analysed at 8 weeks and 2,042 at 16 weeks. PASI-75 was achieved by 819 (34.5%) patients at 8 weeks and 1,034 (50.6%) patients at 16 weeks. The proportion steadily decreased with increased values of BMI, from 41.7% in patients with BMI < 20 to 29.1% in patients with BMI 30 at 8 weeks, and from 59% in patients with BMI <20 to 42.2% in patients with BMI 30, at 16 weeks. Compared to normal-weight (BMI 20-24), obese patients (BMI 30) had a lower chance of improving at 8 and 16 weeks, adjusted ORs for achieving PASI-75 being 0.73 and 0.62, respectively. The impact of BMI on PASI-75 did not present remarkable variations according 195

198 to the drug prescribed at entry. Thus, BMI affects early clinical response to systemic treatment for psoriasis irrespectively of the administered drug. This finding has implications for individual patient care and the design of future randomised clinical trials Nevus count on specific anatomic sites as a predictor of total body count Scanty information is available on the relation between nevus count on specific anatomic areas and the total body surface, particularly in children. We analyzed this issue by using data from a uniquely large study conducted in 1997 on 3,406 schoolchildren (1,746 boys and 1,660 girls) aged years in 13 cities from northern, central, and southern Italy. Children were examined by trained dermatologists who counted melanocytic nevi ( 2 mm in diameter) on 19 different anatomic sites. Overall, the mean number of nevi was 17.3 (18.6 in boys and 15.8 in girls). The adjusted correlation coefficients (r) with number of nevi on the whole body were 0.74 for head and neck, 0.83 for anterior and 0.84 for posterior trunk, and 0.88 for upper and 0.80 for lower limbs. With reference to single anatomic sites, the best predictor of total nevus count was the lateral arms (r=0.80), overall and in strata of sex and pigmentary characteristics. This large study provides definite evidence that examining the upper limbs only, particularly the lateral arms, is a practical and suitable tool for predicting total nevus count in children. Histological types and anatomic sites of cutaneous basal-cell carcinoma Since different clinico-pathologic subtypes and anatomic sites of basal-cell carcinoma (BCC) may display distinct characteristics and mechanisms of development, we analyzed data from an Italian case control study, including 528 subjects with BCC and 512 controls. The risk of nodular (OR=1.53) but not superficial (OR=0.71) BCC was increased for occupational exposure to sunlight. Considering the anatomic site of BCC, the corresponding values were 1.46 for head/neck and 0.74 for truncal location. Direct associations were observed with recreational sunlight exposure, eye color, red hair, and number and early age of severe sunburn episodes, along with some differences in risk between histotypes and anatomic sites. Therefore, this study confirmed the role of (intermittent) sun exposure and phenotypic characteristics as risk factors for BCC, and suggested etiological differences between nodular and superficial histotypes and between head/neck and truncal locations. Anthropometric factors and cutaneous malignant melanoma Several studies have investigated the effect of various anthropometric factors on the risk of cutaneous malignant melanoma (CMM). As the results are controversial, we analyzed the issue in a case-control study conducted in Italy between 1992 and 1994, including 542 patients with CMM and 538 controls. The ORs for the highest versus the lowest quartile were 2.06 for weight, 1.16 for height, 1.90 for BMI and 1.87 for body surface area (BSA). When allowing for BMI and BSA in the same model, the ORs were 1.55 for BMI and 1.41 for BSA. The present findings confirm that obesity increases the risk of CMM. BSA is also related to the risk of CMM. In terms of population attributable risks, overweight and obesity would account for 31% of the cases of CMM in this Italian population, indicating the scope of prevention. Number of naevi and cutaneous malignant melanoma In the same case-control study, we analyzed the association between CMM at a specific anatomical site and number of nevi at the same site. Cases and controls were examined by trained dermatologists who counted the number of melanocytic nevi. The ORs of CMM for the highest versus the lowest tertile of number of nevi at the corresponding site was 1.4 at face and neck, 2.3 at anterior trunk, 4.9 at posterior trunk, 2.9 at upper limbs and 5.0 at lower limbs. In a case-case analysis, comparing CMM cases at a specific site and CMM cases at all other sites, the only excess risk was found for the posterior trunk, the OR being 2.1 for the highest versus 196

199 the lowest tertile of number of nevi. Our data do not support the hypothesis of a specific effect of nevi at each single anatomical site. 197

200 Development of the BPCO.CARE website This web based medical index has been developed by the Laboratory of Medical Informatics in order to collect, classify, evaluate, and describe the most useful medical information on the web related to the chronic obstructive pulmonary disease (COPD). The description associated to each web site included in the BPCO.CARE index illustrates the main contents, the services, and the tools offered to the users, including those related to the web 2.0 technology such as podcasts, RSS feeds, blogs, webcasts, and webinars. A section of the BPCO.CARE website also includes a classified collection of the podcast services available on the net in the COPD and pulmonology area and information on how to subscribe to. The BPCO.CARE website (available at has been developed in collaboration with the Department of Cardiovascular Research. Maintenance of the CARDIO.CARE, ONCO.CARE, GASTRO.CARE, NEURO.CARE, PNEUMO.CARE, PAIN.CARE and DERMA.CARE websites These indexes have been developed by the Laboratory of Medical Informatics in order to collect, classify, evaluate, and describe the most useful medical information on the web, and to provide Internet users with an easy means to surf the net. Several medical areas are covered including oncology ( neurology ( gastroenterology ( cardiology ( pulmonology ( the pain care and management ( and dermatology ( The project is in collaboration with intramural departments (Department of Oncology, Laboratory of Neurological Disorders and Department of Cardiovascular Research, Laboratory of General Practice Research, Laboratory of Translational and Outcome Research in Oncology) and extramural research groups (Italian Group for Epidemiologic Research in Dermatology, GISED). Studies on the typology of the web 2.0 applications in medicine The Laboratory of Medical Informatics is involved in studies and surveys which aim is to describe the typology of the web 2.0 applications and tools (including social networks, podcasts, feed RSS, blogs, and wikis) in medicine available on the net, and how these are perceived by the medical community. Internet as a research and formative tool on the chronic pain in cancer patient This research project was born in the framework of the project "Pain in the patient with cancer", in collaboration with the Laboratory of Medical Research and Consumer Involvement and the Laboratory of Translational and Outcome Research in Oncology. Its aim is to make available for doctors, patients and families correct information about therapies for cancer pain and to produce greater tests on the effectiveness of therapies based on the use of analgesic drugs. This project is articulated in two main activities: - Paincare, set-up a catalogue of selected web pages dedicated to the cancer pain and relative periodical up-to-date, - adaptation of the methods and instruments of Evidence Based Medicine to the resources available on the Internet about chronic pain in patients with cancer. This is done through a specific questionnaire. We are also considering the opportunity to set up a new Italian cancer pain website. Studies on the quality of health information available on the Internet We are conducting an analysis on the quality and reliability of the information available on the Internet related to the management of cancer pain. Through an ad hoc form using worldwide 198

201 applied criteria and principles for the evaluation of health websites and the evidence based medicine tools, an interdisciplinary team of reviewers is involved in the evaluation of the quality of information offered by the websites included in the PAIN.CARE medical index. The same study will also allow to compare the level of the quality of health related information among different classes of websites. The study, in collaboration with the Laboratory of Medical Research and Consumer Involvement, is ongoing and the results will be available in Training activities In 2007, the Laboratory of Medical Informatics continued its training activity on issues related to the use of the Internet in medicine, and extended it to the use of the recent web 2.0 technologies and tools in the medicine area. The members of the laboratory staff activated (or attended as invited teachers) a number of training courses, workshops, and master courses. Public health prevention and information The major products of our activity, mainly related to diet, tobacco and alcohol have also been published in the lay press, in order to increase the project impact on prevention and public health. 199

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203 LABORATORY OF REGULATORY POLICIES STAFF Head Vittorio BERTELE, M.D. 201

204 CURRICULUM VITAE Vittorio Bertele is a clinical pharmacologist. He got his MD degree in 1977 and the specialization in Internal Medicine in 1982, both at the Milan University Medical School. He was research fellow at the Harvard Medical School and then worked at the Milan University and the Mario Negri Institute. His main areas of interest have been clinical pharmacology of drugs active on the hemostatic and vascular system 1,2, epidemiology of interventions in the cardiovascular area, and clinical trials and drug utilization studies in the cardiovascular area 3,4. He was CPMP expert at the EMEA, and member of the Committee for Drug Price Negotiation at the Italian Ministry of Health 5,6. At present he is Head of the Regulatory Policies Laboratory at the "Mario Negri" Institute, and member of the Technical-Scientific Committee at the Italian Drug Agency. Selected publications Bertele' V., Falanga A., Tomasiak M., Dejana E., Cerletti C., De Gaetano G. Platelet thromboxane synthetase inhibitors with low doses of aspirin: Possible resolution of the "aspirin dilemma". Science 1983; 220: Bertele' V., Falanga A., Tomasiak M., Chiabrando C., Cerletti C., De Gaetano G. Pharmacological inhibition of thromboxane synthetase and platelet aggregation: Modulatory role of cyclooxygenase products. Blood 1984; 63: (1984). Bertele' V, Mussoni L, Pintucci G, Del Rosso G, Romano G, de Gaetano G, Libretti A. The inhibitory effect of aspirin on fibrinolysis is reversed by iloprost, a prostacyclin analogue. Thromb Haemost 1989; 61: The i.c.a.i. Group (Gruppo di studio dell'ischemia cronica Critica degli Arti Inferiori). Prostanoids for chronic critical leg ischemia: A randomized, controlled, open-label trial with prostaglandin E 1. Ann Int Med 1999; 130: Collaborative Group of the Primary Prevention Project (PPP). Low-dose aspirin and vitamin E in people at cardiovascular risk: a randomised trial in general practice. Lancet 2001; 357: Garattini S, Bertele V. Adjusting regulatory rules to public health needs. Lancet 2001; 358: Garattini S, Bertele' V. Efficacy, safety, and cost of new anticancer drugs. BMJ 2002; 325: Garattini S, Bertele V, Li Bassi L. How can research ethics committees protect patients better? BMJ 2003; 326: Joppi R, Bertele' V, Garattini S. Disappointing biotech. BMJ 2005; 331: Garattini S, Bertele' V. Non-inferiority trials are unethical because they disregard patients' interests. Lancet 2007; 370 :

205 INTRODUCTION TO THE LABORATORY'S ACTIVITIES Critical appraisal of clinical methodology Optimisation of drug use and healthcare fund stewardship Critical appraisal and recommendations for European Pricing and Reimbursement systems Evaluation of marketing authorization applications submitted to the European regulatory agency (EMEA) and of subsequent variations Evaluation of the appropriateness of drug legislation, institutions, and regulatory procedures with respect to public health needs. Cooperation to the development and to the solution of regulatory issues in developing countries. FINDINGS/MAIN RESULTS Critical appraisal of clinical research methodological aspects as the adoption of equivalence/non-inferiority design in clinical trials Development of Pan-European strategies for rational use of drugs Recommendations for Pan-European pricing policies for generics as well as interchangeable brands in a class once generics are available Raising awareness among interested parties about the deficiencies of the present EU pharmaceutical legislation and about our proposals to improve it in the public health interest. Critical review of drug documentation at the basis of marketing authorizations. Critical review of the criteria to assess pharmaceutical innovation and include new drugs in the national reimbursement scheme. NATIONAL COLLABORATIONS Italian Drug Agency (AIFA) Istituto Superiore di Sanità Department of Health Lombardy RegionItalian Horizon Scanning Project 203

206 European Medicine Agency (EMEA) INTERNATIONAL COLLABORATIONS Karolinska Institutet, Clinical Pharmacology, Department of Laboratory Medicine, SE University of Liverpool Management School, Prescribing Research Group, UK World Health Organisation (Department of Essential Drugs and Medicines Policy) Association of South East Asian Nations (ASEAN) EDITORIAL BOARD MEMBERSHIP Ricerca & Pratica Dialogo sui Farmaci NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Technical Scientific Committee at the Italian Drug Agency (AIFA) Subcommittee of the Community Procedures at the Italian Drug Agency (AIFA) Scientific Committee of the Italian Horizon Scanning Project SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Garattini S, Bertele' V. Non-inferiority trials are unethical because they disregard patients' interests. Lancet 2007; 370: Garattini S, Bertele' V. How can we regulate medicines better? BMJ 2007; 335: Barbui C, Cipriani A, Lintas C, Bertele' V, Garattini S. CNS drugs approved by the centralised European procedure: true innovation or dangerous stagnation? Psychopharmacology (Berl) 2007; 190: Bertele' V, Buonocore C, Michelacci F, Vitocolonna M, Garattini S. Efficacy and safety of immunosuppressive drugs approved in EU through the centralised procedure. Eur J Clin Pharmacol 2007; 63: Bertele' V, Banzi R, Capasso F, Tafuri G, Trotta F, Apolone G, Garattini S. Haematological anticancer drugs in Europe: any added value at the time of approval? Eur J Clin Pharmacol 2007; 63: Bertele' V, Assisi A, Di Muzio V, Renzo D, Garattini S. New antirheumatic drugs: any real added value? A critical overview of regulatory criteria for their marketing approval. Eur J Clin Pharmacol 2007; 63: Godman B, Haycox A, Bertele' V, Schwabe U. UK OFT pricing proposal - a necessity to fund new drugs or new indications. BMJ online 9 May Available in URL from: 204

207 RESEARCH ACTIVITIES Critical appraisal of clinical methodology Raising awareness about potential biases in clinical research Critical evaluation of the EU pharmaceutical legislation Raising awareness among interested parties about the deficiencies of the present EU pharmaceutical legislation and about our proposals to improve it in the public health interest. Critical appraisal of ongoing reforms including pricing reforms in major European countries Evaluation of reforms to enhance generic prescribing rates, drive down the costs of generics and corresponding originator brands as well as potential prices of interchangeable brands once standards become available as generics. Development of Pan-European strategies for rational use of drugs Enhancing rational use in line with an approach that became known as the five Es, namely: evaluation; economics; enforcement; education and engineering Assessment of drug dossiers for regulatory approvals Expert support to the Rapporteurship for marketing authorisation applications and variations to the conditions of marketing authorisation Activities for the Technical Scientific Committee at the AIFA Consultative activities for the Italian Drug Agency regarding regulatory duties with respect to drug quality, safety, efficacy, and cost. Activities for the sub-committee for the European Procedures Assessment of the dossiers for marketing authorisation applications through mutual recognition procedures involving Italy as either Reference or Concerned Member State. 205

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209 LABORATORY FOR MOTHER AND CHILD HEALTH STAFF Head Maurizio BONATI, M.D. 207

210 CURRICULUM VITAE Maurizio Bonati has a Medical School degree at the University of Milan. Areas of interest: Monitoring and epidemiological evaluation of drug utilisation and effects of drugs and vaccines in motherhood and childhood. Research methodology in general hospital and paediatric community practice. Transfer of information to the community. Epidemiology of paediatric and perinatal care. Past and present roles both at the Mario Negri Institute and in other institutions: Research Fellow at the IRFMN, within the Neurochemistry Lab.; Research Assistant at the IRFMN, within the Clinical Pharmacology Lab.; Chief of the Perinatal Clinical Pharmacology Unit at the IRFMN; Advisor to WHO for the Drug Utilization Research Group (pregnancy, paediatrics and breastfeeding); coordinator of the International Cooperative Study of Drug Use in Pregnancy, under the auspices of WHO and the support of EEC; co-editor of The Kangaroo; coordinator of the European Cooperative Study: Development of the European register of clinical trials on medicines for children (DEC-net), under the 5 th Framework Programme s Quality of life and Management of Living Resources; since 1989 he has been director of the Centre for Drug Information; since 1993 head of the Lab. for Mother and Child Health; since 1997 teacher for the Lombardy region s professional training courses; since 2000 teacher for the Lombardy region s professional training courses; since 2002 Editor of the Ricerca & Pratica scientific journal; since 2003 professor of the School of Specialisation in Paediatrics - University of Milan Bicocca; teacher at the annual European course Evaluation of Medicinal Products in Children (promoted by ESDPPP and Eudipharm). Selected publications Impicciatore P, Pandolfini C, Casella N, Bonati M. Reliability of public-oriented health care resources on the World Wide Web the home management of fever in children. BMJ 1997;314: Conroy S, Choonara I, Impicciatore P, Mohn A, Arnell H, Rane A, Knoeppel C, Seyberth H, Pandolfini C, Raffaelli MP, Rocchi F, Bonati M, Jong G, de Hoog M, van den Anker J. Survey of unlicensed and off label drug use in paediatric wards in European countries. BMJ 2000;320: Cazzato T, Pandolfini C, Campi R, Bonati M, and the ACP Puglia-Basilicata Working Group. Drug prescribing in outpatient children in Southern Italy. Eur J Clin Pharmacol 2001;57: Impicciatore P, Choonara I, Clarkson A, Provasi D, Pandolfini C, Bonati M. Incidence of adverse drug reactions in pediatric in/out-patients: a systematic review and meta-analysis of prospective studies. Br J Clin Pharmacol 2001;52: Clavenna A, Pandolfini C, Bonati M. Public disclosure of clinical trials in children. Curr Ther Res 2002;63: Pandolfini C, Bonati M. A literature review on off-label drug use in children. Eur J Ped 2005;164: Bonati M, Clavenna A. The epidemiology of psychotropic drug use in children and adolescents. International Review of Psychiatry 2005;17: Santoro E, Rossi V, Pandolfini C, Bonati M. DEC-net: the development of the European Register of Clinical Trials on Medicines for Children. Clinical Trials 2006;3:

211 INTRODUCTION TO THE LABORATORY'S ACTIVITIES Research, as a multidimensional approach to producing knowledge, characterises the Laboratory s activity. Research provides the basis for planning and carrying out the Laboratory s activity in a critical way and involves the participation of health professionals, social workers, mothers, children, and parents. Special attention is given to activities involving countries in the north and south of the world. The main objective of the Laboratory for Mother and Child Health is to ensure a better mother and child well-being by undertaking interdisciplinary and collaborative work in the field. Four broad areas, or spheres, of research have been selected: - monitoring and epidemiological evaluation of utilisation and effects of drugs and vaccines; - research methodology in general hospital and paediatric community practice; - public health determinants of children s well-being; - transfer of health information to the community. The Drug and Health Information Centre (Centro di Informazione sul Farmaco e la Salute), whose main activity entails promoting rational drug use during breastfeeding and in childhood, is part of the Laboratory. NATIONAL COLLABORATIONS Agenzia Italiana del Farmaco, (AIFA) Associazione Culturale Pediatri, (ACP) Centro Antiveleni-Unità di Tossicologia Clinica-Ospedali Riuniti di Bergamo Centro per la Salute del Bambino, (CSB) Consorzio Interuniversitario, (CINECA) Federfarma Lombardia Istituto Superiore di Sanità (ISS) Osservatorio Italiano Salute Globale (OISG) Il Pensiero Scientifico Editore Unità Operativa di Neuropsichiatria dell'infanzia e dell'adolescenza, Fondazione Policlinico di Milano, (UONPIA) Università degli Studi di Milano-Facoltà di Scienze Politiche Università degli Studi di Milano, Bicocca-Facoltà di Medicina-Clinica Pediatrica 209

212 INTERNATIONAL COLLABORATIONS Agenzia Europea per i Medicinali (EMEA) Centro de Epidemiologia Comunitaria y Medicina Tropical (CECOMET), Ecuador Clinica Infantil Colsubsidio, Bogotà, Colombia European Network Drug Investigation Children (ENDIC) European Society for Developmental Perinatal & Paediatric Pharmacology (ESDPPP) Hôpital Robert Debré, Parigi, Francia International Society of Drug Bulletins (ISDB) Organizzazione Mondiale della Sanità (OMS) Unione Europea (UE) Università di Nottingham - Derbyshire Children's Hospital, Derby, UK EDITORIAL BOARD MEMBERSHIP Dr. Maurizio Bonati, head of the Laboratory, is a member of the following editorial boards: Dialogo sui Farmaci, Disturbi d Attenzione e Iperattività, European Journal of Clinical Pharmacology, Paediatric & Perinatal Drug Therapy, Pediatria (Sao Paulo), Quaderni ACP, Quaderni di Farmacoeconomia, Ricerca & Pratica, Saludarte. PEER REVIEW ACTIVITIES American Journal of Epidemiology, Acta Paediatrica, BMC Pregnancy and Childbirth, British Medical Journal, Dialogo sui Farmaci, Epidemilogia & Prevenzione, European Journal of Clinical Pharmacology, Giornale Italiano di Farmacia Clinica, Journal of Antimicrobial Chemotherapy, Medico e Bambino, Paediatric and Perinatal Drug Therapy, Pediatric Drugs, Pediatric Health, Pediatrics, Prescrire, Quaderni ACP, The Italian Journal of Pediatrics, Vaccine. NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Comitato scientifico ADHD ISS Commissione Nazionale per le Vaccinazioni Ministero della Salute Commissione tecnico-scientifica per la programmazione e verifica delle vaccinazioni - Regione Lombardia Comitato Scientifico del Gruppo di Lavoro Farmaci e Bambini AIFA Commissione tecnica per l'elaborazione, gestione e aggiornamento del Prontuario Terapeutico Regionale (P.T.R.) - Regione Autonoma Valle d'aosta Gruppo di lavoro Network Italiano Promozione Acido Folico ISS Gruppo di Lavoro Pediatrico AIFA Gruppo di Lavoro "Promozione allattamento al seno" - Regione Lombardia Paediatric Expert Group (P.E.G.)- EMEA 210

213 PARTICIPATION IN EVENTS IN WHICH THE LABORATORY WAS INVOLVED January NECESSITA TERAPEUTICHE INSODDISFATE IN PEDIATRIA. 48 Seminario SIAR Attualità Regolatorie: il progetto AIFA per la farmaceutica; l informazione scientifica nel Codice comunitario; esperienze e suggerimenti per le procedure AIFA: il sistema trasparenza; il Regolamento pediatrico. Società Italiana Attività Regolatorie (SIAR); Segrate (MI). EPIDEMIOLOGIA DELLE MALATTIE PSICHIATRICHE DELL ETA EVOLUTIVA E DEI TRATTAMENTI FARMACOLOGICI. Congresso Bambini e Farmaci : tra incertezza scientifica e diritto alla salute. Istituto Superiore di Sanità (ISS) e Istituto di Ricerche Farmacologiche Mario Negri (IRFMN); Roma. February LA FARMACOEPIDEMIOLOGÍA Y EL USO DE LAS MEDICINAS EN EL NIÑO. 2 do Taller International Salud Infanto Juvenil Camagüey Hospital Pediatrico Provincial Docente Dr. Eduardo Agramonte Pina; Camagüey, Cuba. LETTURA CRITICA DELLA LETTERATURA SCIENTIFICA. BANCHE DATI BIBLIOGRAFICHE - PERCORSI DI RICERCA. I DATABASE DELLE AGENZIE INTERNAZIONALI: IL GIOCO DEGLI INDICATORI. Corso di perfezionamento in Medicina Tropicale e Salute Internazionale. Università degli Studi di Brescia; Brescia. March PERCORSO PER LA GESTIONE PSICOMOTORIA IN AREA CRITICA. Congresso Linee guida tra EBM e pratica clinica. Il trattamento del disturbo bipolare e dell agitazione psicomotoria nell adulto e in età evolutiva. Azienda Ospedaliera Ospedale Riguarda Ca Granda; Milano. FARMACI GENERICI IN PEDIATRIA: RISPARMIARE CONVIENE? IV Congresso interregionale della società italiana di pediatria OSTIA 24. Società italiana di pediatria (SIP); Ostia (Roma). LA SALUTE IN UN MONDO GLOBALE, TRA PROGRAMMI E DIRITTI NEGATI. Incontro Pochi più ricchi, tanti sempre più poveri. E questo il mondo che vogliamo?. Centro Socio Culturale COOP; Novate Milanese (MI). April PSICOFARMACI NEI BAMBINI. Corso Orientarsi in salute & sanità per fare scelte IRFMN; Milano. LO SCENARIO: EPIDEMIOLOGIA DELLA MALATTIA MENTALE. Seminario Seminario di avvio della rete ADHD. Istituto Superiore di Sanità (ISS); Roma. TRATTAMENTI PSICOFARMACOLOGICI IN GRAVIDANZA. Corso Patologie e terapie psicofarmacologiche in gravidanza. Azienda Ospedaliera di Melegnano; Vizzolo Predibassi (MI). TRATTAMENTI PSICOFARMACOLOGICI IN GRAVIDANZA. Corso Patologie e terapie psicofarmacologiche in gravidanza. Azienda Ospedaliera di Melegnano; Zizzolo Predibassi (MI). May BAMBINI E FARMACI. Corso. Gli Argonauti VIII. In viaggio per Itaca Associazione Culturale Pediatri (ACP); Messina. 211

214 EPIDEMIOLOGIA DELLE MALATTIE PSICHIATRICHE DELL ETA EVOLUTIVA E DEI TRATTAMENTI FARMACOLOGICI.. Corso I percorsi diagnostici-terapeuticiassistenziali dei bambini e adolescenti con disturbi psichici nella azienda ULSS 20 di Verona. Azienda ULSS 20 di Verona; Marzana (VR). USO DI FARMACI OFF-LABEL IN ETA PEDIATRICA. Conferenza. Ospedale dei Bambini V. Buzzi Istituti Clinici di Perfezionamento; Milano. FARMACI E ALLATTAMENTO. Corso di aggiornamento PLS (Pediatri di Libera Scelta) Udine. REAZIONI AVVERSE IN GRAVIDANZA. REAZIONI AVVERSE IN PEDIATRIA. Master Farmacovigilanza. SEFAP, Milano. June CLINICAL TRIALS OF ANTIDEPRESSANTS IN CHILDREN, WHAT HAVE WE LEARNT? - WHAT IS THE EVIDENCE OF EFFICACY? - WHAT IS THE EVIDENCE OF TOXICITY? WHY TRANSPARENCY IS ESSENTIAL. Corso 5th International Workshop on Paediatric Clinical Trial. The Medical School, Derby City General Hospital; Derby, UK. PRESCRIZIONE FARMACOLOGICA IN PEDIATRIA DI FAMIGLIA E PERCORSI DIAGNOSTICO-TERAPEUTICI. Corso di aggiornamento per Pediatri di Famiglia. Formazione pediatrica ASL Pavia; Strabella (PV). July L IMPORTANZA DEI DATI EPIDEMIOLOGICI NEI PROCESSI DECISIONALI E DI RICERCA IN SANITA. (Tavola Rotonda). Convegno Vent anni di storia dell epidemiologia del farmaco. Osservatorio ARNO, Consorzio Interuniversitario CINECA; Bologna. September IL MONITORAGGIO CRITICO DELLE PRESCRIZIONI FARMACOLOHICHE PER MIGLIORARE LA QUALITA E L APPROPRIATEZZA DELLA TERAPIA. 63 Congresso Nazionale della Società Italiana di Pediatria Difendiamo i diritti dell infanzia e dell adolescenza. Società Italiana di Pediatria (SIP); Pisa. DISUGUAGLIANZE NELLA SALUTE MATERNO-INFANTILE. Corso Health and Health care services, as indicators of Human Rights. Istituto Italiano per gli Studi Filosofici, Fondazione Lelio Basso-Sezione Internazionale; Napoli. Tavola Rotonda: l in-formazione presente e futura. Vaccinare per obbligo o per scelta?. Associazione Culturale Pediatri (ACP); Senato della Repubblica, Roma. October PSICOFARMACI & BAMBINI. Seminario Psicofarmaci e bambini. Gruppo Parlamentare Forza Italia; Milano. ADHD E SALUTE DEL BAMBINO IN ITALIA. Corso: Lo sviluppo delle Linee Guida Europee per le Terapie Farmacologiche dell ADHD: Implementazione nel Mondo Reale ed effetti sulla qualità della vita. Università degli Studi di Cagliari Dipartimento di Neuroscienze, Clinica di Neuropsichiatria Infantile; Cagliari. EPIDEMIOLOGIA. RICONOSCIMENTO DI UNA REAZIONE AVVERSA. REAZIONI AVVERSE PIU FREQUENTI IN ETA PEDIATRICA. Corso la reazione avversa da farmaco: come riconoscerla. L importanza della segnalazione spontanea. Istituti Clinici di Perfezionamento (ICO) Ospedale dei bambini V. Buzzi, Milano. 212

215 ACCESSO AI FARMACI NEI PAESI IN VIA DI SVILUPPO. XXVIII Congresso Nazionale SIFO Innovazione e Salute Pubblica. Efficacia a confronto con: Equità, Economia, Etica. Società Italiana di Farmacia Ospedaliera (SIFO); Rimini, (RN). LO SCENARIO: EPIDEMIOLOGIA DEI DISTURBI PSICHICI DELL ETA EVOLUTIVA. 19 Congresso Nazionale ACP Relazioni per crescere. Il bambino nel contesto relazionale: basi biologiche ed implicazioni cliniche e sociali. Associazione Culturale Pediatri (ACP); Trani (BA). FARMACI E ALLATTAMENTO. Corso di aggiornamento PLS (Pediatri di Libera Scelta) Udine. November LO SCENARIO: EPIDEMIOLOGIA DELLA SALUTE MENTALE IN ETA EVOLUTIVA. Corso. UMANA_MENTE, Milano. INFORMAZIONE E DISINFORMAZIONE NELL ERA DELLA COMUNICAZIONE. Seminario Dialoghi di Bioetica e biodiritto Sintonie e interferenze: le dimensioni della comunicazione in ambito sanitario. Provincia autonoma di Trento, Ordine dei Medici Chirurghi e degli Odontoiatri della Provincia di Trento, Federazione Nazionale Colleghi Infermieri IPASVI, Ordine dei farmacisti della Provincia di Trento; Trento. December METILFENIDATO E ATOMOXETINA. Congresso Confronti in pediatria La Pediatria attraverso i farmaci. Quelli facili e quelli difficili.irccs Burlo Garofano; Trieste. IL REGISTRO NAZIONALE: UNO STRUMENTO DI SALUTE PUBBLICA. Convegno Il Registro Nazionale dell ADHD. SINPIA, Milano. GRANTS AND CONTRACTS AIFA, Agenzia Italiana del Farmaco Boehringer Ingelheim CINECA, Interuniversity Consortium European Union Fondazione Monzino Il Pensiero Scientifico Editore Provincia of Milan Lombardia Region - Assessorato alla Sanità Valle d'aosta Region - Assessorato alla Sanità, Salute e Politiche Sociali 213

216 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Arias G, Palau M, Perez M, Bonati M. Philanthropic prize to remember that Latin American Children s health is a priority. Pan American Journal of Public Health (Revista Panamericana de Salud Publica) 2007;22:294. Campi R, Bonati M. Italian child health statistic review: births and deaths. Ital J Ped 2007;33: Campi R, Bonati M. Italian child health statistic review: morbidity and social habits. Ital J Ped 2007;33: Choonara I, Bonati M. European legislation to improve medicines for children. Paedat Perin Drug Ther 2007;8:2-3. Clavenna A, Bonati M. Antidepressant prescriptions in paediatric outpatients in Europe. Paediatr Perin Drug Ther 2007;8: Clavenna A, Rossi E, De Rosa M, Bonati M. Use of Psychotropic Medications in Italian Children and Adolescents. Eur J Pediatr 2007;166: Marchetti F, Bua J, Ventura A, Notarangelo LDE, Di Maio S, Migliore G, Bonati M. The awareness among paediatricians of off-label prescribing in children: a survey in Italian Hospitals. Eur J Clin Pharmacol 2007;63: Rossignoli A, Clavenna A, Bonati M. Antibiotic prescription and prevalence rate in the outpatient paediatric population: analysis of surveys published during Eur J Clin Pharmacol 2007;63:

217 LAY PRESS SELECTION PUBLISHED IN 2007 Bianchi M, Clavenna A, Labate L, Bortolotti A, Fortino I, Locatelli GW, Giuliani G, Bonati M. Profilo prescrittivo dei farmaci antiasmatici nella popolazione pediatrica della ASL di Lecco. Medico e Bambino 2007;26: Bonaccorsi A. All origine delle disuguaglianze nella salute (traduzione). R&P 2007;23: Bonati M. 2007: l anno dei farmaci per i bambini? Quaderni acp 2007;14:241. Bonati M. Dalla letteratura...pillole di buona pratica prescrittiva. L'utilizzo dei farmaci nell'infanzia: il programma del Ministero della Salute. Newsletter 2007;n.7:3. Bonati M. Disturbi mentali e farmaci: il caso dell ADHD e degli psicostimolanti. Medico e Bambino 2007;XXVI: Bonati M. I disturbi mentali dell età evolutiva: una distorsione tra richiesta e offerta di aiuto. Informazione sui farmaci 2007;3: Bonati M. I farmaci: una delle aree di intervento. R&P 2007;22:270. Bonati M. L anestesia non fa la rete e neppure l equità. Il Sole 24 Ore Sanità 2007;17-23 aprile:3. Bonati M. L utilizzo dei farmaci nell infanzia. Bollettino SIFO 2007;53:109. Bonati M. Quando è l evidenza erariale a sopravanzare quella scientifica. R&P 2007; 23: Bonati M. Una distorsione tra richiesta e offerta di aiuto. SocialNews 2007; ARTICOLIAPRILE2007/Aprile2007Bonati_1.htm. Bonati M, Campi R. Le disuguaglianze tra Nazioni nella salute infantile. Quaderni acp 2007;14: Bonati M, Campi R. La promozione della salute nei Paesi del Sud del mondo. Quaderni acp 2007;14: Bonati M, Gruppo di Lavoro sui Farmaci Pediatrici (Istituito presso AIFA) Decongestionanti nasali: nei bambini i rischi superano i benefici. Medico e Bambino 2007;5: Briantea Paediatric Surveillance Unit. (Bonati M, Calati MG, Clavenna A). Il monitoraggio epidemiologico delle malattie: l esperienza pilota di otto ospedali sentinella. Quaderni acp 2007;14: Campi R, Bonati M. Bambini poveri e disuguali in Italia. R&P 2007;23: Centro di Informazione sul Farmaco e la Salute. Otite media acuta: meglio attendere prima dell antibiotico. Quaderni acp 2007;14:45. Centro di informazione sul Farmaco e la Salute. Decongestionanti nasali: attenzione, pericolo di morte. Quaderni acp 2007;14: 86. Cerzani M, Pasina L, Clavenna A, Nobili A, Garattini L. Revisione critica degli studi italiani di farmacoeconomia sull uso dei farmaci antinfiammatori non steroidei in medicina generale. Quaderni di Farmacoeconomia 2007; 3: Clavenna A. In viaggio tra le evidenze. R&P 2007;23:119. Clavenna A. Società Italiana di Medicina delle Migrazioni. R&P 2007;23:74. Clavenna A, Bonati M. Dieta, Integratori, antibiotici e autismo. Medico e Bambino 2007; 26: Clavenna A, Bonati M, Campi R, Labate L, Nobili A, Pasina L,Tettamanti M, Monesi L, Marzona I, Roncaglioni C, Bortolotti A, Fortino I, Locatelli GW, Giuliani G. La prescrizione di farmaci per i bambini e gli anziani nella ASL di Lecco. R&P 2007;23: Clavenna A, Fortinguerra F. Antibiotici: usare con cautela. Quaderni acp 2007;14:125. Clavenna A, Fortinguerra F. Desmopressina nel trattamento dell enuresi: evitare gli spray. Quaderni acp 2007;14:226. Clavenna A, Fortinguerra F. Farmaci antiraffreddore ancora nella bufera. Quaderni acp 2007;14:272. Clavenna A, partecipanti al corso La risoluzione di scenari clinici con il supporto della EBM come strumento di formazione continua per il pediatra. Esiste una terapia farmacologica per la sindrome delle apnee notturne? Quaderni acp 2007;14: Conti Nibali S, Bonati M, Font M, Toffol G. La prescrizione per DCI in Pediatria ambulatoriale Studio di fattibilità. 215

218 Quaderni acp 2007;14: Conti Nibali S, Bonati M. DCI nella medicina generale: studio di fattibilità. Dialogo sui Farmaci 2007;2: De Rosa M, Rossi E, Bonati M, Clavenna A. L 85% sono antibiotici, antiasmatici e corticosteroidi. Il sole 24 ore Sanità 2007; 30gen-5feb:3. Grandori L, Bonati M, Gangemi M. 8 passi di prevenzione a tutela della salute dei bambini. Medico e Bambino 2007;25: Fortinguerra F, Clavenna A, Baviera M, Cattaneo A, Conti Nibali S, Labate L, Maschi S, Miselli M, Scurti V, Zanfi D, Zermiani G, Bonati M. Le inserzioni pubblicitarie sulle riviste italiane del medico pediatra. R&P 2007;23: Labate L. Dimensione campionaria. R&P 2007;22: Maschi S, Fortinguerra F, Clavenna A, Bonati M. Levotiroxina in età pediatrica. Dialogo sui farmaci 2007;5: OTHER PRODUCTS PUBLISHED IN 2007 Book Chapters Bonati M, Campi R. Le disuguaglianze fra nazioni nella salute infantile. In: Oltre il DNA Scienza, società e cittadinanza. IBIS, Como; Pavia 2007: Clavenna A, Bonati M. Farmacologia e tossicologia in gravidanza e allattamento. In: Ostetrica e Ginecologia. Zanoio, Masson spa, Milano, 2007;Cap.23: Bonati M, Campi R e Gruppo di Lavoro per la Convenzione sui Diritti dell Infanzia e dell Adolescenza. I diritti dell infanzia e dell adolescenza in Italia. 3 rapporto di aggiornamento sul monitoraggio della Convenzione sui diritti dell infanzia e dell adolescenza in Italia Gruppo di lavoro per la CRC c7o Save the Children, Roma, 2007; Book Cirillo B, Bonati M, Campi R, De Campora E, Siani P. Disuguguaglianze nella salute nell infanzia e nell adolescenza in Campagna. Phoebus Edizioni, Casalnuovo, Napoli Schiavetti B, Clavenna A, Fortinguerra F, Miglio D, Bonati M. Psicofarmaci in allattamento. Il Pensiero Scientifico Ed., Roma International Abstracts Bianchi M, Clavenna A, Labate L, Bonati M. Anti-asthmatic drug prescriptions in Italian children and adolescents. Paediatr Perinat Drug Ther 8:72;2007. Pandolfini C, Bonati M, Rossi V, Santoro E, Naylor C, Sammons H, Choonara I, Zarrabian S, Jacqz-Aigrain E, Castel JM, Danés I, Fuentes I, Arnau J. The DEC-net European register of paediatric drug therapy trials: its contents and their context. Paediatr Perinat Drug Ther 8:72; Colombo C, Mosconi P, Bassi C, Bernardi F, Bosisio M, Copelli P, Crotti C, Erroi A, Magri M, Pandolfini C, Pistotti V, Santoro E, Zucchetti E. Evaluating cancer pain websites. MedNet th World Congress on the Internet in Medicine, 7-10 October 2007, Leipzig, Germany 306;

219 RESEARCH ACTIVITIES Psychotropic drugs and breastfeeding The major objective in creating this small formulary is to provide health professionals with a therapeutic guide that not only includes drug monographs, but also acts as an essential tool (that is accurate, effective, and updated) for promoting the rational use of psychotropic drugs. The formulary will be a source of information for current problems in daily clinical practice, especially for psychiatrists, family paediatricians, family physicians, and obstetricgynaecologists. It will be an important tool for psychiatrists for their specialistic pertinence, for family paediatricians for their role in caring for newborns, for family physicians for their prescription activity, and for obstetric-gynaecologists for their role in the pre- and peri-natal periods. The drug information reported in patient leaflets is limited, generic and generally vague in order to protect manufacturers, since little safety (and pharmacokinetic) information is available on the use of the majority of drugs during breastfeeding. In fact, under special precautions we often read: There is no evidence on drug excretion in milk. The benefits for the mother must be evaluated in relation with infant risks. The documentation on real or potential risks, however, is often inexistent. For those drugs on which information is available (often only pharmacokinetic data with respect to the mother s milk/plasma ratio and not with respect to safety) the patient leaflet may report: During breastfeeding, the possibility of early weaning must be considered. Or If the treatment is considered necessary for a breastfeeding mother, she must discontinue breastfeeding to avoid having the breastfed infant ingest even minute drug amounts excreted in breast-milk. The usefulness of these sentences, which should be decisional for physicians and informative for patients, is insufficient. Current medicine is founded on the basing of therapeutic choices on documented safety and efficacy evidence. There is therefore a need to systematically collect and evaluate the available information on the risks of psychotropic drugs during breastfeeding and, briefly, also in pregnancy, as a part of a continuum that cannot be separated. This Guide is unique, and not only at the national level, although its structure reflects the working experience of the Drugs and Human Lactation Working Group founded in 1985 by the European section of WHO. The work behind the creation of this Guide is seen as an on-going, collegial review. It would be wonderful if the work could proceed with the readers participation, including reporting any errors (we hope few) and suggesting any improvements. This would be fundamental for this product to become increasingly useful in clinical practice and to possibly be extended to other drug classes and problems. Child and adolescent health inequalities in the Campania region This book is dedicated to those who provide care to children and families and who don t accept these injustices and fight to reduce and eliminate them. It is dedicated to all those health care operators who believe reducing health inequalities is the right thing to do and who are aware of the change in the epidemiology of family and child health problems (reduction of childhood organic disorders, increase in chronic diseases and in relative mortality, especially that which is determined socially, reduction of comunicable diseases, and increase in psycho-social problems), who want to have a comprehensive view of the causal chains in health and a longitudinal approach to the path of life. It is dedicated to all those who think that today s children are tomorrow s men and women, that childhood inequalities will become inequalities in later phases of life, and that being aware of, and fighting, them can contribute to society in general s health and wellbeing.(carlo Corchia, Dante Baronciani, Epidemiologia della disuguaglianza nell Infanzia, 1995). 217

220 The book is divided into five fundamental parts that address the health determinants in order (from the distal ones socioeconomic, social capital to the proximal ones behaviours and lifestyle habits), access to health services, mortality, and, in conclusion, strategies for reducing health inequalities. In testimony of Italy s, and all industrialised countries, persistence of profound inequalities in children s health, it describes the regional and sub-regional differences in the distal and proximal mortality determinants. Workgroup on the Convention for child and adolescent rights The Laboratory for Mother and Child Health is part of the Workgroup on the Convention for child and adolescent rights (CRC) in Italy. The group was set up in December 2000 with the aim to prepare a report on the status of children in Italy that would act as a supplementary (not alternative) report to that presented by the Italian government, to be submitted to the United Nations Committee on child and adolescent rights. The first report was presented in Rome in 2001 and underwritten by 42 associations and non-governmental organizations. On January 31, 2003, following a public meeting held in Geneva between the United Nations Committee and a large governmental delegation a document was created. This document highlighted Italy s progress in order to enact the Convention, but also reported on worrying matters concerning the lack of adherence to certain principles, and precise recommendations are listed for which the government will have to account in the next United Nations report in 2008 (CRC/C/15/Add.198). While carrying out the monitoring activity, the workgroup decided on the creation of an annual, updated report on the status of children and adolescents in Italy and on the related United Nations Committee s recommendations, focusing on priority issues. The report is not only intended to identify our health system s faults, but also to be an opportunity to prompt a timely and constructive debate among the institutions responsible for the status of children and adolescents in Italy, and, therefore, for the implementation of the rights guaranteed by the convention. The third report, in addition to updating the monitoring of the issues already addressed in the previous report, and to developing additional, detailed discussion, adds to the analysis with two new chapters: the second one dedicated to the CRC s general principles, starting from the monitoring of the principle of participation (art.12 CRC), and the third dedicated to the family environment and alternative measures. The objective of the workgroup is, in fact, to expand on the issues covered until it addresses all eight groupings into which the United Nations Committee has divided the CRC s articles, in the next supplementary United Nations Report. All this is carried out with the hope that it will stimulate, and contribute to, the development of standard procedures and legislative reforms that will bring about a tangible improvement in the condition of all youths in Italy. ARNO paediatric project For over five years one million children and adolescents, and their families, have made up the ARNO-paediatric project s population. The project, a product of the collaboration between the Mario Negri Institute in Milan and the CINECA in Bologna, is unique for its size and duration at both the national and international levels. The initiative is part of the larger ARNO project that monitors out-hospital drug prescriptions and that has been active at the CINECA since 1986 with the participation of numerous local health units across Italy. During 2006, 561,237 children aged less than 14 years (61% of total) were prescribed 620 different drugs (active moieties), using 1,805,521 prescriptions, for a total of 2,697,979 boxes. Each child received an average of 3 prescriptions and 5 drug boxes. The prescription prevalence was highest in 1-4 year old children (76%), decreasing progressively with increasing age and reaching 43% in 13 year olds. It was higher in males than 218

221 females (62 vs 59%). Antibiotics (52%), antiasthmatics (26%), and systemic corticosteroids (8%) were the most prescribed drug classes and corresponded to 84% of prescriptions. The amoxicillin-clavulanic acid association was the most prescribed drug (461,755 boxes to 217,720 children), followed by amoxicillin (256,892 boxes to 129,788 children), and by beclometasone (195,443 boxes to 137,031 children). Beclometasone was the most prescribed drug in children <1 year old (1/5 of the children in this age group received at least one box of this drug). In children 1 year old the amoxicillin-clavulanic acid association was the most prescribed. From these results, it is evident that a total of 15 drugs would suffice to cover the most common needs, independent of age; 6 of these appear among the 10 most prescribed in all age classes. The qualitative and quantitative drug prescription profiles are much different from those of other European countries, implying that there is a need for educational (research) initiatives and for active information-dissemination initiatives aimed at making the use of drugs in children more rational. VI Master s Course An inter-university course aimed at specialising students in the planning and management of cooperation projects for development. The sixth edition of a 10-day Stage in Cooperation and Public Health, part of the Master s Course in Analysis and Management of Projects for Development ( was organised by the Laboratory and held at the Mario Negri Institute in Milan. The Stage involved a in-depth examination of the problems associated with public health in developing countries, taking into consideration their primary importance to the entire world s population in this new millennium. The course was based on both theory and practice. The lessons involved issues such as coping with emergency situations, providing assistance for extreme needs, addressing the problem of access to drugs, health and well-being indexes, analyses of living conditions, and intervention programs, and were complemented by hands-on practice sessions that also involved the basics of epidemiology. National ADHD Register The marketing authorisation for methylphenidate and atomoxetine in Italy has made monitoring the use of these drugs in children with attention deficit hyperactivity disorder (ADHD) necessary. In order to meet this need, a national registry coordinated by the Istituto Superiore di Sanità s drug department, in collaboration with the Italian Drug Agency and the Conferenza permanente degli Assessori alla Sanità delle Regioni and the Province autonome di Trento e Bolzano, was set up and activated on June 18, The Laboratory for Mother and Child Health is part of the scientific committee and participated in writing the protocol, which defines the structure and activities of the national ADHD register and its monitoring. The registry s aims are : to monitor the use of methylphenidate and atomoxetine; to evaluate the safety and compliance of therapies with methylphenidate and atomoxetine, alone or in combination with other therapeutic interventions (pharmacological or not) in the medium and long term; to define the probability of developing ADHD requiring pharmacological treatment in the school-aged population; to define the optimal management strategy for ADHD through the standardisation of the most appropriate diagnostic and therapeutic strategies; to evaluate the effectiveness of psychotropic drugs and/or behavioral therapy in the evolution of ADHD in school-aged children; to calculate the long term risk of persistence of the disorder, of being left behind in school, and of the development of psychosis or other psychiatric disorders. 219

222 The first evaluation of the outcomes will be made after 24 months of the register s activation. FARMACI E BAMBINI workgroup A paediatric group of the Italian Drug Agency (AIFA) was set up on April 4 th, The Laboratory for Mother and Child Health, along with the Associazione Culturale Pediatri (ACP), Centro Salute del Bambino (CSB), Federazione Italiana Medici Pediatri (FIMP), Società Italiana Farmacia Ospedaliera (SIFO), Società Italiana di Pediatria (SIP), and experts in the sector and members of the AIFA, participate. The group s function is to support the AIFA s regulatory activities, informational initiatives, pharmacovigilance work, and promotion of independent clinical research concerning drug use in the paediatric population. The workgroup s activities currently cover three broad areas: Pediatric research: the group s role is to help define the paediatric population s needs and priorities in the research field. The group also aims to suggest the issues that should be reported to the Commissione Ricerca e Sviluppo (CRS) to choose the new independent research grant offers to finance at the Italian level and to eventually transfer to the European level. Pharmacovigilance: to help analyse the reports received by the spontaneous reporting network, thus helping to develop a more active, more efficient pharmacovigilance system. Information/training: to recommend the development of adequate tools for health care workers. More specifically, two of the workgroup s main activities involve the assessment of the offlabel use of drugs and of the adverse drug reactions received by the National Pharmacovigilance Network. During 2007, 3 dear doctor letters concerning the paediatric population were created and distributed: a new contraindication to the use of nasal decongestants containing sympaticomimetics for topical use in children less than 12 years old; new safety information concerning the use of cefaclor and the updating of the Summary of Product Characteristics was issued; and new, important information concerning the use of desmopressin in the nasal spray formulation was issued. A formal article was published on a few national journals describing an evaluation of nasal decongestants. Furthermore, a systemic evaluation of reports concerning the use of antiemetics, especially metoclopramide and domperidon, was carried out. Important safety information is available on the AIFA website Vaccine commission The Laboratory for Mother and Child Health is part of the National Vaccine Commission and the Lombardy Region s Vaccine Commission. Ministero della Salute. The National Vaccine Commission was set up with a Ministerial Law on February 20, The main goal of the commission is to update the National Vaccine Programme, a document whose main objectives are: to maintain a high vaccine coverage rate for the diseases for which the National Health Plans and , as well as the WHO s Regional European Office, reached their goals; to promote appropriate vaccine interventions aimed at recuperating optimal coverage of those vaccines for which a basic working strategy has already been defined, but for which optimal coverage has still not been reached; to provide indications on the new objectives, and on the startup of focused initiatives aimed at prevention through vaccination following the recent availability of new vaccines, as indicated in the National Health Plan ; to increase the safety of immunisation practices; to increase the structural, organisational, training, and communication-based interventions in order to permit the evolution of vaccine policies from mandatory interventions to those characterised by an informed participation on the part of families. Lombardia Region. At the regional level, the group is called the Technical-scientific commission for the planning and verification of vaccinations, based on its assigned tasks. Its main goals are: the elaboration of vaccine strategies, based on epidemiologic profiles; evaluation of the ongoing interventions, in terms of protective efficacy and unwanted 220

223 effects;promotion of the qualitative improvement of vaccine management difficulties, including the vaccine registry. This commission developed a document entitled Vaccines in children and adults: review and reorganisation of vaccine prophylaxis in the Lombardy Region, approved with the 22/12/05 law DGR VIII/1587. The aims of the document are to adapt the national and European vaccine strategies and policies to the community and organisational contexts present in the region, providing guidelines for the improvement of the quality of the vaccination programme offered. Technical commission for the elaboration of the regional therapeutic formulary The Valle d Aosta Autonomous Region set up an agreement with the Mario Negri Pharmacological Research Institute, assigning the Laboratory for Mother and Child Health, as temporary representative of the Mario Negri Institute, the periodic task of providing technical advice on the fulfillment of the following activities: updating and revision of the regional therapeutic formulary (PTR); predisposing opinions on trial protocols on pharmaceutical drug products and other therapeutic remedies; creating documentation on topics on which the collaboration is based. Paediatric pain guidelines The Presidio Ospedaliero degli Spedali Civili di Brescia s health management committee set up 3 work groups, selecting members from among physicians and nurses representing various paediatric wards, in order to create guidelines concerning: pain evaluation and treatment in a paediatric emergency ward; the use of analgesics during venipuncture procedures, positioning of the needle cannula, and capillary blood sampling in neonates; topical analgesic use during venipuncture procedures and positioning of the needle cannula. The scientific co-ordination, necessary for the creation and evaluation of the guidelines, has been assigned to the Laboratory for Mother and Child Health. Health and Drug Information Centre The Health and Drug Information Centre (HDIC), initially created to promote the policy on hospital drug formularies, progressively expanded to cover out-of-hospital practice and to involve different types of users, including professionals and members of the public. The core of the HDIC s activities is the daily telephone service providing information, also to the lay public, and advice on drug therapy and needs during breastfeeding and in childhood. In January 2004, the centre s activity concerning drug information in pregnancy was taken over by Bergamo s Clinical Toxicology Unit s Poison Control Centre (Centro Antiveleni, Unità di Tossicologia Clinica, Ospedali Riuniti di Bergamo). Together, the Poison Control Centre and the Laboratory of Mother and Child Health form the Department of Clinical Pharmacy and Pharmacology. During 2007, 599 people contacted the Health and Drug Information Centre with a total of 1001 questions. In all, 80% were from people who were directly involved (patients or family members), 13% from general/hospital paediatricians, 3% from general practitioners or other non-paediatric specialists, and 4% from other health care workers. A total of 817 requests for information concerned drug use, 184 the use of parapharmaceuticals, general information, and diagnostic exams. In all, 93% of the questions received by the centre concerned the safety of drugs during breastfeeding, while 7% concerned the administration of drugs to children. Although people called from all of Italy, most called from the north of the country (68%), especially from the Milan (228) and Brescia provinces (32). 21% of the requests were received from central Italy and 13% from the south. Drugs and breastfeeding. 221

224 Of the 931 requests for information on breastfeeding, 79% came from 491 mothers and/or relatives, while the remaining 21% from 144 health care workers, of whom 92 were paediatricians. The mothers average age was 34 years (range 22-45) and that of the breastfed babies was 5 months (range 0-42). Almost half (45%) of these babies was 3 months. In all, 81% of the newborns were exclusively breastfed and 12% were fed artificial formula in addition to breast milk. The type of nutrition was still undecided for the remaining 7%. The drug classes for which information during breastfeeding was most often requested were: analgesics/antiinflammatories (11%), systemic antibacterials (9%) and psychotropic drugs (8%). More specifically, psychotropic drugs and antiepileptic agents were the classes for which information was most often requested on the part of health care workers. The indication most frequently associated with drug use was symptomatic pain control (109 queries), followed by allergy (64), and psychiatric disorders (66). A follow-up for 67 mothers whose child turned 6 months old was completed. Of these mothers, 66% continued exclusive breastfeeding, 9% had introduced artificial milk in the baby s diet, and 25% had stopped breastfeeding. In all, 52 mothers had taken the drug for which they had asked information and 15 did not take it. The reasons for not having taken the drug were: because it was no longer necessary (n=13) and for fear (n=2). In all, 48% of the newborns whose mothers were re-contacted for follow-up had been given one or more drugs in the first 6 months of life: of the 83 drugs, 18 were anti-antinfectives, 17 were antiasthmatics, and 10 antacids/proton pump inhibitors. Paracetamol was the most commonly prescribed drug (8), followed by amoxicillin+clavulanic acid (6), beclomethasone (6), and salbutamol+ipratropium bromide (6). The most common reason for prescribing drugs was bronchitis (n=15). Drugs and children The requests for paediatric drug therapy information concerned 39 children aged 1 month to 12 years. In all, 30 mothers/fathers, 7 family or hospital paediatricians, and 1 health care worker contacted the centre. Of the 70 queries received, 62 concerned general drug information, 7 concerned side effects, 1 concerned dosage and 1 efficacy. The most common therapeutic groups concerned psychotropic drugs (10), system antibiotics (8), and nasal products (5). The most common reason for prescribing drugs was respiratory tract infection (16). Folic acid and SIDS As part of the centre s activity involving the monitoring of the general population s level of health awareness, the information the mothers received on the use of folic acid in pregnancy to prevent neural tube defects and on the recommendations concerning sudden infant death syndrome (SIDS) was collected. Concerning folic acid, 91% of the mothers had taken it during pregnancy, but only 21% had begun to take it before conception. Of the 339 newborns for whom information could be collected, 286 were placed in the supine sleep position, 46 on their side, and 7 face down. In 16% of the cases, the position was incorrect for SIDS prevention. Over half of the mothers of those newborns had not received any information concerning the safest sleep position to place their children in. Ricerca & Pratica Ricerca & Pratica was born in January, 1985, as a manifestation of the Mario Negri Institute for Pharmacological Research. Today, the journal is part of the International Society of Drug Bulletins (ISDB), which represents independent journals. For more than twenty years, the journal has represented an arena for all those professionals who collect data and carry out studies in general practice with the aim to increase their knowledge and to improve their practice. Ricerca & Pratica is also appreciated for its ability to go beyond the merely clinical aspect of medicine, without, however, forgetting that it is to this aspect that the readers dedicate most of their time and effort. 222

225 Through its activity, Ricerca & Pratica can therefore represent an exclusive, independent observation point. It is also an area that promotes contemplation, evaluation, and information by applying tools such as data trustworthiness and importance, the balance between benefits and risks and between benefits and costs, independence from conflicts of interest, and the realistic objective to contribute to a progressive, equally distributed improvement in the population s health....and then Offertasociale: Offertasociale runs several social services that protect the weaks within a consortium of 29 counties in the Vimercate and Trezzo areas and entrusted the Laboratory for Mother and Child Health with the carrying out of a course aimed at developing wellness indicators for the young. In order to inform and assess public policies, it is necessary to have local indicators with which to measure community wellness. The population s quality of life is increasingly tied not only to per-capita income, but to a set of multidimensional factors. Among these are life expectancy at birth, literacy and education rates, school drop-out rates, etc. Learning to read the level of cohesion/social exclusion, also through the analysis of data on troubled or hospitalised youth, those living in troubled youth centres, etc, is necessary. All this suggests that the communities, and the operators that provide services, learn the specific tools and methods related to a renewed cultural dimension based on concepts of wellness, lifestyle quality, equal access to services, etc. The course provided the tools for building a community level development index that took into consideration the indicators that could be used to represent the three dimensions of human development (income, education, and health) for the 29 counties in the area: the community level development-wellness index - Human Development Index for the 29 counties starting from the data received and from the proposed indicators. A report on a few determinants of human development of youths residing within the consortium of 29 counties was produced during Nascere in casa: The Associazione Nazionale Culturale Ostetriche Parto a Domicilio (National Cultural Association of Home Birth Obstetricians) aims to increase home births, guaranteeing their quality and safety, verifying and comparing the national and international experiences of care outside the hospital, and defining professional selection and assistance criteria based on scientific evidence from the World Health Organization s recommendations and on women s wishes. The Laboratory for Mother and Child Health collaborates with the association, analysing data collected by the obstetricians in order to improve the quality of care offered to women and children through experience, but also through continuous study, continuing professional training, and debate. These improvements will help protect out-ofhospital births and safeguard the obstetricians specific professional role. Co-operation with countries with limited resources As an expression, test, and original method of expression of the choice to make the Laboratory s research transferable and accessible to all populations, the Laboratory promoted and provided assistance to projects in, and for, the South of the world, also in collaboration with the World Health Organization. The technical and organisational support for local groups and international non-governmental organisations for carrying out socio-sanitary projects in countries with limited resources, especially Colombia, Ecuador, Brazil, Bolivia, Cuba, Vietnam e the Balkans, continues. Bolivia: On April 20, 2007 the laboratory organised a one day meeting at the Mario Negri Institute, entitled: Being born today in Potosì: From Bolivia to the Lombardy Region: a twodirectional, collaborative path. The results of a project, supported by COOPI and the Lombardy Region, for the improvement of the health of both women during pregnancy and of 223

226 young mothers, by facilitating access to health centres though initiatives aimed at integrating the official medical system and the traditional, community level medical system. Reduction of the high maternal mortality in the city of Tinguipaya, Department of Potosí, through the integration of the official and traditional, local medical systems Cuba: 2do Taller International de Salud Infanto-Juvenil. Hospital Pediatrico Provincial Docente Dr. Eduardo Agramante Piña Camagüey The Laboratory actively supported the second International workshop on child health in Cuba, held in Camagüey, aimed at implementing paediatric hospital care, evaluating it, and rendering it informed and up-to-date. The issues addressed highlighted the fact that the family practitioner in Cuba has, in the last 20 years, gained increasing importance. Every practitioner assists about 125 families. The health care system is also supported by a series of polyclinics that mostly manage emergencies. Change is also under way for the paediatric hospitals, and involves continual, significant improvements. Medical students dedicate 15 weeks of study to paediatrics and an additional 6 during the last academic year. In Camagüey there are about 150 students enrolled in medicine, 80 of whom come from Latin America and who have been selected from among the poorest countries. The aim is to train competent staff in order to improve the health of the populations in countries with limited resources. There are, today, 80,000 Cuban physicians, 20,000 of whom work abroad (10,000 just in Venezuela). Ecuador: The Laboratory for Mother and Child Health, as epidemiologic scientific advisor and organisational partner, has participated since 2005 in the project entitled Project for the reduction of maternal and perinatal mortality in the S.Lorenzo District - Esmeraldas -Ecuador Region. The project is co-ordinated by the Bussolengo s (VR) local health unit as part of its work in carrying out decentralised cooperation interventions for international development and solidarity. The Laboratory has two other projects in Ecuador, in collaboration with Cecomet (Centro de Epidemiologia comunitaria y Medicina tropical) in Borbòn s health district, situated north of the Esmeraldas province, Ecuador s rural area. The names of the projects are: Improving vaccine coverage along the Santiago and Cayapas rivers. The aims of the project are to: improve vaccine coverage in the under 5 population, in schoolchildren, in women of child-bearing age, and in pregnant women in order to avoid vaccine-preventable morbidity and mortality; optimise community vigilance of vaccine-preventable diseases so that the programme s advancement can be monitored; and improve the families knowledge of vaccines and their importance. Reducing child mortality through the supplementation of vitamins and nutritional elements. The integration of vitamin A reduces the risk of child mortality. Lack of this vitamin makes children especially vulnerable to infections and leads to greater disease severity. It is also the main cause of blindness in children. According to UNICEF, only 25% of children under 5 receive vitamin A supplementation, which is essential for development and for controlling malaria. The study also involves zinc supplementation: lack of zinc in undernourisced children is among the factors leading to limited physical development and increases predisposition to infections, lack of zinc in pregnant women increases the risk of mortality; iron supplementation: its deficiency leads to severe anemia both in pregnant women and in children; iodine; and vitamin D. 224

227 CENTRE OF COMPUTER SCIENCE ENGINEERING STAFF Head CLIVIO Luca The year 2007 has been for the Centre of Computer Science Engineering the first year of life and at the same time the most challenging from the point of view of works about the networking and server-side infrastructures, that has been heavily revised. About the clinical trials handling software the year 2007 has been the first in witch many of the AIFA sponsored studies have been implemented with case report forms handled by internal made software, while about the area of office automation some of the most important software for administrative tasks have been completed. Also in the area of the multimedia communication this has been an important year: the event of beginning working in the new Institute has carried the necessity (and the opportunity) of preparing a lot of divulgative electronic publishing, such as DVDs, and the new institutional web site. Follows a short list of some of the principal collaborative activities done during the year. Clinical Trials and related eforms (E-CRF) Lab.: Neurological Disorders (Neuroscience Dep.): Extension European Register SLA Trial L-ACETYLCARNITINE Trial ANTIEPILETTICI Trial EPILESSIA E STROKE Trial EPO VS MP IN SPINAL SHOCK Trial VALPROATO Trial Immunoglobuline Lab.: Clinical Trials (Oncology Dep.) Trial FOLFOX Trial MAPS Trial STARPAN Trial HEAD & NECK Trial TOP Trial TAILOR Lab: Clinical Epidemilogy Trial CADASIL Lab.: Quality Assessment of Geriatric Therapies and Services (Neuroscience Dep.) Trial GISAS Registro dei Pazienti per lo studio delle Polipatologie e Politerapie SIMI network Italian Cochrane Centre 225

228 Trial ICEKUBE SINPE (Società Italiana di Nutrizione Artificiale e Metabolismo) DOMUS: Registro Italiano dei Pazienti nutriti per via artificiale IEO (Istituto Europeo di Oncologia) Trial THALDODEX Implementation at IEO of a complete software platform for handling no-profit trials Web based applications related to other research projects Realization of a software for clinical trials users administration Implementation of a software for an assisted analysis of drugs interactions ( in collaboration with Assessment of Geriatric Therapies and Services Lab.) Implementation E-Form for the project PAINCARE-LUVI for an evaluation of the online sources of scientific informations (in collaboration with Medical Research and Consumer Involvement Lab., and LUVI foundation) Implementation E-Form for the project NO-GRAZIE about the spots and promotions of drugs (in collaboration with Mother and Child Health Lab.) E-Form for online registration to workshop SCARLET (in collaboration with Environmental Chemistry and Toxicology Lab.) Electronic board Negri Community Web based application for office automation New booking system for meeting rooms in the Institute New scientific event registration and reports New employed (staff) registration handling system New database for managing the scientific publications New database for donors registration Centralized registration and handling of physician and projects involved in ECM convention Centralized registration for research project related to Animal Care Unit Multimedia communication Realization of networking and software infrastructure for managing the videoconference meeting and a remote assistance VPN between Mario Negri Milano and Mario Negri Bergamo institutions Realization of a DVD about Parkinson syndrome (in collaboration with Geriatric Neuropsychiatry and photographic unit). Video editing and preparation of a DVD for presenting the new Institute (collaboration with photographic unit). Web sites New official institutional web site Web site project GISAS Web site Fondazione Mattioli Web site MANGO group New web site project Rischio & Prevenzione Teaching activities Internal course on standard SQL language for database querying finalized to data analysis of prescriptions in Regione Lombardia. Internal course on E-CRF finalized to Case Report Form preparation for clinical trials. 226

229 ITALIAN COCHRANE CENTRE STAFF Head Alessandro LIBERATI, M.D. 227

230 CURRICULUM VITAE Alessandro Liberati obtained his MD Degree in 1978 at the University of Milano and his post doctoral degree in Hygiene and Preventive Medicine in 1981 at the same university. Teaching activities: Primary responsibility of several academic and non academic training courses on the methodology of clinical research and systematic reviews/metanalyses. He is Director of the advanced Master Evidence based medicine e metodologia della ricerca sanitaria, at the Università degli Studi di Modena e Reggio Emilia. Areas of scientific expertise: methodology of clinical research with particular reference to controlled clinical trials, epidemiological methods for research synthesis (systematic reviews and metanalyses); methods of practice guidelines production and implementation, evaluation of ethical implications of clinical research. Past and current roles at the Mario Negri Institute: Junior researcher at the Laboratory of Clinical pharmacology; Head, Unit of Clinical Epidemiology and Health Services Research; Head Laboratory of Clinical Epidemiology; since 1994 he is Director of the Italian Cochrane Centre; since 1998 he is Associate Professor of Clinical Biostatistics and Epidemiology at the University of Modena and Reggio Emilia; since 1997 he is President of the Associazione per la Ricerca sulla Efficacia della Assistenza Sanitaria - Centro Cochrane Italiano (AREAS-CCI); since 2005 he is President of the Local Ethics Committee of the Local Health Unit of Bologna; since 2004 he is Member of the Commissione Nazionale Ricerca Sanitaria; since 2005 he is Member of the Commissione Ricerca e Sviluppo dell Agenzia Italiana del Farmaco (AIFA). Selected publications Liberati A, Moja LP, Moschetti I.The future of clinical research: why do we need an ecological approach? Recenti Prog Med. 2006; 97: Vigna-Taglianti F, Vineis P, Liberati A, Faggiano F. Quality of systematic reviews used in guidelines for oncology practice. Ann Oncol ;17: Clinical Evidence edizione italiana, IV edizione Zadig, Milano. Moja L., Moschetti I., Liberati A., Gensini G.F., Gusinu R. Understanding systematic reviews: the meta-analysis graph (also called forest plot ). Intern Emerg Med 2007;2: Moja L., Virgili G., Liberati A., Gensini G.F., Gusinu R., Conti A.A. Scale to climb borderline personalities: when science goes nowhere. Intern Emerg Med 2007;2: Moja L., Minozzi S., Liberati A., Gusinu R., Gensini G.F. The drama of cancer pain: when the research abandons patients and reason. Intern Emerg Med 2007;2: Moja L., Moschetti I., Liberati A., Gensini G.F., Gusinu R. Systematic reviews highlight the complex balance between good and harm from screening studies. Intern Emerg Med 2007;2(1):57-9 Moja L., Moschetti I., Liberati A., Manfrini R., Deligant C., Satolli R., Addis A., Martini N., Dri P. Using Clinical Evidence in a national continuing medical education program in Italy. PLoS Medicine (5) e113.doi: /journal Mosconi P., Colombo C., Satolli R., Liberati A. PartecipaSalute, an Italian project to involve lay people, patients' associations and scientific-medical representatives on the health debate. Health Expect 2007 Jun;10(2): Clinical Evidence edizione italiana, V edizione Zadig, Milano. 228

231 INTRODUCTION TO THE CENTRE'S ACTIVITIES The Italian Cochrane Centre (ICC) ( was founded in 1994 and is affiliated to the Cochrane Collaboration (CC). The CC is an international non profit organization that prepares, maintains and promotes systematic reviews of the effects of health care interventions. The main product of the Cochrane Collaboration is the Cochrane Library, a quarterly publication containing Cochrane systematic reviews and other relevant databases of other siblings international organizations. The objectives of the ICC are centered around supporting various activities of the Cochrane Collaboration within Italy. In particular: a) to disseminate the knowledge of CC and CC activities throughout Italy; b) to provide methodological and practical support to all individuals and groups who are interested in collaborating with the CC; c) to contribute to the adoption and dissemination of Evidence-based Medicine in Italy. FINDINGS/MAIN RESULTS The ICC has created a national network with researchers and health care providers who are producing systematic reviews for the Cochrane Collaboration and are actively involved in other activities related to the dissemination of evidence based medicine. NATIONAL COLLABORATIONS Agenzia Italiana del Farmaco (AIFA), Roma Istituto Superiore di Sanità, Roma Ministero della Salute, Roma Centro Valutazione Efficacia Assistenza Sanitaria (CeVEAS), Modena Dipartimento di Epidemiologia della ASL Roma E Istituto Neurologico "Carlo Besta", Milano Agenzia Sanitaria Regionale, Regione Emilia Romagna, Bologna Università degli Studi di Milano Università di Modena e Reggio Emilia Azienda Sanitaria Locale 20, Alessandria INTERNATIONAL COLLABORATIONS The Cochrane Collaboration, Oxford, UK Centre for Reviews and Dissemination, University of York, York, UK British Medical Journal Publishing Group, London, UK Centre for Statistics in Medicine, Oxford, UK Thomas Chalmers Centre for Systematic Reviews, Ottawa, Canada The Campbell Collaboration, Philadelphia, USA 229

232 EDITORIAL BOARD MEMBERSHIP Clinical Evidence (Alessandro Liberati) Evidence Based Health Policy and Research (Alessandro Liberati) Journal of Clinical Epidemiology (Alessandro Liberati) Journal of Health Services Research (Alessandro Liberati) PEER REVIEW ACTIVITIES Annals of Internal Medicine (Alessandro Liberati) British Medical Journal (Alessandro Liberati) JAMA (Alessandro Liberati) Evidence Based Health Policy and Research (Alessandro Liberati) Canadian Medical Association Journal (Alessandro Liberati) EVENT ORGANIZATION V Edition Master on Evidence Based Medicine e metodologia della ricerca sanitaria, March April 2008, Università degli Studi di Modena e Reggio Emilia 1-day Cochrane Workshop in Evidence Based Medicine, Milan Focus in: Cardiology 28 September 2007, Ophthalmology 5 October 2007 Gastroenterology 12 October 2007, General Medicine 23 e 26 November 2007 Workshop on 29 November 2007, Milan: - Identifying and publishing uncertainties about the effects of treatments in DUETs - the Database of Uncertainties about the Effects of Treatments - Transparency in the publication and reporting of research. The Equator project and the importance of reporting guidelines - What s' behind the scene of medical journals? - Transparency in the production of guidelines and recommendation: the WHO experience. XII Annual Meeting of the Italian Cochrane Network "La trasparenza nella produzione e disseminazione delle informazioni scientifiche" November 2007, Milan 230

233 PARTICIPATION IN EVENTS IN WHICH THE CENTRE WAS INVOLVED V Edition Master on Evidence Based Medicine e metodologia della ricerca sanitaria, March April 2008, Università degli Studi di Modena e Reggio Emilia Prove di efficacia e pratica terapeutica: EBM e Cochrane Collaboration, Master per Coordinatori di Unità Operativa e/o di Dipartimento per le professioni infermieristiche e ostetriche, Università degli Studi di Milano, anno October Symposium for Centre and Branch Directors, Amsterdam 9-11 April 2007 Workshop on Evidence-based Medicine e revisioni sistematiche, Ischia 8-10 April 2007 Master Universitario di II livello MeS Management e Sanità-II edizione Elementi di epidemiologia per l analisi dei bisogni sanitari e valutazione sanitaria. Metodi quantitativi e elementi di statistica, Pisa 22 November 2007 Meeting Centri Cochrane e Gruppi di Revisione Europei " Cochrane Continental European Entities" June 2007, Oslo, Norvegia XV Cochrane Colloquium, October 2007, San Paolo, Brasile GRANTS AND CONTRACTS Istituto di Ricerche Farmacologiche Mario Negri, Milano AIFA - Agenzia Italiana del Farmaco, Roma 231

234 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Moja L., Moschetti I., Liberati A., Gensini G.F., Gusinu R. Understanding systematic reviews: the meta-analysis graph (also called forest plot ). Intern Emerg Med 2007;2: Moja L., Virgili G., Liberati A., Gensini G.F., Gusinu R., Conti A.A. Scale to climb borderline personalities: when science goes nowhere. Intern Emerg Med 2007;2: Moja L., Minozzi S., Liberati A., Gusinu R., Gensini G.F. The drama of cancer pain: when the research abandons patients and reason. Intern Emerg Med 2007;2: Moja L., Moschetti I., Liberati A., Gensini G.F., Gusinu R. Systematic reviews highlight the complex balance between good and harm from screening studies. Intern Emerg Med 2007;2(1):57-9 Moja L., Moschetti I., Liberati A., Manfrini R., Deligant C., Satolli R., Addis A., Martini N., Dri P. Using Clinical Evidence in a national continuing medical education program in Italy. PLoS Medicine (5) e113.doi: /journal Mosconi P., Colombo C., Satolli R., Liberati A. PartecipaSalute, an Italian project to involve lay people, patients' associations and scientific-medical representatives on the health debate. Health Expect 2007 Jun;10(2): LAY PRESS SELECTION PUBLISHED IN Liberati A, Mosconi P, Colombo C. Che cosa vuol dire Informati bene PartecipaSalute 2007 Mosconi P, Colombo C, Satolli R, Liberati A. La ricerca clinica risponde ai bisogni dei pazienti? Ricerca & Pratica 2007;137: Mosconi P, Colombo C, Satolli R, Liberati A. Il corso di PartecipaSalute 2007 allo specchio: i commenti di chi ha organizzato. PartecipaSalute 2007 Liberati A. Meno ricerca ma di migliore qualità. La dispensa di PartecipaSalute 2007;59-68 Liberati A, Marsico G. Comitati etici: partecipare alla pari. La dispensa di PartecipaSalute 2007;

235 RESEARCH ACTIVITIES Educational and dissemination activities In 2007 the Italian Cochrane Centre (ICC) participated in the organization of fifth year of the Master s program in Evidence Based Medicine (EBM) and health research methodology in collaboration with the University of Modena and Reggio Emilia in Italy. ICC organized and conducted a series of workshops about basic EBM concepts for healthcare decision-making directed toward general practitioners, specialists and doctors in training. These workshops were held at the Mario Negri Institute in Milan. In collaboration with AIFA and Zadig (a scientific publishing group), ICC has made a significant contributions toward the ECCE project (continuing education centred on evidence based medicine). The ECCE project was established in 2005 and promoted by AIFA as a part of a large educational project for continuing medical education (CME) of all Italian doctors. Within the first year of the project, ECCE attracted 17,000 doctors (2005); by the end of 2007 that number rose to AIFA and Zadig have also been partners of the ICC in the translation, adaptation, distribution and evaluation of Clinical Evidence - an international source of the best available evidence for effective healthcare (published by BMJ publishing group). This past year we have been working on the 6 th Italian edition of Clinical Evidence which corresponds to the 17th English edition. In collaboration with PartecipaSalute project, ICC began to translate and disseminate the Cochrane Collaborations' press releases (plain language summaries of Cochrane Systematic Reviews) to scientific journalists, doctors and consumers Research activities ICC researchers have collaborated in the production of 7 Cochrane Systematic Reviews and 7 protocols currently available in the Cochrane Library. ICC researchers are involved in methodological projects focused on the study of the quality of Systematic Reviews. In 2007 ICC has collaborated with PartecipaSalute project. PartecipaSalute aims to involve consumers and their disease-related associations with follow objectives: PartecipaSalute ("Participate in Health Care") is a project initiated at September 2003 to foster a strategic alliance between patients groups and professional societies with the common goal of promoting better health and shared decision-making. The project s main aims are: - to increase patients associations involvement in healthcare assistance and in decision making processes; - to promote a partnership between patients associations and medical societies soliciting medical societies attention to patients need and perspectives - to develop a partnership between citizens, patients and healthcare system. The project developed a website in order to offer critical tools to read and understand medical and healthcare information, for better healthcare decisions. 233

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237 THE CATULLO AND DANIELA BORGOMAINERIO CENTER One of the buildings on the Mario Negri Institute campus is The Catullo and Daniela Borgomainerio Center built in 1987 thanks to a donation from Mrs. Angela Marchegiano Borgomainerio. This is a Center for the study of rare childhood diseases and even today some of the laboratories housed in the building still conduct this research. For example, the study of new therapies used to treat a very rare form of acute myeloid leukemia, know as acute promyelocytic leukemia. A number of new studies are being done to identify new drugs having different mechanisms able to synergize with trans retinoic acid. Research on epidemiological childhood leukemia is also done at the Borgomainerio and a similar line of research involves testicular cancer in adolescents and young adults. Paediatric research activities done at the Borgomainerio Center are also performed in collaboration with groups located at other Institute locations including, The Aldo and Cele Daccò Center for Clinical Research on Rare Diseases at Ranica in Bergamo, the Regional Centre for Drug Information (CRIF) and the Laboratory for Mother and Child Health which are both located in Milan. 235

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239 G.A. PFEIFFER MEMORIAL LIBRARY STAFF Head Librarian Vanna Pistotti 237

240 The Library, specialised in pharmacology and clinical epidemiology, was founded in 1963 thanks to a generous donation from the Gustavus and Louise Pfeiffer Research Foundation, in Denville, New Jersey, USA. Numerous public and private organisations help keep it operative, through donations in money or books, and subscriptions to periodicals. STAFF One Head and two Assistants plus a clerical worker WHAT THE LIBRARY OFFERS The library has a collection of about 5000 textbooks, monographs and congressional proceedings, and 200 periodicals of which a major part are in an electronic format. The books are classified according to the US National Library of Medicine Classification and the Medical Subject headings of Medline (MeSH). Besides the internal collection, the Library has access to the National Periodical Catalogue and to other Library systems (SBBL, GIDIF, RBM). DATABESES AND ELECTRONIC JOURNALS From every computer in the Institute it is now possible to have access to more than 2000 electronic journals and to five of the most important databases: Medline, Cinhal, the Cochrane Library and Embase. SPECIAL PROJECTS The Library cooperates to the realisation of the Italian Information Specialists (GIDIF, RBM) journal catalog which is updated annually. It collaborates to the Institute web site, particularly taking care of the Publications section, both scientific and lay press. TRAINING Every year courses on the use of the database and electronic journals are organised. These courses are designed for use by those working at the Institute but outsiders who are interested may attend. 238

241 2006 PUBLICATIONS Pistotti V, Colombo C Navigare sulla rete alla ricerca della buona informazione Partecipa Salute 2007 Pistotti V, Santoro E Navigare sulla rete alla ricerca di informazioni di salute In ; La dispensa di Partecipasalute, 2007; Pistotti V PubMed: ma è davvero difficile usarlo al meglio? Ricerca & Pratica 2007 n.138 : CONTRACTS Since 1994 the library has been part of the Lombard Biomedical Library System. Sixteen university and research organisation libraries in Lombardy take part in this project, which allows easy, free access to scientific information to over 140 centres and institutions the Lombardy Region. 239

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243 Negri Bergamo Laboratories ANNUAL REPORT 2007 departments and laboratories 241

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245 DEPARTMENT OF MOLECULAR MEDICINE STAFF Head Ariela BENIGNI, Biol.Sci.D., Ph.D. Unit of Gene Therapy Head Susanna TOMASONI, Biol.Sci.D., Ph.D. Laboratory of Cell Biology and Xenotransplantation Head Marina MORIGI, Biol.Sci.D., Ph.D. Unit of Platelet-Endothelial Cell Interaction Head Miriam GALBUSERA, Biol.Sci.D. Laboratory of Immunology and Genetics of Organ Transplantation and Rare Diseases Head Marina NORIS, Chem.Farm.D., Ph.D. Unit of Cellular biology of Autoimmunity and Transplant Rejection Head Sistiana AIELLO, Biol.Sci.D Unit of Genetics of Rare Diseases Head Jessica CAPRIOLI, Biol.Sci.D Unit of Cellular and Molecular Biology of Transplantation Tolerance Head Federica CASIRAGHI, Chemist Laboratory of Experimental Models of Kidney Diseases Head Carla ZOJA, Biol.Sci.D., Ph.D. Unit of Pathology and Immunophatology Head Mauro ABBATE, M.D. 243

246 CURRICULA VITAE Ariela Benigni got the Biol.Sci. degree in 1979 at the University of Milano, Italy, and the Ph.D. at Maastricht University, Netherlands, in Educational training: in 1979 Post Doctoral Fellow, Istituto di Ricerche Farmacologiche Mario Negri (IRFMN), Laboratory of Cancer Chemotherapy, Milan, Italy; in Post Doctoral Fellow, Associazione Bergamasca per lo Studio delle Malattie Renali, Laboratory of the Division of Nephrology and Dialysis, Ospedali Riuniti di Bergamo, Italy; in 1982 Post Doctoral Fellow, Centre Regional de Transfusion Sanguigne de Strasbourg, France. Areas of interest: vasoactive and inflammatory mediators of progressive renal injury with a particular emphasis on endothelin-1; combined treatment of antipertensive and renoprotective drugs to halt progressive renal injury; use of stem cells for tissue regeneration in acute renal failure in vivo e in vitro gene transfer; prevention of acute graft rejection through gene therapy; induction of kidney transplant tolerance by gene therapy; correction of genetic deficiency in rare diseases. Employement: in 1983 Scientist, IRFMN, Laboratory of Kidney Disease, Bergamo, Italy; in Head Laboratory of Prostaglandin and Leukotriene Metabolism, IRFMN, Bergamo, Italy; from January 1991 Scientific Secretary, IRFMN, Bergamo, Italy; in Head Laboratory of Vasoactive and Inflammatory Mediators of Tissue damage, IRFMN, Bergamo, Italy; from January 2000 Head, Department of Molecular Medicine, IRFMN, Bergamo, Italy; from March 2007 Consultant World Health Organization (WHO) for the multicentre observational study Screening for Pre-eclampsia: evaluation of the predictive ability of angiogenic factors for Pre-eclampsia ; from September 2007 Senior Fellow at the University of Oxford, Nuffield Department of Obstetrics & Gynaecology. Selected publications: G. Remuzzi, A. Benigni, A. Remuzzi. Mechanisms of progression and regression of renal lesions of chronic nephropathies and diabetes. J Clin Invest 2006;116: Azzollini, M. Noris, S. Conti, M. Abbate, M. Giacca, G. Remuzzi. Adeno-Associated Virus-mediated CTLA4Ig gene transfer protects MHC-mismatched renal allografts from chronic rejection. J Am Soc Nephrol 2006;17: B. Imberti, M. Morigi, S. Tomasoni, C. Rota, D. Corna, L. Longaretti, D. Rottoli, F. Valsecchi, A. Benigni, J. Wang, M. Abbate, C. Zoja, G. Remuzzi. Insulin-like growth factor-1 sustains stem cell-mediated renal repair. J Am Soc Nephrol 2007;18: C. Zoja, F. Casiraghi, S. Conti, D. Corna, D. Rottoli, R.A. Cavinato, G. Remuzzi, A. Benigni. Cyclin-dependent kinase inhibition limits glomerulonephritis and extends lifespan of mice with systemic lupus. Arthritis & Rheum 2007;56: Benigni, C. Caroli, L. Longaretti, E. Gagliardini, C. Zoja, M. Galbusera, D. Moioli, P. Romagnani, A. Tincani, L. Andreoli, G. Remuzzi. Involvement of renal tubular toll-like receptor 9 in the development of tubulointerstitial injury in systemic lupus. Arthritis & Rheum 2007;56: Marina Morigi got her Biol.Sci. degree in 1987 at the University of Milano, Milano, Italy and the Ph.D. at Maastricht University, Netherlands, in Educational training: in Research training, IRFMN, Bergamo, Italy; in Post Doctoral Fellow, IRFMN, Bergamo, Italy; in 1991 Stage at Brigham and Women s Hospital, Laboratory of Dr. P. Marsden, Boston, USA. Employement: since 1995 Scientist, IRFMN, Bergamo, Italy; in Head, Unit of Renal and Endothelial Cell Biology; since 2000 Head, Laboratory of Cell Biology and Xenotransplantation, IRFMN, Bergamo, Italy. Areas of interest: role of Shigatoxin in the pathogenesis of endothelial dysfunction and microvascular thrombosis in Hemolytic Uremic Syndrome; in vitro model of hyperacute xenograft rejection (porcine endothelium exposed to human serum as a source of xenoreactive natural antibodies and complement); renal toxicity of the proteins filtered through the capillary barrier. In vitro model to study intracellular signals, gene expression and production of inflammatory mediators in cultured proximal tubular cells and glomerular epithelial cells; cell therapy and tissue regeneration: Capability of adult stem cells to differentiate and to regenerate renal tissue in acute and chronic experimental models of renal disease. Murine embryonic stem cell therapy to correct the genetic defect characteristic of Fabry disease in an experimental mouse model. 244

247 Selected publications M. Morigi, B. Imberti, C. Zoja, D. Corna, S. Tomasoni, M. Abbate, D. Rottoli, S. Angioletti, A. Benigni, N. Perico, M. Alison, G. Remuzzi. Mesenchymal Stem Cells Are Renotropic, Helping to Repair the Kidney and Improve Function in Acute Renal Failure. J Am Soc Nephrol 2004;15: A. Remuzzi, S. Mantero, M. Colombo, M. Morigi, E. Binda, D. Camozzi, B. Imberti. Vascular smooth muscle cells on hyaluronic acid: culture and mechanical characterization of an engineered vascular construct. Tissue Eng 2004;10: M. Morigi, S. Buelli, S. Angioletti, C. Zanchi, L. Longaretti, C. Zoja, M. Galbusera, S. Gastoldi, P. Mundel, G. Remuzzi, A. Benigni. In response to protein load podocytes reorganize cytoskeleton and modulate endothelin-1 gene: implication for permselective dysfunction of chronic nephropathies. Am J Pathol 2005;166: Morigi M, Buelli S, Zanchi C, Longaretti L, Macconi D, Benigni A, Moioli D, Remuzzi G, Zoja C. Shigatoxin-induced endothelin-1 expression in cultured podocytes autocrinally mediates actin remodeling. Am J Pathol Dec; 169(6): Imberti B, Morigi M, Tomasoni S, Rota C, Corna D, Longaretti L, Rottoli D, Valsecchi F, Benigni A, Wang J, Abbate M, Zoja C, Remuzzi G. Insulin-like growth factor-1 sustains stem cell mediated renal repair. J Am Soc Nephrol Nov;18(11): Marina Noris got her degree in Pharmaceutical Chemistry and Technologies in 1986 at the University of Rome La Sapienza) and the Ph.D. at Maastricht University, Netherlands, in Educational training: in Fellow, Istituto di Chimica Farmaceutica e Tossicologica, University of Rome, Italy; in Post Doctoral Fellow, Istituto di Chimica Farmaceutica e Tossicologica, University of Rome, Italy; in September 1987-March 1994 Post Doctoral Fellow, IRFMN, Unit of Mediators of Inflammation and Tissue Damage, Laboratory of Kidney Disease, Bergamo, Italy. Areas of interest: immunology of transplantation, tolerance induction; genetics of hemolytic uremic syndrome, thrombotic thrombocytopenic purpura, focal segmental glomerulosclerosis, diabetic nephropathy, role of nitric oxide and arginine dysfunctions in uremia and in pre-eclampsia. Employment: in Head, Unit of Endothelial Cell Pathophysiology, IRFMN, Bergamo, Italy; Head, Laboratory of Cellular and Molecular Biology of the immune response and autoimmunity, IRFMN, Italy; from January 2000: Head, Laboratory of Immunology and Genetics of Rare Diseases and Organ Transplantation, Department of Molecular Medicine, IRFMN, Bergamo, Italy. Selected publications Caprioli J, Noris M, Brioschi S, Pianetti G, Castelletti F, Bettinaglio P, Mele C, Bresin E, Cassis L, Gamba S, Porrati F, Bucchioni S, Monteferrante G, Fang CJ, Liszewski MK, Kavanagh D, Atkinson JP, Remuzzi G. Genetics of HUS: the impact of MCP, CFH and IF mutations on clinical presentation, response to treatment, and outcome. Blood, 2006; 108(4): Noris M, Casiraghi F, Todeschini M, Cravedi P, Cugini D, Monteferrante G, Aiello S, Cassis L, Gotti E, Gaspari F, Cattaneo D, Perico N, Remuzzi G. Regulatory T Cells and T Cell Depletion: Role of Immunosuppressive Drugs. J Am Soc Nephrol. 2007; 18 (3): Bresin E, Daina E, Noris M, Castelletti F, Stefanov R, Hill P, Goodship TH, Remuzzi G; International Registry of Recurrent and Familial HUS/TTP. Outcome of renal transplantation in patients with non-shiga toxin-associated hemolytic uremic syndrome: prognostic significance of genetic background. Clin J Am Soc Nephrol Jan;1(1):88-99 Noris M, Bucchioni s, Galbusera M, Donadelli R, Bresin E, Castelletti F, Caprioli J, Brioschi S, Scheiflinger F, Remuzzi G and the International Registry of Recurrent and Familial HUS/TTP. Complement factor H mutation in familial thrombotic thrombocytopenic purpura with ADAMTS13 deficency and renal involvement. J Am Soc Nephrol 2005; 16: Manuelian T, Hellwage J, Meri S, Caprioli J, Noris M, Heinen S, Jozsi M, Neumann HP, Remuzzi G, Zipfel PF. Mutations in factor H reduce binding affinity to C3b and heparin and surface attachment to endothelial cells in haemolytic uremic syndrome. J Clin Invest 2003; 111: Noris M, Brioschi S, Caprioli J, Todeschini M, Bresin E, Porrati F, Gamba S, Remuzzi G, on behalf of the International Registry of Familial and Recurrent HUS/TTP. Familial haemolytic uraemic syndrome and an MCP mutation. Lancet 2003; 362: Carlamaria Zoja got her Biol.Sci. degree at the University of Milano, Italy, in 1979 and the Ph.D. at the University of Maastricht, The Netherlands in Educational Training: in Post Doctoral Fellow, Associazione Bergamasca per lo studio delle Malattie Renali, Laboratory of the Division of Nephrology and Dialysis, Ospedali Riuniti di Bergamo, Italy; in Post Doctoral Fellow, Center for Thrombosis and Vascular Research, Department of Research Katholieke Universiteit, Leuven, Belgium; in : Post Doctoral Fellow, IRFMN, Laboratory of Kidney Disease, Bergamo, Italy. Areas of interest: experimental models of kidney diseases of immunological and non immunological origin; vasoactive and inflammatory mediators of renal disease progression; role of proteinuria in progressive kidney damage; protection of renal disease progression by a multidrug approach; novel 245

248 immunosuppressive and anti-inflammatory strategies for the treatment of lupus nephritis; role of Shigatoxin in the pathogenesis of endothelial dysfunction in Hemolytic Uremic Syndrome. Employement: since 1985 Scientist, IRFMN, Bergamo, Italy; in : Head, Unit of Experimental Modelling for Human Renal Diseases, Laboratory of Kidney Diseases, IRFMN, Bergamo, Italy; since 1995: Head, Laboratory of Experimental Models of Kidney Diseases, IRFMN, Bergamo, Italy. Selected publications C.Zoja, S. Angioletti, R. Donadelli, C. Zanchi, S. Tomasoni, E. Binda, B. Imberti, M. te Loo, L. Monnens, G.Remuzzi, M. Morigi. Shiga toxin-2 triggers endothelial leukocyte adhesion and transmigration via NF-kB dependent up-regulation of IL-8 and MCP-1. Kidney Int 2002;62: C. Zoja, D. Corna, D. Camozzi, D. Cattaneo, D. Rottoli, C. Batani, C. Zanchi, M. Abbate, G. Remuzzi. How to fully protect the kidney in a severe model of progressive nephropathy: a multidrug approach. J Am Soc Nephrol 2002;13: A. Benigni, C. Zoja, D. Corna, C. Zatelli, S. Conti, M. Campana, E. Gagliardini, D. Rottoli, C. Zanchi, M. Abbate, S. Ledbetter, G. Remuzzi. Add-on anti-tgf-b antibody to ACE inhibitor arrests progressive diabetic nephropathy in the rat. J Am Soc Nephrol 2003;14: R. Donadelli, C. Zanchi, M. Morigi, S. Buelli, C. Batani, S. Tomasoni, D. Corna, D. Rottoli, A. Benigni, M. Abbate, G. Remuzzi, C. Zoja. Protein overload induces fractalkine upregulation in proximal tubular cells through NF-kB and p38 MAPK dependent pathways. J Am Soc Nephrol 2003; 14: C.Zoja, D. Corna, D. Rottoli, C. Zanchi, M. Abbate and G. Remuzzi. Imatinib ameliorates renal diseases and survival in murine lupus autoimmune diseases. Kidney International 2006; 70; C.Zoja, F.Casiraghi, S.Conti, D.Corna, D.Rottoli, R.A.Cavinato, G.Remuzzi, A.Benigni. Cyclin-Dependent kinase inhibition limits glomerulonephritis and extends lifespan of mice with systemic lupus. Arthritis & Rheumatism 2007; 56; Mauro Abbate obtained his M.D. degree in 1988 at the University of Brescia, Italy. Educational training: in Graduate Student, IRFMN, Bergamo, Italy; in Post Doctoral Fellow, IRFMN, Bergamo, Italy; in Research Fellow, The Renal Unit, Massachusetts General Hospital, HMS, Boston, USA. Areas of interest: renal disease progression: the role of proteinuria, complement, and mediators of injury in progressive kidney damage; mechanisms of glomerular injury; anti-gbm glomerulonephritis; mechanisms of tubular injury; kidney fibrosis; the renal biopsy; membranous nephropathy. Employement: in : Scientist, IRFMN, Bergamo, Italy; from 2000 Head, Unit of Renal Pathology and Immunopathology, IRFMN, Bergamo, Italy. Selected publications Abbate M, Zoja C, Rottoli D, Corna D, Tomasoni S, Remuzzi G: Proximal tubular cells promote fibrogenesis by TGFβ1-mediated induction of peritubular myofibroblasts. Kidney International 2002;61, Abbate M, Zoja C, Morigi M, Rottoli D, Angioletti S, Tomasoni S, Zanchi C, Longaretti L, Donadelli R, Remuzzi G: Transforming Growth Factor-β1 Is Up-Regulated by Podocytes in Response to Excess Intraglomerular Passage of Proteins: A Central Pathway in Progressive Glomerulosclerosis. Am J Pathol.2002;161, Ruggenenti P, Chiurchiu C, Brusegan V, Abbate M, Perna A, Filippi C, Remuzzi G: Rituximab for idiopathic membranous nephropathy: a one year prospective study. J Am Soc Nephrol. 2003; 14, Morigi M, Imberti B, Zoja C, Corna D, Tomasoni S, Abbate M, Rottoli D, Angioletti S, Benigni A, Perico N, Alison M, Remuzzi G.:Mesenchymal stem cells are renotropic, helping to repair the kidney and improve function in acute renal failure. J Am Soc Nephrol. 2004;15: Macconi D, Abbate M, Morigi M, Angioletti S, Mister M, Buelli S, Bonomelli M, Mundel P, Endlich K, Remuzzi A, Remuzzi G.: Permselective dysfunction of podocyte-podocyte contact upon angiotensin II unravels the molecular target for renoprotective intervention. Am J Pathol Apr;168(4): Abbate M, Zoja C, Remuzzi G.: How does proteinuria cause progressive renal damage? J Am Soc Nephrol. 2006; 11: Sistiana Aiello got the Biol.Sci. degree in 1993 at the University of Milano, Italy, and the Specialization in Pharmacology Research in 1996, at IRFMN, Bergamo, Italy. Educational training: in research training, IRFMN, Bergamo; in post doctoral fellow, IRFMN, Bergamo. Areas of interest: transplant immunology with a particular interest on dendritic cell biology and mechanisms by which T regulatory cells arise and work; in vitro and in vivo studies on new compounds with immunosuppressive capacity or capable to prevent ischemia/reperfusion tissue injury; vasoactive and inflammatory mediators of progressive renal injury with a particular emphasis on platelet activating factor (PAF) and nitric oxide (NO). Employement: since 2000 Scientist within Laboratory of Immunology and Genetics of Rare disease and 246

249 Organ Transplantation; IRFMN, Bergamo; since 2006 Head, Unit of Cellular Biology of Autoimmunity and Transplant Rejection, IRFMN, Transplant Research Center Chiara Cucchi de Alessandri e Gilberto Crespi, Ranica. Selected publications: S. Aiello, P. Cassis, L. Cassis, S. Tomasoni, A. Benigni, A. Pezzotta, R.A. Cavinato, D. Cugini, N. Azzollini, M. Mister, L. Longaretti, A.W. Thomson, G. Remuzzi, M. Noris. DnIKK2-transfected dendritic cells induce a novel population of inos-expressing CD4 + CD25 - cells with tolerogenic properties. Transplantation 2007; 83: S. Tomasoni, S. Aiello, L. Cassis, M. Noris, L. Longaretti, R.A. Cavinato, N. Azzollini, A. Pezzotta, G. Remuzzi, A. Benigni. Dendritic cells genetically engineered with adenoviral vector encoding dnikk2 induce the formation of potent CD4 + T regulatory cells. Transplantation 2005;79: L. Cassis, S. Aiello, M. Noris. Natural versus adaptive regulatory T cells. Contrib Nephrol 2005; 146: M. Mister, M. Noris, J. Szymczuk, N. Azzollini, S. Aiello, A. Arduini, L. Trochimowicz, E. Gagliardini, M. Abbate, N. Perico, G. Remuzzi. Propionyl-L-carnitine prevents renal function deterioration due to ischemia/reperfusion in isolated perfused rat kidney and in kidney graft. Kidney Int 2002; 61: S. Aiello, M. Noris, G. Piccinini, S. Tomasoni, F. Casiraghi, S. Bonazzola, M. Mister, M.H. Sayegh, G. Remuzzi. Thymic dendritic cells express inducible nitric oxide synthase and generate nitric oxide in response to self- and alloantigens. J Immunol 2000;164: Jessica Caprioli got the Biological Science degree in 1994 at the University of Pavia, Italy, and the Specialization in Medical Genetics in 2001 at the University of Milano, Italy. Educational training: stage at St. John s College in Cambridge for studies at the Department of Genetics, Medical Research Assistant, Lundbeck Italia S.p.A, Training for Biological Scientist Professional Qualification at the Laboratory of cellular and molecular biology of the immune response and autoimmunity, IRFMN, Negri Bergamo Laboratories (qualification obtained in 1997). School of Professional Education in Pharmacological Research of Regione Lombardia (Specialization obtained in 1998). Specialization School in Medical Genetics (University of Milano), obtained in Areas of interest: Role of the complement system in the pathogenesis of hemolytic uremic sindrome and thrombotic thrombocytopenic purpura. Research of the genes involved in the predisposition to focal segmental glomerulosclerosis. Study of the genes that determine the onset of proteinuria in an experimental animal model. Research of the genes involved in the pathogenesis of uric acid nephrolithiasis. Employement: grant recipient at the Laboratory of cellular and molecular biology of the immune response and autoimmunity, IRFMN, Negri Bergamo Laboratories researcher at the Laboratory of immunology and genetics of rare diseases and transplantation, IRFMN, Negri Bergamo Laboratories. Since 2006 head of the Unit of genetics of renal diseases. Selected publications: M. Galbusera, M. Noris, C. Rossi, S. Orisio, J. Caprioli, Z.M. Ruggeri, B. Amadei, P. Ruggenenti, B. Vasile, G. Casari and G. Remuzzi on behalf of the Italian Registry of Familial and Recurrent HUS/TTP. Increased fragmentation of von Willebrand factor, due to abnormal cleavage of the subunit, parallels disease activity in recurrent hemolytic uremic syndrome and thrombotic thrombocytopenic purpura (HUS/TTP) and discloses genetic predisposition in families. Blood, 1999; 94: J. Caprioli, P. Bettinaglio, PF Zipfel, B. Amadei, E. Daina, S. Gamba, C. Skerka, N. Marziliano, G. Remuzzi and M. Noris on behalf of the Italian Registry of Familial and Recurrent HUS/TTP. The molecular basis of familial hemolytic uremic syndrome: mutation analysis of factor H gene reveals a hot spot in short consensus repeat 20. Journal of the American Society of Nephrology, 2001; 12: J. Caprioli, F. Castelletti, S. Bucchioni, P. Bettinaglio, E. Bresin, G. Pianetti, S. Gamba, S. Brioschi, E. Daina, G. Remuzzi and M. Noris. For the International Registry of Recurrent and Familial HUS/TTP. Complement Factor H mutations and gene polymorphisms in hemolytic uraemic syndrome: the C-257T, the A2089G and the G2881T polymorphisms are strongly associated with the disease Human molecular genetics, 2003; 12:1-11. M. Noris, S. Brioschi, J. Caprioli, M. Todeschini, E. Bresin, F. Porrati, S. Gamba and G. RemuzziFor the International Registry of Recurrent and Familial HUS/TTP. Mutation in membrane cofactor protein of the complement system in familial hemolytic uraemic syndrome. Identification of a second disease-associated gene The Lancet, 2003; 362: Caprioli J, Noris M, Brioschi S, Pianetti G, Castelletti F, Bettinaglio P, Mele C, Bresin E, Cassis L, Gamba S, Porrati F, Bucchioni S, Monteferrante G, Fang CJ, Liszewski MK, Kavanagh D, Atkinson JP, Remuzzi G. Genetics of HUS: the impact of MCP, CFH and IF mutations on clinical presentation, response to treatment, and outcome. Blood, 2006; 108(4):

250 Federica Casiraghi has obtained his degree in Industrial Chemistry in 1988, and the degree in Clinical Monitoring and in Biochemical Research in at IRFMN, Bergamo, Italy. Educational Training: research fellow, IRFMN, Bergamo. Areas of interest: Transplant immunology with particular focus on pharmacological and cellular therapies for induction and maintenance of transplantation tolerance. Characterization of regulatory T cells in renal transplant patients and in experimental models of allograft tolerance. Impact of different immunosuppressive drugs on T cell function in renal transplant patients. Vasoactive and inflammatory mediators of progressive renal injury with a particular emphasis on arachidonic acid metabolites. Employment: since 1994 Scientist within Laboratory of Immunology and Genetics of Rare Disease and Organ Transplantation, IRFM, Bergamo; since 2006 Head, Unit of Cellular and Molecolar Biology of Transplantation Tolerance, Transplant Research Center Chiara Cucchi de Alessandri e Gilberto Crespi, Ranica. Selected Publications: Noris M, Casiraghi F, Todeschini M, Cravedi P, Cugini D, Monteferrante G, Aiello S, Cassis L, Gotti E, Gaspari F, Cattaneo D, Perico N, Remuzzi G. Regulatory T Cells and T Cell Depletion: Role of Immunosuppressive Drugs. J Am Soc Nephrol. 2007; 18 (3): Cavinato RA, Casiraghi F, Azzollini N, Cassis P, Cugini D, Mister M, Pezzotta A, Aiello S, Remuzzi G, Noris M. Pretransplant donor peripheral blood mononuclear cells infusion induces transplantation tolerance by generating regulatory T cells. Transplantation, 2005;79(9): Zoja C, Benigni A, Noris M, Corna D, Casiraghi F, Pagnoncelli M, Rottoli D, Abbate M, Remuzzi G. Mycophenolate mofetil combined with a cyclooxygenase-2 inhibitor ameliorates murine lupus nephritis. Kidney Int. 2001; 60(2): Tomasoni S, Noris M, Zappella S, Gotti E, Casiraghi F, Bonazzola S, Benigni A, Remuzzi G. Upregulation of renal and systemic cyclooxygenase-2 in patients with active lupus nephritis. J Am Soc Nephrol.1998; 9(7): Casiraghi F, Ruggenenti P, Noris M, Locatelli G, Perico N, Perna A, Remuzzi G. Sequential monitoring of urinesoluble interleukin 2 receptor and interleukin 6 predicts acute rejection of human renal allografts before clinical or laboratory signs of renal dysfunction. Transplantation 1997; 63(10): Miriam Galbusera got her Biol.Sci. degree in 1981 at the Università degli Studi di Milano. Educational training: in Post Doctoral Fellow, Istituto di Patologia Speciale Medica dell'università degli Studi di Milano, Italy; in Post Doctoral Fellow, IRFMN, Bergamo, Italy; in Post Doctoral Fellow at Scripps Clinic and Research Foundation, Laboratory of Thrombosis and Hemostasis, La Jolla, CA, USA; in Post Doctoral Fellow, IRFMN, Bergamo, Italy. Areas of interest: ADAMTS-13 and VWF in thrombotic microangiopathies, VWF biochemistry, xenotransplantation, platelet-endothelial cell interaction under flow condition, platelet pathophysiology in uremia, receptor studies in kidney and platelets. Employement: : Scientist, IRFMN, Bergamo, Italy; from 2000 Head, Unit of Platelet- Endothelial Cell Interaction, IRFMN, Bergamo, Italy. Selected publications Tripodi, A., Chantarangkul, V., Bohm, M., Budde, U., Dong, J.-F., Friedman, K.D., Galbusera, M., Girma, J.-P., Moake, J., Rick, M.E., Studt, J.-D., Turecek, P.L., Mannucci, P.M.: Measurement of von Willebrand factor cleaving protease (ADAMTS-13): results of an international collaborative study involving 11 methods testing the same set of coded plasmas. J Thromb Haemost 2004; 2: Galbusera, M., Morigi, M., Buelli, S., Gastoldi, S., Macconi, D., Testa, C., Angioletti, S., Remuzzi, G.: Xenogeneic serum-induced thrombus formation on porcine endothelium is mediated by vitronectin receptor and P-selectin: role of reactive oxygen species. Xenotransplantation 2005;12: Ruiz-Torres, M.P., Casiraghi, F., Galbusera, M., Macconi, D., Gastoldi, S., Todeschini, M., Porrati, F., Belotti, D., Pogliani, E.M., Noris, M., Remuzzi, G.: Complement activation: the missing link between ADAMTS13 deficiency and microvascular thrombosis of thrombotic microangiopathies. Thromb Haemost 2005; 93: Noris, M., Bucchioni, S., Galbusera, M., Donadelli, R., Bresin, E., Castelletti, F., Caprioli, J., Brioschi, S., Scheiflinger, F., Remuzzi, G.: Complement factor H mutation in familial thrombotic thrombocytopenic purpura with ADAMTS13 deficiency and renal involvement. J Am Soc Nephrol 2005; 16: Galbusera, M., Bresin, E., Noris, M., Gastoldi, S., Belotti, D., Capoferri, C., Daina, E., Perseghin, P., Scheiflinger, F., Fakhouri, F., Grunfeld, J-P., Pogliani, E., Remuzzi, G.: Rituximab prevents recurrence of thrombotic thrombocytopenic purpura: a case report. Blood 2005;106: Rieger, M., Mannucci, P.M., Kremer Hovinga, J.A., Herzog, A., Gerstenbauer, G., Konetschny, C., Zimmermann, K., Scharrer, I., Peyvandi, F., Galbusera, M., Remuzzi, G., Böhm, M., Plaimauer, B., Lämmle, B., Scheiflinger, F.: ADAMTS13 autoantibodies in patients with thrombotic microangiopathies and other immunomediated diseases. Blood 2005;106:

251 Susanna Tomasoni got her Biological Science degree in 1991 at the University of Milan. Educational training: in Graduate student, University of Milan; in PhD student, University of Milan; in 1994 Research Fellow, Renal Division, Brigham & Women s Hospital, Harvard Medical School, Boston, USA; in 1995 PhD degree in Physiological Science, University of Bologna; : Post Doctoral Fellow, IRFMN, Bergamo, Italy. Areas of interest: construction of adenoviral vectors for gene therapy; gene transfer to the kidney in the context of transplantation; transfection of dendritic cell for cell therapy; progression of renal disease. Employement: in Scientist, IRFMN, Bergamo, Italy; from 2000 Head, Unit of Gene Therapy, IRFMN, Bergamo, Italy Selected publications Tomasoni S, Azzollini N, Casiraghi F, Capogrossi M C, Remuzzi G, Benigni A. CTLA4Ig gene transfer prolongs survival and induces donor-specific tolerance in a rat renal allograft. J Am Soc Nephrol 2000; 11: Tomasoni S, Benigni A. Gene therapy: How to target the kidney. Promises and pitfalls. Curr Gene Ther 2004; 4: Tomasoni S, Longaretti L, Azzollini N, Gagliardini E, Mister M, Buehler T, Remuzzi G, Benigni A. Favorable effect of cotransfection with TGF-beta and CTLA4Ig of the donor kidney on allograft survival. Am J Nephrol 2004; 24: Morigi M, Imberti B, Zoja C, Corna D, Tomasoni S, Abbate M, Rottoli D, Angioletti S, Benigni A, Perico N, Alison M, Remuzzi G. Mesenchymal stem cells are renotropic, helping to repair the kidney and improve function in acute renal failure. J Am Soc Nephrol 2004; 15: Tomasoni S, Aiello S, Cassis L, Noris M, Longaretti L, Cavinato R, Azzollini N, Pezzotta A, Remuzzi G, Benigni A. Dendritic cells genetically engineered with adenoviral vector encoding dnikk2 induce the formation of potent CD4+ T- regulatory cells. Transplantation 2005; 79: Benigni A, Tomasoni S, Turka LA, Longaretti L, Zentilin L, Mister M, Pezzotta A, Azzollini N, Noris M, Conti S, Abbate M, Giacca M, Remuzzi G, Adeno-associated virus-mediated CTLA4Ig gene transfer protects MHC-mismatched renal allografts from chronic rejection. J Am Soc Nephrol, 2006, 17:

252 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The Department of Molecular Medicine was established in 1999 at the Negri Bergamo laboratories to coordinate the work of three laboratories and three units. The activities of the Department of Molecular Medicine are strictly interrelated with those of the Department of Renal Medicine of the Clinical Research Center for Rare Diseases Aldo e Cele Daccò. The following major objectives have been pursued: 1) identification of mediators and mechanisms responsible for the relentless decline of renal function in kidney diseases and development of therapeutic interventions to slow or even halt the disease progression to end-stage renal failure; 2) understanding the mechanisms underlying endothelial cell dysfunction in thrombotic microangiopathies and hyperacute rejection of xenograft 3) finding new strategies for modulating the immune response and preventing acute and chronic rejection of kidney allograft as well as exploration of immunological pathways leading to donor specific unresponsiveness and tolerance of the graft; 4) investigation of the molecular and genetic basis of rare diseases such as hemolytic uremic syndrome/thrombotic thrombocytopenic purpura and pre-eclampsia and search for diseasesusceptibility genes or gene polymorphisms predicting the patient's response to drug therapy in more common and complex polygenic disorders. Such goals have been pursued using various approaches: 1) experimental models of kidney diseases of immunological and non-immunological origin mimicking human renal diseases to study vasoactive and inflammatory mediators and to test novel antiproteinuric and renoprotective drugs; 2) in vitro cultures of renal cells to address the toxicity of protein overload reproducing the condition of exaggerated protein traffic of proteinuric progressive nephropathies; 3) in vitro models to assess the interaction of vascular endothelial cells with leukocytes and platelets under controlled flow conditions; 4) experimental models of kidney allotransplant to study immunological processes responsible for acute and chronic rejection, the nephrotoxicity of immunosuppressor drugs as well as to explore pathways responsible for accomodation; 5) gene transfer of viral constructs carrying genes encoding immunomodulatory molecules to overcome acute rejection of allotransplantation avoiding immunosuppression; 6) identification of candidate genes with linkage analysis and search for mutations as well as assessment of gene polymorphisms. FINDINGS/MAIN RESULTS Insulin growth factor -1 produced by mesenchymal stem cells induces regeneration of proximal tubular cells in experimental acute kidney injury Bone marrow-derived human mesenchymal stem cells cure acute kidney injury and prolong survival in mice Endothelin -1 mediates renal cell damage induced by Shigatoxin-2, the bacterial toxin implicated in the epidemic form of Hemolytic Uremic Syndrome Complement is responsible for trombus formation on microvascular endothelial cells in response to Shigatoxin 250

253 A multidrug approach that simultaneously interrupts pathways of injury induces regression of renal lesions in experimental diabetes DNA contained in circulating immunocomplexes induces renal Toll-like receptor 9 expression responsible for tubulo-interstitial damage in experimental lupus nephritis Thymic- and graft-dependent mechanisms in tolerance induction to kidney allotransplantation in rats have been identified Seliciclib, an inhibitor of cell cycle, reduces lymphocyte alloreactivity and prolongs kidney allograft survival in rats The R1210C mutation of complement factor H, highly prevalent in atypical HUS patients, is also present as a rare polymorphism in different human populations and predisposes to the development of atypical HUS The characterization of mutants in factor I gene has allowed to better understand the atypical HUS pathogenesis in patients carrying this genetic defect Correction of a protein deficiency by a gene therapy approach in genetically modified animals NATIONAL COLLABORATIONS Dipartimento di Scienze Farmacologiche, Università di Milano Dipartimento di Scienze Biomediche e Tecnologia, Università di Milano Laboratorio di Terapia genica e cellulare, G. Lanzani, Divisione di Ematologia, Ospedali Riuniti di Bergamo Laboratorio di Tecnologie riproduttive, Istituto Sperimentale Lazzaro Spallanzani, Cremona Centro Trasfusionale e di Immunologia dei Trapianti, IRCCS Ospedale Maggiore, Milano Dipartimento di Istologia Microbiologia e Biotecnologie Mediche, Università di Padova International Centre for Genetic Engineering and Biotechnology, Molecular Medicine Group, Trieste U.O. di Ostetricia e Ginecologia, Ospedale San Gerardo di Monza U.O. di Ostetricia e Ginecologia, Azienda Ospedaliera Ospedali Riuniti di Bergamo U.O. di Ostetricia e Ginecologia, Azienda Ospedaliera Spedali Civili di Brescia I.R.C.C.S. Policlinico San Matteo, Pavia Azienda Sanitaria Ospedaliera O.I.R.M. - S. Anna, Torino Istituto Gaslini, Laboratorio di Nefrologia, Genova INTERNATIONAL COLLABORATIONS Academisch Ziekenhuis Maastricht, Interne Geneeskunde, Maastricht, Netherland Beth Israel Deaconess Medical Center and Harvard Medical School, Boston, USA Centro de Investigaciones Biologicas and Centro de Investigacion Biomedica en Enfermedades Raras, Madrid, Spain Children's Hospital and Regional Medical Center, University of Washington, Seattle, USA Departements of Pediatrics and Human Genetics, University of Michigan, Ann Arbor, USA 251

254 Deparment of Medicine, Division of Rheumatology, Washington University School of Medicine, St. Louis, USA Erasmus University of Rotterdam, Netherland Flanders Interuniversity Institute for Biotechnology (VIB), University of Lund, Sweden Hans-Knoll Institute for Natural Products Research, Jena, Germania Inselspital, University of Bern, Switzerland INSERM, Paris, France Insitute of Human Genetics, University of Newcastle upon Tyne, UK Max Delbruck Center for Molecular Medicine, Berlin, Germany Molecular Genetics and Rheumatology Section, Division of Medicine, Imperial College, London, UK Monash Medical Center, Melbourne, Australia National Institute of Health, Bethesda, USA Osaka University School of Medicine, Osaka, Japan Pediatric Nephrology and Hypertension, University of Utah, USA Renal Transplant Unit, Hospital de Bellvitge, Barcelona, Spain Rosalind Franklin University of Medicine and Science, Chicago, USA Service d Immunologie Biologique, Hopital Europeen Georges Pompidou, Paris, France The Scripps Research Institute, La Jolla, USA Universitaet Hamburg, Institut fur Molekulare Neuropathobiologie, Hamburg, Germany University of Colorado Cardiovascular Institute, Denver, USA University of Groningen, Netherland University of Iowa, Department of Internal Medicine and Pediatrics, Iowa City, USA University of Liverpool, School of Biological Sciences, UK University of Oxford, UK University of Pittsburgh School of Medicine, Pittsburgh, USA Vanderbilt University School of Medicine, Nashville, USA Weizman Institute of Science, Rehovot, Israel World Health Organization, Geneva, Switzerland EDITORIAL BOARD MEMBERSHIP Journal of American Society of Nephrology (Carla Zoja, Marina Noris) PEER REVIEW ACTIVITIES American Journal of Pathology American Journal of Physiology American Journal of Physiology-Renal Physiology Blood British Journal of Haematology Circulation Diabetes Febs Letters Hypertension Kidney International Journal of American Society of Nephrology 252

255 Journal of Clinical Investigation Journal of Immunology Journal of Molecular Medicine Journal of Nephrology Nature Genetics Nature Medicine Nephrology Nephrology, Dialysis and Transplantation New England Journal of Medicine Pediatric Nephrology Plos PARTICIPATION IN EVENTS IN WHICH THE DEPARTMENT WAS INVOLVED ISN-Nature Genetics Forefronts Symposium on Nephrogenetics: from development to physiology, Danver, MA, (USA), March International Conference on Rare Diseases and Orphan Drugs, Roma, November World Congress of Nephrology, Rio de Janeiro (Brazil), April European Meeting on Complement in human disease, Cardiff (UK), September Workshop on Thrombotic Microangiopathies, Jena (Germany), October VIP meeting, Lovanio (Belgium), December 3, 2007 Congresso della Società Italiana di Pediatria, Salerno, November GENECURE Meetings, Stoccolma (Sweden), September ; Naarden (Netherland), January 26-27, 2007 Meeting Update Meeting on the Relation of Immunity, its evolution and its effect on Solid, Organ Transplantation, Barcelona (Spain), July Meeting Proteinuria in Renal Transplantation pathophysiology, Diagnosis, Treatment, Firenze, March Tenth International Conference on Endothelin (ET-10), Bergamo, September rd International Workshop on Thrombotic Microangiopathies, Jena (Germany), October XIV Telethon Scientific Convention. Salsomaggiore Terme, Italy, March 12-14, Annual meeting of the American Society of Nephrology, San Francisco, October 31-November 5,

256 10 th International Symposium of Nephrology at Montecatini, Pistoia, March 22-24, Congresso SIF, Cagliari, June 6-9, 2007 Corso Monografico Cellule staminali in medicina rigenerativa e oncologia, Collegio Ghisleri, Pavia, 15 February 2007 Annual EuReGene Meeting, Bruxelles, February 1-4, 2007 WCN 2007, Satellite Symposium, Stem Cells and renal diseases: potential use in kidney injury, Potential of mesenchymal stem cells in the repair of tubular injury, Foz do Iguaço, Paranà, Brazil, April 25-27, 2007 GRANTS AND CONTRACTS Associazione Progetto Alice ONLUS Comitato Telethon Fondazione ONLUS Commissione Europea FP6 Fondazione Aiuti per la Ricerca sulle Malattie Rare (ARMR) Fondazione ART per la Ricerca sui Trapianti ONLUS Fondazione Cariplo Fondazione La Nuova Speranza Lotta alla Sclerosi focale ONLUS Fondazione ROTRF/JDRF Istituto Superiore di Sanità ACRAF (Aziende Chimiche Riunite Angelini Francesco Spa) Encysive Farmaceutici Damor Spa Genzyme Corporation Giuliani Spa Novartis Farma Spa Sanofi-Aventis Spa Speedel Pharma Ltd SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 G. Monteferrante, S. Brioschi, J. Caprioli, G. Pianetti, P. Bettinaglio, E. Bresin, G. Remuzzi, M. Noris. Genetic analysis of the complement factor H related 5 gene in haemolytic uraemic syndrome. Mol Immunol 2007;44: S. Heinen, M. Jozsi, A. Hartmann, M. Noris, G. Remuzzi, C. Skerka, P.F. Zipfel. Hemolytic uremic syndrome: a factor H mutation (E1172Stop) causes defective complement control at the surface of endothelial cells. J Am Soc Nephrol 2007;18: M. Noris, F. Casiraghi, M. Todeschini, P. Cravedi, D. Cugini, G. Monteferrante, S. Aiello, L. Cassis, E. Gotti, F. Gaspari, D. Cattaneo, N. Perico, G. Remuzzi. Regulatory T cells and T cell depletion: role of immunosuppressive drugs. J Am Soc Nephrol 2007;18: S. Aiello, P. Cassis, L. Cassis, S. Tomasoni, A. Benigni, A. Pezzotta, R.A. Cavinato, D. Cugini, N. Azzollini, M. Mister, L. Longaretti, A.W. Thomson, G. Remuzzi, M. Noris. DnIKK2-transfected dendritic cells induce a novel 254

257 population of inducible nitric oxide synthase-expressing CD4 + CD25- cells with tolerogenic properties. Transplantation 2007;83: A. Gianella, E. Nobili, M. Abbate, C. Zoja, P. Gelosa, L. Mussoni, S. Bellosta, M. Canavesi, D. Rottoli, U. Guerrini, M. Brioschi, C. Banfi, E. Tremoli, G. Remuzzi, L. Sironi. Rosuvastatin treatment prevents progressive kidney inflammation and fibrosis in stroke-prone rats. Am J Pathol 2007;170: C. Zoja, F. Casiraghi, S. Conti, D. Corna, D. Rottoli, R.A. Cavinato, G. Remuzzi, A. Benigni. Cyclin-dependent kinase inhibition limits glomerulonephritis and extends lifespan of mice with systemic lupus. Arthritis & Rheum 2007;56: A. Benigni, C. Caroli, L. Longaretti, E. Gagliardini, C. Zoja, M. Galbusera, D. Moioli, P. Romagnani, A. Tincani, L. Andreoli, G. Remuzzi. Involvement of renal tubular toll-like receptor 9 in the development of tubulointerstitial injury in systemic lupus. Arthritis & Rheum 2007;56: N. Perico, M. Abbate, G. Remuzzi. More on renal disease progression: is interstitial inflammation truly protective? (Editorial) J Am Soc Nephrol 2007;18: P. Cravedi, M. Noris, G. Remuzzi. Report of the first World Transplant Congress. (Special Feature) Clin J Am Soc Nephrol 2007;2: D. Corna, F. Sangalli, D. Cattaneo, F. Carrara, F. Gaspari, A. Remuzzi, C. Zoja, A. Benigni, N. Perico, G. Remuzzi. Effects of rosuvastatin on glomerular capillary size-selectivity function in rats with renal mass ablation. Am J Nephrol 2007;27: R.A. Cavinato, F. Casiraghi, N. Azzollini, M. Mister, A. Pezzotta, P. Cassis, D. Cugini, N. Perico, G. Remuzzi, M. Noris. Role of thymic- and graft-dependent mechanisms in tolerance induction to rat kidney transplant by donor PBMC infusion. Kidney Int 2007;71: D. Kavanagh, A. Richards, V. Fremeaux-Bacchi, M. Noris, T. Goodship, G. Remuzzi, J.P. Atkinson. Screening for complement system abnormalities in patients with atypical hemolytic uremic syndrome. Clin J Am Soc Nephrol 2007;2: J. Caprioli, G. Remuzzi. Complement hyperactivation may cause atypical haemolytic uraemic syndrome-gain-offunction mutations in factor B. (Editorial) Nephrol Dial Transplant 2007;22: P. Ruggenenti, N. Perico, E. Gotti, P. Cravedi, V. D Agati, E. Gagliardini, M. Abbate, F. Gaspari, D. Cattaneo, M. Noris, F. Casiraghi, M. Todeschini, D. Cugini, S. Conti, G. Remuzzi. Sirolimus versus cyclosporine therapy increases circulating regulatory T cells, but does not protect renal tranplant patients given alemtuzumbab induction from chronic allograft injury. Transplantation 2007;84: B. Imberti, M. Morigi, S. Tomasoni, C. Rota, D. Corna, L. Longaretti, D. Rottoli, F. Valsecchi, A. Benigni, J. Wang, M. Abbate, C. Zoja, G. Remuzzi. Insulin-like growth factor-1 sustains stem cell-mediated renal repair. J Am Soc Nephrol 2007;18: E. Gagliardini, A. Benigni. Therapeutic potential of TGF-β inhibition in chronic renal failure. Expert Opin Biol Ther 2007;7:

258 RESEARCH ACTIVITIES Laboratory of Cell Biology and Xenotransplantation Insulin-like growth factor-1 sustains stem cell-mediated renal recovery following an acute injury In collaboration with the Laboratory of Experimental Models of Kidney Diseases and with the Unit of Gene Therapy Bone marrow-derived mesenchymal stem cells (MSC) can contribute to renal remodeling. In mice with acute renal failure (ARF) induced by cisplatin, MSC administration restored renal tubular structure and improved renal function, but the underlying mechanism was unclear. We examined the process of kidney cell repair in a setting of MSC co-cultured with cisplatininjured proximal tubular cells (PTEC). Exposure of PTEC to cisplatin led to a marked reduction in cell viability at 4 days. MSC protected PTEC from cisplatin-induced damage by promoting tubular cell proliferation, as assessed by BrdU labeling that marks cells during mitosis, and PKH26 red fluorescence cell linker. This effect was mediated by insulin-like growth factor-1 (IGF-1), a mitogenic and anti-apoptotic factor, highly expressed by MSC as mrna and protein, because functional blocking of the growth factor by specific antibody blocked cell proliferation. At variance, TGF- or IL-10 were not involved in cell mitosis. Knocking down IGF-1 expression in MSC by small interfering(si)-rna resulted in a significant decrease in PTEC proliferation and increased apoptosis. Consistently, in the murine model of cisplatin-induced ARF infusion of MSC transfected with sirna -IGF-1 limited their protective effect on renal function and tubular structure. These findings indicate that MSC exert beneficial effects on tubular cell repair in ARF by virtue of their capability of producing the mitogenic and prosurvival factor IGF-1. Life-saving effect of human bone marrow-mesenchymal stromal cells in experimental acute renal injury In collaboration with the Laboratory of Experimental Models of Kidney Diseases and with the Unit of Gene Therapy Transplantation of bone marrow-mesenchymal stromal/stem cells (BM-MSC) from rodents has been identified as a strategy for renal repair in experimental models of acute kidney injury (AKI), a highly life-threatening clinical setting. The therapeutic potential of BM-MSC of human origin has not been evaluated so far. The aim of the present study was to test whether human BM-MSC treatment could prevent AKI induced by the nephrotoxic anticancer drug cisplatin, and prolong survival in an immunodeficient mouse model. Two groups of NOD-SCID mice were subcutaneously injected with cisplatin (12.7 mg/kg). Twenty-four hours later, the first group was intravenously injected with saline, the second group with 5x10 5 BM-MSC. Results showed that human BM-MSC infusion decreased proximal tubular epithelial cell injury and ameliorated renal function resulting in a reduced recipient mortality. Here, finding PKH-26 fluorescent human BM-MSC in the proximity of tubular profiles and, less frequently, within the context of tubular epithelium supports the concept that MSC contributed locally to tubular regeneration via a paracrine mechanisms. Infused BM-MSC acted to significantly reduce renal apoptotic cells and to increase peritubular capillary diameter and both peritubular endothelial and lumen volume density, as documented by morphometric analysis. These findings indicate that infusion of human MSC of bone marrow origin by preserving the integrity of the tubular epithelium and peritubular microvessels prolongs survival in AKI mice. Human BM-MSC represent an ideal cell population for future cell therapy and hold potential for successful application in human AKI. 256

259 Shigatoxin-2 reduces nephrin gene expression via endothelin (ET)-1/ETA receptor mechanism in cultured glomerular podocytes In collaboration with the Laboratory of Experimental Models of Kidney Diseases and with the Unit of Gene Therapy Shigatoxin (Stx) is the offending agent of post-diarrheal Hemolytic Uremic Syndrome (HUS), a disease characterized by microvascular thrombosis and glomerular ischemic changes that contribute to acute kidney injury. We have previously documented that glomerular podocytes are a functionally relevant target of Stx-2 that, via up-regulation of ET-1 gene and protein, promotes cytoskeletal changes and glomerular permeability dysfunction in an autocrine manner. Here, we studied whether Stx-2 altered in podocytes the expression of nephrin, key component of the slit diaphragm, through an ET-1-dependent pathway. Exposure of murine differentiated podocytes to Stx-2 50 pm and 1 nm for 24 hours significantly reduced nephrin mrna levels in a dose-dependent manner in respect to control cells (Stx-2 50pM: 0.6±0.04, Stx-2 1nM: 0.3±0.06-fold vs control:1). Blockade of ETA receptor with LU prevented Stx-2- dependent decline of nephrin expression, indicating the involvement of ET-1 via ETA receptor (Stx-2 50pM+LU: 0.9±0.1, Stx-2 1nM+LU: 0.9±0.1-fold vs control: 1). Consistent with Stx-2 effect, ET-1 (100nM) added to podocytes decreased by 58% nephrin mrna levels. Inhibition of Rho kinase -crucial for the formation of cytoskeletal stress fibers- partially recovered nephrin mrna levels in respect to Stx-2-treated cells. Injection of Stx-2 in mice caused podocyte changes including focal foot process effacement and increased expression of ET-1 associated with 30% reduction of nephrin protein expression in respect to control mice. In summary, our data show that ET-1 produced by podocytes in response to Stx, is an important mediator of Stxinduced reduction of nephrin expression, and might play a pivotal role in dysfunction of the filtration barrier in HUS. Shiga toxin induces complement deposition on human microvascular endothelial cells. Shiga toxin (Stx) produced by E.coli is the main cause of post-diarrheal HUS (D+ HUS), a disorder of microangiopathic hemolytic anemia, thrombocytopenia and acute renal failure that mainly affects children. We previously documented that Stx directly affected endothelial antithrombogenic properties promoting thrombus formation on human microvascular endothelial cells at high shear stress via upregulation of beta3 integrin and P-selectin. Evidence are available showing that P-selectin expressed on activated cells interacts with complement C3 component. In this study we investigated whether local activation of endothelial cells by Stx might promote complement deposition through P-selectin dependent pathway. Our results showed a marked increase of C3 deposits on cultured human microvascular endothelial cells (HMEC-1) incubated with Stx1 for 24 h and than perfused at high shear stress (60 dynes/cm 2 ) with human serum as a source of complement. Both Stx1 and Stx2 at the concentration of 50 pm induced a strong granular C3 deposits uniformely distributed on endothelial surface. To exclude any possible effect of lipolysaccharide contamination of Stx preparation, the effect of purified Stx1 (Stx1p) on C3 deposits was tested. Results showed that Stx1p induced similar increase in C3 deposits as compared to not purified Stx1. Since Stx induced the upregulation on endothelial surface of P-selectin, an adhesive protein that may directly interact with C3, we studied the role of this ligand in mediating endothelial C3 deposition in response to Stx. Functional blocking of P-selectin with a specific antibody markedly reduced C3 deposits induced by Stx on endothelial surface, indicating that Stx1 induced C3 deposits via P-selectin expression on endothelial cells. C3 deposits induced by Stx on HMEC-1 was completely abolished by treating human serum with the soluble complement receptor 1 (scr- 1), an inhibitor of both the classical and the alternative pathway of complement. Treatment with EGTA, an inhibitor of the classical pathway, did not prevent C3 deposition indicating that C 257

260 activation induced by Stx occurred via the alternative pathway. Further studies will be performed to establish whether there is a causative link between the activation of the alternative complement pathway and microvascular thrombosis in D+ HUS. Laboratory of Experimental Models of Kidney Diseases Involvement of renal tubular Toll-like receptor 9 in the development of tubulointerstitial injury in systemic lupus In collaboration with the Unit of Gene Therapy Toll-like receptor 9 (TLR-9), a receptor for CpG DNA, has been implicated in the activation of immune cells in lupus. We undertook this study to determine whether the expression of TLR-9 in resident renal cells in lupus nephritis is related to the development of tubulointerstitial injury. TLR-9 was analyzed in selectively retrieved renal tissue from (NZB/W) F1 mice at different stages of the disease by laser capture microdissection combined with real-time quantitative reverse transcriptase-polymerase chain reaction, and in renal biopsy specimens from lupus nephritis patients by immunohistochemistry. We investigated for the molecular component responsible for TLR-9 activation by cultured proximal tubular cells in serum from patients with lupus. Renal tissue from NZB/W mice displayed robust TLR-9 expression localized to proximal tubular cells. TLR-9 levels correlated with proteinuria and tubulo-interstitial injury to the extent that seliciclib, a cyclin-dependent kinase inhibitor, while reducing proteinuria and renal structural damage, prevented tubular TLR-9 generation in lupus mice. Consistently, exaggerated TLR-9 staining was found in proximal tubular cells of lupus patients, which correlated with tubulo-interstitial damage. DNA-containing immune complexes purified from sera of patients with lupus induced TLR-9 in cultured proximal tubular cells. This was prevented by CCGG-rich short oligonucleotides, specific antagonists of CpG DNA, indicating that the DNA component of immune complexes was required for TLR9 stimulation. These findings suggest that tubular TLR-9 activation has a pathogenetic role in tubulointerstitial inflammation and damage in experimental and human lupus nephritis, and indicate a novel target for future therapies. Regression of podocyte lesions by a multidrug approach in experimental diabetes The degree of renoprotection afforded by renin-angiotensin system (RAS) inhibitors in progressive nephropathies crucially depends on the time at which therapy is started, with low effectiveness provided by late treatment. Here, we used a rat model of diabetic nephropathy to assess the effect of maximal RAS inhibition by angiotensin converting enzyme inhibitor (ACEi) plus angiotensin II receptor antagonist (ATIIra) combined with statin that displays pleiotropic properties complementing cholesterol lowering effects. Diabetes was induced in uninephrectomized rats by streptozotocin. Animals were orally treated from 4 to 8 months after disease induction with vehicle, lisinopril plus candesartan, lisinopril+candesartan+rosuvastatin or rosuvastatin. Six healthy rats served as control. Blood pressure increased during time in diabetic rats and was significantly reduced by treatment with RAS inhibitors and triple therapy. ACEi+ATIIra significantly reduced proteinuria in respect to vehicle; remission of proteinuria to control levels was achieved by the addition of rosuvastatin. Glomerulosclerosis was ameliorated by ACEi+ATIIra, but totally prevented by triple therapy. Lower podocyte number per glomerulus in diabetic rats on vehicle was partially restored by ACEi+ATIIra while normalized by combining the three drugs. Staining with nephrin, key functional component of the slit diaphragm, was significantly decreased in diabetic rats as compared to controls and normalized by triple therapy. 258

261 This multidrug approach might represent a strategy to induce remission of proteinuria and regression of renal lesions in diabetic patients who do not fully benefit RAS inhibition. Laboratory of Immunology and Genetic of Rare Diseases and Organ Transplantation Role of thymic- and graft-dependent mechanisms in tolerance induction to rat kidney transplant by donor PBMC infusion We previously demonstrated the presence of regulatory T cells (Treg) in lymph nodes (LN) from rats made tolerant to a kidney allograft by donor PBMC infusion. Here we investigated the origin of Treg and characterized their phenotype and mechanisms underlying their suppressive effect. At different points after PBMC infusion, thymus, LN and graft-infiltratinglymphocytes (GIL) alloreactivity was evaluated in MLR, coculture and transwell experiments. GIL phenotype (by FACS and immunohistochemistry) and cytokines mrna expression were analyzed. Before transplantation, CD4 + thymocytes and LN cells from donor PBMC-infused rats showed a reduced anti-donor but a normal anti-third-party proliferation. Anti-donor hyporesponsiveness was reverted by IL-2. CD4 + thymocytes had no regulatory activity on a naive MLR. Treg appeared in LN at 60 days post-transplant. CD4 + -GIL isolated early (5 days) and late post-transplant (days 60-80) were hyporesponsive and suppressed a naïve MLR. IL-10 mrna was up-regulated in GIL and an anti-il-10 mab reverted their inhibitory effect. Cellto-cell contact potentiated the suppressive activity of CD4 + -GIL. We suppose that allograft tolerance in this model is mediated by pre-transplant generation of anergic cells in the thymus, which may have a permissive role to prevent early graft disruption. The healed graft is a source of donor antigens which lead to early selection of Treg. In the late phase tolerance is maintained by appearance of Treg in LN. Effect of seliciclib (CYC202, R-roscovitine) on lymphocyte alloreactivity and acute kidney allograft rejection in rat T cell stimulation by alloantigens is followed by cell cycle progression, an event that is critically dependent on cyclin-dependent kinases (CDKs). We conducted a study to evaluate whether the CDK inhibitor seliciclib affected rat lymph node cell (LNc) activation and proliferation induced by either concanavalin A or allogeneic splenocytes in vitro and studied the mechanisms underlying the suppressive effect. We also investigated the immunosuppressive properties of seliciclib in vivo. Seliciclib completely inhibited in vitro proliferation of LNc and CD8 + Tcells, in response to either concanavalin A or allogeneic splenocytes. The percentage of activated LNc was lower in MLR added with seliciclib than in MLR added with vehicle. The percentages of viable and apoptotic cells at the end of MLR with seliciclib were comparable to those of MLR with vehicle. LNc pre-exposed in MLR to seliciclib did not respond to further stimulation with alloantigens, and neither IL-2 nor IL-15 restored proliferation. These data indicate that the inhibitory effect of seliciclib on T cell alloreactivity is not due to cytotoxic effect but is associated with induction of profound T cell anergy. LNc harvested at the end of the primary MLR with seliciclib did not suppress the proliferation of syngeneic LNc cells toward allogeneic splenocytes, thus excluding that seliciclib induced the formation of regulatory cells. Finally, seliciclib partially prolonged grafted animal survival in a rat model of fully MHC-mismatched kidney transplantation. Altogether these results document that seliciclib regulates lymphocyte reactivity and may exert an immunosuppressive effect in vivo in the setting of transplantation. 259

262 Characterisation of mutations in complement Factor I (CFI) associated with Hemolytic Uremic Syndrome The hemolytic uraemic syndrome (HUS) is characterized by the triad of thrombocytopenia, microangiopathic haemolytic anaemia and acute renal failure. Cases not associated with a preceding Shiga-like toxin producing E.coli are described as atypical HUS (ahus). Recent studies have identified mutations in the complement regulatory proteins factor H (CFH), membrane cofactor protein (MCP) and factor-i (CFI) that predispose to ahus. CFI is a two chain serine protease in which the light chain carries the catalytic domain while the heavy chain s (55 kda) function is unclear. It downregulates the alternative and classical complement pathways by cleaving the α chains of C3b and C4b in the presence of cofactors (known as cofactor activity). Many CFI mutations in ahus result in low CFI levels with a consequent quantitative defect in complement regulation. In others, the mutant protein is present in normal amounts but the presumed functional deficiency has not yet been defined. In this report we examine the nature of the functional defect in HUS-associated CFI mutations. The D501N and D506V mutations reside in the serine protease domain of CFI and result in secreted proteins which lack C3b and C4b cofactor activity. The delttcac ( ) mutant, also in the serine protease domain, leads to a protein which is not secreted. The R299W mutant lies in a region of the CFI heavy chain of no known function. Our assessments demonstrate decreased C3b and C4b cofactor activity, providing evidence that this region is important for cofactor activity. In two other heavy chain mutants, no function deficiency was found. These defective mutant proteins will result in an inability to appropriately control the complement cascade at sites of endothelial cell injury. The excessive complement activation for a given degree of damage may result in generation of a pro-coagulant state and ahus. R1210C, a prevalent mutation in complement factor h associated with atypical Haemolytic Uremic Syndrome Mutations in the gene encoding complement factor H that alter the C3b/polyanions-binding site at the C-terminal region of the protein impair the capacity of factor H to protect host cells and are strongly associated with atypical haemolytic uremic syndrome (ahus). Most of these ahus-associated CFH mutations are unique mutations normally found in only a single patient or family. One exception is the CFH mutation that has been found in several R1210C unrelated ahus patients from distinct geographical origins. Here, 5 ahus pedigrees and 7 individual ahus patients (represented by 13 affected and 16 healthy CFH R1210C mutation carriers) were analyzed to identify potential correlations of the CFH R1210C mutation with particular clinical phenotypes and to characterize the origins of this mutation. Our results showed that the clinical phenotype of ahus patients carrying the CFH R1210C mutation was heterogeneous. Interestingly, 12 out of 13 patients carry at least one additional genetic factor conferring predisposition to ahus. These data are in accord with the low penetrance of ahus (30%) in the CFH R1210C mutation carriers, indicating that the presence of other genetic or environmental risk factors significantly contribute to the manifestation and severity of ahus in CFH R1210C mutation carriers. Genotype analysis of CFH and CFHR3 SNPs in the 12 unrelated carriers suggests that the CFH R1210C mutation has a single origin. In conclusion, CFH R1210C is a prevalent and prototypic ahus mutation that may be present, as a rare polymorphism, in many human populations. 260

263 Unit of Gene Therapy Gene therapy to correct Thrombotic Thrombocytopenic Purpura Thrombotic Thrombocitopoenic Purpura (TTP) is a rare disease with an incidence of 2: per year. TTP is a thrombotic microangiopathy, characterized by anemia and thrombocytopenia caused respectively by red blood cells fragmentation in the microcirculation and by microvascular thrombi formation. TTP patients show prevalent, but not exclusive, neurological symptoms and renal dysfunction. They have plasma accumulation of von Willebrand multimers with a molecular weight higher than normal due to the lack of activity, acquired or familiar, of ADAMTS13, a plasma protein that cleaves vwf secreted by endothelial cells. vwf ultralarge multimers are more adhesive for platelets than the normal counterpart, causing their deposition and thrombi formation. TTP treatment is based on plasma exchange. Although this therapy has increased considerably the survival (from 10% to 80-90%) it may expose patients to the risk of infections or major complications such as hemorrhage due to the insertion of the central venous catheter, venous thrombosis and hypotension equiring dopamine. In patients with TTP, recurrencies of the disease are common and relapsing TTP is a debilitating and life-threatening disease. For these reasons researchers are looking for alternative therapies. Gene therapy could be useful since ADAMTS13 activity levels of 5-6% are protective. In order to transfer genes we took advantage of viruses unable to replicate and to induce disease but still able to infect cells. We constructed two different viral vectors, adenoviral (Ad) and adeno-associated (AAV), containing ADAMTS13 gene. RT-PCR and Western Blot analysis showed that they infect cells efficiently and induce ADAMTS13 mrna and protein expression. Adenoviral vector induces the secretion of the functional active protein as revealed by a Collagen Binding Assay. Recently ADAMTS13 knock out mice were generated. These mice are healthy but, if crossed with mice expressing high levels of vwf and exposed to Shiga-toxin produced by E.Coli, show a syndrome similar to TTP. Studies are now in progress in order to establish the best vector able to induce in vivo expression of appreciable levels of ADAMTS13 (5-6% at least) functionally active and to correct the disease. 261

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265 DEPARTMENT OF BIOMEDICAL ENGINEERING STAFF Head Andrea REMUZZI, Eng. D. Laboratory of Renal Biophysics Head Daniela MACCONI, Biol.Sci.D. Laboratory of Biomedical Technologies Head Bogdan ENE-IORDACHE, Eng.D. Unit of Tissue Engineering Head Marina FIGLIUZZI, Biol.Sci.D. Unit of Medical Imaging Head Luca ANTIGA, Ph.D. 263

266 CURRICULA VITAE Andrea Remuzzi got his degree in Mechanical (Biomedical) Engineering in 1979, Politecnico di Milano. Research experience: 1980 Politecnico di Milano, Dipartimento di Ingegneria Biomedica; 1981 Istituto Mario Negri (Milano), Laboratorio di Farmacologia Cardiovascolare; Massachusetts Institute of Technology, Mechanical Engineering Department, Cambridge, USA. Areas of interest: biological transport phenomena, mathematical models, renal pathophysiology, cellular response to mechanical stimulation, tissue engineering, pancreatic islet transplantation, clinical databases, computational fluid dynamics. Chronology of appointment: From 1984 to 1986 Ricercatore Istituto Mario Negri (Bergamo), Laboratorio di malattie renali, Head, Unità di Bioingegneria, Istituto Mario Negri, Head, Laboratorio di Bioingegneria, Istituto Mario Negri, Head, Dipartimento di Ricerca Renale, Istituto Mario Negri, from 2000 Head, Dipartimento di Bioingegneria, Istituto Mario Negri. Since 1998 contract professor of Bioengineering, Politecnico di Milano. Selected publications Davies PF, Remuzzi A, Gordon EJ, Dewey CF Jr, Gimbrone MA Jr. Turbulent fluid shear stress induces vascular endothelial cell turnover in vitro. Proc Natl Acad Sci U S A Apr;83(7): PMID: Remuzzi A, Puntorieri S, Battaglia C, Bertani T, Remuzzi G. Angiotensin converting enzyme inhibition ameliorates glomerular filtration of macromolecules and water and lessens glomerular injury in the rat. J Clin Invest Feb;85(2): PMID: Morigi M, Micheletti G, Figliuzzi M, Imberti B, Karmali MA, Remuzzi A, Remuzzi G, Zoja C. Verotoxin-1 promotes leukocyte adhesion to cultured endothelial cells under physiologic flow conditions. Blood Dec 15;86(12): PMID: Noris M, Morigi M, Donadelli R, Aiello S, Foppolo M, Todeschini M, Orisio S, Remuzzi G, Remuzzi A. Nitric oxide synthesis by cultured endothelial cells is modulated by flow conditions. Circ Res Apr;76(4): PMID: Giavazzi R, Foppolo M, Dossi R, Remuzzi A. Rolling and adhesion of human tumor cells on vascular endothelium under physiological flow conditions. J Clin Invest Dec;92(6): PMID: Antiga L, Ene-Iordache B, Remuzzi A. Computational geometry for patient-specific reconstruction and meshing of blood vessels from MR and CT angiography. IEEE Trans Med Imaging May;22(5): PMID: Remuzzi A, Gagliardini E, Sangalli F, Bonomelli M, Piccinelli M, Benigni A, Remuzzi G. ACE inhibition reduces glomerulosclerosis and regenerates glomerular tissue in a model of progressive renal disease. Kidney Int Jan 4; PMID: Daniela Macconi got her Biol.Sci.D. degree in Milan in the Research experience: CNR Institute of Neuroscience - Cell Mol Pharmacology - and Department of Medical Pharmacology, University of Milan, Milan, Italy; Laboratory of the Division of Nephrology and Dialysis, Ospedali Riuniti di Bergamo, Bergamo, Italy; University of Michigan, Medical School, Department of Pathology, Medical Science I, Ann Arbor Michigan, USA; Mario Negri Institute for Pharmacological Research, Laboratory of Kidney Disease, Bergamo, Italy. Areas of interest: glomerular permeability, renal disease progression, podocytes, angiotensin II, reactive oxygen species Chronology of appointment: From 2000 Head Laboratory of Renal Biophisics, Department of Biomedical Engineering; Head, Unit of Inflammatory Mediator of Leucocyte Origin; Scientist, post-doctoral fellow Mario Negri Institute for Pharmacological Research, Bergamo, Italy; fellow Laboratory of the Division of Nefrology e Dialysis, Ospedali Riuniti di Bergamo, Bergamo, Italy Selected publications Macconi D, Abbate M, Morigi M, Angioletti S, Mister M, Buelli S, Bonomelli M, Mundel P, Endlich K, Remuzzi A, Remuzzi G: Permselective dysfunction of podocyte-podocyte contact upon angiotensin II unravels the molecular target for renoprotective intervention. Am J Pathol.168: , Macconi D, Bonomelli M, Benigni A, Plati T, Sangalli F, Longaretti L, Conti S, Kawachi H, Hill P, Remuzzi G, Remuzzi A. Pathophysiologic implications of reduced podocyte number in a rat model of progressive glomerular injury. Am J Pathol.168:42-54, Ruiz-Torres MP, Casiraghi F, Galbusera M, Macconi D, Gastoldi S, Todeschini M, Porrati F, Belotti D, Pogliani EM, Noris M, Remuzzi G: Complement activation: the missing link between ADAMTS-13 deficiency and microvascular thrombosis of thrombotic microangiopathies. Thromb Haemost. 93:443-52, Galbusera M, Buelli S, Gastoldi S, Macconi D, Angioletti S, Testa C, Remuzzi G, Morigi M: Activation of porcine endothelium in response to xenogeneic serum causes thrombosis independently of platelet activation. Xenotransplantation. 12:110-20,

267 Morigi M, Macconi D, Zoja C, Donadelli R, Buelli S, Zanchi C, Ghilardi M, Remuzzi G: Protein overload-induced NFkappaB activation in proximal tubular cells requires H(2)O(2) through a PKC-dependent pathway. J Am Soc Nephrol. 13: , 2002 Macconi D, Ghilardi M, Bonassi ME, Mohamed EI, Abbate M, Colombi F, Remuzzi G, Remuzzi A: Effect of angiotensinconverting enzyme inhibition on glomerular basement membrane permeability and distribution of zonula occludens-1 in MWF rats. J Am Soc Nephrol 11:477-89, 2000 Bogdan Ene-Iordache got MSc in Mechanical Engineering in 1990 at the Oil & Gas Institute in Ploiesti (Romania). In 1992 he joined the Bioengineering Laboratory at NegriBERGAMO Laboratories. Main interests: renal research (hemodynamics and remodeling of arteriovenous fistula for vascular access, morfometrical analysis of glomerular capillaries) and controlled clinical trials (data management and data analysis for controlled clinical studies); coordination of IT activities and web sites in the Clinical Research Center for Rare Diseases Aldo e Cele Daccò ; applied clinical informatics (Nephrology, Diabetology and Hematology Units, Bergamo Hospital). Roles: since January 2000 is head of the Biomedical Technologies Laboratory, Department of Biomedical Engineering. Selected publications Ene-Iordache B, Imberti O, Foglieni O, Remuzzi G, Bertani T and Remuzzi A. Effects of angiotensin-converting enzyme inhibition on glomerular capillary wall ultrastructure in MWF/Ztm rats. J Am Soc Nephrol 5: , Ene-Iordache B and Remuzzi A. Numerical analysis of blood flow in reconstructed glomerular capillary segments. Microvasc Res 49: 1-11, Remuzzi A and Ene-Iordache B. Capillary network structure does not affect theoretical analysis of glomerular size selectivity. Am J Physiol 268: F972-F979, Ene-Iordache B, Mosconi L, Remuzzi G, Remuzzi A. Computational fluid dynamics of a vascular access case for hemodialysis. J Biomech Eng 123(3): , Ene-Iordache B, Mosconi L, Antiga L, Bruno S, Anghileri A, Remuzzi G, Remuzzi A. Radial artery remodeling in response to shear stress increase within arteriovenous fistula for hemodialysis access. Endothelium 10(2): , Ruggenenti P, Fassi A, Ilieva AP, Bruno S, Iliev IP, Brusegan V, Rubis N, Gherardi G, Arnoldi F, Ganeva M, Ene-Iordache B, Gaspari F, Perna A, Bossi A, Trevisan R, Dodesini AR, Remuzzi G for the Bergamo Nephrologic Diabetes Complications Trial (BENEDICT) Investigators. Preventing microalbuminuria in type 2 diabetes. NEJM 351(19): , Marina Figliuzzi got her Biol.Sci.D. degree in Milan in the Research experience : Mario Negri Institute for Pharmacological Research, Bergamo, Italy. Areas of interest: isolation of pancreatic islets from human, bovine, pig and rat pancreas, cell culture, immunoisolation devices for pancreatic islets, differentiation of progenitor pancreatic cells in insulin containing cells, immunhistochemistry. Chronology of appointment: From 2000 Head Unit of Tissue Engineering, Department of Biomedical Engineering; fellow laboratory of Renal research, Mario Negri Institute for Pharmacological Research, Bergamo, Italy. Selected publications Figliuzzi M, Cornolti R, plati T, Rajan N, Adobati F, Remuzzi G, Remuzzi A: Subcutaneous xenotransplantation of bovine pancreatic islets. Biomaterials. 26: , Figliuzzi M, Zappella S, morigi M, Rossi P, marchetti P, Remuzzi A: Influence of donor age on bovine pancreatic islet isolation. Transplantation. 70: , 2000 Morigi M, Micheletti G, Figliuzzi M, Imberti B, Karmali MA, Remuzzi A, Remuzzi G, Zoja C. Verotoxin-1 promotes leukocyte adhesion to cultured endothelial cells under physiologic flow conditions. Blood.: , Zoja C, Morigi M, Figliuzzi M, Bruzzi I, Oldroyd S, Benigni A, Ronco P, Remuzzi G. Proximal tubular cell synthesis and secretion of endothelin-1 on challenge with albumin and other proteins. Am J Kidney Dis.26: , Morigi M, Zoja C, Figliuzzi M, Foppolo M, Micheletti G, Bontempelli M, Saronni M, Remuzzi G, Remuzzi A. Fluid shear stress modulates surface expression of adhesion molecules by endothelial cells. Blood. 85: , Morigi M, Zoja C, Figliuzzi M, Remuzzi G, Remuzzi A. Supernatant of endothelial cells exposed to laminar flow inhibits mesangial cell proliferation. Am J Physiol. 264:C1080-3, Luke Antiga graduated in 1999 in Biomedical Engineering and got his PhD in Bioengineering in 2003, Politecnico di Milano, having worked at the research laboratory of Biomedical Technology, Department of Bioengineering Institute Mario Negri. Training activities: 2003 Post-doctoral fellow at Imaging Research Laboratories, Robarts Research Institute, London, Ontario. 265

268 Areas of interest: acquisition and image processing for medical and microscopy applications, numerical modeling of transport phenomena. Roles: from 2000 to 2002 Doctoral Student at the Laboratory of Biomedical Technology, Department of Bioengineering, from 2002 to 2003 visiting scientist Robarts Institute for medical Imaging, London, Ontario, Canada, from 2004 to 2006 resercher at the Laboratory of Biomedical Technology, Department of Bioengineering, from 2007 Head Unit of Medical Imaging, Department of Bioengineering. Selected publications Lee SW, Antiga L, Spence JD and Steinman DA. Geometry of the carotid bifurcation predicts its exposure to disturbed flow. Stroke, in press, Antiga L, Piccinelli M, Fasolini G, Ene-Iordache B, Ondei P, Ruggenenti P, Remuzzi G and Remuzzi A. Computed tomography evaluation of ADPKD progression: a progress report. Clinical Journal of the American Society of Nephrology (CJASN), 1(4): , Jul Thomas JB, Antiga L, Che S, Milner JS, Hangan Steinman DA, Spence JD, Rutt BK and Steinman DA. Variation in the carotid bifurcation geometry of young vs. older adults: Implications for "geometric risk" of atherosclerosis. Stroke, 36(11): , Nov Antiga L, Steinman DA. Robust and objective decomposition and mapping of bifurcating vessels. IEEE Transactions on Medical Imaging, 23(6): , June Antiga L, Ene-Iordache B and Remuzzi A. Computational geometry for patient-specific reconstruction and meshing of blood vessels from MR and CT angiography. IEEE Transactions on Medical Imaging, 22(5): , May Antiga L, Ene-Iordache B, Remuzzi G and Remuzzi A. Automatic generation of glomerular capillary topological organization. Microvascular Research, 62: , June

269 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The research activity of the Department of Biomedical Engineering is aimed to the application of engineering techniques to the study of biological processes, pathological conditions and for the development of innovative therapeutic strategies. Research activities in progress, in basic and applied research, are based on theoretical models, digital histological analysis, medical image processing for the quantification of three-dimensional structures at macro- and microscopic level, direct measure of physical-chemical parameters at experimental and clinical level, cell culture and computer based data management. There are mainly four areas of research at the moment: the study of renal disease progression at experimental and clinical level, investigation of vascular flow dynamics, cell culture for in vitro tissue engineering and the development of information systems for clinical data management in clinical trials and in conventional medical activities. FINDINGS/MAIN RESULTS Evidence has been produced showing that the arterial vessels geometry and haemodynamic conditions influence the localization of cerebral aneurysms. These results open new ground in the understanding of the mechanisms responsible for this vascular disease. Demonstration of a significant relationship between morfometrical parameters of renal parenchyma and the loss of renal function in patients with polycystic kidney disease. Demonstration that the use of mesenchymal cells improves the function of transplanted islands under the kidney capsule. Demonstration that drugs that inhibit angiotensin II allow to regenerate the glomerular capillary with mechanisms that involves podocytes proliferation and endothelial cells remodeling. Implementation of systems for data management of clinical information generated during conduction of controlled clinical trials. Development of an electronic case report form (e-crf) for data handling in randomized clinical trials. NATIONAL COLLABORATIONS Dipartimento di Bioingegneria, Politecnico di Milano, Milano. Fidia Advanced Biopolymers, Abano Terme, Padova. Istituto di Fisiologia Clinica CNR, Pisa. Unità di Diabetologia, Ospedali Riuniti, Bergamo. STMicroelectronics, Agrate Brianza, Milano 267

270 INTERNATIONAL COLLABORATIONS Massachussetts Institute of Technology, Cambridge MA, USA. National Alliance for Medical Imaging Computing, USA. University of Toronto, Ontario, Canada. Ghent University, Ghent, Belgium. Technical University, Eindhoven, The Netherlands. University Hospital, Maastricht, The Netherlands. Centre Européen d Etudes du Diabètes, Strasburg, France EDITORIAL BOARD MEMBERSHIP Drugs of Today, (Andrea Remuzzi) International Journal of Artificial Organs, (Andrea Remuzzi) PEER REVIEW ACTIVITIES Kidney International Journal of the American Society of Nephrology American Journal of Physiology Renal Physiology Physiological Reviewes Medical & Biological Engineering & Computing IEEE Transactions on Medical Imaging IEEE Transactions on Biomedical Ingeneering Medical Physics Journal of Biomechanics Medical Engineering and Physics Artificial Organs International Journal of Artificial Organs Biomaterials Contemporary Clinical Trials 268

271 EVENT ORGANIZATION Seminar: Imaging vascular access: a pre- and post- operative approach, 2 February, Ranica, Bergamo, Italy. Seminar: Openclinica: a web-based software platform for managing multi-site clinical research 12 Luglio, Ranica, Bergamo, Italy. Worshop: Worshop nuove tecnologie: La microscopica elettronica a trasmissione e a scansione", 26 November, Milan, Italy. Seminar: Bogdan Ene-Iordache - Master di Primo Livello in Ricerca Clinica - Gestione dei dati di uno studio clinico: l'informatizzazione. 16 May 2007, Villa Camozzi, Ranica, Italy. PARTICIPATION IN EVENTS IN WHICH THE DEPARTMENT WAS INVOLVED National Alliance for Medical Image Computing, Third All Hands Meeting, Salt Lake City, January National Alliance for Medical Image Computing, Summer Project Week, MIT, Boston, June L'emodinamica nell'evoluzione e nell'instabilità della placca aterotrombotica, Congresso Regionale SIAPAV, Crema, October ESAO, European Society for Artificial Organs, Krems, Austria, September Experimental study of patient-specific kink in arterio-venous graft using particle image velocimetry. ASME Summer Bioengineering Conference, Keystone, CO, June Variation in the hemodynamics of young adult carotid bifurcations. 13 th Symposium on Intracranial Pressure and Brain Monitoring, Mechanism and Treatment, San Francisco, CA, USA, July 22-26, World Congress of Nephrology, Rio de Janeiro, Brazil, April 21-25, Nefropatie croniche e complicanze cardiovascolari: nuove prospettive di prevenzione e trattamento alla luce dei più recenti trials clinici. 30 November Ranica 26 th Workshop of the AIDIPIT Study Group, Montpellier, France, February Characterization of the immunereaction in a bioartificial pancreas in a model of xeno-, alloand syngenic transplantation. 26 th Workshop of the AIDIPIT Study Group, Montpellier, France, February The influence of encapsulation on oxygen consumption rate by bovine pancreatic islets. 269

272 GRANTS AND CONTRACTS Research grant from PKD foundation - Effect of long-acting somatostatin on disease progression in ADPKD: a long-term, three year, follow-up study. Research project FP6 UE-STEPS "A system approach to tissue engineering products and processes". FP DEMAND study: a prospective, randomized, double-blind, parallel-group study aimed at assessing whether the ACE inhibitor delapril alone or in combination with the dccb manidipine slow the rate of GFR decline as compared with placebo plus conventional antihypertensive therapy. Research Project ROTRF/JDRF Mesenchymal stem cells to protect islet allograft in diabetic rats Contract n. ID Reserch Projects: AIFA for contreolled clinical studies (VARIETY, VALID, ATHENA). ISN Research grant for the Kidney Disesease Data Center (COMGAN). Grant from Lombardia Region: development and maintenance of the web-site of Centro di Coordinamento per le Malattie Rare Regione Lombardia SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Remuzzi A, Sangalli F, Fassi A, Remuzzi G. Albumin concentration in the Bowman's capsule: multiphoton microscopy vs micropuncture technique. Kidney Int Dec;72(11):1410-1; Corna D, Sangalli F, Cattaneo D, Carrara F, Gaspari F, Remuzzi A, Zoja C, Benigni A, Perico N, Remuzzi G. Effects of rosuvastatin on glomerular capillary size-selectivity function in rats with renal mass ablation. Am J Nephrol. 2007;27(6): Remuzzi A. Vitamin D, insulin resistance, and renal disease. Kidney Int Jan;71(2):96-8. Luciani D, Cavuto S, Antiga L, Miniati M, Monti S, Pistolesi M, Bertolini G. Bayes pulmonary embolism assisted diagnosis: a new expert system for clinical use. Emerg Med J Mar;24(3): Remuzzi G, Cravedi P, Costantini M, Lesti M, Ganeva M, Gherardi G, Ene-Iordache B, Gotti E, Donati D, Salvadori M, Sandrini S, Segoloni G, Federico S, Rigotti P, Sparacino V, Ruggenenti P. Mycophenolate mofetil versus azathioprine for prevention of chronic allograft dysfunction in renal transplantation: the MYSS follow-up randomized, controlled clinical trial. J Am Soc Nephrol Jun;18(6):

273 RESEARCH ACTIVITIES Laboratory of Renal Biophysics Three dimensional reconstruction of the glomerular capillary network. We have recently documented that angiotensin II blockade not only retards the progression of renal diseases, but also induces partial regression of the glomerular lesions. To quantify the extent of the regression of sclerotic lesions and the potential regeneration of the glomerular capillary induced by a therapy with angiotensin converting enzyme (ACE) inhibitors we are developing three dimensional (3D) reconstruction of tissues. The glomerular capillary structure allows to concentrate in a small space a capillary network that filters high volume of water while retains plasmatic proteins in the circulation. We are setting up a new technique to study the morphology of the glomerular capillary and its wall. The aim is to examine specimens of renal tissue by a new electron microscopy technique, based on a combination of an electron beam with an ionic one, in order to achieve sectioning of samples and acquisition of serial images for 3D reconstruction of the ultrastructure by mathematical algorithms. Role of glomerular podocytes in the regression of glomerular damage induced by angiotensin converting enzyme (ACE) inhibitors. Proteinuria is a key predictor of renal disease progression. Clinical studies have documented that lowering proteinuria by pharmacological interventions retards renal disease progression. As compared to other anti-hypertensive drugs, ACE inhibitors or AII type 1 receptor (AT1R) blockers are more effective in slowing down the decline of glomerular filtration rate and in reducing by 50% the risk of death, dialysis or renal transplantation in both diabetic and non diabetic patients. Prolonged treatment (4 to 7 years) with these drugs can induce remission and in some patients even improve renal function. Three-dimensional reconstruction of the capillary tuft by serial section analysis allowed us to document that administration of a high dose of an ACE inhibitor in MWF rats with advanced nephropathy, not only remarkably reduced sclerosis volume in most glomeruli, but also increased the volume of glomerular tuft occupied by intact capillary, indicating consistent glomerular tuft repair. So far, the therapeutic effect of AII blockade has been mainly attributed to its capability to control extracellular matrix deposition. However the possibility that the treatment can modulate glomerular cell survival and repair is not well explored yet. We have recently show that the ACE inhibitor induces a selective increase in the number of glomerular epithelial cells (podocytes), highly differentiated cells with a limited capability to proliferate. On this basis, we then studied the effect of lisinopril, on the proliferation of podocytes and documented that an increase of it. Our results indicate that the ACE inhibitor allows the glomerular capillary to partially repair the damage in a coordinate way to maintain the podocyte number required for the ultrafiltration process. Studies are in progress to evaluate whether the restoration of podocyte number within the capillary is dependent on progenitor/stem cells of renal or extrarenal origin. We have also studied the role of parietal epithelial cells of the Bowman s capsule in tissue repair process and hypothesized that cells from Bowman s capsule can replace or increase visceral epithelial cells. Our studies on ACE inhibition-induced effects are instrumental for the understanding of the mechanisms responsible for glomerular damage regression and open new therapeutic prospects for chronic progressive nephropathies. 271

274 Application of confocal microscopy to the study cells and tissues. Fluorescent probes to investigate the distribution and co-localization of proteins and other specific antigens in cell monolayers and tissue specimens are commonly used in the majority of experimental applications. Bidimensional images are not of high resolution to provide correct information and the use of 3D reconstruction of fluorescent signal is sometimes mandatory. These problems can not be solved by the classic florescent microscopy that has some limitations. For these reasons we have developed the laser confocal microscopy by using the LSM510 META microscope from Zeiss (Germany). The microscopy is equipped with four laser lines that allow to excite the sample within the whole visible spectrum plus the characteristic META scan head that allow to analyze the emission spectrum of different fluorophores to optimally separate them. The instrument is characterized by and used for its high specificity and for the excellence of the revealed and elaborated signal. Do peptides derived from albumin degradation have a role in the development of renal damage? In proteinuric nephropathies loss of the permselective properties of the glomerular membrane leads to abnormal filtration of proteins, mainly albumin, that damage the proximal tubular cell inducing the release into the interstitium of chemokines responsible for the recruitment and activation of inflammatory cells. The interstitial infiltrates are also characterized by the presence of immune competent cells including dendritic cells (DCs) and T lymphocytes that accumulate within renal parenchyma even in absence of an immune insult. As antigen presenting cells, DCs can initiate immune response or tolerance. Within the kidney, DCs are in close contact with the tubular epithelium and work as immune sentinels to probe renal interstitium in search for foreign antigens. It is conceivable that upon injury, the inflammatory stimuli released from the tubular cell make the DC antigenic favoring the presentation of normally ignored self-antigens and triggering an immune response. To verify our hypothesis we have performed in vitro studies exposing renal proximal tubular cells to albumin concentrations mimicking those present in the ultrafiltrate in proteinuric conditions. We have identified peptides derived from albumin degradation by mass spectrometry and we have evaluated their potential antigenicity, in syngeneic conditions, in mixed leukocyte reaction, using bone marrow-derived DCs (in collaboration with the Department of Molecular Medicine and the Laboratory of Analytic Biochemistry-Department of Environmental Health Sciences). Laboratory of Biomedical Technologies Development of computerized systems for controlled clinical trias. Numerous clinical trials are conducted in the Clinical Research Center for Rare Diseases Aldo e Celè Daccò. These studies must be carried out with respect to the GCP (Good Clinical Practice) guidelines and require high quality in data handling. Every clinical study requires a paper case report form (CRF) for collection of patients clinical observations. These data must be checked for inconsistency by dedicated monitoring staff, and then recorded electronically. In our laboratory we developed applications tailored for data management of clinical studies using relational databases systems (RDBMS) and specific programs aimed to data elaboration, validation and extraction for subsequent statistic analyses. REIN, BENEDICT MYSS REIN2, DKG are only some of the trials concluded successfully and of which results were published in prestigious medical journals. This accomplishment is, in a small part, due to the contribution of our Laboratory. Recently, we have developed an innovative system of data handling using electronic CRFs instead of the usual paper for the DEMAND study. This system is based on the use of laptop computers by the clinical investigators working in eight diabetes outpatient clinics. 272

275 Management of the Registry for Rare Disease in Lombardy. Together with our colleagues from the Information Center for Rare Diseases we are involved in the conduction of the Regional Center for Rare Diseases of Lombardia, Italy. We are directly involved for the development and maintenance of the web site of the center and management of a regional Registry for Rare Diseases. We have set-up the databases and developed related webpages for among centers, associations of patients and congenital rare diseases. These are hosted on the homepage of the center ( The Registry for Rare Diseases was born in 2007 as collaboration between Mario Negri Institute, Lombardia Informatica (LI) and Regione Lombardia. The main idea is to create a regional registry for rare diseases where all medical staff from Lomardia could register information regarding rare diseases. The application (called Sistema Malattie Rare - SMR) was developed by LI in collaboration with our group. SMR application is using web-based technology and can be used jointly with the health patient card. It is actually in use in almost all centers dedicated for rare diseases in Lombardia. Our laboratory is involved in the maintenance of SMR, statistical analyses of registry data and creation of a minimal set of data that are shared at national level by the Istituto Superiore di Sanità (ISS). File download from LI is secured because data are encrypted using an asymmetrical algorithm with public and private key at 2048 bit. Also data transfer at ISS is performed securely by using the htpp Internet protocol. KDDC a coordinating center for data collection and surveillance of prevention programs on non-communicable chronic diseases in emerging countries. Chronic kidney diseases are emerging as a global threat to human health. Prevalence and incidence of renal diseases in developing countries are not known, and this is an obstacle to the adoption of preventive measures. Prevention is the only hope for these countries where treatment options for end stage renal failure are simply not available to the vast majority of the population because of their costs. The International Society of Nephrology (ISN), through the Commission for Global Advancement of Nephrology (COMGAN), has established a research committee in order to face the problems about prevention of kidney diseases in developing countries. The coordination of the team and intervention programs was committed to the Mario Negri Institute for Pharmacological Research at the Clinical Research Center Aldo e Cele Daccò. The general aim of the project is to define programs in developing countries to identify those subjects who are at risk of developing a renal disease later in life, in order to design a prevention strategy on national basis by means of interventions of the local ministries of health to governmental and financial level. The Kidney Disease Data Center (KDDC), headquartered in our Laboratory, is dedicated to data management for the prevention programs underway in emerging countries. We have set up an instrument to collect clinical data from different Centres located world-wide. Data are stored in a dedicated server in our Laboratory. Results of our epidemiological analyses, shared also with medical staff of the center, allow us to have a general overview on the health of population under study. The prevention program is underway and we have already collected data from participating centers from Moldova, Bolivia, Egypt, Nepal, China and Mongolia. First results on screening on several thousands of subjects, confirm the need to proceed with prevention programs in theses countries. Other countries are willing to start their programs, as for example Morocco, Mozambique, Mexico and Argentina. Through the activity of KDDC it will be possible to monitor the course of the prevention programs and to tailor them to fit the needs of each participating country. 273

276 Three-dimensional reconstruction and hemodynamic simulation of vascular segments from CT and MR imaging. Evidence that atherosclerotic plaques form mainly at bifurcations of the arterial system led to the hypothesis that impaired hemodynamic conditions may favour the initiation and progression of atherosclerosis. Furthermore, it has been demonstrated how intimal hyperplasia, cause of failure of grafts and arterovenous fistulas, is also localized at sites of complex hemodynamics. Analogously, recent studies on formation, localization and growth of cerebral aneurysms have underlined the importance of hemodynamics on the development of this pathology. Given the influence of vessel shape on hemodynamics, it is important to dispose of accurate and fast methods for three-dimensional reconstruction of vascular geometry. Modern angiographic techniques, such as computed tomography (CT) and magnetic resonance (MR), allow to obtain detailed information on vascular structures. In our laboratory, we developed techniques for three-dimensional reconstruction, geometric analysis, computational mesh generation and fluiddynamic simulation of vascular tracts from CT and MR angiography. CT and MR images are transferred from clinical scanners on local workstations and are then processed with modeling algorithms to generate three-dimensional surfaces representing the interface between the vascular wall and its lumen. The accuracy of the reconstruction has been experimentally validated by us. Accurate measurements are then performed on the generated model using computational geometry algorithms. These techniques have been applied to several vascular segments, mainly cerebral aneurysms, arterovenous fistulas and grafts for hemodialysis access. The pathologies studied include atherosclerosis, intimal hyperplasia, development of cerebral aneurysms and Takayasu's arteritis, a rare disease affecting the arterial system. Quantification of anatomical structures from CT and MR imaging. The introduction of non-invasive three-dimensional imaging techniques, such as computed tomography (CT) and magnetic resonance (MR), in the clinical setting has opened the possibility to analyze the anatomical structures of organs in high detail. The non-invasiveness of such techniques allows to extend the observation to populations of patients followed over time. The evaluation of natural evolution of diseases or of treatment efficacy often requires a detailed morphological quantification which cannot be uniquely performed by visual inspection. In this context, we developed image-based quantification techniques characterized by high accuracy and reproducibility. Wevare applying these tools to a study on patients affected by autosomal dominant polycystic disease (ADPKD) with the aim of testing the efficacy and safety of some treatments. Volumes of kidneys and their main tissue components (cysts and parenchyma) have been quantified from CT o RM images. Development of devices for the transplantation of immunoisolated islets. The project s main objective was to develop an immunisolation device for pancreatic islets that can be implanted in diabetic patients permitting allo-islet transplantation without the use of pharmacological immunosuppression and avoiding allosensitization of the patient. The study started from the design and characterization of the semi-permeable membrane used for the device construction. The device that we are developing can be implanted with minimally invasive surgical procedures and easy to retry. This system is a device made using polysulfone hollow fibers. The aims of our studies in the next months will be to improve functionality using nanotechnologies for materials characterization. Moreover we will develop new kinds of device and we will test new implantation sites. Differentiation of pancreatic precursor cells. Type 1 diabetes mellitus is an autoimmune disease characterized by distruction of insulinproducing beta cells in pancreatic islets. The damage leads to a deficiency of insulin resulting in a complete relance on infusion of exogenous insulin. The replacement of the beta cell mass is a 274

277 potential cure for type I diabetes, but this procedure suffer from a shortage of available donor tissue in comparison to the number of potential recipients. In our laboratoris we have studied the method to obtain beta cells from progenitor cells extracted by pancreatic tissue. Actually, it is possible to obtain insulin-producing cell in vitro, but the level of insulin is much lower than that contained in pancreatic islet. Our research is oriented towards the development of new strategies to detect and to select progenitors of beta cells in the pancreas. To this purpose we developed a method to incorporate, in vitro, into the pancreatic islets two thymidine analogous (IdU and CldU). This technique permits to identify stem cells into the islets and to evaluate their proliferation capacity. Co-transplantation of mesenchymal stem cells and pancreatic islets to induce immunotolerance. Several recent studies have suggested that pancreatic islet transplantation offers an alternative to pancreas transplantation. In the last year in our laboratories we have defined the optimal conditions for isolation of rat islets using an automated method. Rat pancreatic islets have been transplanted under the kidney capsule of diabetic rats. Diabetes was induced by single injection of streptozotocin. Two models of islet transplantation have been used, syngenic- and allotransplantation. In syngenic transplantation rat islets have been implanted into inbreed rats and normoglycemia conditions were maintained for several months. Different strains of rats were used as donor and recipient in allogenic transplantation. However, immunosuppression is request to prevent islet graft rejection. To avoid the use of immunosuppressive drug therapy, we have planned to study the immunomodulatory properties of mesenchymal stem cells (MSCs). Preliminare experiments have been done using Wistar rats as donors and Lewis rata as recipients. To evaluate if MSCs can suppress the immune response toward allograft, Lewis MSCs were transplanted simultaneously to pancreatic islets. Vascular tissue engineering. Artificial vascular prostheses can be used in patients only if their caliber is larger than 6 mm. A lower caliber prostheses implies a high trombotic risk. For this reason, several investigators are developing bioartificial biological vascular substitutes using tissue engineering technique. In our laboratory we used a biodegradable hialuronic acid matrix as a scaffold to produce cellularized vascular constructs. We set up culture and seeding conditions both for smooth muscle cells and mesenchymal bone marrow cells. To obtain physiological mechanical conditioning of the tubular constructs we developed a new type of rotating wall bioreactor that allows to keep the culture of vascular constructs in dynamic conditions. This simple method allowed to decrease apoptosis incidence and to increase cell matrix depositions. More recently we tested, in collaboration with IFC-CNR in Massa, the possibilityto use innovative elastomeric material to construct cellularized vascular substitutes. 275

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279 LABORATORY OF BIOLOGY AND THERAPY OF METASTASIS* STAFF Head Tumor Angiogenesis Unit Head Molecular Cancer Therapeutics Unit Head Raffaella GIAVAZZI, Biol.Sci.D., Ph.D. Giulia TARABOLETTI, Biol.Sci.D. Maria Rosa BANI, Biol.Sci.D., Ph.D. * Research activities of this Laboratory are listed in the Department of Oncology section (pag. 7) 277

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281 Aldo and Cele Daccò Center Ranica (Bg) ANNUAL REPORT 2007 departments and laboratories 279

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283 DEPARTMENT OF RENAL MEDICINE STAFF Head Piero RUGGENENTI, M.D. Laboratory of Biostatistics Head Annalisa PERNA, Stat.Sci.D. Unit of Drug Monitoring Head Giulia GHERARDI Laboratory of Clinical Chemistry Head Flavio GASPARI, Chem.D. Laboratory of Advanced Development of Drugs Head Norberto PERICO, M.D. Unit of Pharmacology and Pharmacogenetics Head Dario CATTANEO, Chem.Pharm. D., Ph.D Unit of Early Clinical Evaluation of Drugs Head Aneliya PARVANOVA, M.D. 281

284 CURRICULA VITAE Piero Ruggenenti got his Medicine degree in 1983 at the University of Milan, Italy; he got his specialization in Cardiology in 1985 and in Clinical Nephrology in 1989 at the same University; he specialized in Pharmacological Research in 1988 at IRFMN. Educational training: in researcher at "Centro di Fisiologia Clinica ed Ipertensione, Clinica Medica IV", Università degli Studi di Milano; in 1984 Researcher at IRFMN, Bergamo, Italy in Honorary Registrar of the Unit for Metabolic Medicine, Division of Medicine (University of London) of Guy's and St. Thomas's Hospitals, London; in Assistant Professor of the Division of Nephrology and Dialysis of the Ospedali Riuniti di Bergamo. Areas of interest: mechanisms of chronic renal disease progression, diabetes and diabetic complications, clinical transplantation, thrombotic microangiopathies, cardiovascular complications of chronic renal disease, clinical trials, clinical pharmacology. Employment: from 1990 Assistant Professor of the Division of Nephrology and Dialysis of the Ospedali Riuniti di Bergamo; in Head, Unit of Advanced Development of Drugs, Daccò Center, Ranica, Bergamo, Italy; since 2000 Head, Department of Renal Medicine, Daccò Center, Bergamo, Italy. Selected publications P. Ruggenenti, A. Perna, L. Mosconi, M. Matalone, G. Garini, M. Salvadori, C. Zoccali, F. Scolari, Q. Maggiore, G. Tognoni, G. Remuzzi (for The GISEN Group). Randomised placebo-controlled trial of effect of ramipril on decline in glomerular filtration rate and risk of terminal renal failure in proteinuric, non-diabetic nephropathy. Lancet 1997;349: P. Ruggenenti, A. Perna, G. Gherardi, F. Gaspari, R. Benini, G. Remuzzi, on behalf of GISEN. Renal function and requirement for dialysis in chronic nephropathy patients on long-term ramipril: REIN follow-up trial. Lancet 1998;352: P. Ruggenenti, A. Perna, G. Gherardi, G. Garini, C. Zoccali, M. Salvadori, F. Scolari, F.P. Schena, G. Remuzzi. Renoprotective properties of ACE-inhibition in non-diabetic nephropathies with non-nephrotic proteinuria. Lancet 1999;354: G. Remuzzi, C. Chiurchiu, M. Abbate, V. Brusegan, M. Bontempelli, P. Ruggenenti. Rituximab for idiopathic membranous nephropathy. Research Letter. Lancet 2002;360: P. Ruggenenti, A. Fassi, A. Parvanova, S. Bruno, I. Iliev, V. Brusegan, N. Rubis, G. Gherardi, F. Arnoldi, M. Ganeva, B. Ene-Iordache, F. Gaspari, A. Perna, A. Bossi, R. Trevisan, A.R. Dodesini, G. Remuzzi for the Bergamo Nephrologic Diabetes Complications Trial (BENEDICT) Investigators. Preventing microalbuminuria in type 2 diabetes. N Engl J Med 2004;351: G. Remuzzi, P. Cravedi, A. Perna, B.D. Dimitrov, M. Turturro, G. Locatelli, P. Rigotti, N. Baldan, M. Beatini, U. Valente, M. Scalamogna, P. Ruggenenti Dual Kidney Transplant Group. Long-term outcome of renal transplantation from older donors. N Engl J Med 2006;354: Flavio Gaspari got his Chemistry degree in 1977 at the University of Milano, Italy, and the specialization in the same University in Educational training: in Fellow and Researcher at IRFMN, Milan; in at IRFMN, Bergamo, Italy. Areas of interest: pharmacokinetics and the metabolism of xanthines in different animal species; drug pharmacokinetics in uremic patients and in subjects with different degrees of renal function; analytical methods to measure the most important immunosuppressive drugs to determine their pharmacokinetics in kidney, heart, and liver transplant recipients; evaluation of the renal function by using different approaches, in the study of renal disease progression, and in the comparison of different methods for albuminuria determination. Employement: He is Chief of Laboratory of Pharmacokinetics and Clinical Chemistry since January 2000 and he was Chief of this Unit since Selected publications Gotti E, Perico N, Gaspari F, Cattaneo D, Lesti MD, Ruggenenti P, Segoloni G, Salvadori M, Rigotti P, Valente U, Donati D, Sandrini S, Federico S, Sparacino V, Mourad G, Bosmans JL, Dimitrov BD, Iordache BE, Remuzzi G. Blood cyclosporine level soon after kidney transplantation is a major determinant of rejection: insights from the Mycophenolate Steroid-Sparing Trial. Transplant Proc Jun;37(5): Perico N, Gaspari F, Remuzzi G. Assessing renal function by GFR prediction equations in kidney transplantation. Am J Transplant Jun;5(6):

285 D. Cattaneo, F. Gaspari, S. Zanoni, S. Baldelli, E.Gotti, A. Perna, N. Perico, G. Remuzzi. Two-hour post-dose cyclosporine monitoring does not fit all in kidney transplantation. Therapy 2005;2: Ruggenenti P, Fassi A, Ilieva AP, Bruno S, Iliev IP, Brusegan V, Rubis N, Gherardi G, Arnoldi F, Ganeva M, Ene- Iordache B, Gaspari F, Perna A, Bossi A, Trevisan R, Dodesini AR, Remuzzi G; Bergamo Nephrologic Diabetes Complications Trial (BENEDICT) Investigators. Preventing microalbuminuria in type 2 diabetes. N Engl J Med Nov 4;351(19): Epub 2004 Oct 31. Gaspari F, Ferrari S, Stucchi N, Centemeri E, Carrara F, Pellegrino M, Gherardi G, Gotti E, Segoloni G, Salvadori M, Rigotti P, Valente U, Donati D, Sandrini S, Sparacino V, Remuzzi G, Perico N; MY.S.S. Study Investigators. Performance of different prediction equations for estimating renal function in kidney transplantation. Am J Transplant Nov;4(11): Cattaneo D, Merlini S, Pellegrino M, Carrara F, Zenoni S, Murgia S, Baldelli S, Gaspari F, Remuzzi G, Perico N. Related Articles, Links Therapeutic drug monitoring of sirolimus: effect of concomitant immunosuppressive therapy and optimization of drug dosing. Am J Transplant Aug;4(8): Norberto Perico got his Medicine degree in 1983 at the University of Milano, Italy. He got his specialization in Pharmacological Research in 1986 at IRFMN, Bergamo and in Clinical Nephrology in 1989 at the University of Verona, Italy. Educational training: in 1982 Fellow, Department of Pharmacology, New York Medical College, Valhalla, New York, USA; in Post Doctoral Fellow, Laboratory of Kidney Diseases, IRFMN, Bergamo, Italy; in Researcher in the same laboratory. Areas of interest: pathophysiology and pharmacology of cyclosporine nephrotoxicity; new immunosuppressive strategies to prevent renal graft rejection; innovative approach to induce tolerance to organ transplantation; mechanism(s) and management of progression of chronic renal diseases. Employment: in Head, Renal Physiology Unit, Laboratory of Kidney Diseases, IRFMN, Bergamo, Italy; in Assistant Professor, Division of Nephrology and Dialysis, Ospedali Riuniti di Bergamo, Italy; in Head, Laboratory of Transplant Immunology, IRFMN, Bergamo, Italy; from January 2000 Head, Laboratory of Drug Development, Department of Renal Medicine, IRFMN, Bergamo, Italy; from September 2000 Health Director, Daccò Center, IRFMN, Bergamo, Italy. From October 2002 he s Member, ISN-COMGAN Research Committee of the International Society of Nephrology. Selected publications: E. Gotti, N. Perico, A. Perna, F. Gaspari, D. Cattaneo, R. Caruso, S. Ferrari, N. Stucchi, G. Marchetti, M. Abbate, G. Remuzzi. Renal transplantation: can we reduce calcineurin inhibitor/stop steroids? Evidence based on protocol biopsy findings. J Am Soc Nephrol 2003;14: N. Perico, P. Ruggenenti, G. Remuzzi. Losartan in diabetic nephropathy. Expert Rev. Cardiovasc. Ther. 2004; 2(4): Remuzzi G, Lesti M, Gotti E, Ganeva M, Dimitrov BD, Ene-Iordache B, Gherardi G, Donati D, Salvadori M, Sandrini S, Valente U, Segoloni G, Mourad G, Federico S, Rigotti P, Sparacino V, Bosmans JL, Perico N, Ruggenenti P. mofetil versus azathioprine for prevention of acute rejection in renal transplantation (MYSS): a randomised trial. Lancet Aug 7;364(9433): Gaspari F, Ferrari S, Stucchi N, Centemeri E, Carrara F, Pellegrino M, Gherardi G, Gotti E, Segoloni G, Salvadori M, Rigotti P, Valente U, Donati D, Sandrini S, Sparacino V, Remuzzi G, Perico N; MY.S.S. Study Investigators. of different prediction equations for estimating renal function in kidney transplantation. Am J Transplant Nov;4(11): Perico N, Cattaneo D, Sayegh MH, Remuzzi G. Delayed graft function in kidney transplantation. Lancet Nov 13;364(9447): Cattaneo D, Merlini S, Zenoni S, Baldelli S, Gotti E, Remuzzi G, Perico N. Influence of co-medication with sirolimus or cyclosporine on mycophenolic acid pharmacokinetics in kidney transplantation. Am J Transplant Dec;5(12):

286 Annalisa Perna got her Statistical Sciences degree in 1984 at the University of Bologna, Italy. Educational training: She completed her research training at IRFMN, Bergamo Labs. and at the Daccò Center. Areas of interest: statistical methodology of long-term randomised clinical trials in nephrology, statistical methods for calculating sample size and for meta-analytic techniques. She is also involved in performing systematic reviews for the Cochrane Collaboration Renal Review Group. Employment: she is Head of the Laboratory of Biostatistics - Department of Renal Medicine at Daccò Center, Ranica (Bergamo). Principal publications: G. Remuzzi, P. Cravedi, A. Perna, B.D. Dimitrov, M. Turturro, G. Locatelli, P. Rigotti, N. Baldan, M. Beatini, U. Valente, M. Scalamogna, P. Ruggenenti. Dual Kidney Transplant Group. Long-term outcome of renal transplantation from older donors. N Engl J Med 2006;354: Ruggenenti P, Perna A, Loriga G, Ganeva M, Ene-Iordache B, Turturro M, Lesti M, Perticucci E, Chakarski IN, Leonardis D, Garini G, Sessa A, Basile C, Alpa M, Scanziani R, Sorba G, Zoccali C, Remuzzi G for the REIN-2 Study group. Blood-pressure control for renoprotection in patients with non-diabetic chronic renal disease (REIN-2): multicentre, randomised controlled trial. Lancet 365: , Schieppati A, Perna A, Zamora J, Giuliano AG, Braun N, Remuzzi G. Immunosuppressive treatment for idiopathic membranous nephropathy in adults with nephrotic syndrome. Cochrane Database Syst Rev. Oct 18(4):CD004293, 2004 Ruggenenti P, Fassi A, Ilieva AP, Bruno S, Iliev IP, Brusegan V, Rubis N, Gherardi G, Arnoldi F, Ganeva M, Ene- Iordache B, Gaspari F, Perna A, Bossi A, Trevisan R, Dodesini AR, Remuzzi G. Preventing microalbuminuria in type 2 diabetes. N Engl J Med 351(19) , The BENEDICT Group. The Bergamo NEphrologic Diabetes Complications Trial (BENEDICT): design and baseline characteristics. Control Clin Trials 24(4): , Plata R, Cornejo A, Arratia C, Anabaja A, Perna A, Dimitrov BD, Remuzzi G, Ruggenenti P for the Commission on Global Advancement of Nephrology (COMGAN), Research Subcommittee of the International Society of Nephrology. Angiotensin-converting-enzyme inhibition therapy in altitude polycythaemia: a prospective randomised trial. Lancet 359: , Dario Cattaneo got the Pharmacy degree in 1996 at the University of Milan, and the specialisation in Pharmacology (2001) awarded by the same University. In 2000 he got the specialization in Pharmacological Research at the Mario Negri Institute for Pharmacological Research (IRFMN) and in 2005 he has been awarded the PhD degree by the Open University of London, UK. Educational Training: in 1997 Post Doctoral Fellow, IRFMN, Laboratory of Pharmacokinetics and Clinical Chemistry; in 2000 beneficiary of the Fellowship Girola and from 2001 to 2005 recipient of the Monzino Fellowship for his research activity done at the IRFMN. Areas of interest: pharmacology (pharmacokinetics, pharmacodynamics and pharmacogenetics) of immunosuppressants, antiviral agents and hypolipidemic drugs; study of secondary forms of dyslipidemia; polypharmacological approaches for the treatment of chronic kidney diseases; assessment of humoral response as predictor of acute or chronic rejection after organ transplantation. Employement: in 1997 researcher, IRFMN, Laboratory of Pharmacokinetics and Clinical Chemistry,. Since 2006, Head of the Unit of Pharmacology and Pharmacogenetics, IRFMN, Ranica Bergamo. Component of the Ethical Committee (2003) of the Hospital Bolognini (Seriate, Italy) and the Hospital E.Medea (Bosisio Parini, Italy) since Member of the editorial board of Current Clinical Pharmacology and affiliate of the International Association of Therapeutic Drug Monitoring and Clinical Toxicology (IATDMCT) since Selected publications Cattaneo D, Merlini S, Zenoni S, Baldelli S, Gotti E, Remuzzi G, Perico N. Influence of co-medication with sirolimus or cyclosporine on mycophenolic acid pharmacokinetics in kidney transplantation. Am J Transplant Dec;5(12): Cattaneo D, Gotti E, Perico N, Bertolini G, Kainer G, Remuzzi G. Cyclosporine formulation and Kaposi's sarcoma after renal transplantation. Transplantation Sep 27;80(6): Perico N, Cattaneo D, Sayegh MH, Remuzzi G. Delayed graft function in kidney transplantation. Lancet Nov 13-19;364(9447): Cattaneo D, Merlini S, Pellegrino M, Carrara F, Zenoni S, Murgia S, Baldelli S, Gaspari F, Remuzzi G, Perico N. Therapeutic drug monitoring of sirolimus: effect of concomitant immunosuppressive therapy and optimization of drug dosing. Am J Transplant Aug;4(8): Cattaneo D, Perico N, Gaspari F, Gotti E, Remuzzi G. Glucocorticoids interfere with mycophenolate mofetil bioavailability in kidney transplantation. Kidney Int Sep;62(3):

287 Giulia Gherardi got her Scientific High School Diploma on 1989 at the Liceo Scientifico Marie Curie in Zogno (Bergamo), the Nurse Diploma on 1995 at the Scuola per Infermieri Professionali, Ospedali Riuniti, Bergamo. Educational training: Clinical Research Nurse Diploma on 1997 at IRFMN Daccò Center. Areas of interest: statistical methodology of long-term randomised clinical trials in nephrology, diabetology; the organisation and the monitoring of clinical trials. Emplyment: In involved as co-organazing, speaker, co-speaker and tutor for the Clinical Research Course for Nurse at IRFMN Daccò Center (Ranica Bergamo). Several training activities for Nurses in Clinical Research area. In , Clinical Research Monitor at IRFMN Daccò Center; since 2000 Monitoring Unit Chief. Selected pubblications Gaspari F, Ferrari S, Stucchi N, Centemeri E, Carrara F, Pellegrino M, Gherardi G, Gotti E, Segoloni G, Salvadori M, Rigotti P, Valente U, Donati D, Sandrini S, Sparacino V, Remuzzi G and Perico N on the behalf of the MY.S.S. study investigators. Performance of different prediction equations for estimating renal function in kidney transplantation. American Journal of Transplantation. 2004; 4: Ruggenenti P, Fassi A, Parvanova Ilieva A, Bruno S, Petro Iliev I, Brusegan V, Rubis N, Gherardi G, Arnoldi A, Ganeva M, Ene-Iordache B, Gaspari F, Perna A, Bossi A, Trevisan R, Dodesini AR, Remuzzi G, for the Bergamo Nephrologic Diabetes Complications Trial (BENEDICT) Investigators. Preventing microalbuminuria in type 2 diabetes. N Engl J Med. 2004; 351: Remuzzi G, Lesti M, Gotti E, Ganeva M, Dimitrov BD, Ene-Iordache B, Gherardi G, Donati D, Salvadori M, Sandrini S, Valente U, Segoloni G, Mourad G, Federico S, Rigotti P, Sparacino V, Bosmans JL, Perico N, Ruggenenti P, for the MY.S.S. Group. Mycophenolate mofetil versus azathioprine for prevention of acute rejection in renal transplantation (MYSS): a randomized trial. Lancet Aug 7; 364: Ruggenenti P, Perna A, Gherardi G, Benini R, Remuzzi G. Chronic proteinuric nephropathies: outcomes and response to treatment in a prospective cohort of 352 patients with different patterns of renal injury. Am J Kidney Dis Jan; 35 (6): Ruggenenti P, Perna A, Zoccali C, Gherardi G, Benini R, Testa A, Remuzzi G. Chronic proteinuric nephropathies. II. Outcomes and response to treatment in a prospective cohort of 352 patients: differences between women and men in relation to the ACE polymorphism. Gruppo Italiano di Studi Epidemiologici in Nefrologia. J Am Soc Nephrol Jan; 11 (1): Ruggenenti P, Perna A, Gherardi G, Garini G, Zoccali C, Salvadori M, Scolari F, Schena FP, Remuzzi G. Renoprotective properties of ACE-inhibition in non-diabetic nephropathies with non-nephrotic proteinuria. Lancet Jul 31; 354 (9176): Aneliya Parvanova Ilieva got her Medical Doctor degree at the Faculty of Medicine, Thracian University (former Higher Medical Institute), Stara Zagora, Bulgaria in 1988, and the specialisation in Pharmacology in Department of Pharmacology, Faculty of Medicine, of the same university in Educational training: : Teaching of 3 rd, 4 th and 5 th -year medical students and 2 nd and 3 rd -year clinical nurses in a general pharmacology and clinical pharmacology, Thracian University, Stara Zagora, Bulgaria. Examiner of these students in theoretical and practical, oral and written exams and tests. 1993: Course on investigation of isolated organs Bulgarian Academy of Sciences, Sofia. 1998: Visiting scientist, IRFMN, Ranica, Bergamo, Italy. 1998: Proficiency in the methods for insulin sensitivity evaluation (hyperinsulinemic euglicaemic clamp technique) and glomerular filtration rate evaluation (plasma clearance of iohexol). Areas of interest: primary and secondary prevention of the chronic microvascular diabetic complications (diabetic nephropathy, diabetic retinopathy and diabetic neuropathy); role of insulin resistance and hyperhomocysteinemia in these pathologies. Employment: She participated as investigator in several clinical studies. She is Chief Unit of Early Clinical Evaluation of Drugs at IRFMN since She is a member of the Union of Bulgarian Doctors (since 1989), of the Union of Pharmacologists in Bulgaria (since 1990), and member of the Union of Scientists in Bulgaria (since 1991). Selected publications Parvanova A, Chiurchiu C, Ruggenenti P, Remuzzi G. Inhibition of the renin-angiotensin system and cardio-renal protection: focus on losartan and angiotensin receptor blockade. Expert Opinion on Pharmacotherapy 2005 Sep; 6 (11): Ruggenenti P, Fassi A, Parvanova A, Bruno S, Iliev I, Brusegan V, Rubis N, Gherardi G, Arnoldi F, Ganeva M, Ene- Iordache, Gaspari F, Perna A, Bossi A, Trevisan R, Dodesini A, Remuzzi G. Preventing Microalbuminuria in Type 2 Diabetes. NEJM 2004;351(19): Parvanova A, Iliev I, Filipponi M, Dimitrov BD, Vedovato M, Tiengo A, Trevisan R, Remuzzi G, Ruggenenti P. Insulin resistance and proliferative retinopathy: a cross-sectional, case-control study in 115 patients with type 2 diabetes. J Clin Endocrinol Metab 2004 Sep; 89(9):

288 The BENEDICT Group. The Bergamo Nephrologic DIabetes Complications Trial (BENEDICT): design and baseline characteristics. Controlled Clinical Trials 2003; 24: Parvanova A, Iliev I, Dimitrov BD, Arnoldi F, Zaletel J, Remuzzi G, Ruggenenti P. Hyperhomocysteinemia and increased risk of retinopathy: a cross-sectional, case-control study in patients with type 2 diabetes. Diabetes Care 2002; 25 (12):

289 INTRODUCTION TO THE DEPARTMENT'S ACTIVITIES The Department of Renal Medicine was established on 1999 at the Clinical Research Center for Rare Diseases Aldo e Cele Daccò Villa Camozzi, Ranica to coordinate the activities of three Laboratories and two Units. The activities of the Department are mainly focused on the study of the mechanisms of progression of chronic nephropathies, of new prevention and intervention strategies for diabetic nephropathy, non diabetic chronic nephropathies, chronic allograft dysfunction, of cardiovascular complications of diabetes, chronic renal disease, dialysis and transplantation and of thrombotic microangiopathies. The main aims of these activities are: 1. To identify screening and intervention strategies aimed to prevent the onset of nephropathy and of other chronic complications of diabetes and/or hypertension. 2. To define intervention strategies to prevent or slow the progression of chronic nephropathies and eventually obtain remission/regression of renal dysfunction. 3. To optimize immunosuppressive protocols in kidney transplantation and to define new donor selection criteria in order to expand the pool of available organs. These aims will be pursued through the following modalities: 1. Pilot pathophysiology and clinical pharmacology studies fully finalized at the Clinical Research Center to test new pathogenetic hypotheses and new treatment modalities. 2. National and international networks and multicenter trials aimed to verify the efficacy of treatments of potential interest identified as described at point Meta-analyses and probabilistic models to test new risk factors and treatments in large samples of patients and to transfer this information at individual level. Many of these activities rest on the possibility of a tight cooperation with the Department of Molecolar Medicine, the Department of Bioengineering and the Public-Private Department of Specialist and Transplant Medicine. This cooperation allows to plan the research activities of the Department on the basis of new information derived from basic research and of problems of major clinical relevance emerging from routine clinical activities. FINDINGS/MAIN RESULTS Definition and validation of specific treatments aimed to prevent the developing of nephropathies in subjects with type 2 diabetes Definition and validation of new integrated treatment protocols aimed to slow the progression and/or to achieve remission/regression of diabetic and non-diabetic chronic nephropathies Institution of a standardized protocol on line (The Remission Clinics ) finalized to achieve regression/remission of chronic nephropathies in hospital practice in the setting of a multicenter Network Characterization of the antiproteinuric, nephroprotective and cardioprotective effect of maximized and polypharmacologic renin-angiotensin system inhibition Identification of acquired or congenital risk factors for chronic complications of diabetes Definition and validation of new, specific treatments for idiopathic membranous nephropathy and for HUS forms associated with genetic defect of complement factors 287

290 Definition and validation of new laboratory procedures and predictive models to help monitoring and optimizing immunosuppressive therapy in clinical transplantation Definition and validation of selection and allocation criteria of kidneys from marginal donors to increase the donor pool and the transplant activity Optimization of Doppler ultrasound techniques for the diagnosis and monitoring of vascular complication of renal transplant and chronic dialysis patients Finalization and activation of multicenter clinical trials aimed to prevent diabetic nephropathy, the progression of chronic nephropathies, acute and chronic allograft rejection and to identify predictors and promoters of target organ damage Computerization of data acquisition and monitoring procedures for the conduction of controlled clinical trials NATIONAL COLLABORATIONS Lombardia - Ospedale C. Cantù, Abbiategrasso (MI) - Ospedale Civile di Asola, Asola (MN) - Ospedale Fenaroli, Alzano Lombardo (BG) - Azienda Ospedaliera OO.RR., Bergamo - Ospedale Caduti Bollatesi, Bollate (MI) - Azienda Ospedaliera Spedali Civili, Brescia - Ospedale San Biagio, Clusone (BG) - Ospedale S. Anna, Como - Azienda Ospedaliera Istituti Ospedalieri, Cremona - Ospedale di Desio (MI) - Ospedale Briolini, Gazzaniga (BG) - Azienda Ospedaliera di Melegnano, Melegnano Vizzolo Predabissi (MI) - Ospedale San Leopoldo Mandic, Merate (LC) - Ospedale Maggiore Policlinico, Milano - Ospedale Provinciale San Carlo Borromeo, Milano - Azienda Ospedaliera - Polo Universitario L. Sacco, Milano - Ospedale Fatebenefratelli, Milano - Ospedale Niguarda Ca' Granda, Milano - Clinica Pediatrica G. e D. De Marchi, Milano - Ospedale San Raffaele, Milano - Ospedale Pediatrico di Montichiari, Montichiari (BS) - Ospedale San Gerardo, Monza (MI) - Istituti Clinici Zucchi, Monza (MI) - Università degli Studi di Pavia, Dipartimento di Medicina Interna e Terapia Medica, Pavia - Centro Antidiabetico, Ponte San Pietro (BG) - Azienda Ospedaliera Ospedale Treviglio Caravaggio, Romano di Lombardia (BG) - Istituto clinico Humanitas, Rozzano (MI) - Ospedale Bolognini, Seriate (BG) - Azienda Ospedaliera Ospedale Treviglio Caravaggio, Treviglio (BG) - Ospedale Regionale di Circolo Fondazione Macchi, Varese 288

291 - USL 60, Unità Operativa di Nefrologia e Dialisi, Vimercate (LC) Piemonte - Azienda Ospedaliera Santa Croce e Carli, Cuneo - Ospedale Civile, Ivrea - A.S.O. Maggiore della Carità, Novara - Azienda Ospedaliera San Giovanni Battista, Torino - Ospedale Mauriziano Umberto I, Torino - Ospedale Regina Margherita, Torino - Ospedale Martini, Torino - Ospedale Luigi Einaudi, Torino Veneto, Trentino Alto-Adige e Friuli Venezia Giulia - Casa di Cura Abano Terme, Abano Terme (PD) - Ospedale Civile, Belluno - Ospedale S. Giacomo Apostolo, Castelfranco Veneto, Treviso - Ospedale Provinciale Umberto I, Mestre (VE) - Ospedale Giustinianeo, Padova - Università degli Studi di Padova, Istituto di Anatomia Patologica, Padova - Ospedale Civile, Padova - Ospedale S. Camillo dé Lellis, Schio (VI) - Ospedale Regionale Santa Maria dei Battuti, Treviso - Ospedale Ca Fondello, Divisione Nefrologia e Dialisi, Treviso - Ospedale Civile Maggiore Borgo Trento, Verona - Ospedale Policlinico Borgo Roma, Verona - Ospedale Civile San Bortolo, Vicenza - Ospedale Santa Chiara, Trento - Istituto Scientifico per l'infanzia Burlo Garofalo, Trieste - Ospedale S. Antonio, S. Daniele del Friuli, Udine - Università degli Studi di Udine, Centro Trapianti Fegato-Rene-Pancreas, Udine Liguria, Emilia Romagna e Toscana - Azienda Ospedaliera San Martino, Genova - Istituto G. Gaslini, Genova - Ospedale S. Orsola Malpighi, Bologna - Ospedale Policlinico, Modena - Istituto di Clinica Medica e Nefrologia, Parma - Ospedale Santa Maria delle Croci, Ravenna - Arcispedale Santa Maria Nuova, Reggio Emilia - Ospedale Santa Maria Annunziata, Bagno a Ripoli, Firenze - Azienda Ospedaliera Careggi-Monna Tessa, Firenze - Ospedale Nuovo S. Giovanni di Dio, Firenze - Azienda Ospedaliera Meyer, Firenze - Ospedale di S. Miniato, S. Miniato (FI) - Azienda Ospedaliera Cisanello, Pisa - Ospedale di Pistoia, Pistoia Marche - Ospedale Regionale Torrette, Torrette di Ancona, Ancona - Ospedale I.N.R.C.A., Ancona - Azienda Ospedaliera S. Salvatore, Pesaro 289

292 Lazio, Basilicata e Campania - Ospedale Polispecializzato, Anzio, Roma - Ospedale Fatebenefratelli, Roma - Ospedale Pediatrico Bambino Gesù, Roma - Policlinico Gemelli, Roma - Ospedale Policlinico Umberto I, Roma - Ospedale San Camillo Forlanini, Roma - Università Cattolica del Sacro Cuore, Roma - Dipartimento di Biopatologia Umana, Università La Sapienza, Roma - Ospedale Grande degli Infermi, Viterbo - Ospedale Riuniti, Matera - Azienda Ospedaliera Ospedale Civile, Caserta (NA) - Università Federico II di Napoli, Cattedra di Nefrologia, Napoli - Università di Napoli, Policlinico Nuovo, Napoli - Azienda Ospedaliera S. G. di Dio e Ruggi d Aragona, Salerno Abruzzo - Ospedale G. Bernabeo, Ortona, Chieti - Presidio Ospedaliero San Massimo, Penne (PE) - Presidio Ospedaliero "G.Mazzini", Teramo Puglia, Calabria, Sicilia e Sardegna - Ospedale Regionale Miulli, Acquaviva delle Fonti, Bari - Ospedale Pediatrico Giovanni XXIII, Bari - Ospedale Policlinico, Bari - Azienda Ospedaliera V.Fazzi, Lecce - Ospedale Casa Sollievo dalla Sofferenza, S.Giovanni Rotondo (FG) - Presidio Ospedaliero di Martina Franca, Martina Franca, Taranto - A.U.S.L. TA/1 - Presidio Ospedaliero, Taranto - Azienda Ospedaliera Ospedale Pugliese Ciaccio, Catanzaro - Ospedale dell'annunziata, Cosenza - Centro di Fisiologia Clinica del CNR, Divisione di Nefrologia, Reggio Calabria - Azienda Ospedaliera Bianchi-Melacrino-Morelli, Reggio Calabria - Ospedale N. Giannettasio, Rossano Calabro, Cosenza - Nuovo Presidio Ospedaliero, Acireale, Catania - Azienda Ospedaliera "Ferrarotto", Catania - Ospedale Zonale Maggiore, Modica (RG) - Ospedale Civico, Palermo - Ospedale V. Cervello, Palermo - Azienda Ospedaliera "Umberto I", Siracusa - Azienda Sanitaria G. Brotzu, Ospedale San Michele, Cagliari - Istituto di Clinica e Biologia dell Età Evolutiva, Cagliari - Ospedale A. Segni, Ozieri, Sassari - Ospedale SS. Annunziata, Sassari - Istituto di Patologia Speciale Medica dell'università degli Studi di Sassari - Ospedale Policlinico, Sassari Umbria - Azienda Ospedaliera di Perugia, Perugia 290

293 INTERNATIONAL COLLABORATIONS - University Medical Center, Ljubljana Slovenia - University Hospital Ziekenhuius, Edegem Antwerpen, Belgio - Clinique de Nephrologie-Dialyse Chu Brugmann, Bruxelles, Belgio - University Ziekenhuius Gent, Gent, Belgio - U.Z. Gasthuisberg, Leuven, Belgio - General Hospital Maria Middelares, Sint Niklaas, Belgio - University of Groningen, AV Groningen, Olanda - Academisch Ziekenhuis, Maastricht, Olanda - Thracian University, Stara Zagora, Bulgaria - The Birmingham Children's Hospital, Birmingham, Inghilterra - Guy s Hospital, London, Inghilterra - Manchester Children's Hospital, Manchester, Inghilterra - Nottingham City Hospital, Nottingham, Inghilterra - Aalborg Hospital, Aalborg, Denmark - Nephrological Department, University of Copenaghen, Copenaghen, Danimarca - Steno Diabetes Center, Gentofte, Danimarca - Department of Nephrology, Odense University Hospital, Odense, Danimarca - Department of Nephrology, Sahlgrenska University Hospital, Goteborg, Svezia - Ospedale San Giovanni, Bellinzona Svizzera - Department of Nephrology, University of Wien, Wien, Austria - Carl Thiem Klinikum, Cottbus, Germania - Klinikum der Johann Wolfgang, Frankfurt am Main, Germania - Arbeitsgruppe fyr Biomolekulare Medizin, Hamburg, Germania - Univeristatklinik Heidelberg, Heidelberg, Germania - Medizinische Klinik, Mannheim, Germania - Luitpold Krankenhaus Med. Universitatklinik/Dialyse, Wurzburg, Germania - Hospital Ntra Sra. de Sonsoles, Avila, Spagna - Hospitalet de Llobregat, Institut Català de la Salut, Barcellona, Spagna - Fundacion Jimenez Diaz, Madrid, Spagna - Hospital Clinico Martin Logas, Madrid, Spagna - Hospital 12 de Octubre, Madrid, Spagna - Hospital Gregorio Maranon, Madrid, Spagna - Hospital La Paz, Madrid, Spagna - Hospital Puerta de Hierro, Madrid, Spagna - Hospital Ramon y Cajal, Madrid, Spagna - Hospital Severo Ochoa, Leganes, Madrid, Spagna - Hospital Universitario de Tarragona Joan XXIII, Tarragona, Spagna - Hospital Garcia de Orta, Almada, Portogallo - Brigham & Women's Hospital, Boston, USA - Hennepin County Medical Center, Minneapolis, USA - SIU School of Medicine, Springfield, USA - The Toronto Hospital, Toronto, Canada 291

294 - INCUCAI, Buenos Aires, Argentina - Hospital Italiano de Buenos Aires, Buenos Aires, Argentina - Hospital Regional de Valdivia, Valdivia, Cile - Soroka Medical Center, Beer Sheva, Israele EDITORIAL BOARD MEMBERSHIP Journal of the American Society of Nephrology (Piero Ruggenenti) Journal of Nephrology (Piero Ruggenenti) Current Diabetes Reviews (Piero Ruggenenti) Clinical Journal of the American Society of Nephrology (Piero Ruggenenti) Current Pharmacology Reviews (Dario Cattaneo) PEER REVIEW ACTIVITIES American Journal of Physiology-Renal Physiology American Journal of Kidney Diseases American Journal of Transplantation British Journal of Haematology Journal of the American Society of Nephrology (JASN) Clinica Chmica ACTA Clinical Biochemistry Clinical Chemistry Clinical Journal of the American Society of Nephrology (CJASN) Clinical Pharmacology & Therapeutics Current Clinical Pharmacology Current Diabetes Reviews Diabetes Diabetic Medicine Drugs Expert Opinion on Pharmacotherapy Future Medicine Hypertension Journal of Clinical Investigation Journal of Chromatographic Sciences Journal of Clinical Pharmacology Journal of Immunology Journal of Nephrology Journal of PostGraduate Medicine Kidney International Nature Medicine Lancet 292

295 Lupus New England Journal of Medicine Nephrology Dialysis and Transplantation Pediatric Nephrology Pediatric Transplantation Plos Talanta Therapy Transplant Immunology Transplantation WHO Bullettin PARTICIPATION IN EVENTS IN WHICH THE DEPARTMENT WAS INVOLVED C-440T/T-331C polymorphisms in the UGT-1A9 gene affect the pharmacokinetics of mycophenolic acid in renal transplant recipients. All interno dell American Transplant Congress, San Francisco, May, Pharmacokinetics of mycophenolic acid after the conversion from mycophenolate sodium to the mofetil formulation in kidney transplant recipients. All interno dell IATDMCT Meeting, Nizza, September Blood cell count and lipid lowering therapy affect sirolimus exposure in kidney transplant recipients on calcineurin inhibitor-free regimen. All interno dell IATDMCT Meeting, Nizza, September Farmacocinetica degli immunosoppressori: update su acido micofenolico. All interno del corso di aggiornamento Roche, Isola di Capo Rizzuto, 19 September Statine, dislipidemia e nuovi ipolipemizzanti. All interno del convegno: Nefropatie croniche e complicanze cardiovascolari. Ranica 1 December Conference Clinical management of hyperglicemia and metabolic syndrome, Treviglio, Italy, 17 November 2007 Tenth International Conference on Endothelin, Bergamo, Italy, September 2007 International Workshop Endocannabinoids in Endocrinology, Metabolism and Cardiovascular Disease, Padua, Italy, July, Retraining course of BLS-D BASIC LIFE SUPPORT - EARLY DEFIBRILATION. 28 March 2007 Investigator meeting Clinical trial PLANET, Barcelona, March, Society for Clinical trials, Montreal Canada, May Progression of chronic renal disease The state of the art and an update 10 th International Symposium of Nephrology at Montecatini Montecatini Terme, Italy, March 22-24, 2007 Roche Aleglitazar Investigator Meeting, Istanbul, Turkey, April 17-20,

296 Il laboratorio in nefrologia - 5 Incontro Nefrologico a Viterbo, 8 June 2007 VAM Almirall Vascular Disease: A multidisciplinary approach, Nephroprotection, Barcellona, January 26 th -27 th, Aspreva s Renal Strategy Meeting, Amsterdam, February 23 rd, CME Course. Proteinuria in renal transplantation: Pathophysiology, Diagnosis, Treatment, Post transplant HUS: new ways to prevention and treatment, Firenze, March 21 st -22 nd, XLIV ERA-EDTA Congress, Literature Update Diabetic and non-diabetic progressive nephropathies, Barcellona, June 21-24, Congresso Nazionale SIN, Il rene policistico dell adulto: stiamo uscendo dal tunnel?, Fiera del Levante Bari, October 7-10, Cardio Kidney Diabetes (CKD) Global Consensus Conference, Chicago, IL, October 17-19, Corso di Aggiornamento in Nefrologia e Metodiche Dialitiche, Role and Organization of Remission Clinics, Milano, 6-9, 2007 GRANTS AND CONTRACTS AIFA (Agenzia Italiana del Farmaco) PKD Foundation Abbott GmbH & Co. Astrazeneca SPA ACRAF Spa (Aziende Chimiche Riunite Angelini Francesco) Chiesi Farmaceutici Dompè Spa Farmaceutici Damor SpA King Pharmaceuticals Inc. Novartis Farma Roche SpA Sanofi-Aventis Spa Sigma Tau Spa Solvay Pharmaceuticals Speedel Pharma Ltd 294

297 SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Noris M, Casiraghi F, Todeschini M, Cravedi P, Cugini D, Monteferrante G, Aiello S, Cassis L, Gotti E, Gaspari F, Cattaneo D, Perico N, Remuzzi G. Lymphopenia and calcineurin signalling expand T regulatory cells in human recipients of organ transplantation. JASN 2007, 18: Cattaneo D. Does IgA nephropathy affect long-term graft outcome after kidney transplantation? J PostGrad Med, 2007, 53:84. Corna D, Sangalli F, Cattaneo D, Carrara F, Gaspari F, Remuzzi A, Zoja C, Benigni A, Perico N, Remuzzi G. Effects of rosuvastatin on glomerular capillary size-selectivity function in rats with renal mass ablation. Am J Nephrol 2007, 27: Baldelli S, Merlini S, Cattaneo D, Nicastri A, Bartolini B, Perico N, Remuzzi G. C-440T/T-331C polymorphisms in the UGT-1A9 gene affect the pharmacokinetics of mycophenolic acid in renal transplant recipients. Pharmacogenomics, 2007, 8: Carlucci F, Anzini M, Rovini M, Cattaneo D, S. Merlini S, Tabucchi A. Development of a capillary electrophoresis method for the determination of mycophenolic acid in human plasma: a comparison with HPLC. Electrophoresis 2007, 28: Cattaneo D, Cortinovis M, Baldelli S, Bitto A, Gotti E, Remuzzi G, Perico N. Pharmacokinetics of mycophenolate sodium and comparison with the mofetil formulation in stable kidney transplant recipients. CJASN 2007, 2: Ripamonti D, Cattaneo D, Airoldi M, Frigerio L, Bertuletti P, Ruggeri M, Suter F, Maggiolo F. Atazanavir plus lowdose ritonavir in pregnancy: pharmacokinetics and placental transfer. AIDS 2007; 21: Cravedi P, Ruggenenti P, Remuzzi G. Intensified inhibition of renin-angiotensin system: a way to improve renal protection? Curr Hypertens Rep. 2007;9(5): Ruggenenti P, Cravedi P, Remuzzi G. Latest treatment strategies for membranous nephropathy. Expert Opin Pharmacother. 2007;8(18): Ruggenenti P, Perico N, Gotti E, Cravedi P, D'Agati V, Gagliardini E, Abbate M, Gaspari F, Cattaneo D, Noris M, Casiraghi F, Todeschini M, Cugini D, Conti S, Remuzzi G. Sirolimus versus cyclosporine therapy increases circulating regulatory T cells, but does not protect renal transplant patients given alemtuzumab induction from chronic allograft injury. Transplantation. 2007;84(8): Cravedi P, Noris M, Remuzzi G. Report of the first World Transplant Congress. Clin J Am Soc Nephrol. 2007;2(2): Cravedi P, Ruggenenti P, Sghirlanzoni MC, Remuzzi G. Titrating rituximab to circulating B cells to optimize lymphocytolytic therapy in idiopathic membranous nephropathy. Clin J Am Soc Nephrol. 2007;2(5): Remuzzi G, Cravedi P, Costantini M, Lesti M, Ganeva M, Gherardi G, Ene-Iordache B, Gotti E, Donati D, Salvadori M, Sandrini S, Segoloni G, Federico S, Rigotti P, Sparacino V, Ruggenenti P. Mycophenolate mofetil versus azathioprine for prevention of chronic allograft dysfunction in renal transplantation: the MYSS follow-up randomized, controlled clinical trial. J Am Soc Nephrol. 2007;18(6): Cravedi P, Ruggenenti P, Remuzzi G. Does remission of renal disease associated with antihypertensive treatment exist? Curr Hypertens Rep. 2007;9(2): Sanna-Cherchi S, Carnevali ML, Martorana D, Cravedi P, Maggiore U, Alinovi R, Bovino A, Mattei S, Orlandini G, Gatti R, Savi M, Sado Y, Neri TM, Allegri L. Alterations of type IV collagen alpha chains in patients with chronic acquired glomerulopathies: mrna levels, protein expression and urinary loss. Am J Nephrol. 2007;27(2): Noris M, Casiraghi F, Todeschini M, Cravedi P, Cugini D, Monteferrante G, Aiello S, Cassis L, Gotti E, Gaspari F, Cattaneo D, Perico N, Remuzzi G. Regulatory T cells and T cell depletion: role of immunosuppressive drugs. J Am Soc Nephrol. 2007;18(3):

298 Cravedi P, Ruggenenti P, Remuzzi G. Kidney Failure Stabilizes After An Increase over 2 Decades. EDTNA/ERCA Journal M. Alpa, B. Ferrero, R. Cavallo, A. Perna, C. Naretto, M. Gennaro, D. Di Simone, L. Bellizia, M. Mansouri, D. Rossi, V. Modena, O. Giachino, L.M. Sena, D. Roccatello. Anti-GM1 and anti-sulfatide antibodies in patients with systemic lupus erythematosus, Sjogren sindrome, mixed cryoglobulinemia and idiopathic systemic vasculitis. Clin and Experimental Rheumatology 2007, 25(4): A. Perna, G.A. Giuliano, A. Schieppati, M.Costantini, M. Ganeva, E. Daina, R. Stevanov, G. Remuzzi. Controlled trials in rare diseases: how many? How informative? Adequate? 28 th Annual Meeting of the Society for Clinical Trials, Montreal, Canada, May 20-23, RESEARCH ACTIVITIES Laboratory of Biostatistics Mycophenolate Steroid Sparing Study (MYSS) Follow-Up extension study: final analyses MYSS study results did not show any significant difference between Mycophenolate Mofetil (MMF) and Azathioprine (AZA), combined with Cyclosporine Neoral and steroids, in reducing acute rejection in cadaveric kidney allograft recipients. No difference was found even within patients who, after 6 months from transplant, progressively tapered and interrupted steroid treatment. Due to the relatively limited follow-up, however, the above study could not assess the effects of MMF and AZA on the onset and progression of chronic allograft dysfunction, a syndrome of proteinuria and worsening renal function with progressive nephron loss and scarring of the graft (chronic allograft nephropathy). This is a key issue since, after recipient death, chronic allograft dysfunction represents the major cause of graft loss in the long-tem. Moreover, results from registry analyses showed that continued treatment with MMF versus AZA was associated with a protective effect against renal function deterioration beyond 1 year after transplantation and superior graft survival at 4 years. Despite the limitations of the retrospective design of the above analyses, these data further limited the possibility to use results of the MYSS trial to change the practice of most transplant centres to regard MMF as a key component of immunosuppressive drug regimens based on ciclosporine microemulsion. To address this issue, we designed the MYSS follow-up study. This was an extension of the MYSS study, prospectively comparing long-term outcomes of the two cohorts of MYSS patients in the setting of a similar immunosuppressive regimen based on the microemulsion Neoral, according to their original randomization to MMF or AZA. Results showed that long-term outcomes in the two groups were comparable, independently from residual immunosuppression including steroid or not. Thus, in kidney transplantation, the long-term risk/benefit profile of MMF and AZA therapy in combination with cyclosporine Neoral is similar. In view of the cost, standard immunosuppression regimens for kidney transplantation should perhaps include AZA rather than MMF. Remission Clinic Program: final analyses From January 1999 to November 2004, patients referred to the Unit of Nephrology of the Azienda Ospedaliera, Ospedali Riuniti di Bergamo who had a 24 h urinary protein excretion rate of 3 grams or more for at least six months entered the Remission Clinic program. The program included patients who were treated and monitored according to a sequential, stepwise, multimodal protocol titrated to urinary protein excretion. After a period of up-titration during 296

299 the first month, patients were maintained on ramipril (dose range: mg/day) and losartan (dose range: mg/day) doses allowed as deemed appropriate according to blood pressure control and tolerability. At 3 months after inclusion into the Remission Clinic program, those patients who had an heart rate > 60 beats/min and were not receiving a beta-blocker for a specific indication were prescribed a fixed dose (80 mg/day) of a ndccb (verapamil) that, if tolerated, was up-titrated to 120 mg/day. Three months later, a fixed dose (10 mg/day) of a statin (atorvastatin) was prescribed and, if tolerated, was up-titrated to 20 mg/day. Then patients were seen every 3-6 months up to study end. Stopping rules for safety/tolerability reasons were applied. Results of the 56 consecutive patients included in the Remission Clinc program were compared with those of a matched cohort of 56 proteinuric patients who received ACE inhibitor therapy alone. The primary target of treatment was twenty-four hour urinary protein excretion rate. Three categories of response to treatment were a priori defined on the basis of 24 h urinary protein excretion achieved during the follow-up period: 1. Residual 24 h proteinuria in clinical range (i.e. persistently more than 1.0 g); 2. Reduction of 24 h proteinuria to sub-clinical range (1.0 g or less, but more than 0.3 g in 2 consecutive visits); 3. Reduction of 24 h proteinuria to normal range (0.3 g or less in 2 consecutive visits). The primary efficacy variable was the rate of estimated GFR decline (egfr). Hard end-points were all-cause and cardiovascular deaths, non-fatal cardiovascular events and ESRD, considered either individually, either as a composite end point including all the events Preliminary data showed that the Remission Clin approach was more effective in reducing proteinuria than a therapy based only on ACE inhibitor. Reduction of proteinuria translated into improved renal function and decreased cardiovascular events. Preprocessing of hospital clinical data for comparing different immunosuppressive therapies in renal transplanted patients. Since August 2005 the Unit of Nephrology of the Azienda Ospedaliera Ospedali Riuniti di Bergamo outlined an immunosuppressive regimen based on low doses of Rabbit Anti-human Thymocyte Globulin (RATG) in association to basiliximab. This induction therapy attempted to minimize or eliminate steroids, with aim to reduce acute rejections. In order to facilitate statistical evaluation of acute rejection, of adverse drug reactions due to immunosuppressive therapies and of renal and patient survival hospital clinical data were preprocessed, extracted and elaborated by means of dedicated software. For data retrieval we used a relational database, Microsoft Access, where the records of all subjects referring to the Unit of Nephrology are stored.. An ad hoc program coding previous and concomitant diseases was built up. By means of SQL and Visual Basic we facilitated the choice and description of the disease within a predefined list. For previously archived data, an algorithm for an automatic research of key words was implemented, adding the identified code to the corresponding diseases. Finally demographic and clinical data at baseline and on follow up were extracted for data analysis. For follow up visits an ad hoc algorithm searching the nearest visit to a pre-defined time window was implemented. The above data were extracted and subsequently imported in SAS System and submitted to statistical analysis. We compared the outcomes of kidney transplant patients (n=40) who received combined induction with low-dose RATG plus basiliximab with those of a matched-cohort (n=40) of patients who did not receive induction therapy. Beside induction, the first cohort of patients received maintenance immunosuppression with low-dose mycophenolate mofetil and cyclosporine. Methylprednisolone was infused on day 0, 1 and 2 post transplant. Thereafter, patients were free of steroids. Reference patients received standard-dose MMF and CsA In this group, steroids were progressively tapered and withdrawn from month 6 to month 9 after transplantation. Preliminary results showed that the induction strategy with low-dose RATG combined with basiliximab allowed to prevent acute rejection without steroids, and to reduce the doses of maintenance immunosuppression with MMF and CsA. Steroid avoidance improved the blood 297

300 pressure levels and the lipidic profile. Moreover, the use of lower than conventional doses of CsA in patients who received combined induction was associated with a better renal function BENEDICT Phase B: final analyses Phase B of the multicenter double-blind, randomized Bergamo Nephrologic Diabetes Complications Trial (BENEDICT) assessed the effects of angiotensin-converting-enzyme (ACE) inhibitors alone or combined to non-dihydropyridine calcium-channel blockers on regression to normoalbuminuria or progression to macroalbuminuria in type 2 diabetics with microalbuminuria, and evaluated whether these effects were modulated by the ACE insertion(i)/deletion(d) polymorphism. Two-hundred-eighty-one subjects were randomized to receive 4.5 years of treatment with trandolapril (2 mg per day) plus verapamil (180 mg per day) or trandolapril alone (2 mg per day). Target blood pressure was 120/80 mmhg. Main efficacy variables were regression to normoalbuminuria (overnight albuminuria <20 micrograms per minute at two consecutive visits). or progression to macroalbuminuria (overnight albuminuria 200 micrograms per minute at two consecutive visits). In the 2007 we completed data collection and data analyses. A randomized, prospective, double-blind study to evaluate the effects on lipid profile of combined ezetimibe and simvastatin therapy as compared to simvastatin alone in people with type 2 diabetes: final analyses Ezetimibe is the first number of a class of highly selective cholesterol absorption inhibitors that prevent the absorption of cholesterol by blocking the passage across the wall of the small intestine, without affecting absorption of other fat-soluble nutrients. In a prospective clinical trial we studied 108 normoalbuminuric type 2 diabetic patients over a period of 16 weeks that had 4-week wash-out, took simvastatine (40 mg/day) during the run-in phase (8 weeks) and were randomized into 2 groups of a combined therapy with ezetimibe (10 mg/day) or a placebo (simvastatine alone) for 8 weeks. The primary aim was to compare the effect of the combined treatment (simvastatine plus ezetimibe, EZE) versus simvastatine alone (simvastatine plus placebo, PLA) in reducing the low-density (LDL) cholesterol. As secondary aims, we studied the effect on the total cholesterol, apolipoprotein A1 and B and triglycerides as well as the safety profiles of above treatments. We further went to explore the hypothesis of whether the amelioration of dyslipidaemia may also result in a reduction of urinary albumin excretion rate (UAER, μg/min). In the 2007 we completed data collection and data analyses. Predicting individual risk for end-stage kidney disease: the Okinawa Registry Intensified monitoring, prevention and treatment in subjects at risk of end-stage kidney disease (ESKD) based on readily measurable characteristics may help to prevent kidney disease even in low risk populations. We explored the possibility to quantify an individual Screening Renal Risk Index (SRRI) for ESKD by using gender, age, body mass index, dipstick urinary protein (UP) and mean arterial pressure (MAP) in screens of the Okinawa database (Japan), including 420 subjects who progressed to ESKD over the study period. As a first step, in the 70% random training set (74060 screens), the relative risk (RR) over the 18-year study period was defined and cut-off levels for increased probability of developing ESKD for each of the best independent continuous predictors were determined (e.g., UP 1+, MAP 99.3 mmhg). At the second step, in the remaining 30% test set (32111 screens) SRRI for ESKD was separately defined by gender using a decision-tree analysis with Bayes' revision. In the 2007 we completed data analysis. 298

301 Survey on the quality of published randomized controlled trials in rare diseases We conducted a descriptive survey in 71 rare diseases with MESH term selected from a list of 1608 orphan indications in order to evaluate the main characteristics and quality of randomized controlled trials (RCTs) in this neglected area. Through PubMed ( ) we found titles, retrieving 351 abstracts of RCTs, from which we selected 168 full published articles with parallel group design. In the 2007 we completed data collection and data analysis. Laboratory of Clinical Chemistry Reliability of neutrophil gelatinase-associated lipocalin (NGAL) as an early marker in predicting acute renal dysfunction in patients with solid organ cancer given cisplatin as chemotherapeutic agent Acute renal failure represents a very important and potentially devastating disorder in clinical medicine. Despite substantial technical improvements in treatments, mortality and morbidity associated with acute renal failure remain dismally high. Ischemic and nephrotoxic insults to the kidney are the leading cause of acute renal failure and most often manifest as acute tubular necrosis. While several anti-neoplastic agents frequently exhibit nephrotoxicity, the platinum derivatives are among the most frequent compounds leading to renal injury. Since its introduction into clinical trials, cisplatin has had a major impact in cancer medicine, changing the course of therapeutic management of several tumors, such as those of ovary, testes, and the head and neck. Unfortunately, approximately 25-35% of patients develop evidence of nephrotoxicity following an initial dose of cisplatin. Overall these findings indicate that there is a pressing need for way to protect the kidney while administering effective chemotherapeutic agents such as cisplatin. Unfortunately, creatinine is an unreliable indicator during acute changes in kidney function. Among compounds that might serve as a novel biomarkers for the initiation phase of acute renal failure, neutrophil gelatinase-associated lipocalin (NGAL) has been identified as one of the most strikingly upregulated genes and overexpressed proteins in the kidney after ischemia. Markedly increased NGAL concentrations were easily detected in urine early after renal ischemia in mouse and rat models. Recent studies have demonstrated that NGAL is a useful early predictor of acute renal failure also in humans. Thus, it has been shown that the concentration of NGAL in urine and serum is strikingly raised in children with acute renal failure after cardiopulmonary bypass. Interestingly, urine and serum concentrations of NGAL markedly increased 2 h after surgery. Taken altogether these findings indicate that urinary and/or serum NGAL concentration monitoring may represent a sensitive, specific, and highly predictive early biomarker for acute renal failure. So far, however, no data are available on the reliability of urine and/or serum NGAL to predict the development of acute renal dysfunction in cancer patients receiving cisplatin treatment. A study in 46 adult patients with solid tumors is in progress and is aimed to evaluate the suitability of early changes increase in urinary and/or serum NGAL concentrations (determined by ELISA assay) soon after cisplatin infusion, to predict development of subsequent acute renal dysfunction. The study is also aimed to determine, by means of receiver-operating characteristics curve (ROC) analysis, the accuracy of NGAL urinary and/or serum concentrations to predict acute renal injury; to define a cut-off level of NGAL in both urine and serum samples to predict subsequent acute renal dysfunction; and to evaluate a possible correlation between urinary and/or serum NGAL concentrations and the degree of renal function impairment, as the peak of serum creatinine concentration and the nadir of GFR estimated by the Calvert equation. 299

302 Comparison between different analytical methods for albumin determination in urine Although the limit of urinary albumin excretion rate (UAE) of 20 g/min was chosen as a definition of microalbuminuria because 95% of normal individuals had excretion rates below that limit, it is recognized that the risk of cardiovascular events and of progression to overt nephropathy is elevated also in subjects in the high normal range (10 20 g/min). Mounting evidence indicates a continuous relationship between UAE and risk since epidemiological data suggest an inconsistency with the concept of threshold level. Thus, both reliability and sensitivity of the chosen method for urinary albumin determination may play an important role in patients monitoring. Immunochemical assays are the most widely used methods for microalbuminuria measurement. Recent works have demonstrated that intact albumin in urine may exist in two forms, immunoreactive and immuno-unreactive, but only the immunoreactive form can be detected by conventional immunochemical methods. With the purpose of measuring both forms of intact albumin, we recently developed a high performance size-exclusion liquid chromatography (HPLC) method which is capable to determine both immunoreactive and immuno-unreactive albumin moieties. On the other hand, HPLC methods to measure albumin, are not commonly available in clinical laboratories. Other immunochemical methods that use less demanding instruments as been recently proposed as point-of-care testing (POCT) to assess albumin loss resulting from diabetes or other chronic kidney disease states. These instruments offer rapid results and are also good candidates to become the instrument of choice for prevention programs in developing countries due to no need of technical assistance, minimal performance required for the personnel and relatively low cost per test. In a study aimed to evaluate the performance of different analytical methods, we compared the determination of albumin urinary levels in diabetic patients as measured by conventional (nephelometric) and the chromatographic method developed by our laboratory as well as by HemoCue 201, a commercially available POCT instrument, which performs a highly specific turbidymetric assay for human albumin excreted in the urine. Findings indicate that especially for albuminuria <50 g/min, nephelometry underestimated UAE as compared to HPLC. A highly significant correlation was found between albumin concentration determined by conventional nephelometric assay and those determined by HemoCue 201. Data analysis revealed that there was a good agreement between the immunochemical methods over the entire range of linearity of HemoCue g/ml), that the mean bias between methods was 3.2 g/ml and that there was a tendency to overestimate albumin concentrations with increasing levels of the protein. Ficoll analysis by high performance chromatography Ficoll fractional clearance determination is a reliable tool to assess size-selectivity property of the glomerular capillary barrier in animal models. For this purpose, we used a high performance liquid chromatographic technique for measurement of FITC-labeled Ficoll developed by our laboratory. The developed method requires the injection of as low as 5 l of plasma and urine samples onto a TSK G4000 PWXL column and fluorescence detection to measure FITC-Ficoll. It is fast, reliable, simple and allows the evaluation of the FITC-Ficoll fractional clearance for radii at least up to 80 Angstrom. Due to the rapidity of sample handling and the analytical specificity for FITC-Ficoll it has been used in a study aimed to evaluate the effect of different doses statins given alone or in association with ACE inhibitors on renal hemodynamics, permselective properties of glomerular capillary barrier and tubular function in rats with renal mass reduction. The results have shown that the HPLC method to measure FITC-Ficoll can be considered a reference tool for routine assessment of glomerular size-selective function in animal models. 300

303 Reliability of 13 different GFR prediction equations in type 2 diabetic patients Diabetic nephropathy affects 25-40% of diabetic patients, and diabetes is the leading cause of end-stage renal disease. Evaluation of glomerular filtration rate (GFR) is therefore of critical importance in the clinical management of both clinic and outclinic diabetic patients. However, the measurement of the true GFR either by the renal clearance of the gold standard inulin or by plasma clearance of contrast agents is time consuming, difficult to perform and cannot be easily implemented in clinical daily practice. Urinary creatinine clearance is widely used to measure GFR, but it is inconvenient and sometimes inaccurate due to variable creatinine metabolism and tubular secretion. To circumvent these drawbacks, a number of equation has been developed to provide an estimate of renal function from serum creatinine and demographic characteristics. Despite being widely used, their performance and suitability to monitor renal function in patients with type 2 diabetes have not been assessed so far. In 279 normoalbuminuric patients and 134 microalbuminuric subjects enrolled in the BENEDICT and in the DEMAND study we compared with the gold standard plasma iohexol clearance the performance in monitoring renal function of 13 different GFR equations. Data analysis showed that GFR was markedly underestimated in both groups of patients. The Rule and Ibrahim formulas performed slightly better than the other models with about 50% the estimated GFR within +10% error as compared to the reference iohexol. To better investigate the performance of the 13 prediction equations in dependence of the kidney function, the diabetic patients were stratified according in the following GFR ranges: from 60 to 89 ml/min/1.73m 2, from 90 to 119 ml/min/1.73m 2, and > 120 ml/min/1.73m 2. Almost all the tested model underestimated GFR and this behavior was more evident at higher range of renal function, however both Rule and Ibrahim showed a marked overestimation in the low range of GFR. These findings showed that, irrespectively of albumin excretion rate, none of the 13 GFR models seems to safely substitute for the direct measurement of the renal function by means of an affordable and suitable reference methods, such as iohexol plasma clearance. The development of a new robust GFR prediction equation in type 2 diabetic patients is still needed. Development of a sensitive and specific HPLC method for the determination of arginine in plasma samples of patients with preeclampsia Experimental data strongly suggest that nitric oxide (NO), a potent endothelial-derived vasodilator, might be implicated in gestational vasodilatation. NO is synthesized from the aminoacid L-arginine by a family of enzymes, the NO synthases (NOS). In the normal placenta, adequate concentration of L-arginine selectively orients the endothelial isoform of nitric oxide synthase (ecnos) toward NO, which is vital for the low-resistance placenta circulation and also exerts a facilitating role on cytotrophoblast invasion. In the pre-eclamptic placenta, instead, a lower than normal L-arginine concentration re-directs ecnos toward peroxynitrite which is generated in exuberant amounts at the expense of NO. This favors microvascular damage and impairs cytotrophoblast invasion. To investigate whether increasing L-arginine bioavailability after oral administration of the compound might restore physiological NO production in pre-eclamptic placenta we performed, together with the Obstetrics and Gynecology units of the Brescia and Bergamo hospitals, a pilot study aimed to explore the impact of the above treatment on pregnancy outcome. For this purpose we developed a high performance liquid chromatographic (HPLC) method to accurately determine arginine in both pregnant women with pre-eclampsia and healthy control subjects. The developed HPLC method has a good performance and sensitivity and accurately quantifies arginine in plasma. After a simple plasma proteins precipitation step by means of trichloroacetic 301

304 acid, 10 l are injected onto a 3 cm reversed-phase column. A sodium hydroxide solution is pumped and mixed with the column effluent maintained at 75 C to allow post-colum derivatization of eluted arginine with ninhydrin in mobile phase. The reaction is highly specific for guadininic compounds, such as arginine, and the derivative is easily quantified by means of a fluorescence detector. The simplicity and rapidity of sample handling and the analytical specificity for arginine allowed to accurately evaluate the pharmacokinetic profiles of arginine after oral administration in both pregnant women with pre-eclampsia and in healthy pregnant controls. Laboratory of Drug Development Indications for rituximab therapy in patients with membranous nephropathy Rituximab, a chimeric monoclonal antibody able to deplete CD20 positive B cells effectively reduces proteinuria in patients with idiopathic membranous nephropathy (IMN), but response to treatment may vary from patient to patient. Retrospective analyses of heterogeneous patient populations given different immunosuppressive regimens failed to identify determinants of response. Aim of the present study was to assess the possible determinants of response to rituximab, in order to identify those patients who may benefit the best of this therapy To this purpose, we retrospectically evaluated by multivariate analyses the association between baseline clinical, laboratory and histology covariates and proteinuria reduction achieved after 4 weekly rituximab infusions (375 mg/m 2 ) in IMN patients with proteinuria >=3.5 g/24h while on ACE-inhibition for at least 6 months and no previous remissions. Glomerular and tubulointerstitial (TI) scores at baseline biopsy predicted the outcome. Among glomerular and TI score components, tubular atrophy and interstitial fibrosis were significantly associated with threemonth proteinuria. Urinary protein excretion decreased from 9.1±4.0 g/24h to 4.6±3.5 g/24h (p<0.001) in 8 patients with TI score <1.7, but did not change in 6 with a score >1.7. Nine additional IMN patients were then prospectively allocated to rituximab treatment on the basis of a TI score <1.7. Three-month proteinuria decreased in all patients from 8.9±5.3 g/24h to 4.9±3.9 g/24h (p<0.001) and serum albumin increased from 2.2±0.6 mg/dl to 2.8±0.5 mg/dl (p<0.01). Rituximab achieved B cell depletion in all patients. These results suggest that in IMN patients with nephrotic proteinuria despite ACE inhibition therapy, renal biopsy findings may help predicting response to rituximab and defining selection criteria for randomized trials aimed to assess the risk/benefit profile of B cell target therapy as compared to aspecific immunosuppressants and/or conservative therapy alone. Finding the ideal dose for rituximab therapy in patients with membranous nephropathy Rituximab has been shown to reduce proteinuria and suppress symptoms over at least one year follow-up in IMN patients with heavy (>3g/24h) proteinuria that could not be lowered by ACE inhibitor (ACEi) therapy for at least six months. The rationale for this therapeutic approach is that by eliminating B cells, rituximab may prevent the generation of clones involved in the neosynthesis of autoantibodies. With this approach, however, we observed that B cells disappeared from the circulation just after the first Rituximab administration. Conceivably, this should also translate in a complete inhibition of autoantibodies production. Should this be the case, additional administration of Rituximab would not achieve further effect in term of B cells depletion and autoantibodies inhibition, and would unnecessarily expose the patient to the risk of sensitization and reactions. To address this issue, since December 2005 all patients with nephrotic range proteinuria (in spite of at least 6 months of ACEi therapy) referred to our Nephrology Unit were treated with a schedule of Rituximab titrated to circulating B cells. Rituximab was infused weekly until full 302

305 depletion of B cells was achieved for a maximum of 4 doses. To assess the cost/effectiveness of this novel regimen, we designed a matched-cohort analysis comparing the outcome of 12 new incident patients who received a B cell-driven treatment with that of 24 historical reference patients who were given the standard protocol of four weekly doses of 375 mg/m 2. Only one patient needed a second dose to achieve full CD20 cell depletion. At 1 yr, time course of the components of nephrotic syndrome and the proportion of patients who achieved disease remission (25%) was identical in both groups. Persistent CD20 cell depletion was achieved in all patients. Costs for rituximab treatment and hospitalizations totalled euros ($ ) and 13, euros ($18,170.80) with the B cell-driven and the four-dose protocol, respectively. One patient on standard protocol had a severe adverse reaction at second rituximab dose. Thus, B cell titrated as effectively as standard rituximab treatment achieves B cell depletion and idiopathic membranous nephropathy remission but is fourfold less expensive, allowing for more than 10,000 euros, approximately $13,000 in savings per patient. Thus, avoiding unnecessary reexposure to rituximab is extremely cost-saving and may limit the production of antichimeric antibodies that may increase the risk for adverse reactions and prevent re-treatment of disease recurrences. Pharmacokinetics of mycophenolic acid from mycophenolate sodium and comparison with that from mofetil formulation in kidney transplant recipients The introduction of mycophenolate mofetil (MMF) has improved long-term graft survival after organ transplantation. However the use of this agent may be limited by gastrointestinal and hematological side-effects. To overcome these problems, an enteric-coated formulation of mycophenolate sodium (EC-MPS) has been recently developed. Detailed data on the pharmacokinetics of MPA released from EC-MPS are lacking. To compare the profiles of mycophenolic acid (MPA) derived from the two formulations, we have performed pharmacokinetic studies in 20 kidney transplant recipients given EC-MPS (n=10) or MMF (n=10) as a part of their immunosuppressive regimes. At month 6 and 12 post-surgery, despite no differences in mean MPA Cmax and AUC between the two formulations, aberrant and extremely variable pharmacokinetic curves were found in all patients given EC-MPS. Additionally, these patients presented MPA Tmax ranging from 0 to 480 minutes and doseadjusted MPA trough (C0) 4.7-fold higher compared to patients given MMF. Given the emerging strong support for the clinical outcome benefit of MPA monitoring in transplant setting, the manufacturer should face with all the potential problems related with the enteric coating of EC-MPS that limit the application of therapeutic drug monitoring, before promoting an extensive use of this novel formulation. Results of this research have been presented at the American Transplant Congress 2007 (S. Francisco, May 2007) and published in the Clinical Journal of the American Society of Nephrology. Blood cell count and lipid lowering therapy affect sirolimus exposure in kidney transplant recipients on calcineurin inhibitor-free regimen Sirolimus (SRL) is an immunosuppressive agent characterized by a narrow therapeutic index. Studies in organ transplant recipients given this drug in combination with cyclosporine (CsA) have shown that SRL exposure is extremely variable. However, no data are available on the pharmacokinetics of SRL in calcineurin inhibitor-free protocols. Fifty-five pharmacokinetics profiles were collected from 20 kidney transplant patients given SRL over a 36 months follow-up. None of them were on CsA or tacrolimus. Large interindividual variability in the daily distribution of SRL concentrations was documented. Patients with low blood cell count showed a significant decrease in daily SRL exposure, requiring 35% increments in the daily dose to reach drug concentration targets. At variance, concomitant administration of omega-3 polyunsaturated fatty acids increased SRL AUC 0-24 by 25% 303

306 compared to values found in normolipidemic patients. Altogether, these information can be used to optimize SRL dose-adjustments in the routine clinical practice. These results have been presented at the meeting of the International Association of Therapeutic Drug Monitoring and Clinical Toxicology (Nice, September 2007). Genetics partly explains patient variability of mycophenolic acid pharmacokinetics in renal transplant recipients Mycophenolic acid (MPA) is widely used as immunosuppressive agent in renal transplantation. It is mainly metabolized by uridine diphosphate glucuronosyltransferase 1A9 (UGT1A9) to 7- O-glucuronide (MPAG). The UGT1A9 gene that encodes for the UGT protein is localized on chromosome 2q37 and several single nucleotide polymorphisms (SNPs) in the gene have been described. Moreover, excretion in the bile of MPAG occurs through membrane drug efflux protein, the more relevant being MRP2. This protein is encoded by a gene located on the chromosome 10q24 from whom different SNPs have been so far identified that could also affect the expression of the protein. The present study was designed to study the impact of the polymorphisms of UGT1A9 and to investigate the role of polymorphisms in MRP2 in renal transplant patients MPA pharmacokinetics. Thirty-four patients were enrolled in the pharmacogenetic study. Patients had clinical evaluations and laboratory tests at 6 month after transplantation, as well as the assessment of the complete MPA pharmacokinetic profiles and MPAG measurement. They were genotyped for single nucleotide polymorphism (SNPs) in UGT1A9 C-1252T, T-1887G, C-665T, C-440T, T- 331C, T-275A, T98C and for MRP2 SNPs C-24T and G1249A. Association of polymorphisms with MPA and MPAG pharmacokinetic parameters was studied. Great MPA pharmacokinetic variability was found, confirmed by values of MPA/MPAG metabolic ratio ranging from to among the study patients. In particular, a multimodal frequency distribution was documented that highlighted the presence of at least two different phenotypes.. Significant higher MPA trough levels were found in patients carrier of C- 665T polymorphism. MPA and MPAG AUC were significantly associated with the presence of UGT1A9-440/-331 genotype. The presence of MRP2 promoter C-24T and exon 10 SNPs G1294A did not cause any significant variation in any MPA and MPAG pharmacokinetic parameters. The study has demonstrated a significant impact on MPA pharmacokinetics in renal allograft recipients of different polymorphisms in the promoter region of the UGT1A9 in particular C- 440T/T-331C and C-665T. these results were presented at the American Transplant Congress 2007 (S. Francisco, May 2007) and published in Pharmacogenomics. 304

307 LABORATORY OF CLINICAL EPIDEMIOLOGY STAFF Head Guido BERTOLINI, M.D. Clinical Knowledge Engineering Unit Head Davide LUCIANI, M.D. 305

308 CURRICULA VITAE Guido Bertolini got his Medical degree in 1989 at the University of Bologna, and the specialization in Pharmacological Research in 1993 at the Mario Negri Institute and in Gastroenterology in 1994 at the University of Pavia. He founded and chaired from 1997 to 2000 the School of Clinical Methodology and Quality of Care Improvement at the Ospedali Riuniti di Bergamo and the Istituto di Ricerche Farmacologiche Mario Negri. From 1999 to 2003 he was contract professor at the post-doctoral schools in Anaesthesia and Intensive Care, University of Brescia and Milano; from 2002 to 2005 he has been contract professor of Educational Science at the Faculty of Lettere e Filosofia, University of Bergamo. Current research interests: Clinical Research Methodology, Continuous Quality of Care Assessment and Improvement, Health services research and outcome, Medical decision making, Medical Education. These interests are mainly developed within the fields of Intensive Care Medicine and Rare Diseases. Since 1997 he chairs the GiViTI Coordinating Center for research in intensive care medicine. He has been Head of the Unit of Epidemiology and Education for Clinical Practice at the Mario Negri Institute and since 2001 he is the Head of the Laboratory of Clinical Epidemiology. Since 2001 he is Vice-chairman of the Research Group on Cost-effectiveness, Section on Health Services Research and Outcomes European Society of Intensive Care Medicine and, since 2004, he is President of the Scientific Committee of the Ospedale maggiore in Crema. Selected publications Bertolini G, Rossi C, Anghileri A, Livigni S, Addis A, Poole D. Use of drotrecogin alfa (activated) in Italian intensive care units: the results of a nationwide survey. Intensive Care Med 2007; 33: Rossi C, Simini B, Brazzi L, Rossi G, Radrizzani D, Iapichino G, Bertolini G. Variable costs of ICU patients: a multicenter prospective study. Intensive Care Med. 2006;32: Vanoli M, Daina E, Salvarani C, Sabbadini MG, Rossi C, Bacchiani G, Schieppati A, Baldissera E, Bertolini G. Takayasu's arteritis: A study of 104 Italian patients. Arthritis Rheum 2005; 15;53: IF: Malacarne P, Rossi C, Bertolini G; GiViTI Group. Antibiotic usage in intensive care units: a pharmaco-epidemiological multicentre study. J Antimicrob Chemother Jul; 54:221-4 Galli M, Luciani D, Bertolini G, Barbui T. Anti-beta 2-glycoprotein I, antiprothrombin antibodies, and the risk of thrombosis in the antiphospholipid syndrome. Blood 2003;102: Simini B, Bertolini G. Should same anesthetist do preoperative anesthetic visit and give subsequent anesthetic? Questionnaire survey of anesthetists. BMJ 2003; 327: Davide Luciani got his Medical Degree at the University of Bologna in 1995, and the Diploma in "Tropical Medicine and Hygiene" at the University of Liverpool in In 2001, he spent one year at the Department of Statistical Science (University College London). Bayesian probabilistic applications, decision theory and the graphical approach to pathophysiological modeling represent his main interests. Within his research activity, these skills are meant as the main methodological ingredients in the formalization of clinical reasoning, in order to improve its effectiveness and to exploit its educational value. Since 2005 he is responsible of the Unit of Clinical Knowledge Engineering. Selected publications Luciani D, Cavuto S, Antiga L, Miniati M, Monti S, Pistolesi M, Bertolini G Bayes pulmonary embolism assisted diagnosis: a new expert system for clinical use Emerg Med J 2007 ; 24 : M.Cesana, R.Cerutti, E.Grossi, E.Fagiuoli, M.Stabilini, F.Stella, D Luciani.Bayesian Data Mining Techniques: The Evidence Provided by Signals Detected in Single-Company Spontaneous Reports Databases. Drug Information Journal, vol. 41, pp , 2007 Latronico N, Bertolini G, Guarneri B, Botteri M, Peli E, Andreoletti S, Bera P, Luciani D, Nardella A, Vittorielli E, Simini B, Candiani A Simplified electrophysiological evaluation of peripheral nerves in critically ill patients: the Italian multi-centre CRIMYNE study Crit Care. 2007;11(1):R11. Bertolini G, Luciani D, Biolo G Immunonutrition in septic patients: A philosophical view of the current situation Clin Nutr 2007 ; 26 : Luciani D, Marchesi M, Bertolini G. The role of Bayesian Network in the diagnosis of pulmonary embolism. J Thromb Haemost 2003; 1: Galli M, Luciani D, Bertolini G, Barbui T. Anti-beta 2-glycoprotein I, antiprothrombin antibodies, and the risk of thrombosis in the antiphospholipid syndrome. Blood,2003 Oct 15;102(8): Haemostasis, 2003 Apr;1(4):

309 INTRODUCTION TO THE LABORATORY'S ACTIVITIES The general aim of the Laboratory of Clinical Epidemiology is to contribute to the improvement of health care in different medical fields. The guiding principles are mainly two: to help physicians use the available knowledge and resources at their best; to play a role in the growth of useful knowledge for clinical practice. The Laboratory operates particularly in the field of Intensive Care Medicine and Rare Diseases. Within the Laboratory, the Unit of Clinical Knowledge Engineering aims to bring the value of clinical reasoning out, through the implementation of probabilistic models for its formalization, thus favoring the evaluation and the continuous improvement of complex clinical activities. FINDINGS/MAIN RESULTS The participation to the Margherita project on the quality of care assessment in intensive care unit (ICU), which started in 2002, has further increased during this year. The patients recruited in 2007 were more than This allowed the development of a highly accurate model for the prediction of hospital mortality in critically ill patients. Each participating ICU has received both a general and an individual report. In the latter, in addition to a detailed description of the case-mix, we supplied each ICU with the comparison of its own performance with that of others. We monitored, on behalf of the Italian Ministry of Health, the clinical use of Drotrecogin alfa (activated) in ICU. This is a new drug for critically ill patients with severe sepsis or septic shock. In the framework of a general appropriate utilization, we found some problems and, thus, the possibility to further increase the quality of care in this field. We provided each ICU with a detailed report on the results of the project. We realized one of the biggest continuous infection surveillance program in ICU. We identified the major problems of each unit in the diagnosis and treatment of infections. We developed the first version of an electronic clinical record for the ICU: a clinical record shared by many units that will allow to compare their process of care in the framework of a continuous quality of care improvement We realized a survey on end-of-life decisions in ICU which involved 90 centres. Results will be available early in We further developed an expert system to assist physicians in the complex diagnosis of Pulmonary Embolism. The new version showed excellent validity. We eventually contributed to the set-up of a European network for the study of patients with paraneoplastic syndrome. NATIONAL COLLABORATIONS Dipartimento di Neurologia, Ospedale Regionale S. Maria dei Battuti Cà Foncello, Treviso. Dipartimento di Specialità Chirurgiche, Scienze Radiologiche e Medico Forensi, Cattedra di Anestesia dell Università degli Studi di Brescia. Servizio Anestesia e Rianimazione, Osp. San Giovanni Bosco, Torino. Divisione di Medicina Interna, Università di Trieste. Divisione di Fisiopatologia Respiratoria, Università di Firenze. Istituto Mediterraneo per i Trapianti e Terapie ad Alta Specializzazione, Palermo. 307

310 Istituto di Anestesia e Rianimazione, Università di Milano. Servizio Anestesia e Rianimazione, Osp. Civile, Belluno. INTERNATIONAL COLLABORATIONS Bloomsbury Institute of Intensive Care Medicine, Institute of Biomedical Research, University College London, UK. Department of Statistical Science, University College London, UK. Klink fur Anaesthesiologie und Intensivtherapie, Friedrich-Schiller-Universitat, Jena, Germany Machine Intelligence Group, University of Aalborg, Denmark. American Board of Family Medicine, Kentucky, US EDITORIAL BOARD MEMBERSHIP Ricerca & Pratica (Guido Bertolini) Dedalo. Gestire i sistemi complessi in sanità (Guido Bertolini) British Medical Journal Intensive Care Medicine Criticale Care Medicine Canadian Medical Association Journal Ricerca & Pratica PEER REVIEW ACTIVITIES NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Scientific Committee, azienda ospedaliera Ospedale maggiore di Crema. EVENT ORGANIZATION Educational course, Statistica multivariata per la ricerca biomedica, February-May, Ranica (BG). Workshop, Le decisioni di fine-vita in rianimazione, September 10, Ranica (BG). Workshop, Startup Meeting Compact, September 12, Ranica (BG). Educational course, Utilizzo delle banche dati in medicina, September 21-28, Ranica (BG). Congress, GiViTI Annual Meeting, October 18-20, Pesaro. Educational course, Statistica per medici, November 7-14, Ranica (BG). 308

311 PARTICIPATION IN EVENTS IN WHICH THE LABORATORY WAS INVOLVED Meeting, PNS, Additional Outcomes from the Statistical Analysis, February 24, Treviso Workshop, Come raccogliere i dati in medicina, L esperienza del GiViTI: il progetto Margherita Due, March 16, Firenze Congress, BRAIN06, Hot Topic, April 27, Brescia Congress, SMART, Alternative ai trial controllati randomizzati nella ricerca clinica, May 11, Milano Congress, III Meeting di Formazione e Aggiornamento sugli Interventi in TI, Gli aspetti metodologici della valutazione degli interventi nutrizionali nel paziente critico, May 30, Calampiso Meeting, Antibiotici: attualità, riflessioni, Farmacoepidemiologia degli antibiotici: il casomodello delle terapie intensive, June 19, Seriate Meeting, POR Sardegna Seminario finale misura 5.3, La valutazione della performance della TI: quali dati raccogliere e come utilizzarli, July 30, Oristano Meeting, II Meeting GiViTI Regione Veneto, Il SAPS III e i nuovi progetti GiViTI, Come estrarre i dati dalla propria Terapia Intensiva, Presentazione di Margherita, September 29, Treviso Congress, 60 SIAARTI, Le eterne dicotomie della sperimentazione clinica, October 11, Perugia Workshop, Salute e malattia nell era della tecnica, November 10, Spiazzi di Gromo (BG) Meeting, Farmaci: usarli bene per stare meglio. Presentazione di un progetto integrato tra Ospedale e Territorio, Il ruolo tra formazione ed informazione, November 18, Bergamo Meeting, Registro scompenso cardiaco, La metodologia dei registri epidemiologici, November 25, Palermo Workshop, Formazione degli operatori, Presentazione del software Margherita Tre, December 11-12, Livorno Meeting, Progetto Sorveglianza Infezioni, Lettura dei dati della TI di Cesena, December 12, Cesena 309

312 GRANTS AND CONTRACTS Regione Lombardia Regione Toscana Regione Veneto Regione Piemonte ARS Toscana AstraZeneca Italia Sanofi-Synthelabo Italia Draeger Italia Bellco SpA Ministero della Salute Azienda ULSS 16, Padova - Italia Istituto Mediterraneo per i Trapianti e Terapie ad Alta Specializzazione, Palermo Azienda ASL4 Piemonte Private donation SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Luciani D, Cavuto S, Antiga L, Miniati M, Monti S, Pistolesi M, Bertolini G. Bayes pulmonary embolism assisted diagnosis: a new expert system for clinical use. Emerg Med J 2007; 24: IF: Bertolini G, Rossi C, Anghileri A, Livigni S, Addis A, Poole D. Use of drotrecogin alfa (activated) in Italian intensive care units: the results of a nationwide survey. Intensive Care Med 2007; 33: IF: Latronico N, Bertolini G, Guarneri B, Botteri M, Peli E, Andreoletti S, Bera P, Luciani D, Nardella A, Vittorielli E, Simini B, Candiani A. Simplified electrophysiological evaluation of peripheral nerves in critically ill patients: the Italian multi-centre CRIMYNE study. Crit Care 2007; 11(1): R11. IF: Bertolini G, Luciani D, Biolo G. Immunonutrition in septic patients: A philosophical view of the current situation. Clin Nutr 2007; 26(1): IF: Bertolini G, Rossi C, Anghileri A, Livigni S, Addis A, Poole D. Response to the letter by Williams et al. Intensive Care Med 2007; 33: IF: Iapichino G, Radrizzani D, Rossi C, Pezzi A, Anghileri A, Boffelli S, Giardino M, Mistraletti G, Bertolini G. Proposal of a flexible structural-organizing model for the intensive care units. Minerva Anestesiol 2007; 73: Luca A, Angermayr B, Bertolini G, Koenig F, Vizzini G, Ploner M, Peck-Radosavljevic M, Gridelli B, Bosch J. An integrated MELD model including serum sodium and age improves the prediction of early mortality in patients with cirrhosis. Liver Transpl 2007;13: IF: 310

313 OTHER PRODUCTS PUBLISHED IN 2007 Bertolini G (a cura di) Scelte sulla vita. L esperienza di cura nei reparti di terapia intensiva. Milano: Guerini Studio, Boffelli S, Rossi C, Bertolini G. Progetto Margherita. Promuovere la ricerca e la valutazione in Terapia Intensiva. Rapporto Bergamo: Ed. Sestante, RESEARCH ACTIVITIES Appropriateness in Intensive Care Units The main purpose of these research activities is the assessment and improvement of the quality of care in Italian Intensive Care Units (ICUs). It is a multi-annual project promoted on behalf of GiViTI, a collaborative network composed by half of the Italian ICUs and coordinated by the Laboratory. The main focus is the Margherita project. Its aim is the continuous evaluation of the quality of care and it is based on a free software developed by the Laboratory and distributed to all the ICUs adhering to the GiViTI group. The software has been realized on a modular structure, which enables to easily integrate the basic data collection (the core of Margherita) with the data collection of specific research projects (the petals of Margherita). Additionally, in the current year, a software based on a probabilistic model (bayesian network) covering a large set of clinical variables, has been implemented. Such an activity, lead by the Unit of Clinical Knowledge Engineering, and also involving the Department of Statistical Science (University College London) and the Machine Intelligence Group of the University of Aalborg (Denmark), pursues the realization of tools for the retrospective evaluation of specific treatments in ICU. By means of this model, it is meant to overcome the current limits of an overall evaluation of ICU, likewise it happens when the traditional models for mortality prediction are adopted. Within this activity, a bayesian system to monitor the Standardised Mortality Ratio of the single ICU. This system, which resembles the bayesian pharamcovigilance system currently in use at the WHO and FDA, will identify periods during which some problems occurred, allowing the physicians of the Center to promptly and efficacy react. Studies on Multiple Organ Failure pathological processes The Laboratory of Clinical Epidemiology has lead several investigations to clarify which pathophysiological mechanisms induce multiple organ failure, a condition still burdened with high mortality. Among these, the investigation on the neuromuscular impairment in critical patients (the observational study 'Crimyne'), the study on the impact of enteral feeding, and the new treatments proposed for severe sepsis (Xigris and the removal of inflammation mediators through specific filter applied to circuit of plasma-filtration). The value of a strict glycemic control of critical patients has been recently emphasized, given its connection to a drug like insulin that, in spite of its large availability and low cost, induces a relevant reduction of mortality in ICU. The Unit of Clinical Knowledge Engineering has developed a model based on a differential equations system whose aim is to support the physician in dosing both insulin and glucose infusions, in order to extend the possibility of a strict control even to patients with a high risk of hypoglycaemia. This model represents a precious opportunity to investigate the pathophysiological mechanisms behind the benefits of 311

314 insulin already demonstrated at an empirical level, allowing the explanation of the dynamic behaviour of glycemic fluctuations on the basis of the patient's metabolic profile. The reconstruction of clinical reasoning in the medical practice and education This area represents the main concern of the Unit of Clinical Knowledge Engineering, whose objective is the valorization of clinical reasoning in solving complex clinical problems. The diagnosis of pulmonary embolism still represents a relevant clinical challenge, due to the complexity of the patient's clinical presentation and the variability of diagnostic resources among Centres. In this regards, we are conducting an Italian multicenter study, involving mainly Emergency Units, with the aim of prospectively validating the diagnostic software BayPAD (Bayes Pulmonary embolism Assisted Diagnosis). Such a tool, relying on a probabilistic model covering 72 clinical variables and doing without the need to input all the contemplated observations, would overcome the main reasons which prevented ordinary clinical guidelines to be largely accepted. Moreover, the results of the retrospective validation of the system have been obtained. The Unit started a project for the realization of a software assisting the physician in tracing back the basis of his clinical decisions before the description provided by clinical reports, among those that are typical of particular medical specialty. The software has the double target to create specific applications based on probabilistic models representing complex clinical decision problems, and to involve physicians in their construction. The last target is achievable given the strong analogy between the causal structure of the exploited models (bayesian networks) and the pathophysiological structure of medical knowledge. By this, it will be given the chance to adopt this system within medical training projects, with a special attention to e-learning programs. Clinical Epidemiology of rare diseases and orphan medicine Our purpose is to find out and describe clinical and research problems related either to rare diseases or to neglected aspects of well-known diseases. We also focus on the needs of the patients with rare diseases. A specific project is connected with this research activity: PNS Euronetwork. It is a European project on paraneoplastic neurological syndromes that is financed by the Fifth and Sixth Framework Program of the European Community. Its purposes are various: to develop a network of reference centers for these pathologies all sharing a common database; to organize a sample bank of biological fluids and cerebrospinal liquid to point out the best antibody indicators for the diagnosis and prognosis of these patients; to realize some research projects on the treatment of this syndrome. 312

315 LABORATORY OF COORDINATION OF DIAGNOSIS AND INFORMATION ON RARE DISEASES STAFF Head Arrigo SCHIEPPATI, M.D. Information Centre for Rare Diseases Head Erica DAINA, M.D. 313

316 CURRICULA VITAE Arrigo Schieppati got his degree in Medicine at the University of Milan in 1978 and the specialisation in Medical Nephrology in 1984 at the same University. He performed his training at the Mario Negri Bergamo Laboratories with Dr. Remuzzi, and completed it with stages at the laboratories of prof. Patrono (Catholic University in Rome), prof. John Gordon (Cambridge, GB), and at the Division of Renal Diseases - University of Colorado Medical School, directed by Dr. Schrier (Denver, USA). Since 1982 he works at the Division of Nephrology and Dialysis Riuniti Hospital Bergamo, where he is in charge of Outpatients Clinic and Day Hospital. From 1992 to 1995 he was Head of the Information Center for Rare Diseases and since 1996 he is Head of the Laboratory for Coordination of Information and Diagnosis of Rare Disease at the Clinical Research Center for Rare Diseases Aldo e Cele Daccò of the Mario Negri Institute. Areas of interest: diagnosis and therapy of chronic renal diseases, hypertension and rare kidney diseases. Affiliations: ethical committee Riuniti Hospital - Bergamo; member of the working group of the regional network for rare diseases in Lombardy; scientific committee Bolognini Hospital Seriate (BG); member of the Task Force on Rare Diseases (DG Health and Consumer Protection); International Society of Nephrology; American Society of Nephrology; Editorial Board Journal of Nephrology, Editorial Board of JASN - Nephrology Self Assessment Program. Selected publications Schieppati A, Remuzzi G. Chronic renal diseases as a public health problem: epidemiology, social, and economic implications. Kidney Int Suppl Sep;(98):S7-S10. Remuzzi G, Schieppati A, Boissel JP, Garattini S, Horton R. Independent clinical research in Europe. Lancet Nov 6-12;364(9446): Schieppati A, Perna A, Zamora J, Giuliano GA, Braun N, Remuzzi G. Immunosuppressive treatment for idiopathic membranous nephropathy in adults with nephrotic syndrome. Cochrane Database Syst Rev Oct 18;(4):CD Remuzzi G, Schieppati A, Ruggenenti P. Clinical practice. Nephropathy in patients with type 2 diabetes. N Engl J Med Apr 11;346(15): Schieppati A, Remuzzi G, Garattini S. Modulating the profit motive to meet needs of the less-developed world. Lancet Nov 10;358(9293): Ruggenenti P, Schieppati A, Remuzzi G. Progression, remission, regression of chronic renal diseases. Lancet May 19;357(9268): Erica Daina got his degree in Medicine at the University of Milan in 1987 and the specialisation in Medical Nephrology in 1990 at the same University. She performed her training at the II Medical Division - San Raffaele Hospital - Milan, and at the Division of Nephrology and Dialysis - Riuniti Hospital - Bergamo. In March 1988 she started her collaboration with the Mario Negri Institute and since June 1993 she works as full-time clinical researcher at the Clinical Research Center for Rare Diseases Aldo e Cele Daccò. Since 1996 she is Unit Head Information Center for Rare Diseases. Areas of interest: rare diseases, Takayasu arteritis, Hemolytic Uremic Syndrome/Thrombotic Thrombocytopenic Purpura, Fabry s disease, Alport s syndrome. Since January 2002 she is representative of Coordinating Centre - Regional Network for Rare Diseases - and collaborates to didactics activity at the University of Turin (Clinical Pathology of Rare Diseases - School of Clinical Pathology Specialization). Selected publications Vanoli M, Daina E, Salvarani C, Sabbadini MG, Rossi C, Bacchiani G, Schieppati A, Baldissera E, Bertolini G; Itaka Study Group. Takayasu's arteritis: A study of 104 Italian patients. Arthritis Rheum Feb 15;53(1): Remuzzi G, Galbusera M, Noris M, Canciani MT, Daina E, Bresin E, Contaretti S, Caprioli J, Gamba S, Ruggenenti P, Perico N, Mannucci PM; Italian Registry of Recurrent and Familial HUS/TTP. von Willebrand factor cleaving protease (ADAMTS13) is deficient in recurrent and familial thrombotic thrombocytopenic purpura and hemolytic uremic syndrome. Blood Aug 1;100(3): Noris M, Daina E, Gamba S, Bonazzola S, Remuzzi G. Interleukin-6 and RANTES in Takayasu arteritis: a guide for therapeutic decisions? Circulation Jul 6;100(1): Daina E, Schieppati A, Remuzzi G. Mycophenolate mofetil for the treatment of Takayasu arteritis: report of three cases. Ann Intern Med Mar 2;130(5): Bresin E, Daina E, Noris M, Castelletti F, Stefanov R, Hill P, Goodship T H J, Remuzzi G, International Registry Recurrent Familial HUS/TTP. Outcome of renal transplantation in patients with non-shiga toxin-associated Hemolytic Uremic Syndrome: Prognostic significance of genetic background. Clinical Journal American Society Nephrology 2006; 1:

317 INTRODUCTION TO THE LABORATORY S ACTIVITIES Rare Diseases (RD) represent about ten percent of all human medical illnesses and infirmities. It is difficult to define what exactly is intended as a RD. The US Congress in the Orphan Drug Act has given the first definition in Under this law it is considered rare a disease that affects less than Americans (prevalence 0.75 per 1 000). Recently, the European Parliament adopted a more strict definition; they consider rare a condition that affects not more than five individuals per in the European Community (prevalence 0.5 per 1 000). Besides the epidemiological parameter, RD have certain characteristics in common: 1) most of them are of genetic origin; 2) rarity often brings a difficult and/or late diagnosis; 3) generally, RD are heavy social burdens both to the family and community and cause early mortality. The greatest barrier to prevention, diagnosis and treatment of RD is inadequate knowledge. Once a diagnosis of RD is put, a major complaint of patients and those involved in their care is the difficulty to obtain pertinent information about causes, symptoms and either established or experimental treatments. Often, patients with RD are willing to participate in clinical studies, but they do not know where and how, and physicians or health authorities are seldom able to help them. RD is not a very attracting field for basic and clinical investigators for several reasons: it is difficult to find adequate animal models for many rare disorders; clinical trials may require more patients than available; financial support is insufficient. Few countries have a central body or system to disseminate information on RD. Accurate information on the incidence and prevalence of RD is extremely important for both basic and clinical investigators. Invaluable help to research advances in RD would come from the availability of registries and databases containing diagnostic, clinical and biological data of patients with rare disorders. A pilot experience has been established at the Clinical Research Centre for Rare Diseases Aldo and Cele Daccò since This Centre is unique with its integration of an Information Centre for Rare Diseases, clinical facilities for the implementation and development of clinical studies, educational activities for physicians, nurses and patients. In 2001 it has been nominated Coordinating Centre of the Regional Network for Rare Diseases in the Lombardy Region, an area of 9 million people in Northern Italy. As Coordinating Centre is also working with the National Centre of Rare Diseases at Istituto Superiore di Sanità. All the up-to-date information regarding the activities of the Coordinating Centre are available at the web site: 315

318 FINDINGS/MAIN RESULTS The database of the Information Centre for Rare Diseases contains data about patients affected by 782 different rare disorders. In the Bank of biological materials, samples from 1241 patients with rare conditions and their families have been collected. The Centre has established contacts with more than 310 Italian Associations for rare diseases. It was even possible that patients with more than 81 different rare diseases - for which no Associations have been established in Italy yet - to meet among themselves. In July 2000, the European Commission recognized the Laboratory as a site for "Postgraduate training on rare diseases" (contract No. QLK ). In December 2001 (Delibera della Giunta Lombarda N. 7328), the Centre was identified as "Coordinating Centre of the Regional Network for Rare Diseases". The Laboratory coordinates the International Registry of Recurrent and Familial Hemolytic Uremic Syndrome (HUS) and Thrombotic Thrombocytopenic Purpura (TTP), since The research projects developed in collaboration with the Laboratory of Immunology and Genetic of Rare Diseases and Organ Transplantation have allowed to better comprehend the pathogenesis of these diseases and to identify some genetically determined forms of HUS, associated to defects of complement regulatory factors, and of TTP, associated to congenital deficiency of ADAMTS13 enzyme. NATIONAL COLLABORATIONS Italian National Institute of Health G. Bosco Hospital, Turin, Regional Coordinating Center for Rare Diseases Riuniti Hospital, Bergamo Italian Takayasu's Arteritis Study Group - ITAKA Group Biotechnology Laboratory, IRCCS Policlinico San Matteo, Pavia Assessorato alla Sanità, Lombardia Region University of Torino, School of Clinical Pathology, Faculty of Medicine and Surgery Italian Network for Promotion of Folic Acid to Prevent Birth Defects University of Turin, Department of Experimental Medicine and Oncology, 2 nd Level Master in Rare Diseases Italian Society of Neonatology (Lombary section), Rare Congenital Respiratory Diseases Study Group University of Milan, 1 st Level Master in Clinical Research Department of Molecular Biology, Unit of Medical Genetics, Policlinico Le Scotte, Siena Unit of Cytogenetics and Genetics, Careggi Hospital, Firenze Laboratory of Metabolic Diseases, National Neurological Institute Carlo Besta, Milano Laboratory of Biochemistry and Genetics, National Neurological Institute Carlo Besta, Milano BergamoScienza Association 316

319 INTERNATIONAL COLLABORATIONS European Community International Registry of Recurrent and Familial Hemolytic Uremic Syndrome and Thrombotic Thrombocytopenic Purpura Information Centre for Rare Diseases and Orphan Drugs ICRDOD, Bulgaria EDITORIAL COMMITTEE MEMBERSHIP Journal of American Society of Nephrology - Nephrology Self Assessment Program (Arrigo Schieppati) Journal of Nephrology (Arrigo Schieppati) Quaderni di Farmacoeconomia (Erica Daina) NATIONAL AND INTERNATIONAL COMMITTEE MEMBERSHIP Network for Rare Diseases Lombardy Region (Delibera Regione Lombardia N 7328, 11/12/2001). Task Force on Rare Diseases (established by DG Health and Consumer Protection on 21 January 2004). Scientific Committee A.O. Bologni di Seriate. Ethical Committe, A.O. Ospedali Riuniti di Bergamo EVENT ORGANIZATION Consensus Conference: Kidney and Liver Transplantation in Hemolitic Uremic Syndrome Clinical Research Centre for Rare Diseases Aldo and Cele Daccò Mario Negri Institute Ranica (Bergamo), December 15 th 2007 PARTICIPATION IN EVENTS IN WHICH THE LABORATORY WAS INVOLVED 10 Convegno Patologia Immune e Malattie Orfane Torino, January 25-27, 2007 Workshop: Epidemiological methods to define incidence, prevalence anda mortality or rare diseases affecting the young Rome, February 16, 2007 Clinical and Experimental Rheumatology II Corso teorico-pratico. La Settimana delle malattie rare in Reumatologia Pisa/Buti, march 14-20, 2007 Diagnosticare le malattie rare in Reumatologia 317

320 Milan, March 17, 2007 Workshop: Progetto RAPSODY Rome, March 24, 2007 Malattie metaboliche rare: la diagnosi e la gestione del paziente Monza, may Rare Diseases Task Force: Working Group on Coding and Classification Paris, May Partecipasalute: Orientarsi in salute e sanità per fare scelte consapevoli Milan, May 17, 2007 La malattia di Anderson Fabry Bergamo, May 28, Convegno Nazionale: Medicina del trapianto: responsabilità e nuove opportunità per gli infermieri Bergamo, June 8-9, 2007 INSIEME Giornate di socializzazione per malatti affetti da patologie rare Milan, June 29-july 1, 2007 CAPOIRA Seminari di formazione: Comprendere i protocolli di ricerca Milan, July 2, 2007 and Rome, September 20, 2007 I Test Genetici nella pratica clinica Tivoli Terme (Rome), July 5-6, th European Meeting Complement in Human Disease Cardiff UK, September 8-11, 2007 Progetto di formazione sul campo: Registro Regionale Malattie Rare Presidi Rete Regionale, 2007 Comunicazione e Malattie Rare Focus Group Rome, October 4, rd International Workshop on Thrombotic Microangiopathies Jena, October 4-6, 2007 Tavola Rotonda: Geni, proteine, malattie Genoa, October 26, 2007 La gestione pratica della malattia di Pompe: aspetti diagnostici e terapia Pavia, November 21, 2007 La Federazione Lombarda Malattie Rare si presenta alle Associazioni di Malattie Rare Milan, November 24, 2007 European Conference on Rare Diseases 318

321 Lisbon, November 27-28, 2007 European Commission "Fondazione Ricerca Malattie Rare", Bergamo Lombardy Region GRANTS AND CONTRACTS SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Monteferrante G, Brioschi S, Caprioli J, Pianetti G, Bettinaglio P, Bresin E, Remuzzi G, Noris M Genetic analysis of the complement factor H related 5 gene in haemolytic uraemic syndrome Mol Immunol; 2007; 44: OTHER PRODUCTS PUBLISHED IN 2007 Noris M, Bresin E, Mele C, Remuzzi G, Caprioli J (November 2007) Atypical Hemolytic-Uremic Syndrome in: GeneReviews at GeneTests: Medical Genetics Information Resource [database online]. Copyright, University of Washington, Seattle, Available at 319

322 RESEARCH ACTIVITIES Information Centre for Rare Diseases In 1992, in the frame of the Clinical Research Centre for Rare Diseases Aldo e Cele Daccò was established an Information Centre for Rare Diseases. The Information Service is available to patients with rare diseases, their families and doctors. It offers information, update and useful addresses free of charge with particular emphasis on etiology, pathogenesis, genetics, treatments and availability of referral research centres. One of the most difficult issues to address when studying rare diseases are those, related to the recruitment of a sufficient number of patients with a given disease. The database of the Information Centre for Rare Diseases has become a useful tool for identification of potentially eligible patients for clinical studies. Till now, the Information Centre for Rare Diseases has collected patients clinical data for about 200 different rare conditions with 10 or more cases. Furthermore, intensive collaboration with groups with the same interest from Italy and abroad provides many possibilities for starting of clinical projects with a multicentral design. Bank of biological samples and description of hereditary nephropathies The aim of this project is to collect clinical data and biological samples from patients and their families with rare genetic conditions. A database with clinical data and a Bank for biological samples collection and preservation has been created. The availability of clinical data and biological samples is useful to perform new biochemical and genetic tests within specific research projects aimed to better reveal the mechanisms of the diseases, their manifestation and therapeutic opportunities. In particular, the attention is focused on rare genetic disorders of the kidney. A thorough clinical evaluation, including clinical data collection, medical physical examination, renal ultrasonography, laboratory tests of blood and urine is offered to patients, affected by hereditary nephropathies (Alport syndrome, Fabry disease, Focal Segmental Glomerulosclerosis, Glomerulopathy with Fibronectin deposits, Membranoproliferative glomerulonephritis, Medullary Cystic Kidney disease, Cystinuria), who are addressing our Centre. After obtaining a written informed consent, biological samples from patients and their relatives are collected, labelled with specific codes to assure the anonymity and conserved in the Bank for biological samples. In case the responsible gene for a hereditary nephropathy is known (Alport syndrome, Fabry disease, Cystinuria), the blood samples are redirected to the relevant Laboratory of reference. For other nephropathies, where the identification of the gene mutation is unknown or still in course, the blood samples are conserved with the aim to be used in specific future research projects. Evaluation of the long term efficacy of enzyme replacement therapy in Fabry disease Fabry disease is an X-linked disorder of the glycosphingolipid catabolism caused by the deficient activity of the lysosomal hydrolase alfa-galactosidase A (A-gal A) in tissues and fluids of affected hemizygous males. Most heterozygous females have an intermediate level of enzymatic activity. The enzymatic defect leads to a systemic deposition of glycosphingolipid in the heart, kidneys, eyes and other organs and tissues. Preliminary studies of enzyme replacement therapy demonstrate that periodic infusions of recombinant human Alfa-galactosidase A are safe (in terms of infusion reactions) and effective in patients with Fabry disease. The purpose of this study is to evaluate the long term efficacy of enzyme replacement therapy in patients with Fabry disease and renal involvement. Twenty patients (males and females) referred to the SMIMAF (Studio Multicentrico Italiano sulla Malattia di Anderson-Fabry) will be enrolled. 320

323 International Registry of Recurrent and Familial Hemolytic Uremic Syndrome and Thrombotic Thrombocytopenic Purpura Hemolytic Uremic Syndrome (HUS) and Thrombotic Thrombocytopenic Purpura (TTP) are rare diseases of microangiopathic haemolytic anemia and thrombocytopenia with signs of renal (most prevalent in HUS) and cerebral (most prevalent in TTP) damage. There are extremely rare forms of HUS and TTP, often occurring in families, in which the patients relapse even after complete recovery of the first episode with permanent renal and neurological sequelae. In the familial and recurrent forms of HUS and TTP, the attention is concentrated mainly on the genetic predisposition to the disease. The Laboratory coordinates the International Registry of Recurrent and Familial HUS/TTP, since The Registry has collected more than 460 cases of HUS/TTP referred from 95 Italian and 73 European and extra-european Centres. Clinical and laboratory data of all patients referred to the Registry are collected by a uniform data extraction form. The family history and also the personal data of the unaffected relatives are collected, when possible. Biological samples are collected from all patients and available relatives, for the biochemical and genetic analyses. Many research projects in collaboration with the Laboratory of Immunology and Genetic of Rare Diseases and Organ Transplantation and the Laboratory of Cellular Biology Negri Bergamo are developed and have still allowed to identify some genetically determined forms of HUS and TTP. The maintenance of a centralised bank of biological samples ensure the availability of clinical material for new investigative approaches as they will be developed. Rituximab in relapsing and chronic thrombotic thrombocytopenic purpura Thrombotic thrombocytopenic purpura (TTP) is a rare disease characterized by thrombocytopenia, microangiopathic hemolytic anemia and neurological signs. In most cases, TTP is related to an acquired deficiency of ADAMTS13 activity, due to the presence of autoantibodies anti-adamts13. The absence of ADAMTS13 activity leads to the accumulation of von Willebrand Factor ultralarge multimers (normally cleaved to smaller molecular forms by ADAMTS13 enzyme), which probably induces platelets aggregation and the ensuing process of thrombotic microangiopathy. The purpose of the project is to evaluate the use of a monoclonal anti-cd20 antibody (Rituximab) in patients with relapsing and chronic thrombotic thrombocytopenic purpura, related to the presence of anti-adamts13 antibodies. Rituximab is a humanized monoclonal antibody that targets the CD20 molecule on B cells (white cells producing antibodies). Administration of Rituximab leads to a selective B cell depletion that usually lasts 6 to 9 months. Inhibition of B cell activation or growth by rituximab treatment limit the uncontrolled synthesis of autoantibodies, that may predispose to chronic relapsing TTP. Rituximab is infused intravenously once weekly for a total of four infusions. After Rituximab treatment, the patients are periodically controlled with medical examination, blood and urine exams, and with ADAMTS13 plasma activity assay and research of anti-adamts13 autoantibodies, in collaboration with the Laboratory of Cellular Biology Negri Bergamo. Observational period lasts 12 months. Evaluation of urinary podocyte excretion Recent evidence recognize an important role for podocytes in the progression of renal diseases. Podocytes contribute to glomerular permeability and are important target cell for renal disease progression. Glomerular injury is usually associated with the leakage of protein across the filter into the urine and with the disappearance of podocyte foot process. Injuried podocytes either undergo apoptosis or detach from the glomerular basement membrane, with subsequent appearance in the urine. Urinary excretion of podocytes has been reported as possible predictor of disease progression in a number of progressive glomerular diseases, such as IgA 321

324 nephropathy, focal segmental glomerulosclerosis, diabetic nephropathy. During the past years the Clinical Research Center for Rare Diseases, in collaboration with the Department of Molecular Medicine at Mario Negri Institute, has established an indirect immunohistochemistry method for the detection of podocytes in urinary sediments. This method has allowed to identify and quantitate the extent of urinary podocyte loss in patients with glomerular disease. The present study has the aim to evaluate excretion of urinary podocytes in rare genetic renal diseases. This methodology, applied on a larger number of patients, could provide a non invasive way of indexing the extent of glomerular damage and a useful instrument to evaluate the response to treatment. Effects of an intensified treatment with ACE-inhibitors, Angiotensin II receptor antagonists and Statins in Alport syndrome Alport syndrome (AS) represents a form of progressive hereditary nephritis in which the genetic defect resides in the synthesis of one of several subunits of type IV collagen, the predominant constituent of basement membranes in renal glomeruli. Renal impairment occurs with time and severe renal failure with hypertension and uremia represent the end stage of the disease, even if a high variability in the rate of progression is described. The prognosis is variable. Males are usually affected by a progressive form of the disease. Affected females with X-linked syndrome usually have a good prognosis with a mild renal impairment. Other clinical manifestations are anterior lenticonus which occur in about one third of patients. Other findings are myopia, lens opacities and retinal pigment abnormalities and corneal vesicles or erosions. The disease is also associated to a sensor neural deafness which can occur in approximately half of the patient affected and usually correlates with renal impairment. No definite treatment exists in order to delay the time of dialysis or a kidney transplant. Many studies showed that Angiotensin converting enzyme (ACE) inhibitors slow glomerular filtration rate (GFR) decline and limit progression to end stage renal disease (ERDS) and dialysis in several chronic nephropathies associated with proteinuria. The combination of ACE-I with Angiotensin II receptor antagonists may reduce proteinuria more effectively than the two drugs alone. Moreover the addition of Statins may synergize the antiproteinuric effects of ACE-I and ATAII antagonists in experimental models of chronic renal diseases. The purpose of this study is to evaluate the effect of a standardized multimodal nephroprotection intervention (Remission Clinic) in Alport patients with renal involvement. Nine patients with Alport syndrome and macroalbuminuria will be enrolled in this study. Recruitment of normo or microalbuminuric patients will be stopped and the analysis will be performed on the patients that will be available when the recruitment phase of macroalbuminuric patients will be completed. Update of the Congenital Respiratory Malformations database This Project originated from the collaboration between the Study Group on Respiratory Disease of the newborn and the Coordinating Centre for Rare Diseases. The Congenital Respiratory malformations database was conceived to facilitate the diagnostic and assistance procedures for paediatricians facing with more frequent respiratory malformations. The update consisted of a review of the lists of malformation categories and of all syndromes previously included; more syndromes have been added. Furthermore, in the brief description, more detailed than the previous version, the Code for Rare Disease was added, allowing to address the patient to the Referral Centres for Lombardy. A link with the OMIM database is provided for each syndrome for a better description of the disease. 322

325 Identification of new genes associated to the Autosomal Dominant Familial form of Focal Segmental Glomerulosclerosis Focal segmental glomerulosclerosis (FSGS) is a pathological entity, and is a significant cause of end-stage renal disease (ESRD). Glomerular disease is the third leading cause of ESRD, and FSGS comprises a significant proportion of this subgroup: up to 5% of adults and 20% of children with ESRD. The diagnosis of FSGS is based on renal pathology. The clinical hallmarks include proteinuria, nephrotic syndrome, occasional hematuria and frequent progression to ESRD. Hypertension is also a common finding. At present, there are no consistently reliable treatments for FSGS and response rates to available treatments have been estimated at <30-50%. Because FSGS has become an important cause of ESRD, it is essential to understand the molecular basis and pathogenesis of this disease process. There are various subtypes of FSGS, which include primary (idiopathic or sporadic), secondary, familial and FSGS associated with congenital syndromes. Among familial forms of FSGS, both autosomal recessive and dominant inheritance patterns have been reported. The study, in collaboration with the Laboratory of Immunology and Genetic of Rare Diseases and Organ Transplantation, is aimed at identifying the genetic causes of the autosomal dominant familial forms of FSGS, in order to help in the clinical management. Patients with an autosomal dominant familial form of FSGS (at least two affected subjects in two different generations within a family) and their available relatives, will be enrolled. Clinical data and blood samples will be collected and used to perform routine laboratory exams and to obtain DNA for the genetic analyses. Genetic counselling will be provided to all patients and to their relatives. 323

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327 CESAV A. and A. Valenti Health Economics Center STAFF Head Livio GARATTINI, Econ.D. 325

328 CURRICULUM VITAE Livio Garattini: got his degree in Economics in March 1983 at the Bocconi University in Milan. Educational activities: King s Fund College, London: courses of health care management; Centre for Health Economics, York: review of publications on the English NHS; Ecole Nationale de la Santé Publique, Rennes: courses of health policy. Areas of interest: Health Economics and health Policy Analysis. At present he is the Director of CESAV (Centre of Health Economics A. e A. Valenti - M. Negri Institute); : researcher at M. Negri Institute; : clerk at Banca Commerciale Italiana in Milan; : junior consultant at Sogess srl in Milan; : researcher at Bocconi University in Milan. Selected publications: Garattini L, De Compadri P, Clemente R, Cornago D (2003) Economic Evaluations in Italy: a Review of the Literature International Journal of Technology Assessment in Health Care 19(4): Garattini L, Ghislandi S (2006) Off-patent drugs in Italy- A short-sighted view? The European Journal of Health Economics 7(1): Garattini L, Ghislandi S (2007) Should we really worry about launch delays of new drugs in OECD countries? (Editoriale) The European Journal of Health Economics 8(1): 1-3. Cornago D, Li Bassi L, De Compadri P, Garattini L (2007) Pharmacoeconomic studies in Italy: a critical review of the literature The European Journal of Health Economics 8(2): Garattini L, Cornago D De Compadri P (2007) Pricing and reimbursement of in-patent drugs in seven European countries: A comparative analysis Health Policy 82: Koleva D, Motterlini N, Banfi P, Garattini L (2007) Healthcare costs of COPD in Italian referral centres: A prospective study Respiratory Medicine 101:

329 INTRODUCTION TO THE CENTER'S ACTIVITIES The "Angelo e Angela Valenti" Centre for Health Economics (CESAV) was established in 1992 at the "M. Negri Institute" and based at Villa Camozzi- Ranica (Bergamo)-Italy. CESAV is primarily a research centre, but also does educational work. The centre is involved in health economics and health policy research. The main areas of research are: Economic Evaluation of Health Care Programs (i.e. assessment of costs and benefits of alternative health care treatments and services) and Comparative Health Policy Analysis (i.e. study of foreign health care systems, in particular aimed at identifying possible innovations for European countries). FINDINGS/MAIN RESULTS In 2007 economic evaluations regarded the costs of the following pathologies: glaucoma, multiple myeloma and therapies for hepatitis C. The studies of comparative analysis were focused on the management of vaccination campaigns in health authorities in Lombardy and on the management of drug expenditure in a National sample of health authorities. NATIONAL COLLABORATIONS Public and private institutions, other health care organizations (Ministry of Health, Regional and Local Health Authorities, Hospital Trusts). INTERNATIONAL COLLABORATIONS CES (Collège des Economistes de la Santé) of Paris Corvinus University of Budapest Global Fund of Geneva WidO of Bonn University Carlos III of Madrid University of Hannover University of York University Pompeu Fabra of Barcelona University Erasmus of Rotterdam 327

330 EDITORIAL BOARD MEMBERSHIP Acta Bio Medica (Livio Garattini) Biomedical Statistics and Clinical Epidemiology (Livio Garattini) Economia e Politica del Farmaco (Livio Garattini) FarmacoEconomia News (Livio Garattini) Farmeconomia e Percorsi Terapeutici (Livio Garattini) Health Policy (Livio Garattini) L'Internista (Livio Garattini) Journal of Medical Economics (Livio Garattini) PharmacoEconomics Italian Research Articles (Livio Garattini) Quaderni di FarmacoEconomia (Livio Garattini) The European Journal of Health Economics (Livio Garattini) PEER REVIEW ACTIVITIES BJCP (British Journal of Clinical Pharmacology) Health Policy PharmacoEconomics Social Science & Medicine The European Journal of Health Economics EVENT ORGANIZATION Congress: Convegno Nazionale CESAV di Farmacoeconomia: Economia del farmaco: fra soluzioni tecniche e decisioni politiche. Prezzi e rimborsabilità dei farmaci innovativi. Educazione Continua Medica : L ECM in Europa: esperienze a confronto. Gli aspetti economici del glaucoma. Informazione Medico-Scientifica sui farmaci : La normativa sull informazione medico scientifica in Italia: la situazione a livello nazionale. La normativa sull informazione medico scientifica in Italia: i risultati di un inchiesta a livello regionale. May 29-30, Ranica (BG) Congress: La farmacoeconomia nelle terapie per il CCR. Gli studi di farmacoeconomia sui trattamenti chemioterapici nel CCR : Una revisione critica degli studi di farmacoeconomia. Metodologie di analisi a confronto: i risultati di una simulazione. Opinioni a confronto: Gli studi di farmacoeconomia: l utilità percepita in oncologia. December 11, Ranica (BG). 328

331 PARTICIPATION IN EVENTS IN WHICH THE CENTER WAS INVOLVED Congress: Global Pricing and Reimbursement Congress European Pricing and Reimbursement for Generic Medicines Country By Country Pricing And Reimbursement Policies Current pricing policies in Italy. Reimbursement processes and timelines. Identifying critical cost containment measures. (Global Pricing and Reimbursement-Congress 2007) Pricing Models and Systems-A Country by Country Breakdown The current pricing policies in Italy and the impact on the European generic industry. Reimbursement processes and timelines for generic medicines. Reference pricing and pharmacists incentive schemes. (European Pricing and Reimbursement for Generic Medicines) January, Prague. Congress: Congreso Europeo de Oficina de Farmacia. Los genéricos en Europa: vision farmacoeconomica. 28 February-2 March, Barcellona. Course: Le maculopatie e la degenerazione maculare legata all età: gestione clinico-terapeutica ed economica. L impatto socioeconomico delle maculopatie:farmacoeconomia e Qualità di Vita. Discussione. 15 May, Milan. Workshop: European Advisory Panel meeting to discuss the positioning of temsirolimus for use in the treatment of renal cell carcinoma. 13 July, London. Workshop: ADHD (Attention Deficit and Hyperactivity Disorder) Advisory Board Meeting July, Frankfurt on Maine. Congress: Acomplia Focus Meeting. Round Table on Knowing our customers. 4 October, Paris. Congress: Le maculopatie e la degenerazione maculare legata all età: gestione clinicoterapeutica ed economica. L impatto socioeconomico delle maculopatie: farmacoeconomia e Qualità di Vita. 10 October, Genoa. Congress: Il corretto management terapeutico ed assistenziale nella terapia del dolore. Impatto economico della terapia del dolore. 7 November, Milan. Congress: Le maculopatie e la degenerazione maculare legata all età: gestione clinicoterapeutica ed economica. Analisi farmaco-economiche applicate alla CNV. 7 December, Naples. 329

332 GRANTS AND CONTRACTS Abbott EG Grunenthal-Prodotti Formenti Jannsen Cilag Novartis Farm Ratiopharm Sanofi Aventis Sanofi Pasteur MSD Schering Plough Vivisol SELECTION OF SCIENTIFIC PUBLICATIONS FROM 2007 Garattini L, Ghislandi S Should we really worry about launch delays of new drugs in OECD countries? (Editorial) The European Journal of Health Economics 8(1): 1-3. Cornago D, Li Bassi L, De Compadri P, Garattini L Pharmacoeconomic studies in Italy: a critical review of the literature The European Journal of Health Economics 8(2): Garattini L, Koleva D, Motterlini N, Cornago D Medical Costs of Chronic Musculoskeletal Pain in Italy Clinical Drug Investigation 27(2): Garattini L, Cornago D, De Compadri P Pricing and reimbursement of in-patent drugs in seven European countries: A comparative analysis Health Policy 82: Koleva D, Motterlini N, Schiavone M, Garattini L Medical Costs of Glaucoma and Ocular Hypertension in Italian Referral Centres: A Prospective Study Ophthalmologica 221(5): Koleva D, Motterlini N, Banfi P, Garattini L Healthcare costs of COPD in Italian referral centres: A prospective study Respiratory Medicine 101: Garattini L Pricing and Reimbursement Policies in Italy: Current and Future Trends in Garau M, Mestre-Ferrandiz J (edited by) European Medicines Pricing and Reimbursement. OHE. Oxon, Radcliffe Publishing. 330

333 LAY PRESS SELECTION PUBLISHED IN 2007 Motterlini N, Garattini L I costi di struttura dei reparti di pneumologia in Italia FarmacoEconomia News 1: 3-7. Cerzani M, Barbui C, Garattini L Le valutazioni economiche del trattamento farmacologico con antipsicotici nella schizofrenia: una rassegna degli studi italiani Quaderni di Farmaco Economia 2: Mc Nee, England S (Translation by: De Compadri P, Garattini L, Li Bassi L) Pharmac: un esempio di sfide e successi Quaderni di Farmaco Economia 3:6-15. Cerzani M, Pasina L; Clavenna A, Nobili A, Garattini L Revisione critica degli studi italiani di farmacoeconomia sull uso dei farmaci antinfiammatori non steroidei in medicina generale Quaderni di Farmaco Economia 3: Gritti S, De Compadri P, Garattini L L informazione medico scientifica fra Stato e Regioni Quaderni di Farmaco Economia 4: De Compadri P, Koleva D, Zaniboni A, Garattini L Modalità di somministrazione per terapie del CCR a confronto: un esercizio di valutazione economica Quaderni di Farmaco Economia 4: Garattini L Finanziaria 2007: vere e false novità Dialogo sui farmaci 1: RESEARCH ACTIVITIES Educational activity Educational activities are developed only if related to research studies, in order to offer original contributions which naturally reinforce the research aims. Economic Evaluation of Health Care Programs The aim of this research area is to assess the costs of illnesses and the cost-effectiveness ratios of the diagnostic/therapeutic existing alternatives. In general, analyses can be classified into two groups: partial economic evaluations (e.g. observational studies on the costs of pathologies) and full economic evaluations (e.g. cost-effectiveness analyses). Comparative Health Policy Analysis The aim of this research area is to study the organization of health care systems, in order to draw lessons from international comparisons. This is particularly important in a "market" like health care where economic competition lacks by definition and therefore public regulation plays a crucial role. 331

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335 The Transplant Research Center Chiara Cucchi De Alessandri e Gilberto Crespi The Transplant Research Center (CRT) was set up in 2002 to support and promote the work of outstanding research scientists throughout the world and to carry out major organ transplant research programs. The Center is housed in the Villa Camozzi, at Ranica, under the same roof as the Mario Negri Institute in Bergamo and is managed in collaboration with the Institute. The Center s staff is mainly made up of senior and junior researchers that were trained in the laboratories of the Mario Negri Institute in Bergamo, focusing on transplant immunology, research for less toxic immunosuppressant drugs, and new gene therapy techniques to prevent acute rejection of transplanted organs. Information on the Center s activities can be found in the sections addressed to the Department of Molecular Medicine (Laboratory of Immunology and Genetics of Organ Transplantation and Rare Diseases) and the Department of Renal Medicine (Laboratory of Pharmacokinetics and Clinical Chemistry). 333

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337 EDUCATIONAL ACTIVITIES Dean, Mario Salmona, PH.D. The Institute's training programs fall under the heading of biomedical science, and are part of the Lombardy Region's professional training schemes. There are courses for specialized laboratory technicians, and for graduates intending to do research. Special training for clinical trial nurses is provided at the Aldo and Cele Daccò Clinical Research Center for Rare Diseases, at Ranica, near Bergamo. In 1999 the Institute has set up a Ph.D. course, in collaboration with the Open University of London. This degree is recognised throughout Europe and in the USA. In 2006 the Institute also set up a First Level master Course in Clinical Research, in collaboration with the Milan University and a Second Level master Course in rare Diseases, in collaboration with the University of Torino. Students enrolled in formal courses receive three months' preparatory training, after which they are assigned study grants. Between 1963 and 2004 the Mario Negri Institute awarded 6,123 grants, 668 of them to foreign researchers who came to the Institute for special training. Everything possible is done to help students find work once they finish the course. The main feature of these courses is that technicians and researchers receive their training "on site". They work full-time in research programs of a high scientific standard, using advanced equipment and learning the latest methods, in regular contact with colleagues in different countries. Besides its scientific value, this approach provides an excellent preparation on the human and personal scale. Students are usually assigned to one of the Institute s Laboratories, where they gradually gain specialized skills by working on specific research projects. They are expected to attend lessons, seminars, courses and congresses and learn to make full use of the Institute s well-stocked library. Students all have access to the internet and to biomedical database and can print out copies of articles they need to consult, from major international journals. Should the opportunity arise, students are expected to be available for trips abroad, to participate in conferences or courses. The Pharmacological Research Specialists and Biochemical Research Technicians will receive diplomas issued officially by the Lombardy Region and the Mario Negri Institute for Pharmacological Research. These have legal value throughout Italy, and are recognised in competitions for public posts, where they are worth a certain number of points. Mario Negri Institute diplomas are widely considered a guarantee of an excellent theoretical and practical training. The Open University of London Ph.D. degree earned at the Institute has legal value throughout Europe and in the USA. Once they have their diploma or Phd., graduates who want to continue doing research at the Institute may be offered a chance to spend a year or two abroad. At the moment these courses are available: 335

338 Three-year course for graduates, in Milan or Bergamo, leading to a diploma as Pharmacological Research Specialist. Three-year course for diploma-holders, in Milan or Bergamo, leading to a diploma as Biochemical Research Technician. Research doctorates (Ph.D.), run under an agreement with the Open University of London. Two-year courses are run at the Daccò Center, in Ranica, near Bergamo, for nurses intending to work in clinical trials. These give a diploma as Professional Clinical Trial Nurse. Other training opportunities PREPARING A THESIS FOR A DEGREE: Students can prepare their thesis in scientific subjects at the Institute, with the approval of their university faculty. These students must work at the Institute for at least two years. SUMMER STUDENTS In June and July each year the Institute accepts a certain number of students in their last two years at high school, to give them experience as part of school/work programs. 336

339 STAFF Executive Offices Services and Offices 337

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341 Prof. Silvio Garattini Silvio Garattini was born in Bergamo (Italy) on 12th Nov Diploma in Chemistry. Degree in Medicine. Lecturer in Chemotherapy and Pharmacology. Assistant then Deputy Professor at the Milan University Institute of Pharmacology until Founder in 1963 and director of the Mario Negri Institute for Pharmacological Research. The Institute has now four locations (Milan, Bergamo, Ranica (Bg), S. Maria Imbaro (Ch)) and more than 850 people. He is a member of the Gruppo 2003 (a group of the most cited Italian scientists in international scientific literature). Garattini's publications in Italian and in English in international scientific journals, and texts on pharmacology, run into the hundreds. Founder of the European Organisation for Research and Treatment of Cancer (EORTC). During the last ten years professor Garattini has been a member of various organizations, among which: the Italian National Research Council (CNR) - Committee on Biology and Medicine; the National Health Council, the Committee for Italian Research Policy, set up by the Presidency of the Council of Ministers; the Commissione Unica del Farmaco (CUF) of the Ministry of Health. He has held the following posts: President of the UICC Committee on Antitumoral Chemotherapy, President of the European Organisation for Research and Treatment of Cancer (EORTC), Consultant to the World Health Organisation. He was President of the European Society of Biochemical Pharmacology; member of the Committee for Proprietary Medicinal Products (CPMP) of the European Agency for the Evaluation of Medicinal Products (EMEA), Member of the CEPR (Committee of Experts of Research Policy) at the Ministry for University and Scientific and Technological Research; Member Scientific Committee of the Lega Italiana per la Lotta Contro i Tumori; member of the Board of the Istituto Superiore di Sanità. Vice-president of the Consiglio Superiore di Sanità; President of the research and Development Commission of the Agenzia Italiana del Farmaco (AIFA); President of the Angelo and Angela Valenti Foundation and of the "Via di Natale" Foundation; President of the Technical Commission for Pharmaceutical Assistance of the Regione Autonoma of Sardegna; Member of the Strategic Committee for Welfare, Regione Lombardia. Member of the Consiglio Superiore di Sanità and the Comitato Nazionale di Bioetica. Silvio Garattini is a Fellow of the New York Academy of Sciences, the American Association for the Advancement of Science, Honorary Fellow of the Royal College of Physicians (Pharmaceutical Medicine), London, and a member of numerous other Italian and international scientific societies. He has received many awards for his work, including the French Legion d'honneur for scientific merit, and the Grand Ufficiale della Repubblica Italiana, and holds honorary degrees from the Universities of Bialystok in Poland, and Barcelona in Spain. Most recent prizes: Prize Ippocrate, 2003; Prize Mens Sana in Corpore Sano; Prize Nuova Spoleto, 2003; Prize Angelo dell anno; Alkmeon International Prize; International Prize Sant Agostino Città di Bergamo; Prize Il Campione della Scienza; Medal Natta; Prize Coppola. In its 40-plus years, the Mario Negri Institute for Pharmacological Research, under Professor Garattini's leadership, has published more than scientific papers and about 200 books, on topics ranging from cancer and its treatment to tumour immunology, neuropsychopharmacology, and cardiovascular and renal pharmacology. More than 4000 young Italian and 600 foreigner graduates and technicians have obtained specialist qualifications at the Institute. 339

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343 Prof. Giuseppe Remuzzi Giuseppe Remuzzi was born in Bergamo, Italy in Upon completion of his medical training at the University of Pavia in 1974, he received specialty training in Hematology and Nephrology at the University of Milan. Since 1975, he has pursued his academic career at the Ospedali Riuniti of Bergamo, where he was appointed Professor of Nephrology and Director of the Department of Medicine and Transplantation in Since 1999, he is Director of the Department of Nephrology and Dialysis of the same hospital. The Negri Bergamo Laboratories, which he has directed since 1984, is a group of basic scientists, physiologists, pharmacologists, molecular and cellular biologists, pathologists and clinicians devoted to the study of renal disease. As Research Coordinator of the Negri Bergamo Laboratories and the affiliated Clinical Research Center for Rare Diseases "Aldo e Cele Dacco", both divisions of the Mario Negri Institute for Pharmacological Research, he conduct a diverse team of researchers studying human renal diseases and their corresponding experimental models from the perspective of pathophysiology and therapeutic intervention. He touched major advances in many areas of nephrology, with particular focus on platelet endothelial interactions, vascular prostaglandin biology, coagulation and renal disease, progression of renal disease, experimental models of glomerular damage, and transplant immunology and tolerance. Particularly his contributions to the understanding of the pathophysiology of hemolytic uremic syndrome, prostaglandin metabolism in pregnancy, renal vascular biology in uremia, the role of protein trafficking in renal disease progression, the induction of graft tolerance by intrathymic injection of donor antigens, the role of the co-stimulatory CD28-B67 pathway in transplant rejection and the prevention of renal and cardiovascular damage in diabetes. Prof. Remuzzi serves on editorial boards of numerous journals including the prestigious New England Journal of Medicine and is member of the International Advisory Board of The Lancet. In recognition of his achievements, he has been awarded in 1998 honorary memberships of the Association of American Physicians and the British Royal College of Physicians. In 2001 he was nominated Chairman of the Research Committee of the COMGAN (Commission on Global Advancement of Nephrology) of the International Society of Nephrology (ISN). He received an Honorary Professorship at the University of Maastricht in In 2005 during the World Congress of Nephrology in Singapore he received the ISN Jean Hamburger Award and in November 2007 he received during the annual congress of the American Society of Nephrology the precious John P. Peters Award. Prof. Remuzzi has authored and co-authored more than 885 scientific articles, reviews and monographs. 341

344 Mario Negri Institute Milan Executive Office Director Prof. Silvio GARATTINI, M.D. Administrative Office Maria Grazia PEZZONI, Chief Technical Office Fabio BRIGHENTI, D. Arch. General Maintenance Emanuele SINELLI Studies Office Armanda JORI, Pharm.D. Press Office Isabella BORDOGNA, Phil. D. Public Relations Office Claudio PANTAROTTO, Chemist Prevention and Safety Office Emilio BENFENATI, Chem.D. Annamaria SEGALINI, Phys.D. English Style Editor Judi BAGGOTT Photography and Audio-Visual Service Felice DE CEGLIE Purchasing Office Eufrasia COVIELLO Director s Office Rosanna MAPELLI, Chief General Secretariat Elena POZZOLI 342

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346 Negri Bergamo Laboratories Executive Offices Director Research Coordinator Scientific Secretariat Prof. Silvio GARATTINI, M.D. Giuseppe REMUZZI, M.D. Ariela BENIGNI, Biol.Sci.D., Ph.D. Administration of Research Projects/Admn. Assistance Daniela MELACINI, Biol.Sci.D. Press and Communication Office Francesca Di Fronzo, Mod. Lit. D. Audio-Visual Service Antonella PICCINELLI, Biol.Sci.D. Prevention and Safety Office Chief Annamaria SEGALINI, Phys.D. Library Chief Anna BOZZALE Valeria MIGLIOLI General Maintenance Giancarlo GASPARI Director s Office Antoinette van ENGELEN, Chief 344

347 Centro Aldo e Cele Daccò Ranica (Bg) Executive Offices Director Research Coordinator Scientific Secretariat Health Director Prof. Silvio GARATTINI, M.D. Giuseppe REMUZZI, M.D. Ariela BENIGNI, Biol.Sci.D., Ph.D. Norberto PERICO,M.D. Press and Communication Office Francesca Di Fronzo, Mod. Lit. D. Prevention and Protection Office Chief Annamaria SEGALINI, Phys.D. Paola BOCCARDO, Biol.Sci.D. Library Mansueto Astori Chief Anna BOZZALE Monica MINALI Director s Office Daniela RICEPUTI, Chief Clinical Trials Office Paola BOCCARDO, Biol.Sci.D. Custodian/general maintenace Giampiero CUGUSI 345