Annual Report

Transcription

1 Annual Report

2 Annual Report 2015/2016 Deutsches Rheuma-Forschungszentrum Berlin A Leibniz Institute This Annual Report covers the research activities of the Deutsches Rheuma-Forschungszentrum Berlin (DRFZ) during the years 2015 and The research activities of the DRFZ are made possible through financial support from the Senate Administration for Economy, Technology and Research of the Land Berlin, the Leibniz Association, the German Research Foundation (DFG), the Federal Ministry of Education and Research (BMBF), the European Commission, and through various other third parties, as mentioned in the text. Thanks are due to all of them. Please visit our web site: Photographs: Poster session during the on-site visit of the Scientific Advisory Committee, 2016

3 Indroduction Content Content Content Introduction Organisational Chart...3 Word of Welcome...4 Research Highlights 2015/ Research Projects and Networks Leibniz ScienceCampus Berlin Chronic Inflammation at a Glance...22 Working at the DRFZ...26 Public Outreach Programme Area Pathophysiology of Rheumatic Inflammation Baumgrass, Signal Transduction...32 Kubagawa, Humoral Immune Regulation...48 Buttgereit, Glucocorticoids and Bioenergetics Nedospasov, Inflammation Biology Dörner, B Cell Memory...36 Niesner, Biophysical Analytics Hamann, Experimental Rheumatology...38 Radbruch, Cell Biology Hauser, Immune Dynamics...40 Romagnani, Innate Immunity...56 Hiepe, Autoimmunology...42 Scheffold, Cellular Immunology...58 Hutloff, Chronic Immune Reactions...44 Tokoyoda, Osteoimmunology...60 Kruglov, Chronic inflammation...46 Worm, Allergology...62 Programme Area Regenerative Rheumatology Löhning, Pitzer Laboratory of Osteoarthritis Research...66 Mashreghi, Therapeutic Gene Regulation...68 Programme Area Epidemiology of Rheumatic Diseases Minden, Paediatric rheumatology and health services research Poddubnyy, Spondyloarthritiden Strangfeld, Pharmacoepidemiology...76 Zink, Health services research in adult rheumatology...78 Core Facilities,Technical Units & Central Services Central Laboratory for Microscopy...82 Mass cytometry...90 Central Laboratory...84 Regine v. Ramin-Lab for Molecular Rheumatology...92 Flow Cytometry & Cell Sorting (FCCF)...86 Animal Facility...94 Immune Monitoring...88 Embryo Technology Laboratory...95 Appendix Third party funded projects - DRFZ Groups...98 Third party funding projects - Liaison Groups Third party funds Qualifications Qualifications Sitemap Sitemap Imprint

4 Organisational Chart Introduction Organisational Chart Organisational Chart Data Security, Animal Welfare, Job Safety, Ombudsman, Equal Opportunities Scientific Director Radbruch Board of Trustees Herrhausen Administrative Director Starke Scientific Advisory Committee Meuer Programme Area Central Units Pathophysiology of Rheumatic Inflammation Radbruch Regenerative Rheumatology Radbruch (temporary) Epidemiology of Rheumatic Diseases Zink Core Facilities and Technical Units DRFZ groups Baumgrass Signal Transduction Fillatreau (until ) Immune Regulation Hamann Experimental Rheumatology Kruglov (since ) Chronic Inflammation Kubagawa Humoral Immune Regulation Nedospasov Inflammation Biology Radbruch Cell Biology Tokoyoda Osteoimmunology DRFZ group Mashreghi (since ) Therapeutic Gene Regulation Liaison group Löhning (since ) Pitzer Laboratory of Osteoarthritis Research DRFZ groups Listing (until ) Statics and clinical studies Minden Paediatric Rheumatology Strangfeld Pharmacoepidemiology Zink Health Services Research Liaison group Poddubnyy (since ) Spondyloarthritides Flow Cytometry & Cell Sorting (FCCF) Chang, Kaiser Mass Cytometry (CyTOF) Mei, Grützkau Central Laboratory for Microscopy (Cinima) Hauser, Niesner Immune Monitoring Grützkau Ramin Labor for Molecular Rheumatology Grützkau, Häupl Animal Facility Schulz, Noe Central Laboratory Hecker-Kia 3 Liaison groups Buttgereit Glucocorticoids & Bioenergetics Diefenbach Developmental Immunology Dörner B Cell Memory Hauser Immunodynamics Hiepe Autoimmunology Hutloff Chronic Immune Reactions Löhning (until ) Experimental Immunology Riemekasten (until ) Cell Autoimmunity Niesner Biophysical Analytics Romagnani Innate Immunity Scheffold Cellular Immunology Sieper (until ) Spondyloarthritides Worm Allergology Charité -Rheumatology and Clinical Immunology, Burmester Charité - Gastroenterology, Infectiology and Rheumatology, Siegmund Cooperation Partners Charité - Dermatology, Venerology and Allergology, Blume-Peytavi (acting director) Charité - Institute of Microbiology and Hygiene, Diefenbach Charité - Cluster of Excellence NeuroCure, Schmitz Robert Koch Institute, Wieler Infrastructure Administration, Scientific Coordination, IT Service, Public Relations, Lab Kitchen, Library, Reception Desk, Store March 2017

5 Introduction Word of Welcome Word of Welcome Traudl Herrhausen President of Board of Trustees Dear friends of the German Rheumatism Research Centre, 4 as President of the Board of Trustees of the German Rheumatism Research Centre (Deutsches Rheuma- Forschungszentrum Berlin, DRFZ, a Leibniz Institute), it is a pleasure for me to welcome and to invite you to study the annual report for 2015/2016. As laid down in the statutes of the DRFZ Foundation, one of the most important responsibilities of the Board of Trustees is to appoint the Scientific Director and the Administrative Director. I am delighted that both Andreas Radbruch and Petra Starke chose to extend their contracts beyond The recommendation by the Scientific Advisory Committee and the support of the responsible Senate Administration and the Charité- Universitätsmedizin Berlin were of substantial help in this matter. Maintaining continuity in the management of the DRFZ is a good basis for the upcoming evaluation by the Leibniz Association in Another essential task of the Board of Trustees is to ensure the quality of the research. First and foremost, the directorate of the DRFZ bears this responsibility. The Board of Trustees must also approve decisions about permanent employment of the scientists. We thus guide and support our scientists in order to promote their careers. Cutting-edge research on an international level is impossible without additional funding. We appreciate that the Federal German Government and the State Government of Berlin raised their financial support for the DRFZ by 2.2 million Euros per year, thus increasing the core budget to 9.5 million Euros. This allows us to attract private sponsors to support our basic research, in addition to the third-party funding from various public sources. Several private foundations have recently made possible new initiatives at the DRFZ. We are especially grateful for their trust and support. In the past two years, three professorships orchestrated by the DRFZ have been established at the Charité: in 2015, Kirsten Minden was appointed endowed Professor for Health Service Research, supported by the Rheumastiftung. In Spring 2016, Max Löhning was appointed to an endowed Professorship for Osteoarthritis Research, funded by the Willy Robert Pitzer Foundation. Chiara Romagnani was appointed Professor for Innate Mechanisms of Chronic Inflammation at the Charité within the Heisenberg programme of the German Research Council (DFG) at the end of These joint professorships document the sound and close longterm cooperation between the DRFZ and the Charité. To this effect, the Board of Trustees is grateful to the authours of this cooperation: Andreas Radbruch, the Scientific Director of the DRFZ and Gerd-Rüdiger Burmester, Director of the Medical Department, of Rheumatology and Clincal Immunology at the Charité. In establishing the concept of the Liaison Groups, they paved the way for the close integration of basic research and clinical applications. In the name of the Board of Trustees, I thank all DRFZ staff for their dedication, shown in this annual report. Enjoy reading. Yours, Traudl Herrhausen

6 Word of Welcome Introduction Word of Welcome Gerd R. Burmester, MD Professor of Medicine Director, Department of Rheumatology and Clinical Immunology Charité - University Medicine Berlin Dear friends of the German Rheumatism Research Centre, as a clinical and scientific partner of the German Rheumatism Research Centre (DRFZ), I would like to express a warm welcome to all the readers of the Annual Report. Rheumatic diseases comprise musculoskeletal and systemic autoimmune diseases (RMDs). These diseases are a massive burden for society; they are, to a high degree, responsible for work loss and premature invalidity. They are also one of the three leading reasons why a patient consults a general practitioner or a specialist. Despite major progress in the diagnosis and treatment of RMDs, such as cytokine directed approaches and cell depleting therapies, the need for new treatment is huge. The Department of Rheumatology and Clinical Immunology of the Charité, together with the Rheumatology Unit of the Campus Benjamin Franklin, joined forces with the DRFZ to meet these challenges. The collaboration among these institutions is based on the very successful interaction both in the clinic and the research laboratory. Our primary focus lies on a patient centered approach for finding new strategies in innovative therapy. Molecular and cellular disease mechanisms lead to clinical manifestations that result in a comprehensive pattern of molecular and clinical profiles, which are essential for early diagnosis and the most appropriate treatment. Another important focus is the autoreactive immunological memory that drives chronic inflammation and maintains autoimmunity. This autoreactive memory is refractory to immunosuppressive therapies. So far, it can be only eliminated by unselective immunoablative regimens followed by autologous stem cell transplantation that may lead to long-term treatmentfree remissions in refractory autoimmune diseases. Therefore, the research aims to identify targets for a selective depletion of autoreactive memory. Other fields of common research include bioenergetics of immune functions, glucocorticoid therapy, osteoporosis, and vasculitis. What do we need to do to achieve our goals? Of course, the close collaboration between basic and clinical research must continue. The joint efforts of scientists and physicians, covering all aspects of rheumatology research at the Charité and the DRFZ, serve as an excellent model for such collaborations, working hard to achieve our common ultimate goal - research to benefit the patients. Yours sincerely, Gerd-Rüdiger Burmester 5 Foto: Charité

7 Introduction Welcome Welcome Prof. Dr. rer. nat. Andreas Radbruch Scientific Director Petra Starke Administrative Director Our aim is simple: We want to cure rheumatic diseases 6 Fotos: G. Rottmann The DRFZ, a Leibniz Institute, is the only extra-academic research institution in Germany focusing on rheumatic diseases. It meets an urgent medical need. Although available treatment has improved over the last years, there is no cure for most of the rheumatic diseases yet. They still can cause considerable morbidity and mortality and most of the patients are on lifelong medication. Our aim is challenging: we want to develop curative therapies, to induce therapy-free remission. The last two years brought us a little bit closer to this aim, again. 17 of our research groups are working in the Programme Area Pathophysiology of Rheumatic Inflammation. They focus on the characteristic chronic inflammations that occur in joints, bones, muscles, tendons as well as in organs like the kidney, the skin and blood vessels. Most of the international research in rheumatology tries to understand how these inflammations develop and how they spread. We, in contrast, focus on the cells that are involved in driving the inflammation. These are mostly cells of the immune system that are actually supposed to fight pathogens. Apparently, in rheumatic diseases, some cells start to fight the own body and become pathogenic themselves, instead. Protective and pathogenic cells are constantly competing to gain control. Our goal is to identify the cell types involved and to describe their function in detail. This knowledge helps to develop new therapeutic approaches. Here, we are pursuing two strategies at the same time: one is to strengthen the protective cells so that they are able to suppress the pathogenic ones. The other strategy is to target the pathogenic cells directly and selectively while sparing the protective cells. To date, such selective therapies are not available. Lately, we have been able to identify major players. One is the pathogenic long-lived memory plasma cell which is targeted in new anti-rheumatic therapies. And even more recently, we have identified and characterised pathogenic innate lymphocytes and pathogenic T lymphocytes as targets of interest. The counterparts of these cells might also be an attractive starting point for novel therapies: Regulatory T and B cells which can keep pathogenic lymphocytes in check. Our studies show that, in order to survive, protective and pathogenic lymphocytes rely on support by cells of the surrounding tissue. We investigate these survival niches on the molecular level. Understanding the communication between lymphocytes and their environment gives us new options for therapies, not only for rheumatic diseases but also for other chronic inflammatory diseases like inflammation of the gut, the skin, or the nervous system. We are pleased that two new research groups joined the Programme Area Pathophysiology of Rheumatic Inflammation in 2016: the group Chronic Inflammation, headed by junior scientist Andrey Kruglov, and a new liaison group Developmental and Mucosal Immunology, headed by Andreas Diefenbach who strengthens our research on innate immunology. This topic has so far been mainly addressed by Chiara Romagnani, who in 2016 was appointed to a professorship at the Charité within the Heisenberg programme of the German Research Council (DFG). Two group leaders left the DRFZ in 2015: Gabriela Riemekasten was appointed full professor and director of the Clinic for Rheumatology at the University Medicine in Lübeck. Simon Fillatreau was appointed to a professorship at the Institute Necker-Enfants Malades in Paris, France. Before he moved, he had been awarded a prestigious ERC Consolidator Grant, based on his research at the DRFZ. Since 2015, the new Programme Area Regenerative Rheumatology complements our biomedical research in focusing on the chronic wear and tear joint disease osteoarthritis. This condition affects joint and bone tissue. However, what is happening on the molecular level is still enigmatic and no curative therapies are available. Furthermore, there are no therapeutic options to regenerate damaged or lost tissue. The research group Pitzer Lab Osteoarthritis Research headed by Max Löhning will meet the challenge to advance research on osteoarthritis and to improve treatments. This endowed professorship is supported by the Willy Robert Pitzer Foundation. Junior group leader Mir-Farzin Mashreghi

8 Welcome Introduction started a second research group in 2016 that will focus on therapeutic gene regulation. The Programme Area Epidemiology of Rheumatic Diseases has the mission to discover determinants of the prognosis and outcome of rheumatic diseases, to understand the complex interaction of disease-related and treatment-related risks, to assess the adequacy of health care provision and to evaluate new therapeutic options. The overarching aim is to improve the quality and the outcomes of care and thus the quality of life of people with rheumatic diseases. One example of the clinical impact of our research is the RABBIT risk score calculator for infection which is available online. It helps physicians and patients to assess how different therapies impact on the risk of infection in order to make informed treatment decisions. Since 2015, the Rheumastiftung, a joint foundation of the German Society for Rheumatology and the patients organization Deutsche Rheuma-Liga, has been supporting an endowed professorship at the Charité. Kirsten Minden has been appointed to this position and head of the research group on Paediatric Rheumatology and Health Services Research. Since 2016, Denis Poddubnyy and his liaison group Spondyloarthritides complement the Programme Area, replacing the internationally reknown research group of Jochen Sieper, who retired. Together, four research groups cover the fields of pharmacoepidemiology, health services research in paediatric and adult rheumatology and spondyloarthritides. With the longterm cohort studies established over the years, the tremendous impact of disease activity on adverse outcomes such as growth retardation in children, anterior uveitis, myocardial infarction, stroke, infections, and mortality has been verified. Changes in patient profiles reflecting constantly improved outcomes over the last decades have been shown for adult and juvenile patients. However, the burden of disease is still high. The data are essential for health policy making and supply planning in Germany. This work would not be possible without Rheumatic and musculoskeletal diseases in Germany in a nutshell: More than 100 different rheumatic and musculoskeletal diseases are described. About 1.5 million adult persons and children and young adolescents suffer from inflammatory rheumatic diseases such as rheumatoid arthritis, spondyloarthritides or vasculitis. An additional 5 million persons suffer from symptomatic, medically treated osteoarthritis. Biologic disease-modifying antirheumatic drugs that are used to treat inflammatory rheumatic diseases are the close collaboration with a network comprising more than 500 adult and paediatric rheumatologists all over Germany. A hallmark of our work here at the DRFZ is the integration into the Charité medical campus. Not only do we share joint laboratories, we also have established 10 liaison research groups jointly supported by the Charité and the DRFZ. Clinical scientists of the Charité thus directly impact on the research at the DRFZ, and also translate new scientific concepts into novel diagnostic strategies and experimental therapies. Since July 2016, a new cornerstone of this collaboration is the Leibniz ScienceCampus for Chronic Inflammatory Diseases (C3ID). The Leibniz Association is funding this integrative research centre for 4 years, initially. We think that similar mechanisms drive inflammatory rheumatic diseases and chronic inflammatory diseases of the brain, the vasculature, the skin, the gut and other organs. Comparing the diseases and translating therapeutic concepts from one disease to another will offer on top of the health expenditures of the statutory health insurances. Rheumatic and musculoskeletal diseases are the main causes of permanent work disability and medical rehabilitation measures. unique chances to identify the drivers of inflammation in each disease and to tailor therapies to individual patients. Our research would not be possible without the support of competitive research grants and private foundations, in particular the Willy Robert Pitzer Foundation and the Rheumastiftung. Our groups are part of national and international networks funded by the German Research Council (DFG), the Federal Ministry of Education and Research (BMBF), the Leibniz Association, the European Commission and the European Research Council. In this annual report 2015/2016, we introduce some of these networks and we give insight into how research works at the DRFZ. And, needless to say, we describe our recent research highlights. Enjoy reading. Andreas Radbruch and Petra Starke 7

9 Introduction Research Highlights 2015/2016 Research Highlights How the immune system gains experience. Memory CD8(+) T cells colocalize with IL-7(+) stromal cells in bone marrow and rest in terms of proliferation and transcription. Sercan Alp Ö, Durlanik S, Schulz D, McGrath M, Grün JR, Bardua M, Ikuta K, Sgouroudis E, Riedel R, Zehentmeier S, Hauser AE, Tsuneto M, Melchers F, Tokoyoda K, Chang HD, Thiel A, Radbruch A. Eur J Immunol. 2015;45(4): Open access. Research at the DRFZ has defined the bone marrow as the home of memory lymphocytes protecting us from systemic pathogens for a lifetime. Resting in terms of activation and proliferation, these memory lymphocytes are maintained individually in survival niches organized by mesenchymal stroma cells. Epigenomic Profiling of Human CD4+ T Cells Supports a Linear Differentiation Model and Highlights Molecular Regulators of Memory Development. Durek P, Nordström K, Gasparoni G, Salhab A, Kressler C, de Almeida M, Bassler K, Ulas T, Schmidt F, Xiong J, Glažar P, Klironomos F, Sinha A, Kinkley S, Yang X, Arrigoni L, Amirabad AD, Ardakani FB, Feuerbach L, Gorka O, Ebert P, Müller F, Li N, Frischbutter S, Schlickeiser S, Cendon C, Fröhler S, Felder B, Gasparoni N, Imbusch CD, Hutter B, Zipprich G, Tauchmann Y, Reinke S, Wassilew G, Hoffmann U, Richter AS, Sieverling L; DEEP Consortium., Chang HD, Syrbe U, Kalus U, Eils J, Brors B, Manke T, Ruland J, Lengauer T, Rajewsky N, Chen W, Dong J, Sawitzki B, Chung HR, Rosenstiel P, Schulz MH, Schultze JL, Radbruch A, Walter J, Hamann A, Polansky JK. Immunity Nov 15;45(5): and how a fatally experienced immune system drives chronic inflammation. SIGLEC1 is a biomarker of disease activity and indicates extraglandular manifestation in primary Sjögren s syndrome. Rose T, Szelinski F, Lisney A, Reiter K, Fleischer SJ, Burmester GR, Radbruch A, Hiepe F, Grützkau A, Biesen R, Dörner T. RMD Open Dec 30;2(2):e Siglec-1 as a surrogate marker for type I IFN responses in monocytes in a few microliters of peripheral blood outperforms gold standards in lupus diagnostics, such as autoantibody titres or the consumption of complement factors. Siglec-1 can be used for a longitudinal monitoring of disease activity to predict lupus flares. The quantitative measurement of Siglec-1 was included in the service portfolio of the common diagnostic lab Labor Berlin of the Charité. The genes of memory T lymphocytes are epigenetically imprinted, reflecting their experiences and determining their functions in recall immune responses. The imprinting also shows that memory T cells from blood and bone keep different memories. Individual T helper cells have a quantitative cytokine memory. Helmstetter C, Flossdorf M, Peine M, Kupz A, Zhu J, Hegazy AN, Duque- Correa MA, Zhang Q, Vainshtein Y, Radbruch A, Kaufmann SH, Paul WE, Höfer T, Löhning M. Immunity Jan 20;42(1): The transcription factor T-bet controls imprinting and expression of the gene for the proinflammatory cytokine interferon-g in memory T lymphocytes in a quantitative way, thus controlling the extent of inflammation. Toralf Kaiser

10 Research Highlights Introduction Qingyu Cheng T-bet expression by Th cells promotes type 1 inflammation but is dispensable for colitis. Zimmermann J, Kühl AA, Weber M, Grün JR, Löffler J, Haftmann C, Riedel R, Maschmeyer P, Lehmann K, Westendorf K, Mashreghi MF, Löhning M, Mack M, Radbruch A, Chang HD. Mucosal Immunol Nov;9(6): In a murine model of inflammatory bowel disease, experienced T lymphocytes of the Th1 type are driving the chronic inflammation, mainly by expression of chemokines attracting proinflammatory M1 macrophages, which in turn attract more Th1 lymphocytes. The inflammation is initiated by bacteria of the microbiota. While some bacteria only can induce Th1 lymphocytes, others can activate all Th lymphocytes. mir-148a is upregulated by Twist1 and T-bet and promotes Th1-cell survival by regulating the proapoptotic gene Bim. Haftmann C, Stittrich AB, Zimmermann J, Fang Z, Hradilkova K, Bardua M, Westendorf K, Heinz GA, Riedel R, Siede J, Lehmann K, Weinberger EE, Zimmel D, Lauer U, Häupl T, Sieper J, Backhaus M, Neumann C, Hoffmann U, Porstner M, Chen W, Grün JR, Baumgrass R, Matz M, Löhning M, Scheffold A, Wittmann J, Chang HD, Rajewsky N, Jäck HM, Radbruch A, Mashreghi MF. Eur J Immunol Apr;45(4): The transcription factors T-bet and Twist1, the latter a hallmark of experienced T lymphocytes from chronically inflamed tissue, are essential for survival of these cells in inflammation. They induce expression of microrna-148a, which downregulates expression of Bim, a critical death factor. The Twist1- microrna-148a-bim axis could be used to target selectively T lymphocytes driving chronic inflammation. Therapeutic ablation of pathogenic cells driving chronic inflammation The proteasome inhibitior bortezomib depletes plasma cells and ameliorates clinical manifestations of refractory systemic lupus erythematosus. Alexander T, Sarfert R, Klotsche J, Kühl AA, Rubbert-Roth A, Lorenz HM, Rech J, Hoyer BF, Cheng Q, Waka A, Taddeo A, Wiesener M, Schett G, Burmester GR, Radbruch A, Hiepe F, Voll RE. Ann Rheum Dis. 2015;74(7): Open access. The multicentric clinical trial TAVAB (Therapy of Antibody-mediated Autoimmune Diseases by Bortezomib), reflects the collaboration of the DRFZ, the Charité and the University of Freiburg. It shows drastic efficacy of the drug Bortezomib in the treatment of SLE-patients. Bortezomib is a proteasome inhibitor, licensed for treatment of multiple myeloma, which targets plasma cells. In 12 SLE-patients, refractory to conventional treatment, and treated with Bortezomib, disease activity significantly declined and remained stable for 6 months on maintenance therapies. This trial defines autoantibody-secreting memory plasma cells as a novel, critical therapeutic target in rheumatic diseases driven by pathogenic autoantibodies, and proteasome inhibition as a therapy to eliminate plasma cells. Selection and depletion of plasma cells based on the specificity of the secreted antibody. Taddeo A, Gerl V, Hoyer BF, Chang HD, Kohler S, Schaffert H, Thiel A, Radbruch A, Hiepe F. Eur J Immunol Jan;45(1): This is the first approach to target plasma cells according to the specificity of the (auto)antibodies they secrete. Plasma cells are stained in vitro or in vivo with a conjugate of an anti-plasma cell antibody fragment and an antigen of interest. Plasma cells secreting antibodies specific for the antigen decorate themselves with their antibodies, and they are killed by complement-mediated lysis or antibody-dependent cellular cytotoxicity (ADCC). 9

11 Introduction Research Highlights Can we regenerate immunregulation in chronic inflammation and allergy? Thymus-Derived Regulatory T Cells Are Positively Selected on Natural Self-Antigen through Cognate Interactions of High Functional Avidity. Kieback E*, Hilgenberg E*, Stervbo U, Lampropoulou V, Shen P, Bunse M, Jaimes Y, Boudinot P, Radbruch A, Klemm U, Kühl AA, Liblau R, Hoevelmeyer N, Anderton SM, Uckert W, Fillatreau S. * equally contributed Immunity May 17;44(5): The development of regulatory T lymphocytes, required to prevent chronic autoimmune inflammation, critically depends on the expression of their endogenous cognate self-antigen in the thymus. T cells with strong recognition of autoantigen become regulatory T lymphocytes, T cells with weaker affinity become conventional CD4+ T cells, which are kept in check by the regulatory T lymphocytes. Regulatory T Cell Specificity Directs Tolerance versus Allergy against Aeroantigens in Humans. Bacher P, Heinrich F, Stervbo U, Nienen M, Vahldieck M, Iwert C, Vogt K, Kollet J, Babel N, Sawitzki B, Schwarz C, Bereswill S, Heimesaat MM, Heine G, Gadermaier G, Asam C, Assenmacher M, Kniemeyer O, Brakhage AA, Ferreira F, Wallner M, Worm M, Scheffold A. Cell Nov 3;167(4): e16. With a new technology developed at the DRFZ, regulatory and effector T lymphocytes reacting to a given antigen can efficiently be isolated together from tissue or blood. In healthy persons regulatory and effector T lymphocytes see the same proteins of airborne antigens, in allergic persons they see different ones, allowing the effector T lymphocytes to trigger allergy. Adequate immune response ensured by binary IL-2 and graded CD25 expression in a murine transfer model. F, Lischke T, Gross F, Scheel T, Bauer L, Kalim KW, Radbruch A, Herzel H, Hutloff A, Baumgrass R. Elife Dec 30;5. Regulatory T lymphocytes require Interleukin-2 from activated effector T lymphocytes, in order to become activeted themselves and to suppress the effector T lymphocytes in turn. To prevent systemic immunosuppression, expression of Interleukin-2 is regulated in an all-or-nothing fashion, on the level of individual effector T lymphocytes. Their antigen-receptor signal is converted into a digital decision to express or not Interleukin-2, at the level of the transcription factor NFAT (nuclear factor of activated T cells). Low-dose interleukin-2 selectively corrects regulatory T cell defects in patients with systemic lupus erythematosus. von Spee-Mayer C, Siegert E, Abdirama D, Rose A, Klaus A, Alexander T, Enghard P, Sawitzki B, Hiepe F, Radbruch A, Burmester GR, Riemekasten G, Humrich JY. Ann Rheum Dis Aug 31. Based on the observation that effector T lymphocytes driving chronic inflammation do not express Interleukin-2, and thus do not activate regulatory T lymphocytes, subcutaneous low-dose IL-2 therapy was tried for patients with Systemic Lupus Erythematosus. 10 patients refractory to conventional treatment were included in the phase I/IIa study PRO- IMMUN. A remarkable expansion and activation of the Treg population was observed and disease activity decreased. Mir-Farzin Mashreghi

12 Research Highlights Introduction The impact of state-of-the-art therapies Outcome and Trends in Treatment of Systemic Juvenile Idiopathic Arthritis in the German National Pediatric Rheumatologic Database, Klotsche J, Raab A, Niewerth M, Sengler C, Ganser G, Kallinich T, Niehues T, Hufnagel M, Thon A, Hospach T, Horneff G, Minden K. Arthritis Rheumatol 2016;68(12): Disease severity and burden of illness in systemic juvenile idiopathic arthritis have improved significantly between the years 2000 to The median disease activity (JADAS-10) declined from 7 in 2000 to 2 in 2013, while the proportion of patients with inactive disease increased from 19% to 41%. At 3-year follow-up, 72% of patients with systemic JIA had inactive disease, and 77% had no functional limitations. Risk for lower intestinal perforations in patients with rheumatoid arthritis treated with tocilizumab in comparison to treatment with other biologic or conventional synthetic DMARDs. Strangfeld A, Richter A, Siegmund B, Herzer P, Rockwitz K, Demary W, Aringer M, Meissner Y, Zink A, Listing J. Ann Rheum Dis. 2017; 76(3): The risk of lower gastrointestinal perforations is significantly increased in patients treated with the IL-6 inhibitor tocilizumab. Further, clinical presentation of this rare but dangerous complication is altered, making diagnosis difficult. Association Between the Use of Oral Contraceptives and Patient-Reported Outcomes in an Early Arthritis Cohort. Albrecht K, Callhoff J, Buttgereit F, Straub RH, Westhoff G, Zink A. Arthritis Care Res (Hoboken) 2016;68(3): The Prevalence of Dental Implants and Related Factors in Patients with Sjogren Syndrome: Results from a Cohort Study. Albrecht K, Callhoff J, Westhoff G, Dietrich T, Dorner T, Zink A. J Rheumatol 2016;43(7): Patients with Sjogren s syndrome who often have problems with their teeth can safely use dental implants. 11 Impact of treatment with biologic DMARDs on the risk of sepsis or mortality after serious infection in patients with rheumatoid arthritis. Richter A, Listing J, Schneider M, Klopsch T, Kapelle A, Kaufmann J, Zink A, Strangfeld A. Ann Rheum Dis 2016;75(9): Current and even past use of oral contraceptives moderated patientreported outcomes in inflammatory arthritis. Protective effects may be induced via central nervous pathways rather than through the suppression of peripheral inflammation. Patients who experience a serious infection have a significantly lower risk to develop a subsequent sepsis or to die from the infection if they are exposed to biologic DMARDs at the time of the infection. This has the potential to change clinical practice. Anja Weiss

13 Introduction Research Projects and Networks 12 Research Projects and Networks ERC Advanced Grant Protective and pathogenic immunological memory and its organisation by stroma cells (IMMEMO) Hidden in the bone marrow: Longlived memory cells are driving chronic inflammation. We found new targets to eliminate them. Immunological memory protects us from recurrent infections and provides the basis for the use of vaccines. But the dark side is its ability to drive chronic inflammatory diseases and allergies, some of which are refractory to conventional immunosuppressive therapies. Despite its relevance, little is known about how immunological memory is generated and maintained. In the ERC-IMMEMO project, we investigated the organization and role of immunological memory both in protective immunity and in diseases mediated by the immune system. We had originally described antibodysecreting memory plasma cells that reside and rest in survival niches in the bone marrow for very long times. We now extended this concept to memory T helper lymphocytes, cytotoxic memory lymphocytes and memory B lymphocytes. We identified prominent populations of bone marrow resident resting memory lymphocytes that most likely maintain lifelong memory to systemic antigens, like measles and mumps. We identified signals involved in homing of memory cell precursors to the bone marrow, their differentiation into memory cells and their survival as resting cells. This led to the entirely novel concept of maintenance of immunological memory by persistence of memory cells in dedicated stromal niches, resting in terms of activation, proliferation and mobility, and surviving as long as they receive survival signals from their environment. The results of IMMEMO have thus fundamentally challenged the previous dogma that immunological memory is maintained by homeostatic proliferation of circulating memory lymphocytes and by memory lymphocytes residing in the epithelial tissues. We have identified molecular adaptations of memory-effector T helper cells (Th1) to chronic antigenexposure and inflammation that qualify as original and novel targets for the selective therapeutic ablation of proinflammatory Th1 cells. Above all, we have identified autoreactive memory plasma cells as the driving force and unmet therapeutic challenge of refractory inflammatory autoimmune and allergic diseases. We have developed and translated into the clinics novel concepts for the selective depletion of these plasma cells. This has already resulted in experimental clinical trials breaking the refraction of therapyrefractory SLE patients. Our results have considerable clinical impact for the treatment of immune-diseases like allergies and chronic inflammatory diseases, most of which are still not curable today. Output: 5 PhDs complete, 1 patent, >30 publications Most relevant publications: Okhrimenko et al., 2014 Sercan Alp et al., 2015 Westendorf et al., 2014 Haftmann et al., 2015 Zehentmeier et al., 2014 Taddeo et al., 2015 ERC Advanced Grant IMMEMO PI s at DRFZ Funded by Radbruch, Chang European Research Council Funding period erc-advanced-grant-immemo/ Be the Cure for Rheumatoid Arthritis (BTCure) BTCure is the largest European program aiming at the development of new therapies against RA. It brings together pharmaceutical and academic rheumatology research. A better understanding of the molecular mechanisms causing RA will help to develop new diagnostic tools. The goal is to identify new targets and biomarkers that will benefit patients in the future. Our scientists focus on the exploration of the mechanisms responsible for the pathogenic memory of T- and B-cell subsets, most probably driving the chronicity of autoinflammatory rheumatic diseases. The main goal is to identify and characterize particular cell subsets responsible for a pathogenic immunological memory and those involved in the chronification of rheumatic diseases by global molecular approaches. New biomarkers for monitoring and therapeutical targeting of these cell subsets will be identified, aiming to reset the dysregulated immunological tolerance. The DRFZ also plays an important role in the development of new techniques and standard operating procedures for cellular and translational medicine. So far, major achievements of the DRFZ within BTCure are:

14 Research Projects and Networks Introduction Identification of an inhibitory micro RNA (mir-148a) that is involved in the longevity of pathological T helper lymphocytes (Haftmann et al., 2015) Development of a strategy to deplete autoantibody-secreting plasma cells (Taddeo et al., 2015) Development of a standardized procedure for a genome-wide characterization of effector T cell subsets (Westendorf et al., 2014) Identification of protective and pathogenic type I interferon signatures in viral infection and autoimmunity (Kyogoku et al., 2013) Characterization of a new biomarker (SIGLEC-1) for monitoring disease activity in SLE (Rose et al., 2013) Identification of disease-associated gene signatures in monocytes of RA and SLE patients that are potential biomarkers (Smiljanovic et al, 2012) Establishment of mass cytometry (CyTOF) to identify cellular biosensors for therapy monitoring and diagnostics in blood, synovial fluid and urine samples (Baumgart et al., 2017; Schulz et al., 2017). BT Cure will be completed in March 2017, but a new successor IMI consortium has been already formed, called RTCure (Rheuma Tolerance for Cure), which focuses on the development of immune tolerance therapies for the treatment of rheumatic diseases. The RTCure application has been already successfully evaluated and will be probably start in May The DRFZ is heading a work package aiming to elucidate critical immune reactions driving chronic rheumatic inflammation. This knowledge will be used to define novel targets and pathways that can be addressed therapeutically to achieve immune tolerance. In addition, we will be responsible for the development of new tools, which will allow immune monitoring of tolerance therapies and identifying cellular signatures for companion prognostics. BT Cure PI s at DRFZ/ Charité Coordinator Partner Funded by Funding period Radbruch, Burmester, Chang, Häupl, Grützkau UCB; Karolinska Institutet, Sweden; Leiden University Medical Centre 24 academic and 14 industrial partners Innovative Medicines Initiative (IMI) 13 Since 2013 the German Research Council (DFG)-funded Transregional Collaborative Research Center (TRR) 130 B cells: Immunity and Autoimmunity brings together experts in the field of B cell immunology from 5 different places in Germany (Erlangen Nuremberg, Berlin, Freiburg, Göttingen, Ulm). Research within this program focuses on a better understanding of the role of B lymphocytes during physiologic immune responses as well as chronic inflammatory pathologies. The DRFZ contributes to this consortium with its expertise in the field of memory B lymphocytes, especially memory plasma cells, in biomedical and clinical context. Furthermore, it provides optical technology development, i.e. flow cytometry and intravital microscopy. The main achievements of the DRFZ groups are summarized below: B cells: Immunity and Autoimmunity Collaborative Research Center 130 In the project Selective plasma cell targeting an affinity matrix to selectively target pathogenic (auto) antigen-specific plasma cells was developed. The method has potential relevance in the treatment of systemic lupus erythematosus since in contrast to existing therapies such as bortezomib protective humoral memory remains unaffected (Taddeo et al 2015). The project Dynamics of memory plasma cell niches characterizes stable and transient components of memory plasma cell niches in the bone marrow and their evolution (Zehentmeier et al 2014). Furthermore we started to identify the mechanisms influencing the survival and function of memory plasma cells. In order to better understand these mechanistic aspects, an artificial plasma cell survival niche mimicking the in vivo conditions has been developed. In the project Analysis of B cell dynamics in chronic neuroinflammation we for the first time described that memory plasma cells are maintained in the central nervous system of mice suffering from Experimental Autoimmune Encephalo-myelitis, a mouse model of chronic multiple sclerosis. Moreover, the concept of tissue oxidative stress memory was defined, i.e. tissue oxidative stress persisting even in the recovery phase of the disease (Radbruch et al 2016). Currently the link between possibly pathogenic longlived plasma cells and oxidative distress in chronic neuroinflammation is investigated. In the project Deciphering the antibody-independent functions of antibody-producing cells in autoimmune diseases the effector role of cytokines produced by B lymphocytes was elucidated in various autoimmune pathologies

15 Introduction Research Projects and Networks Andrey Kruglov 14 with a special focus on chronic neuroinflammatory diseases (Shen and Fillatreau 2015). A key role of vitamin D in triggering distinct differentiation pathways of B lymphocytes determining pathogenic or regulatory functions in chronic inflammatory diseases was determined in the project Regulation of B cell differentiation by vitamin A and D (Hallau et al 2016 and Heine et al 2014). The central project Dynamic multi-photon microscopy and microendoscopy quantifying communication and function of B lymphocytes in vivo develops new techniques and provides expertise in intravital microscopy. This allows the consortium to better study the role of B lymphocytes in their natural environment the living organism (Mossakowski et al 2015 and Keller et al 2017). Currently, the application for a second funding period is under review. SFB TRR 130 PI s at DRFZ/ Charité Coordinator Partner Funded by Hiepe, Radbruch (vice speaker), Hauser, Fillatreau, Worm, Niesner Lars Nitschke, University of Erlangen-Nurenberg 6 academic partners German Research Council (DFG) Funding period de/ Molecular adaptations of pathogenic memory T helper cells in chronic inflammation - Ideenwettbewerb Rheumastiftung The Rheumastiftung, funded by the German Society for Rheumatology and the patient organization Deutsche Rheuma-Liga, supports innovative scientific projects aiming at the development of novel therapeutical approaches for rheumatic diseases. In this project we are investigating effector/memory T helper (Th) cells in the context of chronic inflammatory diseases. Th cells control and enhance immune reactions through the production of interleukins and chemokines, the molecules by which cells of the immune system communicate with each other and with cells of the tissue. They are essential for protective immunity but can also initiate and maintain chronic inflammation. Clinical trials aiming at the depletion of all Th cells in chronic inflammatory diseases were unsuccessful so far, as they did not distinguish between anti- and proinflammatory Th cells and were also accompanied by loss of protective immunity. We want to understand how protective and anti-inflammatory memory T helper cells can be distinguished from memory T helper cells that are driving rheumatic diseases with the ultimate goal to selectively deplete pathogenic T helper memory cells. Previous studies from our group have indicated that Th cells involved in chronic inflammation, i.e. cells which have a history of chronic reactivation, undergo changes in gene expression, which promote their persistence and function in the inflamed tissue. In order to find biomarkers that are specifically expressed by pro-inflammatory and pathogenic memory Th cells, we are comparing the gene expression profiles of memory Th cells isolated directly from inflamed joints of patients with rheumatic diseases to those of protective memory Th cells isolated from the blood of healthy individuals. Candidate biomarkers identified in this comparison will be tested whether they qualify as targets to selectively deplete pro-inflammatory memory Th cells in preclinical models of chronic inflammation. By this approach we hope to develop new drugs to treat rheumatic diseases. Ideenwettbewerb Rheumastiftung PI s at DRFZ Funded by Chang Rheumastiftung/ Rheuma-Liga Funding period

16 Research Projects and Networks Introduction DEEP- The German epigenome programme DEEP is the German contribution to the International Human Epigenome Consortium which aims at generating genome-wide epigenetic maps for numerous cell types. Although all cells in the human body share the identical genome, the cells differentiate into an array of diverse cell types expressing a distinct and characteristic set of genes. Epigenetic signatures - based on modifications of the DNA and of DNAbinding proteins - structure the genome and license or restrict the transcriptional activity of genes. Analyzing the epigenetic landscape of a cell population reveals substantial information on its life-style and functional capabilities and highlights essential genes characteristic for a given phenotype. This allows insights into cellular physiology of unprecedented depth. DEEP aims at supplying and interpreting up to 100 epigenomes of primary human cells relevant in metabolic and in inflammatory diseases. It brings together experts on the generation of epigenomic data, on bioinformatic analysis of large and complex data-sets and scientists and clinicians investigating metabolic diseases and inflammatory diseases. At the DRFZ, we focus on the epigenetic regulation of memory T helper (Th) lymphocytes under healthy conditions as well as in rheumatoid arthritis (RA). The project has 3 main goals: How do epigenetics shape the development of CD4+ T memory cells under healthy conditions? What are critical regulators? How do effector/memory T cells alter to chronically activated cells in inflammatory diseases? What is the role of epigenetic imprinting? Which critical regulators under epigenetic control might identify novel therapeutic targets? Together with partners at the Charité, we have developed standardized workflows for sample acquisition, cell sorting and sample processing and have delivered highly purified T cell populations from healthy individuals and rheumatic patients for the generation of the epigenomic datasets. We completed the analyses of the T memory fractions from healthy donors and made several exciting findings which we published in a recent study in IMMUNITY (Durek et al, IMMUNITY, 2016): CD4+ T memory circulating subsets are generated in a consecutive order TN-TCM-TEM- TEMRA while tissue-resident subsets branch-off early and display a specific epigenomic imprint Several known molecular players involved in memory differentiation are under epigenetic expression control The comparative analyses of the epigenomic signatures revealed new candidate genes which might be controlling T memory formation A progressive heterochromatinassociated loss of DNA methylation accompanies memory T cell formation, which might qualify as a cellular senescence marker. In addition to this comprehensive study, we also utilized our datasets for the analysis of molecules which are involved in guiding the migration E p i g e n o m e Integrative analyses enable the identification of lineage regulators for CD4 + memory T cell subsets (e.g FOXP1). behavior of T cells (Pink et al, J. Immunol, 2016) and contributed to a DEEP-study analyzing the interesting class of bivalent promoters (Kinkley et al, Nature Communications, 2016). Currently, we are analyzing the datasets of T cells from rheumatic patients, to find mechanisms and molecular players which might be involved in the alternative differentiation of memory T cells under chronic inflammatory conditions. DEEP PI s at DRFZ Partners at DRFZ/ Charite Coordinator Partner Funded by Funding period Tn Tcm Tem Temra FOXP1 FOXP1 Gene X BM- Trm Gene X Consecutive changes in the epigenome and transcriptome govern differentiation of CD4 + memory T cells. Polansky, Hamann Chang, Dong, Radbruch, Sawitzki, Siegmund, Sieper, Syrbe Jörn Walter, Saarland University, Saarbrücken, Germany 19 academic and 3 industrial partners Federal Ministry of Education and Research (BMBF) Gene Y Gene Z Gene X Gene Y Tr a nscript ome 15

17 Introduction Research Projects and Networks 16 Sysinflame - A Systems Approach to Chronic Inflammatory Diseases sysinflame is a national research network and part of the program e:med, founded to accelerate the system-oriented research of diseases and preventive measures by linking life and information sciences. SysINFLAME is focusing on chronic inflammatory disorders (CID), which comprise a group of indications with fast growing expenses due to rising incidence and large unmet need. The exploration process for a systemsbased medicine in CID has three main directions: Disease manifestation: The mechanism by which disease susceptibility precipitates health into a particular phenotype at a particular time point is unclear. Following high risk populations into manifestation events and describing molecular network events associated with the shift from sub-clinical to overt disease, will enable the design of early health maintenance or pre-emptive therapeutic interventions on an individual level. Disease progression and comorbidities: Multidimensional molecular data, biospecimen and high-resolution phenotype data will lead to a better understanding of regulatory networks and events that drive complicated disease behavior and comorbidities and enable the clinicians to develop a better use of existing drugs. Therapy Response: In a timeresolved study the response to targeted anti-cytokine therapies will be analyzed. Understanding kinetics of transcriptional network changes associated with response and non-response to different biologics will generate insights into pathways that are modulated by the different compounds and define the quality of a molecular remission as an important future target. This will also provide a good starting point for developing sound biomarkers that may predict therapy response and steer clinical therapy. The main contribution of the DRFZ is to provide high dimensional immunophenotyping data generated by multiparametric flow and mass cytometry (CyTOF) technologies on peripheral leukocytes responding to targeted therapies in c h r o n i c inflammatory conditions. This approach will provide an exceptional opportunity to provide new immunophenotypic data on anti- TNF- studies in rheumatoid arthritis, ankylosing spondylitis and Crohn s disease, which offer a new dimension in systems-oriented clinical research. The main achievements of the first funding period were (1) identification of a NK-cell subpopulation that correlates with responsiveness to TNF-a blockers in ankylosing spondylitis; (2) establishment of a mass cytometry antibody panel (CyTOF OMIP) to analyze peripheral blood cells by 30 cellular parameters, (3) introduction of silver nanoparticles as high sensitivity probes in mass cytometry and (4) establishment of a protocol to stabilize whole blood samples for long-term storage at -80 C. The first funding period of sysinflame has been finished at the end of Fortunately, the BMBF has agreed to support a second funding period, sysinflame phase 2, which has started in February of sysinflame PI s at DRFZ Radbruch, Grützkau Stefan Schreiber, Coordinator University Hospital Kiel Partner 7 academic partners e:med - Maßnahmen zur Etablierung der Funded by Systemmedizin; BMBF Funding period sysinflame/tp-6/ Ute Hoffmann

18 Research Projects and Networks Introduction T-sys Systems biology of T helper cells: immunomodulation not just immunosuppression T cells are key mediators of immunity and tolerance. Dysregulation of T cell development and functions can lead to autoimmune diseases and allergies affecting a large part of the world s population. A comprehensive understanding of T cell processes by applying a systems biology approach will help to develop more sophisticated pharmacological intervention strategies. Our network developed out of an already existing nucleus for systems biology of immune cells and focuses on identification of decision-making processes in Th cells. Our aim is to identify hubs, autoregulatory loops, and limiting processes in Th cell activation and cell fate decision to discover potential sensitive targets for effective and specific manipulation of Th cells. We concentrate not only on immunosuppression, but also on immunomodulation of Th cells. This could provide an opportunity not only to block the immune system but in addition to interfere with its dysregulation in diseases. The ultimate goal of such interference will be to balance the homeostasis of the immune system. In close cooperation four interdisciplinary work packages, containing at least one experimental work group and one theoretical work group each, examine transcription factor networks, digital decision making, microrna (mirna) pro-files and functional imprinting of gene expression during Th cell activation as well as differentiation. So far we have obtained the following main results: Identification of T helper cell subtype-specific enhancers and super-enhancers (Fang et al. 2015) Discovery that mir-148a controls survival of Th1 cells by regulating Bim expression (Haftmann et al. EJI 2015). Identification of a quantitative IFN-g memory in individual Th1 cells (Helmstetter et al. 2015) Development of a protocol for delivery of antagomirs into primary B and T-lymphocytes (Haftmann C et al. JIM 2015) Construction of a STAT6- associated Th2 network (Jargosch et al. 2016) Identification of T cell-mediated vascular rejection after kidney transplantation by the combined measurement of 5 specific micrornas as biomarkers in blood of patients. (Matz et al. 2016) Discovery that binary IL-2 expression ensures a wide linear antigen response range in vivo (Fuhrmann et al. 2016) T-sys PI s at DRFZ/ Charité Coordinator Partner Funded by Baumgrass, Radbruch, Löhning Baumgrass 8 academic and 1 industrial partner BMBF, e:bio - Innovationswettbewerb Systembiologie Funding period netzwerke/t-sys-systembiologie-von-thelferzellen/ 17 Targeting neuromodulators expressed in proinflammatory T cells - NEUROIMPA Musculoskeletal disorders are debilitating diseases drastically reducing the quality of life of affected patients. The pain these pathologies are causing can often not be managed adequately. The Neuroimmunology and Pain consortium (short: NEUROIMPA), being part of the Forschungsnetz Muskuloskelettale Erkrankungen, aims at an understanding of how the interaction of cells of the immune system and the nervous system regulates the generation and maintenance of inflammation associated pain. At the DRFZ, we are particularly focussing on the role of effector/ memory Th lymphocytes in the generation and maintenance of pain in chronic inflammation. We have already collected evidence which demonstrate that different types of Th lymphocytes differ in their capacity to induce inflammatory pain. Based on transcriptome analyses we have identified several candidate molecules which could mediate the sensitisation of sensory neurons in inflammation. It is the aim of this project to better understand on a molecular level the interaction of Th lymphocytes with neurons in the inflamed tissue. We do this by interfering with the expression or action of candidate molecules and testing the consequence on pain. The results of this project and the consortium as a whole will lead to a better understanding of the mechanisms of inflammatory pain generation and the development of therapeutic strategies for pain management in inflammatory rheumatic diseases. NEUROIMPA PI s at DRFZ Chang, Radbruch Coordinator Hans-Georg Schaible, Universitätsklinikum Jena Partner 7 academic partners Funded by BMBF Funding period

19 Introduction Research Projects and Networks Bimba Hoyer Proclair and Metarthros 18 The Federal Ministry of Education and Research has been funding eight research networks in Germany since The epidemiologists coordinate the research network PROCLAIR (linking patient-reported outcomes with claims data for health services research in rheumatology) and participate in METARTHROS (metabolic impact on joint and bone diseases). In PROCLAIR, persons insured in the BARMER GEK health insurance with claims diagnoses of rheumatoid arthritis, ankylosing spondylitis or osteoarthritis of the hips, knees or hands answered written questionnaires pertaining to burden of illness, health behaviour and current treatment. The questionnaire reports were linked to the claims data on treatment, co-morbidity, and health care utilization. First results show significant discrepancies in access to rheumatological care and in preventive measures such as vaccination depending on the region of residence and sociodemographic factors, as well as a high burden of disease. With collaborators from the universities of Oldenburg, Dresden and Berlin, socioeconomic aspects of ankylosing spondylitis and outcomes of joint surgery in osteoarthritis are currently investigated. In METARTHROS the impact of overweight and diabetes on the outcome of juvenile idiopathic arthritis and rheumatoid arthritis has been investigated with data from five different cohort studies. PROCLAIR PI s at DRFZ Zink Coordinator Angela Zink, DRFZ University Oldenburg, Partner BARMER GEK, Charité, University Dresden Forschungsnetz Funded by Muskuloskelettale Erkrankungen, BMBF Funding period METARTHROS PI s at DRFZ Coordinator Partner Funded by Zink, Sieper Georg Schett, University of Erlangen-Nuremberg, Erlangen Charité, Deutsches Diabetes Zentrum Düsseldorf, University Medicine Hamburg, University Gießen, Kerckhoff Klinik Naunburg Forschungsnetz Muskuloskelettale Erkrankungen, BMBF Funding period Kirsten Minden

20 Research Projects and Networks Introduction Leibniz Research Alliance Healthy Ageing Healthy ageing living free of disease, functional decline, and disability as long as possible while remaining active until late in life should be an achievable goal for most older people in modern industrialized societies. Nonetheless advanced age still means an increased likelihood of serious illness. Only very few enjoy full health and independence in the later phases of their lives. The increased percentage of senior citizens in the population also presents its own social and economic challenges. That is why we are searching for new, effective, and practical approaches to make the life of seniors as healthy as possible. Who is the Alliance Healthy Ageing To address the multiple challenges of healthy ageing and demographic change, the LRA Healthy Ageing was formed in The LRA consists of 20 Leibniz Institutes with scientists from many different disciplines: Biology, medicine, psychology, education science, sociology, and economics. The goal of the LRA is to illuminate the biological and social factors behind the ageing process and its effects in order to develop novel strategies for intervention and adaption capable of sustainable promoting healthy ageing. It sees itself as a competent contact for politics and media concerning all biomedical and social-economic questions of ageing and demographic change. Research The main topics are: Characterizing the biological basics of ageing, understanding intrinsic and extrinsic ageing, molecular understanding of age related diseases, cognitive dysfunctions in ageing and its social impacts, interdependency between socio-economic and psycho-social life circumstances and its impact on biological ageing. Within the LRA, the DRFZ initiated the focus group Immunology and Ageing, bringing together all member institutes working in the field of immunology. Leibniz Research Alliance Healthy Ageing PI s at DRFZ: Speakers Partner: Funding Grützkau, Radbruch Prof. Dr. Jean Krutmann, Leibniz Research Institute for Environmental Medicine (IUF) and Prof. Dr. K. Lenhard Rudolph, Leibniz Institute for Age Research Fritz Lipmann Institute (FLI) 19 Institutes Leibniz 19 Leibniz Research Alliance Bioactive compounds and Biotechnology Bioactive compounds are the basis of everyday medical treatments based on pharmaceuticals. While part of them are designed by chemists, the majority was rather designed by nature in the evolution of organisms that interact with their living environment. Thus, microbes or even higher organisms might be a rich source of bioactive compounds with effects on physiology, metabolism, the immune system, or with antibiotic action. The alliance strives to exploit this treasure for application in a variety of biomedical fields. The research alliance bundles activities within the Leibniz Association focusing on: the collection of strains and biological materials as potential sources of novel compounds, the isolation, analysis, and chemical modification of bioactive compounds, the testing of potential fields of application as anti-infectives, antiinflammatory or other medical uses, or application in health products, nutrition and agriculture. Two groups of the DRFZ participate in the activities of the research alliance: AG Experimental Rheumatology (Hamann) and AG Immune Regulation (Fillatreau). Their aim is to find novel compounds that might modulate the immune system, notably those that down-regulate inflammation and upregulate inhibitory pathways involving e.g. suppressive cytokines such as IL- 10, IL-27, IL-35 or suppressive cells such as Tregs or Bregs. The alliance will be instrumental both, in finding novel sources of potential immunomodulators and in developing first hits for potential drug candidates. By this, the alliance is strengthening the translational activities within the DRFZ that aim at converting the cutting edge knowledge in immune regulation into novel tools for therapy of chronic inflammatory and autoimmune diseases. Leibniz Research Alliance Bioactive compounds and Biotechnology PI s at DRFZ: Speakers Partner: Hamann, Fillatreau Prof. Dr. Ludger Wessjohann, Leibniz Institute of Plant Biochemistry (IPB), Halle. 17 Institutes research/leibniz-research-alliances/ bioactive-compounds-and-biotechnology/

21 Introduction Leibniz ScienceCampus Berlin Chronic Inflammation Leibniz ScienceCampus Berlin Chronic Inflammation CHRONISCHE ENTZÜNDUNG Leibniz WissenschaftsCampus Berlin Leibniz ScienceCampus Berlin Chronic Inflammation 20 It is Friday noon and the seminar room at the DRFZ is filled to the very last seat. The last attendees rush through the door and grab their lunchboxes. Right on time the newly established interdisciplinary Campus Inflammation Lecture starts. The innovation of the new lecture format is, that a physician and a basic researcher give the lecture together by addressing chronic inflammatory diseases (CID) from a clinical and an experimental point of view. Physicians can register for CME points of the Berliner Ärztekammer. We will come again! the researchers note on the evaluation form which is collected at the end of the Campus Inflammation Lecture. For nearly 30 years, the Leibniz- Institute DRFZ and the Charité Universitätsmedizin Berlin have established an intimate cooperation for basic research on rheumatic diseases and its translation into clinical reality. This successful cooperation set the ground for the comprehensive center for Chronic Inflammatory Diseases, the Leibniz ScienceCampus Berlin. Scientific Challenge The ScienceCampus started in Summer 2016 and is based on the scientific understanding that highly similar cellular mechanisms drive inflammatory rheumatic diseases and CID of the brain, the vasculature, the skin, the gut, the pancreas, and other organs. Today, for most of these diseases no cure is available. In the ScienceCampus we compare these diseases in one integrative research center. We aim to translate therapeutic concepts from one disease to another and monitor their efficacy with cuttingedge biomedical technology. This offers unique chances to identify the drivers of inflammation in each disease, and define innovative ways to block them. Milestones Milestones of research are the identification of the cells and mechanisms controlling the chronicity of inflammation in inflammatory diseases of the gut and the skin, neuroinflammation, and inflammatory rheumatic diseases. Further goals are the development of biomarkers and biosignatures identifying the key players, and predicting the response to therapy for a tailored, individualized treatment based on a precision medicine approach. Beyond the scientific perspective, the ScienceCampus raises public awareness of CID as a major unmet medical entity and challenge of utmost public relevance, inflicting morbidity and mortality on 5-10% of the population. Structural Elements The Inflammation Lecture is only one of the many structural elements of the ScienceCampus. Others are e.g. the joint governance shared by the DRFZ and the Charité and joint appointments of principal investigators in Liaison Research Groups - a unique tool originally developed by the DRFZ and the Charité. Furthermore, joint grantbased research networks, joint educational programs, like the Leibniz Graduate School on Chronic Inflammatory Diseases and the Leibniz Postdoc College strengthen the scientific cooperation. Cutting-edge biomedical technology platforms and epidemiological expertise is located at the DRFZ. Several weekly scientific discussion clubs are organized under the umbrella of the Chronic Inflammation Forum. The first international symposium of the ScienceCampus Chronic Inflammation - one Disease, a thousand Manifestations will take place on November 20-21, 2017 in Berlin. Comprehensive consultation hour of CID patients for their individualized treatment For physicians in practice and their patients the Entzündungssprechstunde, an original consultation platform for joint patient management by specialists from the Departments of Rheumatology, Gastroenterology, Neurology and Dermatology, was installed. Patients suffering from CID, especially those with multiple organ manifestations, are counseled jointly by clinical specialists of inflammatory diseases of the different organs, who on the background of the ScienceCampus, have developed a comprehensive scientific and clinical understanding of CID. First general assembly of the Leibniz ScienceCampus Chronic Inflammation, September 2016, Berlin, DRFZ

22 Leibniz ScienceCampus Berlin Chronic Inflammation Introduction Scientists & Clinicians scientific discussion clubs education, lectures, talks access to core facilities gain of knowledge joint publications investigator driven trials access to patients material Patients access to the comprehensive consultation hour,,entzündungssprechstunde access to state of the art knowledge access to an interdisciplinary pool of experts increased chance of cure The strength of the ScienceCampus The benefit of the ScienceCampus is multi-layered: scientists and clinicians, students, graduates and PostDocs as well as patients and physicians profit from the tight interaction. Physicians in private practice discussion of problematic cases training in quality circles CME points contact to basic scientists and state of the art science gain of knowledge increased chances of a cure access to a pool of experts Students, Graduates, PostDocs scientific discussion clubs work in a translational environment soft skills credit points for graduates graduate school for chronic inflammation PostDoc mentoring WissenschaftsCampus Office: Dr. Elke Luger Charitéplatz 1, Berlin luger@drfz.de 21 Leibniz ScienceCampi - the underlying idea The Leibniz ScienceCampus model is the Leibniz Association s answer to the often lamented parallel existence of university and non-university research in Germany. Leibniz ScienceCampi promote cooperation on an equal footing between Leibniz institutions and universities in the form of thematically focused, complementary regional partnerships. The objective is to create networks to drive the respective research field and to strengthen the scientific environment for the relevant themes. The networks conduct strategic research, encourage interdisciplinarity in their topics, projects and methods, enhance the visibility of the respective location and sharpen its research profile. The Campus Charité Mitte in the heart of Berlin We team up with our local partners Charité Universitätsmedizin Berlin: Klinik für Rheumatologie und Klinische Immunologie, Med. Klinik mit Schwerpunkt Rheumatologie u. Klinische Immunologie Medizinische Klinik I, Medizinische Klinik für Gastroenterologie, Infektiologie, Rheumatologie Experimental and Clinical Research Center (ECRC) and Exzellenzcluster Neurocure Institut für Neuropathologie Charité Centrum Innere Medizin und Dermatologie, Klinik für Dermatologie, Venerologie und Allergologie Charité Centrum Innere Medizin mit Gastroenterologie und Nephrologie, Medizinische Klinik mit Schwerpunkt Nephrologie u. Internist. Intensivmedizin, CVK Charité Centrum Audiologie/Phoniatrie, Augen- und HNO- Heilkunde, CVK Max Planck Institute for Infection Biology

23 Introduction 2015/2016 at a Glance 2015/2016 at a Glance at a Glance About 90 Patients visting the DRFZ on the World Arthritis Day 2015 February Simon Fillatreau is awarded an ERC consolidator grant September Andreas Radbruch is elected incoming president of the European Federation of Immunological Societies (EFIS, and president ) Max Löhning is appointed endowed Professor for Osteoarthritis Research Opening of the Program Area Regenerative Rheumatology Simon Fillatreau is appointed group leader at the Institut Necker Enfants Malades, Paris Bimba Hoyer is awarded the Rudolf- Schön-Preis of the German Society for Rheumatology Advanced Cytometry Course at the 4 th European Congress of Immunology, Vienna, Austria, H.-D. Chang, A.Grützkau, A. Scheffold 22 March Gabriela Riemekasten is appointed director of the Clinic for Rheumatology in Lübeck Joachim Sieper enters retirement April Habilitation Hildrun Haibel from the Sieper group May Kirsten Minden is appointed endowed Professor for Health Services Research June Retreat of the Leibniz Graduate School for Rheumatology The Long Night of Science attracts more than visitors October 25 th Annual Meeting of the German Society for Cytometry at the DRFZ and Charité, Hyun-Dong Chang, president November Andreas Radbruch is elected spokesman of the Leibniz Section C (Life Sciences) Tobias Alexander and Bimba Hoyer win the Ideenwettbewerb Rheumastiftung December Albrecht Hasinger Lecture by Michael L. Dustin (Oxford, UK) and Award of Avrion Mitchison Prize for Rheumatology to Caroline von Spee-Mayer and Christian Neumann (both DRFZ) Andreas Radbruch is appointed member of the Advisory Board of the Research Institute for Biomedical Sciences, Tokyo University, Japan July 4 th EULAR Advanced Epidemiology Course Simon Fillatreau is awarded a research award of the GlaxoSmithKline Foundation 2 nd Symposium of the German-Chinese Association of Molecular Medicine and Molecular Pharmacology Annual on-site visit of the Scientific Advisory Board International Symposium What s Immunological Memory, organized by K. Tokoyoda and A. Radbruch together with T. Nakayama (Chiba, JP) and H. Kiyono (Tokyo, JP) from the International Immunological Memory and Vaccine Forum 2016 January Habilitation Paula Hoff from the Buttgereit group February International Meeting Immunology and Microbiology - building a bridge between Paris andberlin, organized by P. van Endert, S. Fillatreau (both Paris, FR), S.H.E. Kaufmann (MPI-IB Berlin) and A. Radbruch (DRFZ) Habilitation Bimba Hoyer from the Hiepe group Joachim Listing enters retirement

24 2015/2016 at a Glance Introduction April Denis Poddubnyy, appointed director of the Rheumatology Department at the Campus Benjamin Franklin, Charité, starts his liaison group Spondyloarthritides Andrey Kruglov starts his junior research group Chronic Inflammation at the DRFZ Opening of the Pitzer Lab Osteoarthritis Research July Mir-Farzin Mashreghi starts his junior research group Therapeutic Generegulation The Leibniz Association funds the Leibniz Science Campus for Chronic Inflammation International Symposium The Bone Marrow with 200 participants Angela Zink awarded the Carol-Nachman-Medaille for Rheumatology of Wiesbaden, Germany Retreat of the Leibniz Graduate School for Rheumatology September Visit of 22 international Postdocs Postdoc-NeT 2016, organized by the German Academic Exchange Service (DAAD) 2 nd EMBO Conference Innate Lymphoid Cells, organized by C. Romagnani, A. Diefenbach and M. Colonna (Washington, US) with 300 participants 23 Day of Immunology: 6 school classes visit the DRFZ May Chiara Romagnani appointed to a professorship at the Charité within the Heisenberg programme of the DFG June The Long Night of Science attracts visitors October 26 th Annual Meeting of the German Society for Cytometry at the DRFZ and Charité, Hyun-Dong Chang, president Annual on-site visit of the Scientific Advisory Board November Andreas Diefenbach starts his liaison group Developmental and Mucosal Immunology International Symposium: New Scales: Immune and Neuronal Diversity and Disease with 200 participants December Albrecht Hasinger Lecture by Mary Goldring (New York, US) and Award of Avrion Mitchison Prize for Rheumatology to Corinna Wehmeyer (Birmingham, UK)

25 Introduction 2015/2016 at a Glance 6 book chapters 17 others open access reviews peer reviewed Publications 2015 joint publications DRFZ - Charité 174 first authorships 56 last authorships 63 Qualifications Bachelor 5 4 Magister/Diploma/ Master Dr. med. (MD) 8 6 Dr. rer. med. 0 1 Dr. rer. nat. (PhD) Habilitation 2 1

26 2015/2016 at a Glance Introduction Publications book chapters 48 reviews 20 others 238 open access peer reviewed 25 first authorships 51 last authorships 62 joint publications DRFZ - Charité 182 Presentations talks at national meetings talks at international meetings basic funding (in million ) third party funds DRFZ groups (in million ) third party funds Liaison groups (in million )

27 Introduction Working at the DRFZ Working at the DRFZ Working at the DRFZ We are......international: About 15% of the scientists come from abroad, from 18 different countries. Excellent work life balance flexible working hours virtual desktops for home office support of parental leave 26...interdisciplinary: Physicians, biologists, chemists, physicists, veterinarians, sociologists, bioinformaticians and mathematicians join their expertise at the DRFZ....interactive: In 13 different scientific discussion clubs and seminar series the scientists discuss their work. scholarships for guest scientists 66 % female employees streams of selected talks available online 3 places in the nearby INA. Kindergarten in 2016, the DRFZ has received the Total E-Quality award for the third time Promotion of Young Researchers Since 2013, we run the Leibniz Graduate School on Rheumatology (LGRh), offering translational training for MD and PhD students. about 110 PhD and MD students (including liaison groups) 22 students enrolled in 9 graduate programmes teaching activities at several universities Photograph: U. Hoffmann, LGRh Retreat 2016 Networking with the European League against Rheumatism Together with the Division of Rheumatology and Clinical Immunology of the Charité, the DRFZ is one out of 25 European Centers of Excellence in Rheumatology. DRFZ group leaders participate in 8 Task Forces of EULAR and two FOREUM calls. Networking with the German Society of Immunology The DRFZ is a certified training center of the German Society for Immunology (DGfI). It is involved in organizing the DGfI Academy on Immunology with its Spring School, Autumn School and Translational Immunology School. (See:

28 Public Outreach Introduction Public Outreach Public Outreach Public events Regularly, the DRFZ organizes events for the public, in particular for patients. During the Berlin Long Night of Science, taking place every year in Summer, about guests visit the DRFZ and communicate with the researchers. With hands-on experiments and a guided tour they get an insight into the research at the DRFZ. Our partners from the Charité and rheumatology departments of other local clinics as well as patient organisations also contribute with information and activities on rheumatic diseases. About 80 different scientific intitutions all over Berlin participate in this event. 27 Every year in Autumn, the DRFZ opens its doors to patients on the World Arthritis Day. This event is jointly organised with the Charité and the Rheuma-Liga Berlin. The Leibniz Association The Leibniz Association connects 91 independent research institutions that range in focus from the natural, engineering and environmental sciences via economics, spatial and social sciences to the humanities. Leibniz Institutes address issues of social, economic and ecological relevance. They conduct knowledgedriven and applied basic research, maintain scientific infrastructure and provide research-based services. The Leibniz Association identifies focus areas for knowledge transfer to policy-makers, academia, business and the public. Leibniz institutions collaborate intensively with universities in the form of Leibniz ScienceCampi (thematic partnerships between university and non-university research Day of Immunology in the DRFZ, 2016 and 2017 institutes), for example as well as with industry and other partners at home and abroad. They are subject to an independent evaluation procedure that is unparalleled in its transparency. Due to the importance of the institutions for the country as a whole, they are funded jointly by the Federation and the Länder, employing some 18,600 individuals, including 9,500 researchers. The entire budget of all the institutes is approximately 1.7 billion Euros. as of May 2017 On the Day of Immunology in April, we invite school classes to our labs. The children get insight in the world of science and are doing some hands-on experiments themselves. Publishing in journals for physicians and patients For many years, Andreas Radbruch has been a co-editor of the Zeitschrift für Rheumatologie, a journal for physicians. The section Forschung aktuell updates on new findings in basic and clinical research. In addition, Andreas Radbruch edits a section in Mobil, a journal for patients published by the Deutsche Rheuma- Liga. Since 2014, this section has been informing more than readers about research highlights and new therapeutic concepts. Media Coverage Activities and research results of the DRFZ are frequently covered in the media, with a recorded average of about 300 hits a year.

29 Introduction Organisation Organisation 28 Organisation The DRFZ was founded 1988 as a foundation of the civil law is a member of the Leibniz Association since 2009 ( has 207 employees (163 scientists and 44 infrastructure staff) has 6 endowed professorships with the Charité: - Andreas Radbruch, Experimental Rheumatology (1998) - Angela Zink, Epidemiology of Rheumatic Diseases (2003) - Falk Hiepe, Rheumatology (1997) - Anja Hauser, In Vivo Imaging and Immuno Dynamics (2012) - Kirsten Minden, Health Services Research (2015), funded by the Rheumastiftung - Max Löhning, Osteoarthritis Research (2015), funded by the Willy Robert Pitzer Foundation DRFZ Bodies Board of Trustees The Board of Trustees supervises the management of the DRFZ and ensures the maintenance of research quality. It determines the framework concerning the realisation of the foundation s purpose. Permanent members: Immanuel-Krankenhaus GmbH, Berlin, Germany, Prof. Dr. Andreas Krause Chairman of Charité - Universitätsmedizin, Berlin, Germany Prof. Dr. Karl-Max Einhäupl Federal Ministery of Education and Research (BMBF), Germany - represented by Dr. Julia Dahlkamp The Mayor of the City of Berlin, Senate Department for Natural, Material and Life Sciences, Germany - represented by Dr. Björn Maul Elected members: Traudl Herrhausen, Bad Homburg, Germany, President of the Board of Trustees Univ. Prof. Dr. Georg Stingl, Leiter der klin. Abt. für Immundermatologie und Infektiöse Hautkrankheiten, Allgemeines Krankenhaus Universitätskliniken, Vienna, Austria Prof. Dr. med. Reinhold E. Schmidt, Geschäftsführender Direktor des Zentrums Innere Medizin, Klinik für Immunologie und Rheumatologie, Medizinische Hochschule Hannover, Germany Prof. Dr. Elisabeth Märker- Hermann, Klinik für Innere Medizin IV der HSK Dr. Horst Schmidt Kliniken GmbH, Wiesbaden, Germany Prof. Dr. Erika Gromnica-Ihle, President Deutsche Rheuma-Liga, Berlin, Germany Prof. Dr. Jörg Hacker, President Leopoldina, Halle/Berlin, Germany Dr. Helmut Häuser, Willy Robert Pitzer Stiftung, Frankfurt am Main, Germany Scientific Advisory Committee The principal task of the Scientific Advisory Committee (SAC) is to advise the Board of Trustees regarding scientific questions. The SAC is made up of internationally renowned scientists acknowledged for their research in the field of rheumatism and in the related basic research. Members Professor Iain McInnes, Instiute of Immunology, Infection and Inflammation College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, Scotland, UK Professor Brigitta Stockinger, The Francis Crick Institute, London, UK Professor Helen Foster, Institute of Cellular Medicine, Musculoskeletal Research Group, The Medical School, Newcastle University, Newcastle upon Tyne, UK Professor John Isaacs, Institute of Cellular Medicine, Medical School, Newcastle University, Newcastle upon Tyne, UK Professor Stefan Meuer, Institute of Immunology, Ruprecht Karls- Universität Heidelberg, Germany Professor Johan Askling, Enheten för klinisk epidemiologi, Institutionen för medicin (Solna), Karolinska institutet, Stockholm, Sweden

30 Organisation Introduction 29 Campus Charité Mitte, left: the building of the Max-Planck Institute for Infection Biology and the DRFZ. Right: the Charité Campus housing the Epidemiology Unit. Administration and Technical Staff Administration Assistant to the Business Director Holger Schnurre Travel costs/ Secretary Marion Nowak Personnel Department Teifel Büroservice, Sabine Wernhard Finance Department Norbert Adametz, Manuela Ljunggren Third Party Money Corinna Krüger, Katharina Horn, Dirk Schlienz, Annette Bastigkeit, Miriam Stephan, Alexander Hinz Purchasing Department Catja Wendt, Alexander Hinz Scientific Services Scientific Director s Office Christine Raulfs, Tanja Durez Science Management Dr. Elke Luger, Dr. Eva-Juliane Kreiß Dr. Katrin Moser Public Relations Jacqueline Hirscher, Hilmar Fünning Library Beate Löhr Veterinarians Dr. Anja Schulz, Astrid Puppe (on parental leave), Dr. Elena Noe Technical Services Animal Facilities Sabine Gruczek, Manuela Ohde, Maria Hoffmann, Patrick Thiemann, Christiana Kasprzak, Vivien Theißig, Animal Caretaker Trainees Claudia Arndt, Julian Frey Stores Carsten Tressin Lab Kitchen Regina Schuck, Angela Lindner, Birgit Füßel IT Services Antje Klauß, Falk Neumann, Hilmar Frank, Hilmar Fünning, Andreas Chochrjakow Reception Desk Wolfgang Höhn Officers Job Safety Robert Günther Equal Opportunity Dr. Elke Luger, Heike Hirseland Data Security Dina Schreckeis Ombudsman Fritz Melchers Animal Welfare Dr. Elena Noe As of April 2017

31 DRFZ groups Ria Baumgrass 32 Alf Hamann 38 Andrey Kruglov 46 Hiromi Kubagawa Sergei Nedospasov 50 Raluca Niesner 52 Andreas Radbruch 54 Koji Tokoyoda 60 Liaison groups Frank Buttgereit 34 Thomas Dörner 36 Anja Hauser 40 Falk Hiepe 42 Andreas Hutloff 44 Chiara Romagnani 56 Alexander Scheffold 58 Margitta Worm 62

32 Programme Area Pathophysiology of Rheumatic Inflammation Baumgrass, Signal Transduction Buttgereit, Glucocorticoids and Bioenergetics...34 Dörner, B Cell Memory Hamann, Experimental Rheumatology Hauser, Immune Dynamics Hiepe, Autoimmunology Hutloff, Chronic Immune Reactions...44 Kruglov, Chronic inflammation Kubagawa, Humoral Immune Regulation Nedospasov, Inflammation Biology...50 Niesner, Biophysical Analytics...52 Radbruch, Cell Biology...54 Romagnani, Innate Immunity Scheffold, Cellular Immunology Tokoyoda, Osteoimmunology Worm, Allergology... 62

33 Pathophysiology of Rheumatic Inflammation Baumgrass, Signal Transduction Regulation of cytokine expression in different T cell subsets Baumgrass, DRFZ group Prof. Dr. rer. nat. Ria Baumgrass Signal Transduction 32 Keywords T cell activation, Cytokines, Transcription factors, T cell subsets, Multiparametric flow cytometry data analysis Group Member Group leader Prof. Dr. rer. nat. Ria Baumgrass Scientists Dr. Timo Lischke, Dr. Melanie Venzke PhD students Anna Abajyan, Christian Gabriel, Manja Jargosch, Katharina Hecklau, Martin Karl, Stefanie Gryzik, Yen Hoang Bachelor/Diploma/ Master students Julia Schubert, Janet Pubanz, Markus Göthel, Mykola Ivanov, Merce Conill-Cortes, Isabelle Kadner, Matthias Sikora, Anna-Floriane Hennig, Ela Gralinska T lymphocytes are a crucial part of the adaptive immune system and play an important role in pathogen defense. However, their malfunction is related to various autoimmune and allergic disorders. Our group is investigating molecular mechanisms of lymphocyte activation and activation-induced cell fate decisions to understand the role of T cells in chronic inflammatory diseases (such as Rheumatoid Arthritis and Systemic Lupus Erythematosus). We focus on the regulation of cytokine expression in different T cell subsets, digital (all-or-none responses as opposed to analog/graded) decision-making, and transcription factor networks under physiological and pathological conditions. In order to identify so far unknown or to characterize already known hubs playing a role in chronic inflammation we characterized IL-2 action in vivo, identified NFAT-associated proteins by SILACbased mass spectrometry, created STAT6- and STAT4-associated TF- networks, discovered T helper cell subtype-specific enhancers and super-enhancers, and analyzed the interplay of cytokine expression in disease-associated Th cell subpopulations. Furthermore, we are working on novel approaches for analysis and visualization of multi-parametric flow cytometry data. Our findings will contribute to a better understanding of physiological and pathophysiological processes of Th cell subsets. Moreover, the characterization of decisive steps and identification of critical molecules during Th cell activation as well as discovery of certain T cell subsets with a specific cytokine pattern are of potential value for diagnostic/ prognostic purposes and therapeutic manipulations in autoimmune and allergic diseases.

34 Baumgrass, Signal Transduction Pathophysiology of Rheumatic Inflammation 33 Cooperation Partners Dr. Wei Chen; Max-Delbrück-Centrum für Molekulare Medizin, Berlin Prof. Hanspeter Herzel, Dr. Fridolin Gross; Humboldt-Universität, Institut für Theoretische Biologie, Berlin Selected Publications Fuhrmann, F., T. Lischke, F. Gross, T. Scheel, L. Bauer, K.W. Kalim, A. Radbruch, H. Herzel, A. Hutloff, and R. Baumgrass Adequate immune response ensured by binary IL-2 and graded CD25 expression in a murine transfer model. Elife 5: elife Walter, A. Hamann, and U. Syrbe Imprinting of Skin/Inflammation Homing in CD4+ T Cells Is Controlled by DNA Methylation within the Fucosyltransferase 7 Gene. J Immunol 197: Dr. Eberhard Krauise, Dr. Heike Stephanovitz, Leibniz-Institut für Molekulare Pharmakologie (FMP), Berlin Prof. Ulf Leser; Stefan Kröger; Humboldt-Universität, Institut für Informatik, Berlin Dr. Detlef Groth; Universität Potsdam Dr. Johannes Schuchard, Ivo Bachmann; Microdiscovery GmbH, Berlin Prof. Margitta Worm, Prof. Gabriela Riemekasten, Prof. Britta Siegmund, Dr. Uta Syrbe; Charité Berlin Gabriel, C.H., F. Gross, M. Karl, H. Stephanowitz, A.F. Hennig, M. Weber, S. Gryzik, I. Bachmann, K. Hecklau, J. Wienands, J. Schuchhardt, H. Herzel, A. Radbruch, E. Krause, and R. Baumgrass Identification of Novel Nuclear Factor of Activated T Cell (NFAT)-associated Proteins in T Cells. J Biol Chem 291: Fang, Z., K. Hecklau, F. Gross, I. Bachmann, M. Venzke, M. Karl, J. Schuchhardt, A. Radbruch, H. Herzel, and R. Baumgrass Transcription factor co-occupied regions in the murine genome constitute T-helper-cell subtype-specific enhancers. Eur J Immunol 45: Jargosch, M., S. Kroger, E. Gralinska, U. Klotz, Z. Fang, W. Chen, U. Leser, J. Selbig, D. Groth, and R. Baumgrass Data integration for identification of important transcription factors of STAT6- mediated cell fate decisions. Genet Mol Res 15: Pink, M., B.A. Ratsch, M. Mardahl, P. Durek, J.K. Polansky, M. Karl, R. Baumgrass, S. Wallner, C. Cadenas, K. Gianmoena, S. Floess, W. Chen, K. Nordstroem, S. Tierling, S. Olek, J.

35 Pathophysiology of Rheumatic Inflammation Buttgereit, Glucocorticoids and Bioenergetics How do cells adapt to oxygen deficiency and a lack of nutrients in the inflamed tissue? Liaison group with Prof. Dr. med. Frank Buttgereit Glucocorticoids and Bioenergetics 34 Keywords Bioenergetics of immune functions, Hypoxia and angiogenesis, Glucocorticoids, Autoimmune Diseases, Fracture healing Group Member Group leader Prof. Dr. med. Frank Buttgereit Scientists Timo Gaber, Dr. rer. nat. (Laboratory head, scientific), Cindy Strehl, Dr. rer. nat. (Laboratory head, administrative), Paula Hoff, PD. Dr. med., Sandra Hermann, Dr. med., Robert Biesen, Dr. med. PhD students Annemarie Lang, Dr. med. vet., Moritz Pfeiffenberger. M.Sc., Johannes Neuhaus, Dipl.-Ing. (FH) MD students Sarah Fügener, Edgar Wiebe, Marie-Christin Weber, Lisa Ehlers, Pierre-Louis Krauß, Yannick Palmowski,Peter Chefalo, Ph.D., Susanne Schillo Bachelor/Diploma/ Master students Peggy Kunath, Dipl.-Ing., Erik Schröder, M.Sc.,Lisa Deichgräber, M.Sc.,Janika Bartsch, Sc., Alexandra Damerau, B.Sc.,Sema Basaran, B.Sc. Technicians Manuela Jakstadt, Gabriele May Students Antje-Maria Lapschies, B.Sc., Wiktor Burdzinski Others Yuling Chen (visiting scholar) The bioenergetic balance of cellular systems determines their survival, development and functionality. In acute and chronic inflammation (e.g. rheumatoid arthritis: RA), as well as in injuries to the blood vessel system (e.g. fractures), the cellular need for nutrient and oxygen may exceed the supply. This results in local nutrient and oxygen deficiency (hypoxia). To maintain their functions, immune cells, mesenchymal stem cells (MSC) and endothelial cells that accumulate in inflammatory regions or in the fracture hematoma must own appropriate adaptation mechanisms. Important elements of cellular adaptation to hypoxia are the isoforms of the transcription factor hypoxia-inducible factor (HIF). HIF regulates cellular functionality and differentiation and plays a critical role in inflammatory and regenerative processes. Our research focuses on cellular adaptation mechanisms during inflammation. We aim to investigate the influence of hypoxia as a key factor in acute and chronic inflammation. Furthermore, we are interested in the cellular mechanisms of the inflammatory phase of tissue regeneration and their potential manipulation by therapeutics such as glucocorticoids and DMARDs (disease-modifying anti-rheumatic drugs). We demonstrated that growth and energy state of human CD4+ T helper cells are influenced differently depending on the degree of hypoxia. Physiological O2-deficiency does not influence the growth of T helper cells. However, pathophysiological hypoxia leads to a disruption of IL2-mediated signaling and the intracellular signaling pathways mediated by oxygen radicals. In addition, we demonstrated that IL-6 receptor blockade in human neutrophils causes different cellular responses depending on the availability of cellular oxygen. Neutrophils show a significantly higher activity in the uptake of pathogens under pathophysiological hypoxia than under optimal oxygen availability. A blockade of the IL-6 receptor inhibits this activity under hypoxia but not under normal levels of oxygen. This demonstrates that (I) blocking the IL-6 receptor is especially efficient in chronic inflammation which is usually characterized by hypoxia and (II) the mode of action of antiinflammatory drugs depends on the cellular microenvironment. Adaptation mechanisms of immune cells to a change in the inflammatory microenvironment differ from species to species. In particular, animal models do not reflect the exact mechanisms of inflammatory joint disease and of fracture healing in humans. Thus, an extrapolation of data from animal models to the human is hardly possible. We are developing human in vitro 3D models for musculoskeletal research. This allows us to better understand the pathogenesis of inflammatory joint disease. Replacing research on animal models with human in vitro models avoids painful interventions and stress on animals and may even lead to a better understanding of the mechanism of disease in humans.

36 Buttgereit, Glucocorticoids and Bioenergetics Pathophysiology of Rheumatic Inflammation 35 Cooperation Partners Gerd-Rüdiger Burmester, Thomas Häupl, Alexander Scheffold, Jacqueline Detert, Eugen Feist, Charité Universitätsmedizin Berlin, Department of Rheumatology and Clinical Immunology, Berlin, Germany Georg N. Duda, Katharina Schmidt-Bleek, Charité Universitätsmedizin Berlin, Berlin Brandenburg Center for Regenerative Therapies/Julius- Wolff Institut, Berlin, Germany Carsten Perka, Charité Universitätsmedizin Berlin, Center for Musculoskeletal Surgery, Berlin, Germany Yvonne Dörffel, Charité Universitätsmedizin Berlin, Department of Gastroenterology and Hepatology, Berlin, Germany Nicole Pischon, Katharina Gurzawska, Charité - Universitätsmedizin Berlin, Centrum für Zahn-, Mund- und Kieferheilkunde, Abteilung für Parodontologie und Synoptische Zahnmedizin, Berlin, Germany Andreas Radbruch, Max Löhning, Deutsches Rheuma-Forschungszentrum, Cell Biology, Berlin, Germany Roland Lauster, TU-Berlin, Medical Biotechnology, Berlin, Germany Markus Seibel, Hong Zhou, University of Sydney, ANZAC Institute, Sydney, Australia Dr. Stefan Krauss, Beth Israel Deaconess Medical Center, Harvard Medical School, and Merck Research Laboratories, Boston, USA Jon Yewdell, National Institute of Allergy and Infectious Diseases, Cellular Biology Section, Bethesda, USA Heinrich Hoffmann, Ècole polytechnique fédérale de Lausanne, Department of Materials science and engineering, Lausanne, Switzerland Magarethe Hofmann-Amtenbrink, Alessandra Hool, Mat Search Consulting, Dr.-Ing. Lausanne, Switzerland Patrik Önnerfjord, Viveka Tilgren, Lund University, Section of Rheumatology, Department of Clinical Sciences, Lund, Sweden Joachim Koitzsch, Andreas Brandt, medac GmbH, Wedel, Deutschland Douglas J. Veale, University College Dublin, The Conway Institute for Biomedical and Biomolecular Research, Dublin, Ireland A Robin Poole, McGill University, Department of Surgery, Montreal, QC, Canada Prof. Susanna Röblitz und Dr. Rainald Ehrig, Konrad-Zuse-Zentrum Berlin Dr. Dirk Barnwitz und Dr. Igor Ponomarev, fzmb GmbH Bad Langensalza Dr. Paulin Jirkof, Institut für Experimentelle Chirurgie, UniversitätsSpital Zürich, Universität Zürich Selected Publications Strehl C, Maurizi L, Gaber T, Hoff P, Broschard T, Poole AR, Hofmann H, Buttgereit F. Modification of the surface of superparamagnetic iron oxide nanoparticles to enable their safe application in humans. Int J Nanomedicine Nov 8;11: PubMed PMID: ; PubMed Central PMCID: PMC Hoff P, Gaber T, Strehl C, Schmidt-Bleek K, Lang A, Huscher D, Burmester GR, Schmidmaier G, Perka C, Duda GN, Buttgereit F. Immunological characterization of the early human fracture hematoma. Immunol Res Dec;64(5-6): PubMed PMID: Buttgereit F, Dejaco C, Matteson EL, Dasgupta B. Polymyalgia Rheumatica and Giant Cell Arteritis: A Systematic Review. JAMA Jun 14;315(22): doi: /jama Review. PubMed PMID: Gaber T, Hahne M, Strehl C, Hoff P, Dörffel Y, Feist E, Burmester GR, Buttgereit F. Disentangling the effects of tocilizumab on neutrophil survival and function. Immunol Res Jun;64(3): doi: /s x. PubMed PMID: Buttgereit F, Smolen JS, Coogan AN, Cajochen C. Clocking in: chronobiology in rheumatoid arthritis. Nat Rev Rheumatol Jun;11(6): doi: /nrrheum Review. PubMed PMID:

37 Pathophysiology of Rheumatic Inflammation Dörner, B Cell Memory Innovative insights into B Cell Memory with impact on protective and autoreactive immunity Dörner, Liaison group with Prof. Dr. med. Thomas Dörner B Cell Memory 36 Keywords antigen-specific memory B cells, human plasma cells, bone marrow, immunization, autoimmunity, SLE, BCR signaling Group Member Group leader Prof. Dr. med. Thomas Dörner Scientists Dr. rer. nat. Sarah J. Fleischer, Dr. rer. nat. Claudia Giesecke, Dr. med. Thomas Rose, Dr. med. Luisa Stefanski PhD students Dipl. mol.biol. Ina Wirries, Dipl. Ing. Annika Wiedemann, Sarah Weißenberg, M. Sc. MD students Finn Moritz Wilkens, Maximilian Koenig (derzeit John s Hopkins Univ., Baltimore, MD), Julia Sieber, Marie Lettau, Nadja Nomovi, Armin Aue, Anna Lisney Bachelor/Diploma/ Master students Franziska Szelinski, Alexander Daum Vanessa Fleischer Technicians Karin Reiter Our group has a focus on basic principles of the induction and maintenance of human memory B cells (BC) and plasma cells (PC). In particular, the humoral memory plays an important role in long-term protection against pathogens, but unfortunately also in the pathology of autoimmune diseases. We believe that the autoimmune BC memory represents a key challenge for successful therapeutic immunotargeting. Understanding the differential processes that ultimately lead to the induction and maintenance of protective versus autoreactive immunoglobulins is a central topic of our research. Therefore we address three main areas: 1) basic mechanisms in the induction and maintenance of protective, antigen-specific memory BC upon primary and secondary vaccination, 2) maintenance of distinct memory PC subsets in human bone marrow and 3) specific characteristics of memory BC in different autoimmune disorders, such as rheumatoid arthritis, primary Sjögren s syndrome and systemic lupus erythematosus. First, kinetics and characteristics of memory BC differentiation into PC in human are studied during active disease, upon immunotherapy and immunizations. In these studies we apply antigen-specific detection systems using tetanus toxoid as well as autoantigens. The identification and isolation of specific BC permit comprehensive phenotypic, molecular and functional studies. A second line of research comprises studies of various PC subsets residing in the bone marrow and extending previous findings of CD19 positive and negative human bone marrow PC. On preclinical grounds (in vitro activity and mechanism of action) and during phase 1-3 of clinical development, we study a number of monoclonal antibodies considered for therapeutic use. We analyze in detail the impact of certain therapeutics on the B cell receptor (BCR) signaling network as well as the general differences of BCR related kinases in health and disease with a special focus on the equilibrium of kinases and phosphatases. Our current hypothesis is that disturbances of signaling pathways and interrelated survival mechanisms are key to understand abnormalities in autoimmunity. The modulation of BCR responses will help us to better understand how memory BC are regulated during steady state or after activation. We aim to get further insights into the differential regulation and development of protective versus autoimmune memory BC and PC to clear the way for the development of innovative and selective therapeutics targeting BC functions.

38 Dörner, B Cell Memory Pathophysiology of Rheumatic Inflammation 37 Cooperation Partners Selected Publications M. Brisslert, PhD, Sahlgrenska Academy at Gothenburg university Hospital, Gothenburg, Sweden Prof. S. Fillatreau, PhD, Inserm, Paris, France Dr. A. A Kühl and Spieckermann S, Charité - Universitätsmedizin, Research Center Immuno- Science (RCIS) and Department of Gastroenterology, Infectiology and Rheumatology, Berlin, Germany Prof. C. Perka, Department of Orthopedics, Charité Universitätsmedizin, Berlin, Germany Prof. Dr. M. Wahren-Herlenius, Karolinska University and Institute, Stockholm, Sweden Prof. Dr. Hans-Peter Brezinschek, Medical Faculty, Univ. Graz, Austria Fleischer SJ, Daridon C, Fleischer V, Lipsky PE, Dörner T. Enhanced Tyrosine Phosphatase Activity Underlies Dysregulated B Cell Receptor Signaling and Promotes Survival of Human Lupus B Cells. Arthritis Rheumatol (5): Fleischer V, Sieber J, Fleischer SJ, Shock A, Heine G, Daridon C, Dörner T. Epratuzumab inhibits the production of the proinflammatory cytokines IL-6 and TNF-alpha, but not the regulatory cytokine IL-10, by B cells from healthy donors and SLE patients. Arthritis Res Ther :185. Mei HE, Wirries I, Frölich D, Brisslert M, Giesecke C, Grün JR, Alexander T, Schmidt S, Luda K, Kühl AA, Engelmann R, Dürr M, Scheel T, Bokarewa M, Perka C, Radbruch A, Dörner T. A unique population of IgG-expressing plasma cells lacking CD19 is enriched in human bone marrow. Blood (11): Sieber J, Daridon C, Fleischer SJ, Fleischer V, Hiepe F, Alexander T, Heine G, Burmester GR, Fillatreau S, Dörner T. Active systemic lupus erythematosus is associated with a reduced cytokine production by B cells in response to TLR9 stimulation. Arthritis Res Ther Nov 11;16(6):477. Fleischer V, Sieber J, Fleischer SJ, Shock A, Heine G, Daridon C, Dörner T. Epratuzumab inhibits the production of the proinflammatory cytokines IL-6 and TNF-α, but not the regulatory cytokine IL-10, by B cells from healthy donors and SLE patients. Arthritis Res Ther Jul 17;17:185.

39 Pathophysiology of Rheumatic Inflammation Hamann, Experimental Rheumatology Prof. Dr. rer. nat. Alf Hamann Epigenetics, molecular regulation of T cell differentiation, and novel concepts for the therapy of autoimmune/chronic inflammatory diseases Hamann, Experimental Rheumatology DRFZ group 38 Keywords Epigenetics, T memory cells, regulatory T cells, Immunoregulation, Immunotherapy Group Member Group leader Prof. Dr. rer. nat. Alf Hamann Scientists Dr. Pawel Durek Dr. Stefan Frischbutter Dr. Ute Hoffmann Dr. Matthias Pink Dr. Julia Polansky-Biskup PD Dr. Uta Syrbe* *Med. Klinik m. S. Gastroenterologie, Infektiologie und Rheumatologie Charité CBF Berlin PhD students Dipl. Biol. Elisabeth Kenngott (2016: Dr.rer.nat) M. Sci. Biochem. Christopher Kressler, Dipl. Biol. Matthias Kröger, M. Sci. Mol. Biol. Maibritt Mardahl (2016: Dr.rer. nat), Dipl. Biol. Jennifer Pfeil (2016: Dr.rer.nat), Dipl. Biochem. Julia Triebus (2015: Dr.rer.nat) Bachelor/Diploma/ Master students Melanie de Almeida Siska Wilantri Sabrina Schüngel Olga Trupp Wojciech Leszczynski Lydia Verlaat Technicians Uta Lauer René Maier Our group is focusing on two areas: I) epigenetics and II) immune regulation/translation. I) Here we study the epigenetic impact on CD4+ T cell differentiation in health and under chronic inflammatory conditions, to understand how long-term shifts in the phenotype and functional profile of T cells are maintained on the molecular level. Our work has documented the impact of epigenetic regulation for the stable expression of the master transcription factor Foxp3 in regulatory T cells and for several homing-related molecules. More recently, as members of the DEEP consortium, we established full epigenomes of human T cells and studied the role of epigenetics for T memory cell differentiation. This work: unravelled a major loss of genome-wide DNA methylation with memory development; supported a linear model of differentiation in circulating T cells in the order naïve -> Tcm (T central memory cells) -> Tem (T effector memory re-expressing cells) -> Temra (T effector memory re-expressing CD45RA cells), highlighted various known or novel transcription factors regulating the memory differentiation process and which are under epigenetic expression control. We expect that this research will help to better understand the features of T cells fuelling chronic inflammation. Moreover, it might lead to the identification of novel targets and tools for the therapeutic modulation of T cells in inflammatory diseases. II) Here we tested several approaches to strengthen self-tolerance: a) by vaccination with chemically modified peptides and, b) by vaccination with peptides coupled to a transporter module targeting it into the (tolerogenic) compartment of mucosal tissues. In addition, we searched by high throughput screening for novel compounds able to induce inhibitory cytokines of the immune system such as IL-27, IL-35, IL-10, or to induce Foxp3. Ad a) We found that coupling of peptides to synthetic carriers (PEG) or nanobeads is enhancing their tolerogenic effect and is able to inhibit the development of experimental autoimmune encephalomyelitis, a murine model for Multiple Sclerosis; yet application at the peak of disease was so far not effective. Ad b) Use of a transporter peptide improved the tolerogenic effect of local vaccination, but in total, the mucosal pathway was not as tolerogenic as predicted. Our screenings for compounds targeting inhibitory pathways delivered a number of candidates that increase EBI3 (a component of the inhibitory cytokines IL-27 and IL-35) and simultaneously down-regulated effector cytokines. In addition, one substance was found that induces Foxp3 in mouse T cells. In cooperation with S. Fillatreau, several hits were found that induced IL-10 in B cells. Thus, exploiting the portfolio of natural tolerance and inhibitory mechanisms of the immune system might provide novel approaches for the therapy of autoimmunity and chronic inflammation in diseases such as rheumatoid arthritis.

40 Pathophysiology of Rheumatic Inflammation Hamann, Experimental Rheumatology group picture from Cooperation Partners Germany Celares GmbH, Berlin Dr. Ho-Ryun Chung - Max Planck Institut für Molekulare Genetik, Berlin Dr. Thomas Hanke, Evotec AG, Hamburg Prof. Rainer Haag, Freie Universität, Institut für Chemie und Biochemie, Berlin Prof. Susanne Hartmann, Dr. Svenja Steinfelder, Institut für Immunologie, Veterinärmedizin, FU Berlin Prof. Andreas Herrmann, Humboldt-Universität, Inst. für Biologie; Molekulare Biophysik, Berlin Prof. Jochen Hühn, Dr. Stefan Flöß, Helmholtz- Zentrum für Infektionsforschung, Braunschweig Dr. Jens Peter von Kries, Dr. Marc Nazaré, Dr. Edgar Specker, Screening Unit, Medicinal Chemistry, Leibniz-Institut für Molekulare Pharmakologie (FMP); Berlin Dr. Anja Kühl, Dr. Ulrike Erben, Charité, ipath Berlin and RCIS, Berlin Dr. Thomas Manke, Einheit f. Informatik, MPI f. Immunbiologie und Epigenetik, Freiburg Prof. Thomas Schneider, Dr. Verena Moos, Prof. Dr. Britta Siegmund, Charité, Inst. f. Gastroenterologie, Infektiologie und Rheumatologie, Berlin Prof. Tim Sparwasser and Dr. Matthias Lochner, Twin Core, Institut für Infektionsimmunologie, MH Hannover Dr. Marcel Schulz, High-throughput Genomics & Systems Biology, MPI f. Informatik, Saarbrücken Prof. Joachim L. Schultze, Genomforschung und Immunregulation. LIMES-Institut, Universität Bonn Prof. Jörn Walter, Universität des Saarlandes, FB Genetik, Saarbrücken Others Prof. Gudrun Debes, Prof. Chris Hunter, University of Pennsylvania, Dept. Pathobiology, Philadelphia, USA Dr. Sara Atkisons, Dept. Diabetes Complications Res., Global Research, Novo Nordisk A/S, and Dept. Vet. Disease Biol., Univ. Copenhagen, Denmark. Dr. Anton Pernthaner, AG Research Ltd., Hopkirk Research Institute, Neuseeland Dr. Olaf Rötzschke, Singapore Immunology Network, Agency for Science, Technology and Research, Singapore Weitere Partner des DEEP Konsortiums (www. deutsches-epigenom-programm.de) Selected Publications Gupta, S., J. Pfeil, S. Kumar, C. Poulsen, U. Lauer, A. Hamann, U. Hoffmann, and R. Haag Tolerogenic modulation of the immune response by oligoglycerol- and polyglycerol-peptide conjugates. Bioconjugate chemistry 26: Pink, M., B. A. Ratsch, M. Mardahl, P. Durek, J. K. Polansky, M. Karl, R. Baumgrass, S. Wallner, C. Cadenas, K. Gianmoena, S. Floess, W. Chen, K. Nordstroem, S. Tierling, S. Olek, J. Walter, A. Hamann, and U. Syrbe Imprinting of Skin/Inflammation Homing in CD4+ T Cells Is Controlled by DNA Methylation within the Fucosyltransferase 7 Gene. J Immunol 197: Durek, P., K. Nordstrom, G. Gasparoni, A. Salhab, C. Kressler, M. de Almeida, S. Frischbutter, the DEEP Consortium, U. Syrbe, A. Radbruch, J. Walter, A. Hamann, and J. K. Polansky Epigenomic Profiling of Human CD4+ T Cells Supports a Linear Differentiation Model and Highlights Molecular Regulators of Memory Development. Immunity 45: Szilagyi, B. A., J. Triebus, C. Kressler, M. d. Almeida, S. Tierling, P. Durek, M. Mardahl, A. Szilagyi, S. Floess, J. Huehn, U. Syrbe, J. Walter, J. K. Polansky, and A. Hamann Gut memories do not fade: Epigenetic regulation of lasting gut homing receptor expression in CD4+ memory T cell. Mucosal Immunol. Advance online publication 15 February doi: /mi Kenngott, E. E., J. Pfeil, U. Hoffmann, U. Lauer, A. A. Kuehl, A. Rigby, A. Pernthaner, and A. Hamann Facilitated Peptide Transport via the Mucosal Epithelium: Impact on Tolerance induction. Front Immunol. 8:216. doi: / mmu

41 Pathophysiology of Rheumatic Inflammation Hauser, Immune Dynamics Using advanced microscopy to understand the interplay of immune cells in the living organism Hauser, Liaison group with Prof. Dr. med. vet. Anja E. Hauser Immune Dynamics 40 Keywords multiphoton microscopy, plasma cells, confocal microscopy, histocytometry, chronic inflammation Group Member Group leader Prof. Dr. med. vet. Anja E. Hauser Scientists Dr. Sandra Zehentmeier, Dr. Randall Lindquist, Dr. Karolin Pollok PhD students Carolin Ulbricht, Jonathan Stefanowski, Karolin Holzwarth, Laura Tech, Jannike Bayat Sarmadi, Anna Pascual MD students Agata Mossakowski, Magdalena Kraft, Dominik Lammerding, Julian Pohlan, Lennard Ostendorf Bachelor/Diploma/ Master students Veronica Raba, Sylvia Uhlmann, Sven Flatow, Alina Liebheit, Fabian Kriegel Technicians Robert Günther, Ralf Uecker Microscope Operator Ralf Köhler Student Markus Köhler We are interested in determining how immune cells behave in the tissue context, how they interact with other immune cells as well as with various cell types in the tissues they reside in, and how that shapes immune responses. We aim to dissect how the behavior of various immune cell subsets in various tissues affects processes such as chronic inflammation, which occurs in the course of rheumatic diseases. One focus of our lab is analyzing the biology of long lived plasma cells. As the producers of antibodies, they play a crucial role in immunological memory, securing life-long protective immune responses. The longevity of plasma cells depends on their microenvironment, which provides survival factors in tissue niches of the bone marrow. We could identify bone marrow stromal cells acting as stable components of these niches. They attract plasma cells to the bone marrow niches by secretion of CXCL12. The numbers of these stromal niche organizers remain stable during the course of an immune response, thereby limiting the number of available niches to the plasma cells, which migrate from secondary lymphoid organs to the bone marrow to become long lived. In contrast, eosinophils, which provide the survival factor APRIL to plasma cells, are transient niche inhabitants. In order to further characterize the cellular dynamics in the bone marrow niches, we have recently developed a microendoscopic implant which allows us to analyze cellular migration and interactions in the bone marrow over months by intravital microscopy. Furthermore, we could show that plasma cells in the small intestine can also become long lived, and that the microenvironment of the gut provides similar survival signals to them as the bone marrow, although the cell types that produce these factors differ between tissues. Interestingly, plasma blasts generated in mucosal immune responses can also contribute to the long-lived plasma cell pool in the bone marrow. Besides their crucial functions in protective immunity, long lived plasma cells also contribute to autoimmune diseases by secreting autoreactive antibodies that can mediate tissue destruction and the perpetuation of chronic inflammation. Using experimental autoimmune encephalomyelitis, a mouse model of multiple sclerosis, we recently demonstrated that the chronically inflamed central nervous system also provides niches for long lived plasma cells. In line with that, non-proliferative plasma cells could be found in biopsies derived from the CNS of multiple sclerosis patients. Hence, under the conditions of chronic inflammation, niches for immune memory can form even in organs that are void of immune cells in healthy individuals. A better understanding of the mechanisms underlying the formation and maintenance of those niches is crucial to therapeutically target pathogenic plasma cells.

42 Hauser, Immune Dynamics Pathophysiology of Rheumatic Inflammation 41 Cooperation Partners Selected Publications Dr. G. Duda, Charité, Justus-Wolff-Institut, Berlin, Germany Dr. H. Radbruch, Charité, Institut für Neuropathologie, Berlin, Germany Dr. E. Esplugues, Yale University School of Medicine, New Haven, CT, USA Prof. M.-T. Figge, Applied Systems Biology, HKI Jena, Germany PD Dr. U. Höpken, MDC Berlin, Germany Prof. K.-M. Toellner, University of Birmingham, Institute of Immunology and Immunotherapy, Birmingham, United Kingdom Dr. A. Triantafyllopoulou, Klinik für Rheumatologie und Klinische Immunologie, Universitätsklinikum Freiburg, Germany TH17 cells express ST2 and are controlled by the alarmin IL-33 in the small intestine. Pascual-Reguant A, Bayat Sarmadi J, Baumann C, Noster R, Cirera-Salinas D, Curato C, Pelczar P, Huber S, Zielinski CE, Löhning M, Hauser AE, Esplugues E. *equally contributing senior authors. Mucosal Immunol Feb 15. doi: /mi Herrtwich, L., I. Nanda, K. Evangelou, T. Nikolova, V. Horn, Sagar, D. Erny, J. Stefanowski, L. Rogell, C. Klein, K. Gharun, M. Follo, M. Seidl, B. Kremer, N. Munke, J. Senges, M. Fliegauf, T. Aschman, D. Pfeifer, S. Sarrazin, M. H. Sieweke, D. Wagner, C. Dierks, T. Haaf, T. Ness, M. M. Zaiss, R. E. Voll, S. D. Deshmukh, M. Prinz, T. Goldmann, C. Holscher, A. E. Hauser, A. J. Lopez-Contreras, D. Grun, V. Gorgoulis, A. Diefenbach, P. Henneke, and A. Triantafyllopoulou DNA Damage Signaling Instructs Polyploid Macrophage Fate in Granulomas. Cell 167: e1218. Lemke, A., M. Kraft, K. Roth, R. Riedel, D. Lammerding, and A. E. Hauser Long-lived plasma cells are generated in mucosal immune responses and contribute to the bone marrow plasma cell pool in mice. Mucosal Immunol 9: Automated quantification of hematopoietic cell - stromal cell interactions in histological images of undecalcified bone. Zehentmeier S, Cseresnyes Z, Escribano Navarro J, Niesner RA, Hauser AE. J Vis Exp Apr 8;(98). doi: / Radbruch, H., D. Bremer, R. Guenther, Z. Cseresnyes, R. Lindquist, A. E. Hauser, and R. Niesner Ongoing Oxidative Stress Causes Subclinical Neuronal Dysfunction in the Recovery Phase of EAE. Frontiers in immunology 7: 92

43 Pathophysiology of Rheumatic Inflammation Hiepe, Autoimmunology On new ways against autoimmune diseases Hiepe, Liaison group with Prof. Dr. med. Falk Hiepe Autoimmunology 42 Keywords Memory plasma cells, Autoantibodies, Systemic autoimmune diseases, Cellular therapies, Autologous hematopoietic stem cell transplantation Group Member Group leader Prof. Dr. med. Falk Hiepe Scientists Dr. med. Tobias Alexander, Dr. med. Robert Biesen, Dr. med. Anne Bruns, Dr. Qingyu Cheng, Dr. rer. nat. Velia Gerl, PD Dr. med. Bimba F. Hoyer, Laleh Khodadadi, PhD Adriano Taddeo, PhD PhD students Anne von Bülow, Julia-Carmen von Glischinski, Katrin Kozeck, Dominik Lammerding, Jonas Martin, Sarnai Naran, Lennard Ostendorf, Andreas Pelz, Veronika Scholz, Georg Tilgner, Theresa Wilhelm Our research group is fundamentally involved in analyzing the mechanisms which contribute to and help maintain severe autoimmune diseases, in order to develop new therapeutic concepts. Our main focus is on the role of long-lived autoreactive plasma cells in autoimmune diseases. Long-lived plasma cells reside in niches in the bone marrow and inflamed tissues, where they are resistant to immunosuppressive/ cytotoxic drugs or therapies targeting B cells. In collaboration with the research group from Andreas Radbruch, we look for new therapeutic strategies targeting the autoreactive memory. We introduced the proteasome inhibitor bortezomib that depletes plasma cells in the treatment of refractory autoimmune diseases. We also learned that selective plasma cell depletion has to be combined with a therapy targeting the plasma cell precursors. Since all these therapies unselectively deplete plasma cells regardless whether they secrete protective of pathogenic antibodies, we developed an affinity matrix technology for antigen-specific plasma cell depletion. Very recently, we could show for the first time that long-lived memory plasma cells can be depleted in an antigen-specific manner using this technology in a murine model. Together with the Unit for Bone Marrow Transplantation (Renate Arnold) at the Charité Universitätsmedizin Berlin and the research group of Andreas Thiel (BCRT), we demonstrated that the autoreactive memory could be eliminated by immunoablation followed by autologous hematopoietic stem cell transplantation, in patients with severe autoimmune diseases that are refractory to conventional immunosuppression. In most cases this provided the fundament for the subsequent regeneration of an intact immune system. In some patients, however, the disease relapsed or secondary autoimmune disorders occurred. We investigate the reasons for this in a controlled clinical trial in systemic lupus erythematosus (SLE). In another project, we study the role of dendritic cells in SLE. These cells, in their function as antigen-presenting cells and producers of cytokines, play a significant role in the pathogenesis of SLE. As they are a potential target in the development of new therapies, their characterization is of major relevance. Several cytokines are involved in the pathogenesis of SLE and other systemic autoimmune diseases. In past and future clinical trials, we have studied biologics selectively targeting different cytokines or cells (e.g. BAFF/BLys, APRIL, type I interferon, IL-10, B cells, PDC, co-stimulatory molecules). We expect that these different therapeutic approaches will allow us to develop personalized therapies. To identify the optimal therapeutic target and/or characterize disease activity adequate biomarkers are required. We have already identified several serologic and cellular biomarkers (autoantibodies, Siglec1 expression on monocytes, B and T cell subpopulations). Their relevance for the aforementioned aims is being studied in the clinic.

44 Hiepe, Autoimmunology Pathophysiology of Rheumatic Inflammation 43 Cooperation Partners Univ.-Prof. Dr. med. Renate Arnold, Charité Universitätsmedizin Berlin, CVK, Med. Klinik m. S. Hämatologie, Onkologie und Tumorimmunologie, Berlin, Germany Prof. Dr. Klemens Budde and Dr. Michael Dürr, Charité Universitätsmedizin Berlin, CCM, Medizinische Klinik m.s. Nephrologie, Berlin, Germany Prof. Dr. Richard Burt, Northwestern University, Department of Medicine, Division of Immunotherapy, Chicago, USA Univ.-Prof. Dr. med. Thomas Dörner, Charité Universitätsmedizin Berlin, CCM, Medizinische Klinik m.s. Rheumatologie und Klin. Immunologie, Berlin Univ.-Prof. Dr. Peter M. Kloetzel, Charité Universitätsmedizin Berlin, Institut für Biochemie, Berlin, Germany Univ.-Prof. Dr. Rudolf Manz, Universität zu Lübeck, Institut für Systemische Entzündungsforschung ISEF, Lübeck, Germany Prof. Dr. Edgar Meinl, Institut für Klinische Neuroimmunologie, Ludwig Maximilian Universität München, Germany Prof. Dr. Andreas Meisel and Dr. Siegfried Kohler, Klinik für Neurologie, Charité Universitätsmedizin Berlin, Germany Prof. Dr. med. Thomas Porstmann, Seramun Diagnostica GmbH, Heidesee, Germany Prof. Dr. Michael Reth, Max-Planck-Institut für Immunbiologie, Freiburg. Germany Dr. Jürgen Schmitz, Dr. Andrzej Dzionek, Dr. Barbara Behle, Miltenyi Biotech, Bergisch Gladbach, Germany Dr. Wolfgang Schlumberger, EUROIMMUN Medizinische Labordiagnostika AG, Lübeck, Germany Prof. Dr. David Tarlinton, Walter und Eliza-Hall- Institut für Medizinische Forschung, Melbourne, Australia Univ.-Prof. Dr. med. Reinhard E. Voll, Universitätsklinikum Freiburg, Med. Klinik, Abteilung Rheumatologie und Klinische Immunologie, Freiburg, Germany Prof. Dr. rer. nat. Hedda Wardemann, DKFZ, Heidelberg, Germany Univ.-Prof. Dr. med. Margitta Worm, Charité Universitätsmedizin Berlin, CCM, Klinik für Dermatologie, Venerologie und Allergologie, Berlin, Germany Selected Publications 1. Scheibe, F., H. Pruss, A. M. Mengel, S. Kohler, A. Numann, M. Kohnlein, K. Ruprecht, T. Alexander, F. Hiepe, and A. Meisel Bortezomib for treatment of therapy-refractory anti-nmda receptor encephalitis. Neurology 88: Wilhelm, T. R., A. Taddeo, O. Winter, A. R. Schulz, J. N. Malzer, C. Domingo, R. Biesen, T. Alexander, A. Thiel, A. Radbruch, F. Hiepe, and V. Gerl Siglec-1-positive plasmacytoid dendritic cells (pdcs) in human peripheral blood: A semi-mature and myeloid-like subset imbalanced during protective and autoimmune responses. Clin Immunol 163: Hiepe, F., and A. Radbruch Plasma cells as an innovative target in autoimmune disease with renal manifestations. Nat Rev Nephrol 12: Taddeo, A., V. Gerl, B. F. Hoyer, H. D. Chang, S. Kohler, H. Schaffert, A. Thiel, A. Radbruch, and F. Hiepe Selection and depletion of plasma cells based on the specificity of the secreted antibody. Eur J Immunol 45: Khodadadi, L., Q. Cheng, T. Alexander, O. Sercan-Alp, J. Klotsche, A. Radbruch, F. Hiepe, B. F. Hoyer, and A. Taddeo Bortezomib Plus Continuous B Cell Depletion Results in Sustained Plasma Cell Depletion and Amelioration of Lupus Nephritis in NZB/W F1 Mice. PloS one 10: e Alexander, T., R. Sarfert, J. Klotsche, A. A. Kuhl, A. Rubbert-Roth, H. M. Lorenz, J. Rech, B. F. Hoyer, Q. Cheng, A. Waka, A. Taddeo, M. Wiesener, G. Schett, G. R. Burmester, A. Radbruch, F. Hiepe, and R. E. Voll The proteasome inhibitior bortezomib depletes plasma cells and ameliorates clinical manifestations of refractory systemic lupus erythematosus. Ann Rheum Dis 74:

45 Pathophysiology of Rheumatic Inflammation Hutloff, Chronic Immune Reactions When T cell / B cell crosstalk fails Hutloff, Liaison group with Dr. rer. nat. Andreas Hutloff Chronic Immune Reactions 44 Keywords T/B cooperation, T follicular helper cells, Chronic inflammation, ICOS costimulation Group Member Group leader Dr. rer. nat. Andreas Hutloff PhD students Laura Bauer (M.Sc.), Julia Kuhrau (M.Sc.), Annette Lahmann (Dipl. Humanbiol.), Josephine Scholz (M.Sc.) MD students Lisa Jasmin Müller Master students Josefine Graffunder Successful interaction of antigen-specific T and B cells is the key for an effective adaptive immune response. T follicular helper (Tfh) cells are the T cell subset providing help for B cells during the germinal center reaction. They are the prerequisite for the generation of high affinity memory B cells and long-lived plasma cells. Therefore, manipulation of the Tfh response is of particular clinical interest to either promote the generation of protective antibodies during vaccination or to eliminate harmful antibodies in autoimmune diseases or allergy. Our group is interested in the generation of Tfh cells, their maintenance, and dysregulated germinal center responses resulting in autoimmunity. One focus of our group are costimulatory receptors which are important for fine-tuning of the immune response. Absence of costimulation can result in severe immunodeficiency, whereas an overexpression of costimulatory receptors is often associated with exaggerated immune responses, resulting in autoimmunity or allergy (Grimbacher et al, Nat Immunol 2003; Hutloff et al, Arthr Rheum 2004). Consequently, costimulatory receptors are an attractive target for therapeutic intervention. Blocking reagents against various costimulatory receptors are currently in clinical trials or already in use. We recently demonstrated that the costimulatory molecule ICOS, which was originally identified in our group (Hutloff et al, Nature 1999), has a very specific role for maintenance of Tfh cells in late phases of the germinal center response. Blockade of ICOS signaling leads to upregulation of the transcription factor KLF2 in Tfh cells. Via regulation of several migratory receptors, this results in relocation of Tfh cells from the B cell back to the T cell zone and subsequent loss of the Tfh phenotype (Weber et al, J Exp Med 2015). This mechanism exemplifies the importance of migratory receptors and illustrates that location can dictate cell fate. Another focus of our group is T/B interaction outside lymphoid tissues. Lymphocytic infiltrates are frequently found in inflamed tissues where they substantially contribute to tissue destruction. Using a mouse airway inflammation model we showed that a population of tissue-resident Tfhlike cells drives the differentiation of antigenspecific B cells into germinal center like cells and plasma cells, thereby contributing to the local generation of pathogenic antibodies (Vu Van et al, Nat Commun 2016). Our findings that the majority of antigen-specific T and B cells does not reside in secondary lymphoid organs but the inflamed tissue and that active T/B cooperation can also take place in non-organized tissue infiltrates has important implications for the treatment of autoimmune diseases.

46 Hutloff, Chronic Immune Reactions Pathophysiology of Rheumatic Inflammation 45 Cooperation Partners Selected Publications Ashraful Haque, QIMR Berghofer Medical Research Institute, Herston, Queensland, Australia Richard Kroczek, Hans-Werner Mages, Robert Koch Institute, Berlin, Germany Anke Osterloh, Bernhard Fleischer, Bernhard Nocht Institute for Tropical Medicine, Hamburg, Germany Carola Vinuesa, Australian National University, John Curtin School of Medical Research, Immunology and Genetics, Canberra, Australia Weber JP,* Fuhrmann F,* Feist RK,* Lahmann A,* Al Baz MS, Gentz LJ, Vu Van D, Mages HW, Haftmann C, Riedel R, Grün JR, Schuh W, Kroczek RA, Radbruch A, Mashreghi MF, Hutloff A. ICOS maintains the T follicular helper cell phenotype by downregulating Krüppel-like factor 2. J Exp Med (2015) 212: Pratama A, Srivastava M, Williams NJ, Papa I, Lee SK, Dinh XT, Hutloff A, Jordan MA, Zhao JL, Casellas R, Athanasopoulos V, Vinuesa CG. MicroR- NA-146a regulates ICOS-ICOSL signaling to limit accumulation of T follicular helper cells and germinal centers. Nat Commun (2015) 6:6436. Paclik D, Stehle C, Lahmann A, Hutloff A,* Romagnani C.* ICOS regulates the pool of group 2 innate lymphoid cells under homeostatic and inflammatory conditions in mice. Eur J Immunol (2015) 45: Vu Van D, Beier KC, Pietzke LJ, Al Baz MS, Feist RK, Gurka S, Hamelmann E, Kroczek RA, Hutloff A. Local T/B cooperation in inflamed tissues is supported by T follicular helper-like cells. Nat Commun (2016) 7: Fuhrmann F,* Lischke T,* Gross F,* Scheel T, Bauer L, Kalim KW, Radbruch A, Herzel H, Hutloff A, Baumgrass R. Adequate immune response ensured by binary IL-2 and graded CD25 expression in a murine transfer model. elife (2016) 5: e

47 Pathophysiology of Rheumatic Inflammation Kruglov, Chronic inflammation Interaction of immune system and microbiota in chronic inflammation Kruglov, DRFZ group Dr. rer. nat. Andrey Kruglov Chronic inflammation 46 Keywords microbiota, chronic inflammation, cytokines Group Member Group leader Dr. rer. nat. Andrey Kruglov PhD students Justus Ninnemann Chronic inflammation in humans is strongly associated with dramatic changes in microbiota in various compartments. There is a clear indication that rheumatic diseases, such as rheumatoid arthritis and autoimmune colitis, can be quantitatively and qualitatively influenced by commensal microorganisms, but mechanistic aspects of microbiota contribution to the disease development, pathogenesis and chronicity remain to be determined. We are dissecting interactions of microbiota and the immune system in the development of chronic rheumatic diseases. Tissue repair during chronic immune reactions is one of the key aspects of successful recovery from disease. We have uncovered that TNF, one of the key proinflammatory mediators, blocks tissue repair via enhancement of IL-22BP expression by epithelial cells during autoimmune colitis. Increase in IL-22BP expression by TNF downregulates IL-22 biological activity and, thereby, dampens epithelial cell proliferation, microbial peptide synthesis and alters microbiota composition during chronic intestinal inflammation. One of the mechanisms of immune-mediated control of microbiota composition is the production of microbiota-specific IgA antibodies by intestinal plasma B cells. The population of IgA producing plasma cells in the intestine is heterogeneous with respect to the expression of surface molecule. However, there is lack of data describing the relation of these cell subsets to each other and their specificity. We took advantage of the differential expression of CD11b, Ly6C and Ly6G molecules on IgA producing B cells in the lamina propria. We have found that these molecules are upregulated after homing to the gut. Subsequent in vivo and in vitro analysis revealed that Ly6C-expressing IgA cells seemed to represent a terminal stage of CD45+IgA+ plasma cells in the small intestine. Finally, IgA antibodies produced by Ly6C- and Ly6C+ plasma cell subsets showed distinct recognition pattern of Gram-positive and Gramnegative commensal bacteria. Altogether, our data suggest the presence of distinct stages of IgA producing cells in the lamina propria, their developmental pathways and specificity towards various microbial species. Moreover, we have generated a set of microbiotaspecific IgA antibodies specific to distinct microbiota species. These antibodies can be further used for selective targeting of single microbiota species during rheumatic diseases. As an example, we have tested IgA antibody against Lachnospiraceae, bacteria that are increased in the intestine during chronic rheumatic diseases. We showed that administration of these antibodies during colitis ameliorated disease, suggesting that targeting distinct microbes can modulate chronic inflammation. Altogether, our data reveal new pathways in tissue repair during chronic inflammation and dissect the mechanisms of microbiota control by immune system and suggest new ways of treating rheumatic diseases by modulating intestinal microbiota.

48 Kruglov, Chronic inflammation Pathophysiology of Rheumatic Inflammation 47 Cooperation Partners Selected Publications Prof. Dr. Mathias Heikenwälder DKFZ, Heidelberg, Germany Dr. Anja Kühl Charite, Campus Benjamin Franklin, Berlin, Germany Prof. Dr. Britta Siegmund (Charite, Campus Benjamin Franklin, Berlin, Germany Prof. Dr. Markus Heimesaat, Charite, Campus Benjamin Franklin, Berlin, Germany Prof. Dr. Bastian Opitz, Charite, Campus Benjamin Franklin, Berlin, Germany Prof. Dr. Ivaylo Ivanov, Columbia University, New York, USA 1: Yuzhakova DV, Shirmanova MV, Bocharov AA, Astrakhantseva IV, Vasilenko EA, Gorshkova EN, Drutskaya MS, Zagaynova EV, Nedospasov SA, Kruglov AA. Microbiota. Induces Expression of Tumor Necrosis Factor in Postnatal Mouse Skin. Biochemistry, (Mosc) Nov;81(11): PubMed PMID: : Efimov GA, Kruglov AA, Khlopchatnikova ZV, Rozov FN, Mokhonov VV, Rose-John S, Scheller J, Gordon S, Stacey M, Drutskaya MS, Tillib SV, Nedospasov SA. Cell-type-restricted anti-cytokine therapy: TNF inhibition from one pathogenic source. Proc Natl Acad Sci U S A Mar 15;113(11): doi: /pnas PubMed PMID: ; PubMed Central PM- CID: PMC : Winsauer C, Prepens S, Schlienz D, Nedospasov S, Kruglov AA. Novel mouse model to study T cell-dependent IgA induction in vivo. J Immunol Methods Jun;421: doi: /j. jim PubMed PMID: : Van Praet JT, Donovan E, Vanassche I, Drennan MB, Windels F, Dendooven A, Allais L, Cuvelier CA, van de Loo F, Norris PS, Kruglov AA, Nedospasov SA, Rabot S, Tito R, Raes J, Gaboriau- Routhiau V, Cerf-Bensussan N, Van de Wiele T, Eberl G, Ware CF, Elewaut D. Commensal microbiota influence systemic autoimmune responses. EMBO J Feb 12;34(4): doi: / embj PubMed. PMID: ; PubMed Central PMCID: PMC Prof. Maxim Artomov, Washington University, Saint Louis, USA Prof. Sergei Grivennikov, Fox Chase Cancer Center, Philadelphia, USA 3: Benezech C, Luu NT, Walker JA, Kruglov AA, Loo Y, Nakamura K, Zhang Y, Nayar, S, Jones LH, Flores-Langarica A, McIntosh A, Marshall J, Barone F, Besra G, Miles, K, Allen JE, Gray M, Kollias G, Cunningham AF, Withers DR, Toellner KM, Jones ND, Veldhoen M, Nedospasov SA, McKenzie AN, Caamaño JH. Inflammation-induced formation of fat-associated lymphoid clusters. Nat Immunol Aug;16(8): doi: / ni PubMed PMID: ; PubMed Central PMCID: PMC

49 Pathophysiology of Rheumatic Inflammation Kubagawa, Humoral Immune Regulation What is the functional composition of IgM Fc receptor (FcµR) and how does FcµR function in lymphocytes. Kubagawa, DRFZ group Hiromi Kubagawa, MD Humoral Immune Regulation 48 Keywords natural IgM antibody; IgM effector function; lymphocyte Fc receptor; humoral immune regulation; tolerance. Group Member Group leader Hiromi Kubagawa, MD Bachelor/Diploma/ Master students Christopher M. Skopnik, M.S., Anika Oehme, M.S., Marie Urbicht, M.S. Antibody, a key player in humoral immunity, has dual binding activities: to antigens via its amino terminal variable regions, called Fab, and to effector molecules via its carboxyl terminal constant region, called Fc. One of the effector molecules is a family of Fc receptors (FcRs). FcRs for switched antibodies (i.e., IgG, IgE, IgA) are expressed by many different immune cells and are extensively characterized as central mediators coupling innate and adaptive immune responses. Much of the gained knowledge from these studies has been translated to clinical fields. By contrast, FcR for IgM, the first antibody appearing in phylogeny, ontogeny and immune responses, has defied genetic identification until our successful cloning eight years ago. IgM FcR (FcµR) is only expressed by adaptive immune lymphocytes: B, T, and, to a lesser extent, NK cells in humans and only B cells in mice, suggesting the distinct function of FcµR compared to other FcRs as well as the species difference in cellular distribution. FcµR-bearing cells bind secreted IgM antibody with strikingly high avidity of ~10 nm (trans interaction). In our recent analyses, however, FcµR binds more efficiently the Fc portion of IgM antibody when it is attached to a membrane component via its Fab region on the same lymphocyte surface (cis interaction). This preferential cis engagement of FcµR implies that FcµR can modulate the functional activity of lymphocyte surface receptors or components recognized by either natural or immune IgM antibody. Based on mutational analyses, we have defined several key residues in the transmembrane and cytoplasmic tail of the human FcµR molecule that involve in the receptor function: (i) the transmembrane His residue anchors the receptor in the plasma membrane; (ii) the membraneproximal cytoplasmic Tyr residue mediates protection from apoptosis when the cis engagement occurs between Fas death receptor and FcµR; and (iii) two membrane-distal cytoplasmic Tyr residues involve the receptormediated endocytosis. Cell surface levels of FcµR are sensitive to the extracellular IgM concentration, tissue milieu and cellular activation status. Since the serum IgM concentration in humans is ~1 µm, 100-fold above the FcµR avidity for IgM ligands, FcµR on lymphocytes must be occupied with IgM in vivo and the IgM-bound FcµR should be internalized, rapidly retrieved from early endosomes and returned to the cell surface. Brief pre-incubation of freshly-prepared lymphocytes in IgM-free media is thus required for the assessment of the full cell surface expression of FcµR by flow cytometry using either IgM ligands or receptorspecific monoclonal antibodies. Given these findings, we predict that cell surface FcµR is not vulnerable in patients with selective IgM immunodeficiency. Contrary to this assumption, the surface FcµR levels on blood B cells, especially marginal zone-like B cells, in these patients are significantly diminished irrespective of preincubation, as compared to those in healthy controls. This suggests a different, not yet found, mechanism for reduced FcµR expression in this immunodeficiency. The existing nomenclature issue of Toso, Fas apoptosis inhibitory molecule 3 (FAIM3) versus FcµR now reaches a general consensus that Toso/ FAIM3 is an authentic IgM Fc-binding protein and that the initial designated FAIM3 (for

50 Kubagawa, Humoral Immune Regulation Pathophysiology of Rheumatic Inflammation 49 humans) and Faim3 (for mice) genes are thus officially renamed to FCMR and Fcmr, respectively. The outcomes from these investigations are expected to confirm the importance of IgM and FcµR in immune protection and regulation in rheumatic diseases. We are currently exploring another membrane protein non-covalently associating with FcµR. Cooperation Partners Gupta, Sudhir, M.D., Ph.D., Professor, University of California at Irvine, Division of Basic and Clinical Immunology, USA Honjo, Kazuhito, D.V.M., University of Alabama at Birmingham, Department of Medicine, USA Kearney, John F., D.M.D, Ph.D., Professor, UAB, Department of Microbiology, USA Vivarelli, Marina, M.D., Bombino Gesù Children s Hospital-Scientific Institute, Division of Nephrology and Dialysis, Italy Walter, Mark, R., Ph.D., Professor, UAB, Department of Microbiology, USA Selected Publications Honjo, K., Y. Kubagawa, J. F. Kearney, and H. Kubagawa Unique ligand-binding property of the human IgM Fc receptor. J. Immunol. 194: Kubagawa, H., M. C. Carroll, C. O. Jacob, K. S. Lang, K. H. Lee, T. Mak, M. McAndrews, H. C. Morse, G. P. Nolan, H. Ohno, G. H. Richter, R. Seal, J. Y. Wang, A. Wiestner, and J. E. Coligan Nomenclature of Toso, Fas apoptosis inhibitory molecule 3, and IgM FcR. J. Immunol. 194: Gupta, S., S. Agrawal, S. Gollapudi, and H. Kubagawa FcµR in human B cell subsets in primary selective IgM deficiency, and regulation of FcµR and production of natural IgM antibodies by IGIV. Hum. Immunol. 77:

51 Pathophysiology of Rheumatic Inflammation Nedospasov, Inflammation Biology Cytokines in inflammation and anti-cytokine therapy Nedospasov, DRFZ group Prof. Dr. Sergei Nedospasov Inflammation Biology 50 Keywords Autoimmune disease, reverse genetics, Anti-cytokine therapy Group Member Group leader Prof. Dr. Sergei Nedospasov Scientists Dr. Andrei Kruglov (till 04/2016 ) Dr. Josefine Dunst PhD students Maxim Nosenko Kama Atretkhany Others Marina Drutskaya The main focus of the group is to develop optical technologies and methodologies allowing to visualize and quantify dynamic processes at cellular and sub-cellular level in the living organism in order to provide new insights into pathophysiologic phenomena relevant in chronic inflammation. In tight cooperation with clinicians (Dr. Helena Radbruch, Prof. Dr. Friedemann Paul and Prof. Dr. Frank Heppner) and immunologists (Prof. Dr. Anja Hauser), we set a special focus on developing methods for better understanding chronic neuroinflammation. Thereby, we designed and used NADH and NADPH fluorescence lifetime imaging (FLIM) applied to intravital as well as to live cell imaging to identify over-activated NADPH oxidases (NOX1-4, DUOX1,2), which is the source of massive O2 production the initiator of reactive oxygen species. If this process is controlled, it represents the oxidative burst and thus oxidative eustress (e.g. in phagocytosis), but if it is random, it leads to oxidative stress. We found in both the brain stem tissue of a mouse model of Multiple Sclerosis (MS) and in blood-derived monocytes of MS patients a persisting over-activation of NADPH oxidases and defined the concept of oxidative stress memory (Mossakowski et al, 2015, Acta Neuropath.). By combining our technology with fluorescent reporter mice we were further able to identify astrocytes and microglia as main contributors of tissue oxidative stress especially in the recovery phase of the disease (Radbruch et al, 2016, Front. Immunol.). Currently, we are translating NAD(P) H-FLIM to imaging in mouse lupus models as well as to SLE patients, to characterize the oxidative stress behavior in the kidney as compared to blood-derived cells. In order to better understand inflammatory processes in chronic neuroinflammation models, we developed an intravital imaging approach of the retina which allowed us to longitudinally monitor immune infiltration of this part of the central nervous system and to correlate this with the degeneration of its microglial network in one and the same animal (Bremer et al, 2016, Front. Immunol.). To further develop our intravital imaging, we designed a microendoscopic implant to longitudinally image femoral marrow both during bone healing and homeostasis. Moreover, we developed a combined imaging strategy for multiplexed imaging in living organisms allowing for dynamic visualization of complex phenomena involving up to 8 cellular and tissue compartments.

52 Nedospasov, Inflammation Biology Pathophysiology of Rheumatic Inflammation 51 Cooperation Partners Selected Publications Prof. S. Rose-John, Institute of Biochemistry, University of Kiel, Germany. Prof. M. Heikenwalder, German Cancer Research Center (DKFZ), Heidelberg, Germany. Prof. H. Wajant, University of Wurzburg, Germany Prof. D.V. Kuprash, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia. Prof. S.V. Tillib, Institute of Gene Biology, Russian Academy of Sciences, Moscow, Russia. Prof. V.V. Novikov, Lobachevsky University of Nizhni Novgorod, N. Novgorod, Russia. Prof. W.J. Murphy, University of California at Davis, Sacramento, CA, USA. Dr. A.V. Tumanov, University of Texas, San Antonio, TX, USA. Prof. S.-S. Bolz, University of Toronto, Canada. Efimov, G.A., Kruglov, A.A., Khlopchatnikova, Z.V., Rozov, F.N., Mokhonov, V.V., Rose-John, S., Scheller, J., Gordon, S., Stacey, M., Drutskaya, M.S., Tillib, S.V., Nedospasov, S.A. "Cell-type-restricted anti-cytokine therapy: TNF inhibition from one pathogenic source" Proc Natl Acad Sci USA (2016), 113, 11, Atretkhany, K.-S.N., Nosenko, M.A., Gogoleva, V.S., Zvartsev, R.V., Qin, Z., Nedospasov, S.A., Drutskaya, M.S."TNF neutralization results in the delay of transplantable tumor growth and reduced MDSC accumulation" Front Immunol (2016), 7, 147, DOI: /fimmu Nosenko, M.A., Drutskaya, M.S., Moisenovich, M.M., Nedospasov, S.A. "Bioengineering of artificial lymphoid organs" Acta Naturae (2016), 8, 2, Dudeck, J., Ghouse, S.M., Lehmann, C.H., Hoppe, A., Schubert, N., Nedospasov, S.A., Dudziak, D., Dudeck, A. Mast-Cell-Derived TNF Amplifies CD8(+) Dendritic Cell Functionality and CD8(+) T Cell Priming. Cell Rep. (2015), 13(2): Bénézech, C., Walker, J.A., Kruglov, A.A., Loo, Y, Nayar, S., Jones, L.H., Marshall, J., Barone, F.,, Besra, G., Miles, K., Gray, M., Kollias, G., Cunningham, A.F., Toellner, K.M., Jones, N.D., Veldhoen, M., Nedospasov, S.A., McKenzie, A.N.J., and Caamaño, J.H. inkt cell activation and TNFR signaling drive the formation of Fat Associated Lymphoid Clusters. Nat. Immunol. 2015, 16(8):

53 Pathophysiology of Rheumatic Inflammation Niesner, Biophysical Analytics We develop technologies for intravital imaging and live cell imaging to achieve new insights into chronic inflammatory processes Niesner, DRFZ group Dr. rer. nat. Raluca A. Niesner Biophysical Analytics 52 Keywords intravital multi-photon imaging, fluorescence lifetime imaging, NADPH oxidases, microendoscopy Group Member Group leader Dr. rer. nat. Raluca A. Niesner Scientists Dr. rer. nat. Asylkhan Rakhymzhan PhD students Daniel Bremer, MSc engineering/medical physics David Reismann, MSc biotechnology Ruth Leben, MSc engineering, medical physics MD students Ronja Mothes Master students Judith Platzer, BSc engineering, medical physics Technicians Peggy Mex The main focus of the group is to develop optical technologies and methodologies allowing to visualize and quantify dynamic processes at cellular and sub-cellular level in the living organism in order to provide new insights into pathophysiologic phenomena relevant in chronic inflammation. In tight cooperation with clinicians (Dr. Helena Radbruch, Prof. Dr. Friedemann Paul and Prof. Dr. Frank Heppner) and immunologists (Prof. Dr. Anja Hauser), we set a special focus on developing methods for better understanding chronic neuroinflammation. Thereby, we designed and used NADH and NADPH fluorescence lifetime imaging (FLIM) applied to intravital as well as to live cell imaging to identify over-activated NADPH oxidases (NOX1-4, DUOX1,2), which is the source of massive O2 production the initiator of reactive oxygen species. If this process is controlled, it represents the oxidative burst and thus oxidative eustress (e.g. in phagocytosis), but if it is random, it leads to oxidative stress. We found in both the brain stem tissue of a mouse model of Multiple Sclerosis (MS) and in blood-derived monocytes of MS patients a persisting over-activation of NADPH oxidases and defined the concept of oxidative stress memory (Mossakowski et al, 2015, Acta Neuropath.). By combining our technology with fluorescent reporter mice we were further able to identify astrocytes and microglia as main contributors of tissue oxidative stress especially in the recovery phase of the disease (Radbruch et al, 2016, Front. Immunol.). Currently, we are translating NAD(P) H-FLIM to imaging in mouse lupus models as well as to SLE patients, to characterize the oxidative stress behavior in the kidney as compared to blood-derived cells. In order to better understand inflammatory processes in chronic neuroinflammation models, we developed an intravital imaging approach of the retina which allowed us to longitudinally monitor immune infiltration of this part of the central nervous system and to correlate this with the degeneration of its microglial network in one and the same animal (Bremer et al, 2016, Front. Immunol.). To further develop our intravital imaging, we designed a microendoscopic implant to longitudinally image femoral marrow both during bone healing and homeostasis. Moreover, we developed a combined imaging strategy for multiplexed imaging in living organisms allowing for dynamic visualization of complex phenomena involving up to 8 cellular and tissue compartments.

54 Niesner, Biophysical Analytics Pathophysiology of Rheumatic Inflammation 53 Cooperation Partners Prof. Dr. Salim Seyfried University Potsdam/ Golm/ Institute for Physiology, Potsdam, Germany Prof. Dr. Rainer Heintzmann, University of Jena/ Institute for Physical Chemistry and IPHT, Jena, Germany Prof. Dr. Reinhard Voll, University of Medicine, Freiburg/ Rheumathology, Freiburg, Germany PD Dr. Dirk Mielenz, University Nurmberg- Erlangen/ Fibiger Institute, Erlangen, Germany Prof. Dr. Klaus Warnatz, University of Medicine, Freiburg/Center for Immunodeficiency/Freiburg, Germany Prof. Dr. Georg Duda and Dr. Katharina Schmidt-Bleek, Julius-Wolf-Institute, Charitè Universitätsmedizin, Berlin PD Dr. Gereon Hüttmann, MLL, University Lübeck, Germany Prof. Dr. Vajcozy and Dr. Simon Bayerl, Charité Prof. Dr. Friedemann Paul and Dr. Alexander Brandt, NCRC, Charité Universitätsmedizin, Berlin Selected Publications Intravital FRET: Probing Cellular and Tissue Function in Vivo. Radbruch H, Bremer D, Mothes R, Günther R, Rinnenthal JL, Pohlan J, Ulbricht C, Hauser AE, Niesner R. Int J Mol Sci. 2015;16(5): doi: / ijms [IF: 3.257] Tracking CNS and systemic sources of oxidative stress during the course of chronic neuroinflammation. Mossakowski AA, Pohlan J, Bremer D, Lindquist R, Millward JM, Bock M, Pollok K, Mothes R, Viohl L, Radbruch M, Gerhard J, Bellmann-Strobl J, Behrens J, Infante-Duarte C, Mähler A, Boschmann M, Rinnenthal JL, Füchtemeier M, Herz J, Pache FC, Bardua M, Priller J, Hauser AE, Paul F, Niesner R*, Radbruch H*. Acta Neuropathol. 2015;130(6): doi: /s x. [IF: ] Ongoing Oxidative Stress Causes Subclinical Neuronal Dysfunction in the Recovery Phase of EAE. Radbruch H, Bremer D, Guenther R, Cseresnyes Z, Lindquist R, Hauser AE, Niesner R. Front Immunol. 2016;7:92. doi: /fimmu [IF: 5.695] Disturbed canonical NF-kB signaling in B cells of CVID patients. Keller B, Cseresnyes Z, Stumpf I, Wehr C, Fliegauf M, Bulashevska A, Usadel S, Grimbacher B, Rizzi M, Eibel H, Niesner R, Warnatz K. Journal of Allergy and Clinical Immunology, 2017, 139(1): [IF: ] Longitudinal intravital imaging of the retina reveals long-term dynamics of immune infiltrate and its effects on the glial network in experimental autoimmune uveoretinitis, without evident signs of neuronal dysfunction in the ganglion cell layer. Bremer D, Pache F, Günther R, Hornow J, Andresen V, Leben R, Mothes R, Zimmermann H, Brandt AU, Paul F, Hauser AE, Radbruch H, Niesner R. Front. Immunol, 2016, 7:642. [IF: 5.695]

55 Pathophysiology of Rheumatic Inflammation Radbruch, Cell Biology Immunological memory as driver of rheumatic inflammation Prof. Dr. rer. nat. Andreas Radbruch Radbruch, Cell Biology DRFZ group 54 Keywords Immunological memory, chronic inflammation, T lymphocytes, B lymphocytes, plasma cells Group Member Group leader Prof. Dr. rer. nat. Andreas Radbruch Scientists Dr. rer. nat. Hyun-Dong Chang, Dr. rer. nat. Jun Dong, Mairi McGrath PhD, Dr. rer. nat. Özen Sercan, Dr. rer. nat. Claudia Giesecke, Dr. rer nat. Pawel Durek, Elodie Mohr PhD PhD students Daniel Schulz, Richard Addo, René Riedel, Francesco Siracusa, Kerstin Westendorf, Melanie Weber, Jakob Zimmermann, Carla Cendon, Haider Abid, Stefanie Schmidt, Weiji Du, Alexander Beller, Kristyna Hradilkova, Sandra Naundorf Bachelor/Diploma/ Master students Cora Claeden, Katharina Werner, Darya Malko, Cassandra Steinkraus, Marlen Schulze, Sarah Schiele, Jessica Dysarz, Melanie Klingauf, Alexander Schmidt, Josefine Spalding Technicians Katrin Lehmann Others Sarah Schimmelpfennig Darya Malko Chronic immune reactions are a key feature of inflammatory rheumatic diseases. Current stateof-the-art immunosuppressive therapies are able to stop disease progression and maintain remission. However these therapies do not induce a therapy-free remission, i.e. discontinuation of treatment leads to relapse of disease. From the experimental therapy of immunoablation and regeneration of a patient s immune system from their own progenitor cells, also termed autologous stem cell transplantation, with about 20 years of experience at the Charité and DRFZ, we know that most if not all patients have an immunological memory driving the rheumatic inflammation, and this memory is refractory to conventional therapy. It is the aim of our group to identify the cells impersonating this pathogenic immunological memory, understand their development, their persistence, and their resistance to current therapies. From such an understanding new options should emerge for their selective therapeutic targeting and the induction of therapy-free remission. In a basic approach, we have revisited the current concepts on how immunological memory is organized. We have developed the novel concept of a compartmentalized immunological memory in which the long-term memory to systemic antigens is maintained in the bone marrow, while the memory cells found in circulation are rather short-lived. We have identified a new type of memory cell, the memory plasma cell, secreting protective or pathogenic antibodies. We could show that reticular mesenchymal stromal cells of the bone marrow organize dedicated survival niches in which antigen-experienced lymphocytes differentiate into memory cells and persist as cells resting in terms of activation, proliferation, and mobility, as long as they are provided with survival signals. In such niches, memory cells are also protected from conventional therapies targeting activated immune cells. We are identifying the specific survival signals for different types of memory cells. Such signals and their receptors qualify as novel and selective therapeutic targets. At present we are collaborating with the group of Falk Hiepe to identify and target (pathogenic) memory plasma cells. How can we distinguish pathogenic from protective memory cells? For memory T helper (Th) lymphocytes we could show that pathogenic memory Th lymphocytes initiate and drive inflammation through chemokines recruiting inflammatory effector cells, like macrophages and granulocytes. In chronic inflammation, pathogenic memory Th lymphocytes adapt to repeated reactivation. We have identified such molecular adaptations, e.g. expression of the genes Hopx, Twist1 and the microrna mir- 148a. Together with the group of Mir-Farzin Mashreghi we are now testing synthetic, membrane-permeable oligonucleotides for the inhibition of these vital adaptations, and thus the selective targeting of pathogenic memory Th lymphocytes.

56 Radbruch, Cell Biology Pathophysiology of Rheumatic Inflammation 55 Cooperation Partners Miltenyi Biotec GmbH, Bergisch Gladbach: Dr. Andreas Bosio, Dr. Ute Bissels, Dr. Anne Richter Charité Universitätsmedizin Berlin, Klinik für Rheumatologie und klinische Immunologie: Prof. Dr. Gerd-Rüdiger Burmester, Dr. Tobias Alexander, Prof. Dr. Thomas Dörner, Dr. Thomas Häupl,Prof. Dr. Falk Hiepe, Silvia Pade, Prof. Dr. Alexander Scheffold, Dr. Anja Kühl, Medizinische Klinik I Prof. Dr. Georg Duda, Julius Wolff Inst. für Biomechanik und Muskuloskeletale Regeneration Prof. Dr. Britta Siegmund, Prof. Dr. Jochen Sieper/ Denis Poddubnyy, Charité Univeristätsmedizin, Medizinische Klinik I Prof. Dr. Klemens Budde, Dr. Mareen Matz, Charité Univeristätsmedizin, Nephrologie Weitere Prof. Dr. Fritz Melchers, Max Planck Institut für Infektionsbiologie, Berlin Dr. Wei Chen, Max-Delbrück-Centrum für Molekulare Medizin und Berlin Institute for Medical Systems Biology, Berlin Prof. Dr. Steffen Gay, Zentrum für Experimentelle Rheumatologie, Universitätspital Zürich Prof. Dr. Hans-Martin Jäck, Molekulare Immunologie, Universitätsklinikum Erlangen Prof. Dr. Thomas Kamradt, Institut für Immunologie, Universitätsklinikum, Friedrich-Schiller- Universität, Jena Prof. Dr. Susann Müller, Helmholtz Institut für Umweltforschung, Leipzig Prof. Dr. Toshinori Nakayama, Department of Immunology, Chiba University, Chiba, Japan Prof. Dr. Nikolaus Rajewsky, Max-Delbrück-Centrum für Molekulare Medizin und Berlin Institute for Medical Systems Biology, Berlin Prof. Dr. Andreas Thiel, Berlin-Brandenburg Center for Regenerative Therapies BCRT, Berlin Prof. Dr. Kai Wucherpfennig, Dana-Farber Cancer Institute, Boston, MA, USA Prof. Dr. Michael Lohoff, Institut für Medizinische Mikrobiologie, Philipps University Marburg Selected Publications 1. Zimmermann J, Hübschmann T, Schattenberg F, Schumann J, Durek P, Riedel R, Friedrich M, Glauben R, Siegmund B, Radbruch A, Müller S, Chang HD. High-resolution microbiota flow cytometry reveals dynamic colitis-associated changes in fecal bacterial composition. Eur J Immunol May;46(5): doi: / eji PubMed PMID: Zimmermann J, Kühl AA, Weber M, Grün JR, Löffler J, Haftmann C, Riedel R, Maschmeyer P, Lehmann K, Westendorf K, Mashreghi MF, Löhning M, Mack M, Radbruch A, Chang HD. T-bet expression by Th cells promotes type 1 inflammation but is dispensable for colitis. Mucosal Immunol Feb 17. doi: /mi [Epub ahead of print] PubMed PMID: Alp ÖS, Radbruch A. The lifestyle of memory CD8(+) T cells. Nat Rev Immunol Apr;16(4):271. doi: /nri PubMed PMID: Sercan Alp Ö, Durlanik S, Schulz D, McGrath M, Grün JR, Bardua M, Ikuta K, Sgouroudis E, Riedel R, Zehentmeier S, Hauser AE, Tsuneto M, Melchers F, Tokoyoda K, Chang HD, Thiel A, Radbruch A. Memory CD8(+) T cells colocalize with IL-7(+) stromal cells in bone marrow and rest in terms of proliferation and transcription. Eur J Immunol Apr;45(4): doi: / eji Epub 2015 Feb 27. PubMed PMID: ; PubMed Central PMCID: PMC Haftmann C, Stittrich AB, Zimmermann J, Fang Z, Hradilkova K, Bardua M, Westendorf K, Heinz GA, Riedel R, Siede J, Lehmann K, Weinberger EE, Zimmel D, Lauer U, Häupl T, Sieper J, Backhaus M, Neumann C, Hoffmann U, Porstner M, Chen W, Grün JR, Baumgrass R, Matz M, Löhning M, Scheffold A, Wittmann J, Chang HD, Rajewsky N, Jäck HM, Radbruch A, Mashreghi MF. mir- 148a is upregulated by Twist1 and T-bet and promotes Th1-cell survival by regulating the proapoptotic gene Bim. Eur J Immunol Apr;45(4): doi: /eji PubMed PMID: ; PubMed Central PM- CID: PMC

57 Pathophysiology of Rheumatic Inflammation Romagnani, Innate Immunity Identifying innate signals initiating and perpetuating chronic inflammation Romagnani, Liaison group with Prof. Chiara Romagnani, MD PhD Innate Immunity 56 Heisenberg Professorship Innate mechanisms of chronic inflammation Keywords chronic inflammation, innate lymphoid cells, innate receptors, epigenetic imprinting Group Member Group leader Prof. Chiara Romagnani MD PhD Scientists Daniela Paclik, Kerstin Juelke, Marina Babic PhD students Christina Stehle, Quirin Hammer, Timo Rückert MD students Andre Haubner Bachelor/Diploma/ Master students Almut Eisele Student assistant Yan Dyck Chronic inflammatory disorders, especially rheumatic diseases, are triggered and maintained by effector mediators produced by the adaptive immune system, such as T cells and B cells. The adaptive immune system employs three major effector modules, namely type 1 (IFN-γ and TNF), type 2 (IL-4, IL-5 and IL-13) and type 3 or type 17 (IL-17, IL-22), which give rise to distinct inflammatory tissue responses. In T cells, such inflammatory programs are induced by the T cell receptor (TCR) in conjunction with distinct cytokines and/or environmental signals and can be epigenetically imprinted and memorized. While the imprinting of inflammatory signatures enables a faster and stronger response during secondary infection, it facilitates on the other hand the induction and perpetuation of chronic inflammation, a hallmark of rheumatic diseases. Recently, it appeared evident that emerging innate cell subsets lacking the TCR and collectively known as innate lymphoid cells (ILCs), exhibit a similar heterogeneity of effector modules, which can be activated in the course of inflammation. The ILC family includes three main groups of cells: group 1 ILCs, including cytotoxic Natural Killer (NK) cells and the IFN- γ producing ILC1; ILC2 producing IL-13/IL-5, and ILC3 secreting IL-22/IL-17. The signals and innate receptors instructing the different effector programs and their execution in ILCs remain largely unknown. Such innate sensors could also enhance effector functions in T cells, thus promoting inflammation in a TCR-independent fashion. Moreover, it is still unclear whether these inflammatory programs can be also memorized and imprinted in ILCs, thus possibly sustaining chronic inflammation. Therefore, our main research focus is devoted to study the innate modules and triggers employed by ILCs and T cells to initiate and maintain inflammation in a TCR-independent fashion and to understand whether distinct inflammatory programs can be imprinted in ILCs to promote rheumatic diseases. The identification of alternative triggering and perpetuators of inflammation will provide us with new potential targets for the treatment of chronic rheumatic diseases.

58 Romagnani, Innate Immunity Pathophysiology of Rheumatic Inflammation 57 Cooperation Partners Selected Publications Bluethgen N, Medical Systems Biology, Charité Universitätsmedizin, Berlin, Germany Cerwenka A, German Cancer Research Center (DKFZ), Innate Immunity, Heidelberg, Germany Geginat J, Istituto Nazionale di Genetica Molecolare, "Romeo ed Enrica Invernizzi", Milan, Italy. Malmberg KJ, Béziat V, Center for Infectious Medicine, Department of Medicine Huddinge, Karolinska Institutet, Stockholm, Sweden. Moretta A, Pietra G, University of Genova, Italy Ullrich E, Division of Stem Cell Transplantation and Immunology, Department for Children and Adolescents Medicine, University Hospital Frankfurt, Goethe University, Frankfurt, Germany. Vivier E, Centre d'immunologie de Marseille- Luminy, Aix Marseille Université, INSERM, CNRS, Marseille, France Walter J, Genetics/Epigenetics, University of Saarland, Saarbrücken, Germany Differentiation of human innate lymphoid cells (ILCs). Juelke K, Romagnani C. Curr Opin Immunol Feb;38: doi: /j. coi Epub 2015 Dec 17. Review. Putting the brakes on ILC2 cells. Stehle C, Saikali P, Romagnani C. Nat Immunol Jan;17(1):43-4. doi: /ni No abstract available. ICOS regulates the pool of group 2 innate lymphoid cells under homeostatic and inflammatory conditions in mice. Paclik D, Stehle C, Lahmann A, Hutloff A, Romagnani C. Eur J Immunol Oct;45(10): doi: /eji Epub 2015 Aug 31. Schüler T, Institute of Molecular and Clinical Immunology, Medical Faculty, Otto von Guericke University, Magdeburg, Germany Schwab JM, Department of Neurology and Experimental Neurology, Charité - Universitätsmedizin Berlin, Germany Seidl R, HNO-Klinik, Unfallkrankenhaus, Berlin, Germany Stölzel K, Hals-Nasen-Ohrenklinik und Poliklinik, Charité Universitätsmedizin CBF, Berlin Group 3 innate lymphoid cells (ILC3s): Origin, differentiation, and plasticity in humans and mice. Montaldo E, Juelke K, Romagnani C. Eur J Immunol Aug;45(8): doi: / eji Epub 2015 Jun 18. Review. The 3 major types of innate and adaptive cell-mediated effector immunity. Annunziato F, Romagnani C, Romagnani S. J Allergy Clin Immunol Mar;135(3): doi: /j. jaci Review.

59 Pathophysiology of Rheumatic Inflammation Scheffold, Cellular Immunology Suppression and self-control: How T cells fight autoimmunity. Scheffold, Liaison group with Prof. Dr. rer. nat. Alexander Scheffold Cellular Immunology 58 Keywords Regulatory T cells, antigen-specific T cells, autoimmunity, allergy, notch Group Member Group leader Prof. Dr. rer. nat. Alexander Scheffold Scientists Dr. rer. nat. Petra Bacher, Dr. rer. nat. Marko Janke, Dr. rer. nat. Christian Neumann PhD students Jonas Ahlers M.Sc., Anna Nowak, M.Sc., Dipl. human. biol. Stefanie Gamradt Bachelor/Diploma/ Master students Lukas Heiberger, B.Sc Technicians Thordis Hohnstein Others Dr. rer. nat. Tim Meyer, Dr. rer. nat. Andrej Mantei (guest scientists) Immune-mediated diseases such as Rheumatoid Arthritis, Diabetes or Multiple Sclerosis, but also inflammatory bowel diseases or allergies are thought to be initiated and maintained by T and B lymphocytes, which react against own body components (self-antigens) or harmless foreign substances (allergens, commensal microbiota). Current therapeutic approaches focus on general immune suppression instead of targeting those few disease-relevant lymphocytes. In contrast to murine models, major roadblocks towards specific therapies in humans are technical difficulties A) to identify and characterize the antigen-specific T cells and their target antigens in patients and B) to specifically approach them therapeutically. Our group has established sensitive technologies to study antigen-specific T cells involved in immunopathology directly from patients. We characterize T cells contributing to pathologic or healthy immune reactions against autoantigens or harmless foreign substances and describe the alterations in immunopathology. Understanding antigen-specific T cells as the drivers of disease and how these T cells can control or prevent autoimmunity or allergy has both diagnostic and therapeutic relevance. For development of therapeutic approaches, we are trying to utilize the physiological population of regulatory T cells (Treg), known to prevent autoimmunity. We are trying to identify their physiologic target antigens and to specifically modify their suppressive potential in vitro, i.e. by genetic engineering. Besides professional immune-suppressive Treg, we are also interested which mechanisms of selfcontrol inflammatory T cells employ to limit overt inflammation. Amplifying these autoregulatory circuits would be a valuable therapeutic strategy, building on re-establishment of the body s physiological control mechanisms. Here we are focusing on the regulation of the anti-inflammatory or tissue-protective cytokines IL-10 and IL-22. Using gene expression profiling and functional in vitro and in vivo assays, we are studying the inducing signals and the transcriptional network regulating their expression. We have identified Notch as a switch to activate IL-10 as well as IL-22 in inflammatory T cells and defined additional critical transcription factors. We are also trying to define the physiological niche for T cell modulation by Notch and additional environmental signals, which shall be translated into in vitro T cell modulation strategies.

60 Scheffold, Cellular Immunology Pathophysiology of Rheumatic Inflammation 59 Cooperation Partners Germany Prof. Hinrich Abken, Prof. Oliver Cornely, Angela Steinbach, Klinik I für Innere Medizin, Uniklinik Köln Dr. Mario Assenmacher, Dr. Michael Apel, Miltenyi Biotec GmbH, Bergisch Prof. Axel Brakhage, Dr. Olaf Kniemeyer, Leibniz-Institut für Naturstoff-Forschung und Infektionsbiologie (HKI), Jena Dr. Uta Düesberg, Mukoviszidose Institut ggmbh, Bonn Prof. Hermann Einsele, Dr. Michael Hudecek, Medizinische Klinik und Poliklinik II, Zentrum Innere Medizin, Uniklinik Würzburg Prof. Charlotte Esser, Leibniz-Institut für umwelt-medizinische Forschung, Düsseldorf Germany, Berlin Prof. Nina Babel, Marien Hospital Herne, Universitätsklinikum der Ruhr-Universität Bochum/Charité Universitätsmedizin Berlin Prof. Susanne Hartmann, FU Berlin, Fachbereich Veterinärmedizin, Institut für Immunologie Prof. Dr. Dr. h.c. Stefan H. E. Kaufmann, Max-Planck-Institut für Infektionsbiologie Prof. Wolfgang Uckert, Max-Delbrück-Centrum für Molekulare Medizin, Berlin Germany, Berlin, Charité Universitätsmedizin Berlin Prof. Stephan Bereswil, Dr. Markus Heimesaat, Institut für Mikrobiologie und Hygiene Dr. Eugen Feist, PD Dr. Thomas Häupl, Med. Klinik m. S. Rheumatologie und Klinische Immunologie Dr. Anja Kühl, Med. Klinik für Gastroenterologie, Infektiologie und Rheumatologie CBF Dr. Berit Rosche, Klinik und Hochschulambulanz für Neurologie Dr. Carsten Schwarz, Klinik für Pädiatrie m.s. Pneumologie und Immunologie Prof. Britta Siegmund, Dr. Jochen Maul, Medizinische Klinik für Gastroenterologie, Infektiologie und Rheumatologie CBF Prof. Margitta Worm, Dr. Guido Heine, Kl.für Dermatologie, Venerologie und Allergologie Others Dr. Sascha Rutz, Genentech, San Francisco, USA Dr. Simon Fillatreau, Université de Paris 5 - René Descartes, Hopital Necker Enfants Malades, Paris, Frankreich Selected Publications Bacher P, Heinrich F, Stervbo U, Nienen M, Vahldieck M, Iwert C, Vogt K, Kollet J, Babel N, Sawitzki B, Schwarz C, Bereswill S, Heimesaat MM, Heine G, Gadermaier G, Asam C, Assenmacher M, Kniemeyer O, Brakhage AA, Ferreira F, Wallner M, Worm M, Scheffold A. Regulatory T Cell Specificity Directs Tolerance versus Allergy against Aeroantigens in Humans. Cell (4): tbacher P, Steinbach A, Kniemeyer O, Hamprecht A, Assenmacher M, Vehreschild MJGT, Vehreschild JJ, Brakhage AA, Cornely OA, Scheffold A. Fungus-specific CD4+ T cells for rapid identification of invasive pulmonary mold infection. Am J Respir Crit Care Med (3): Bacher P, Kniemeyer O, Teutschbein J, Thön M, Vödisch M, Wartenberg D, Scharf DH, Koester- Eiserfunke N, Schütte M, Dübel S, Assenmacher M, Brakhage AA*, and Scheffold A*. Identification of immunogenic antigens from Aspergillus fumigatus by direct multi-parameter characterization of specific conventional and regulatory CD4+ T cells J Immunol (7): Neumann C, Heinrich F, Neumann K, Junghans V, Mashreghi MF, Ahlers J, Janke M, Mockel-Tenbrinck N, Kühl A, Heimesaat M, Esser C, Im SH, Radbruch A, Rutz S and Scheffold A. Role of Blimp-1 in programing Th effector cells into IL-10 producers. J Exp Med (9): Bacher P, Kniemeyer O, Schönbrunn A, Sawitzki B, Assenmacher M, Rietschel E, Steinbach A, Cornely OA, Brakhage AA, Thiel A, Scheffold A. Antigen-specific expansion of human regulatory T cells as a major tolerance mechanism against mucosal fungi. Mucosal Immunol (4):

61 Pathophysiology of Rheumatic Inflammation Tokoyoda, Osteoimmunology Roles of memory lymphocytes in protective and auto-reactive immunological memory Tokoyoda, DRFZ group Koji Tokoyoda, PhD Osteoimmunology 60 Keywords Immunological memory, T helper cells, Bone marrow, Secondary immune response Group Member Group leader Koji Tokoyoda, PhD Scientists Shintaro Hojyo, PhD PhD students Jana Sarkander, Christian Männe, Mathias Mursell Students Tsung Yen Wu Memory T helper (Th) cells are critical for longlasting immune memory to infectious pathogens and autoantigens. Despite their central role, the generation, maintenance and reactivation of memory Th cells in the body still remained unclear. Recently, we have found and further described the microenvironment niches for survival of memory Th cells in the bone marrow (BM). We have also reported that CD69 and CD49b (integrin alpha2) are required as homing receptors for the generation and maintenance of memory Th cells in the BM. These molecules can be target molecules to block the generation and maintenance of pathogenic memory Th cells in autoimmune diseases. We have identified the precursors of BM memory Th cells as CD49b+T-bet+ CD4 T cells, generated during the primary immune response. It has been previously reported that B cells contribute to the generation of effector Th cells (Tfh cells). However, we determined that the generation of the precursors of BM memory Th cells was negatively controlled by B cells. These findings suggest that B cells play a role in the bifurcation of activated effector and resting memory Th cell lineages. This finding alerts that, if only inflammation is blocked, pathogenic memory can be inversely enhanced. We should consider blocking both effector and memory T cells to treat autoimmune diseases. We have identified a novel functional ligand of CD69, myosin light chain (Myl) 9 and 12. Myl9/12 is expressed in megakaryocytes and platelets. The migratory precursors of BM memory Th cells adhere to BM sinusoidal endothelial cells via the interaction of CD69 and most likely Myl9/12. In the pathogenesis of allergic airway inflammation, the interaction also functions to recruit inflammatory T cells into inflamed tissues. The role of the interaction of CD69 and Myl9/12 in autoimmune diseases will be investigated. There is still much to discover about the reactivation of memory Th cells, i.e. the recall response. To determine how memory Th cells reactivate memory B cells, we currently study the cellular and molecular mechanisms of humoral memory response in the body, focusing on memory Th cells. Based on the knowledge of the protective antigen-specific memory Th cells in vivo, we now compare the localization and dynamics of protective and auto-reactive memory Th cells and then will determine how auto-reactive memory Th cells are generated, maintained and reactivated in the body, finding out the cellular and molecular targets to treat autoimmune diseases.

62 Tokoyoda, Osteoimmunology Pathophysiology of Rheumatic Inflammation 61 Cooperation Partners Selected Publications Prof. Dr. Stefan H.E. Kaufmann, Immunology, Max Planck Institute for Infection Biology, Berlin, Germany Sarkander J, Hojyo S, Tokoyoda K. Vaccination to gain humoral immune memory. Clin. Transl. Immunology. 5:e120, 2016 Prof. Dr. Toshinori Nakayama, Immunology, Graduate School of Medicine, Chiba University, Chiba, Japan Prof. Dr. Tomoko Yamamoto, Dr. Akiko Takaya, Microbiology, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, Japan Hayashizaki K*, Kimura MY*, Tokoyoda K*, Hosokawa H, Shinoda K, Hirahara K, Ichikawa T, Onodera A, Hanazawa A, Iwamura C, Kakuta J, Muramoto K, Motohashi S, Tumes DJ, Iinuma T, Yamamoto H, Ikehara Y, Okamoto Y, Nakayama T. Myosin light chain 9 and 12 are functional ligands for CD69 that regulate airway inflammation. Science Immunol. 1:eaaf9154, 2016 *equal contribution Hojyo S, Sarkander J, Männe C, Mursell M, Hanazawa A, Zimmel D, Zhu J, Paul WE, Fillatreau S, Löhning M, Radbruch A, Tokoyoda K. B cells negatively regulate the establishment of CD49b+T-bet+ resting memory T helper cells in the bone marrow. Front. Immunol. 7:26, 2016 Dong J, Chang HD, Tokoyoda K, Radbruch A. Immunological memory of the bone marrow. Z. Rheumatol. 74:527-8, 2015 Sercan Alp Ö, Durlanik S, Schulz D, McGrath M, Grün JR, Bardua M, Ikuta K, Sgouroudis E, Riedel R, Zehentmeier S, Hauser AE, Tsuneto M, Melchers F, Tokoyoda K, Chang HD, Thiel A, Radbruch A. Memory CD8(+) T cells colocalize with IL- 7(+) stromal cells in bone marrow and rest in terms of proliferation and transcription. Eur. J. Immunol. 45: ,

63 Pathophysiology of Rheumatic Inflammation Worm, Allergology Novel therapeutic approaches of type I-allergy by immunomodulation Worm, Liaison group with Prof. Dr. med. Margitta Worm Allergology 62 Keywords Allergy B-cells Immune modulation Anaphylaxis Group Member Group leader Prof. Dr. med. Margitta Worm Scientists Dr. Guido Heine, Dr. Kristin Franke, Dr. Sabine Dölle, Dr. Magda Babina, Dr. Vandana Kumari, Dr. Maria Nassiri PhD students Davender Davender, Tina Krause, Juliane Lindner Tarek Hazzan, Shruti Rastogi, Wojciech Francuzik, Maria Michaelidou, Josephine Scholz, Aida Abadpour Bachelor/Diploma/ Master students Nora Schumacher, Jana Hallau, Bianca Lötzsch, Nadine Hielscher, Elvira Ruppel, Oliver Eckermann, Juliane Schulz, Helen Schlachta, Theresa Bartholomäus, Kathleen Winkel, Annika Lahl, Kristina Heins, Aida Abadpour, Wojciech Francuzik, Tarek, Mandana Schindler, Marcel Wittenberg Technicians Dennis Ernst Students Sebastian Knitter, Ronja Ramien Others Study Unit Ariane Lungwitz, Kathrin Wiesenack, Katharina Wylon, Rabea Reinert, Hannah Ahrens The Allergology group is focussed on immunomodulation of IgE-dependent type I allergies. Currently, approx. 20% of the German population are affected by hay fever, allergic asthma or atopic dermatitis. Our research is focussed on the understanding of molecular and cellular key events for the development and maintenance of IgE production. Special interests among our group include immunomodulatory functions of nuclear receptor ligands mechanisms of specific immunotherapy anaphylaxis and regulation of skin homeostasis. Nuclear receptor ligands include molecules like vitamin D and retinoids. We have shown that vitamin D modulates profoundly B cell activation and hampers the IgE response. Currently, we study in detail molecular and cellular signalling and also initiated a clinical translation program. In these studies, we target vitamin D receptors in immune cells in vivo and determine the impact on specific allergic symptoms. Our overall aim is to achieve long-term tolerance to allergens. The studies on specific immunotherapy anaphylaxis and regulation of skin homeostasis include the investigation of patient cohorts upon treatment or reaction (bedside). However, the application of targeted mouse models enables us to set up models with targeted deletion of specific mediators and their receptors to unravel mechanistic functions upon defined pathophysiological conditions. The overall perspective of our research is to develop novel strategies for innovative treatment protocols in allergy. We transfer our experimental data into clinical trials (ad hoc translation).

64 Worm, Allergology Pathophysiology of Rheumatic Inflammation 63 Cooperation Partners Prof. René St. Arnaud, Shriners Hospital for Children, Genetics Unit, Montréal, Canada Prof. Dr. rer. nat. Ria Baumgrass, Deutsches Rheuma-Forschungszentrum Berlin, Cell signaling group, Berlin Prof. Dr. med. Thomas Dörner, Med. Klinik mit Schw. Rheumatologie und Klinische Immunologie, Charité Universitätsmedizin Berlin, Germany Prof. Dr. phil. nat. Hermann Eibel, Center for Chronic Immunodeficiency, Universitätsklinikum Freiburg, Germany Prof. Dr. vet. Reinhold Erben, Universität für Tiermedizin, Institut für Pathophysiologie, Wien, Österreich PD Dr. rer. nat. Lutz Hamann, Charité Universitätsmedizin Berlin, Institut für Mikrobiologie und Hygiene CCM, Germany Prof. Dr. rer. nat. Susanne Hartmann, Institut für Immunologie, FB Veterinärmedizin, Freie Universität Berlin, Germany Dr. rer. nat. Andreas Hutloff, PhD, Deutsches Rheuma-Forschungszentrum Berlin, Germany Prof. Dr. med. Bastian Opitz, Prof. Dr. med. Martin Witzenrath, Christoph Tabeling, Charité Universitätsmedizin Berlin, Med. Klinik mit Schw. Infektiologie und Pneumologie, Germany Prof. Dr. med. Harald Renz, Philipps-University of Marburg, Dept of Clinical Chemistry and Molecular Diagnostics, Marburg, Germany Prof. Dr. rer. nat. Michael Reth, MPI Immunbiology and Epigenetics, Freiburg, Germany Prof. Dr. med. Birgit Sawitzki, Charité Universitätsmedizin Berlin, Institut für Medizinische Immunologie, Germany Prof. Dr. rer. nat. Alexander Scheffold, Deutsches Rheuma-Forschungszentrum Berlin, Germany Prof. Dr. med. Ralf Schumann, PD Lutz Hamann, Charité Universitätsmedizin Berlin, Institut für Mikrobiologie und Hygiene, Berlink, Germany Prof. Dr. med. Hans-Dieter Volk, Charité Universitätsmedizin Berlin, Klinik für Medizinische Immunologie, Germany Selected Publications Moinzadeh P; Aberer E; Ahmadi-Simab K; Blank N; Distler JH; Fierlbeck G; Genth E; Guenther C; Hein R; Henes J; Herich L; Herrgott I; Koetter I; Kreuter A; Krieg T; Kuhr K; Lorenz HM; Meier F; Melchers I; Mensing H; Mueller-Ladner U; Pfeiffer C; Riemekasten G; Sárdy M; Schmalzing M; Sunderkoetter C; Susok L; Tarner IH; Vaith P; Worm M; Wozel G; Zeidler G; Hunzelmann N; and all participating DNSS centers (2015). Disease progression in systemic sclerosis-overlap syndrome is significantly different from limited and diffuse cutaneous systemic sclerosis. Ann Rheum Dis. 74(4): Nassiri M, Babina M, Dölle S, Edenharter G, Ruëff F, Worm M (2015). Ramipril and metoprolol intake aggravate human and murine anaphylaxis: evidence for direct mast cell priming. J Allergy Clin Immunol. 135(2): Grabenhenrich LB, Dölle S, Moneret-Vautrin A, Köhli A, Lange L, Spindler T, Ruëff F, Nemat K, Maris I, Roumpedaki E, Scherer K, Ott H, Reese T, Mustakov T, Lang R, Fernandez-Rivas M, Kowalski ML, Bilò MB, Hourihane JO, Papadopoulos NG, Beyer K, Muraro A, Worm M (2016). Anaphylaxis in children and adolescents: The European Anaphylaxis Registry. J Allergy Clin Immunol. Apr;137(4): e1. Bacher P, Heinrich F, Stervbo U, Nienen M, Vahldieck M, Iwert C, Vogt K, Kollet J, Babel N, Sawitzki B, Schwarz C, Bereswill S, Heimesaat MM, Heine G, Gadermaier G, Asam C, Assenmacher M, Kniemeyer O, Brakhage AA, Ferreira F, Wallner M, Worm M, Scheffold (2016). Regulatory T Cell Specificity Directs Tolerance versus Allergy against Aeroantigens in Humans. Cell. Nov 3;167(4): e16. Thaçi D; Simpson EL; Beck LA; Bieber T; Blauvelt A; Papp K; Soong W; Worm M; Szepietowski JC; Sofen H; Kawashima M; Wu R; Weinstein SP; Graham NM; Pirozzi G; Teper A; Sutherland ER; Mastey V; Stahl N; Yancopoulos GD; Ardeleanu M (2016). Efficacy and safety of dupilumab in adults with moderate-to-severe atopic dermatitis inadequately controlled by topical treatments: a randomised, placebo-controlled, dose-ranging phase 2b trial. Lancet. Jan 2;387(10013):40-52.t

65 DRFZ group Mir Farzin Mashreghi Liaison group Max Löhning 66

66 Programme Area Regenerative Rheumatology Löhning, Pitzer Laboratory of Osteoarthritis Research Mashreghi, Therapeutic Gene Regulation... 68

67 Regenerative Rheumatology Löhning, Pitzer Laboratory of Osteoarthritis Research Biological regeneration of cartilage by reprogramming of chondrocytes Prof. Dr. rer. nat. Max Löhning Löhning, Pitzer Laboratory of Osteoarthritis Research Liaison group with 66 Keywords Osteoarthritis pathogenesis, Chondrocyte differentiation, Bone cell development, Bone angiogenesis, Lymphocyte differentiation, Immunological memory Group Member Group leader Prof. Dr. rer. nat. Max Löhning Scientists Dr. Claudia Baumann, Dr. Caroline Helmstetter, Dr. Roman Marek, Dr. Melba Muñoz, Dr. Michael Peine, Dr. Philippe Saikali, Dr. Ping Shen, Dr. David Zimmel PhD students Tobias Brunner, Angeliki Datsi, Maria Dzamukova, Dominik Niesen, Julia Siede, Domonkos Vince Varga Technicians Sabine Ebel, Christiane Golz-Wehner, Vivien Holecska, Sabine Iden, Isabel Panse, Carola Rüster, Peihua Wu The Pitzer Laboratory of Osteoarthritis Research investigates the cellular and molecular mechanisms leading to the development of osteoarthritis (OA). Our goal is biological regeneration of cartilage in affected joints. OA is the most frequently occurring joint disease among adults worldwide and the most important cause of disability among elderly people in Germany. This chronic degenerative disease leads to progressive cartilage loss and is often accompanied by inflammatory processes. Recent studies indicate dysfunctional molecular signaling within the cartilage-producing cells of the joints, the chondrocytes. This applies particularly to differentiation and apoptosis programs of these cells. So far, the chondrocytes in situ biology is understood only to some degree. Here, the biggest obstacles for further advancement in the research of these rare cells are (1.) their difficult accessibility and (2.) the limited possibilities to isolate and culture them without changing their phenotypes. Chondrocytes reside in the joint cartilage layers in different arrangements and with varying types of metabolism. It is not clear yet whether this points to a homogenous population or to differentiated subtypes. It is known that central transcription factors like SOX9 and RUNX2 regulate the chondrocytes development right up to hypertrophic and/or degenerative stages. Comparable processes take place in the immune system that can be regarded as a prototypical model of cell differentiation in a complex network responding to its environment. Here, our group's accumulated knowledge on the (re-) programming of T cells will be transferred to chondrocytes and the field of OA research. In previous studies, we identified central cytokines and key transcription factors controlling the differentiation of T cells into subtypes (Bonilla et al., Science 2012; Peine et al., PLoS Biol. 2013; Baumann et al., PNAS 2015; Peine et al., Trends Immunol. 2016). In addition, we used this understanding of molecular processes to reprogram mature T cells into new stable phenotypes with additional functions (Hegazy et al., Immunity 2010). More recently, we showed a quantitative cytokine memory in individual cells. This means that a cell memorizes and stably maintains its individual production amount of a given cytokine (Helmstetter et al., Immunity 2015). We suggest that chondrocytes feature similar subtypes, differentiation programs, and possibilities of reprogramming. Ultimately, we want to reprogram the chondrocyte phenotypes that lead to the development of OA in patients in such a way as to achieve a longlasting cartilage build-up.

68 Löhning, Pitzer Laboratory of Osteoarthritis Research Regenerative Rheumatology 67 Cooperation Partners Selected Publications University of Colorado, Denver, CO, USA: Prof. Dr. Charles A. Dinarello NIH, NIAID, Bethesda, MD, USA: Dr. Jinfang Zhu, Dr. Dragana Jankovic University of Tokyo, Japan: Prof. Dr. Susumu Nakae Chiba University, Japan: Prof. Dr. Toshinori Nakayama University of Oxford, UK: Prof. Dr. Fiona Powrie, Dr. Dr. Ahmed N. Hegazy ETH & Universitätsspital Zürich, Schweiz: Prof. Dr. Rolf M. Zinkernagel, Prof. Dr. Hans Hengartner Universität Basel, Schweiz: Dr. Weldy Bonilla, Prof. Dr. Daniel D. Pinschewer, Prof. Dr. Mike Recher Universitätsklinikum Essen: Prof. Dr. Karl S. Lang Universität Düsseldorf: Prof. Dr. Philipp A. Lang Deutsches Krebsforschungszentrum (DKFZ), Heidelberg: Prof. Dr. Thomas Höfer, Dr. Michael Floßdorf, Dr. Yevgeniy Vainshtein, Dr. Qin Zhang Universität Marburg: Prof. Dr. Michael Lohoff Universität Gießen: Prof. Dr. Christoph G. Grevelding FU Berlin, Veterinärmedizin: Prof. Dr. Susanne Hartmann, Dr. Sebastian Rausch MPI für Infektionsbiologie, Berlin: Prof. Dr. Stefan Kaufmann, Prof. Dr. Arturo Zychlinsky, Dr. Nikolaus Deigendesch, Dr. Andreas Kupz Helmstetter C, Flossdorf M, Peine M, Kupz A, Zhu J, Hegazy AN, Duque-Correa MA, Zhang Q, Vainshtein Y, Radbruch A, Kaufmann SH, Paul WE, Höfer T, Löhning M. Individual T helper cells have a quantitative cytokine memory. Immunity Jan 20; 42(1): Baumann C, Bonilla WV, Fröhlich A, Helmstetter C, Peine M, Hegazy AN, Pinschewer DD, Löhning M. T-bet- and STAT4-dependent IL-33 receptor expression directly promotes antiviral Th1 cell responses. Proc Natl Acad Sci USA Mar 31; 112(13): Bhattacharya* A, Hegazy* AN, Deigendesch N, Kosack L, Cupovic J, Kandasamy RK, Hildebrandt A, Merkler D, Kühl AA, Vilagos B, Schliehe C, Panse I, Khamina K, Baazim H, Arnold I, Flatz L, Xu HC, Lang PA, Aderem A, Takaoka A, Superti- Furga G, Colinge J, Ludewig B, Löhning# M, Bergthaler# A. Superoxide dismutase 1 protects hepatocytes from type I interferon-driven oxidative damage. Immunity Nov 17; 43(5): */# equal contributions. Universität Genf, Schweiz: Prof. Dr. Doron Merkler Kantonsspital St. Gallen, Schweiz: Prof. Dr. Lukas Flatz Universität Lausanne, Schweiz: Prof. Dr. Sanjiv Luther Research Center for Molecular Medicine of the Austrian Academy of Science, Wien, Österreich: Dr. Andreas Bergthaler Charité, Berlin: Prof. Dr. Gerd-Rüdiger Burmester, Prof. Dr. Frank Buttgereit, Prof. Dr. Georg Duda, Dr. Michael Fuchs, Dr. Anja A. Kühl, Prof. Dr. Marcus Maurer, Prof. Dr. Martin Metz, Prof. Dr. Carsten Perka, Dr. Philipp von Roth, Prof. Dr. Alexander Scheffold, PD Dr. Uta Syrbe, Prof. Dr. Hans-Dieter Volk, Prof. Dr. Margitta Worm Miltenyi Biotec, Bergisch Gladbach: Dr. Mario Assenmacher, Dr. Andreas Bosio, Dr. Anne Richter Peine M, Marek RM, Löhning M. IL-33 in T cell differentiation, function, and immune homeostasis. Trends Immunol May; 37(5): Siede J, Fröhlich A, Datsi A, Hegazy AN, Varga DV, Holecska V, Saito H, Nakae S, Löhning M. IL-33 receptor-expressing regulatory T cells are highly activated, Th2 biased and suppress CD4 T cell proliferation through IL-10 and TGFβ release. PLoS One Aug 22; 11(8):e

69 Regenerative Rheumatology Mashreghi, Therapeutic Gene Regulation Targeting of molecular factors mediating inflammatory processes in osteoarthritis Mashreghi, DRFZ group Dr. rer. nat. Mir-Farzin Mashreghi Therapeutic Gene Regulation 68 Keywords Regulatory RNA, Oligonucleotide therapy, Gene Regulation, Next Generation Sequencing, Chronic Inflammation Group Member Group leader Dr. rer. nat. Mir-Farzin Mashreghi Scientists Dr. rer. nat. Gitta Anne Heinz, Dr. rer nat. Patrick Maschmeyer, Dr. rer. nat. Frederik Heinrich PhD students Markus Bardua, Cam Loan Tran Bachelor/Diploma/ Master students Vanessa Treffner, Sarah Schimmelpfennig, Melanie Klingauf Chronic inflammation is not only a key feature of autoimmune disorders including rheumatoid arthritis, inflammatory bowel disease and multiple sclerosis, but is currently discussed to be a trigger for the initiation and propagation of osteoarthritis (OA), formerly considered as a non-inflammatory arthritis. Many immune cell types contribute to the persistence of chronic inflammation, either by promoting proinflammatory responses or by interfering with biological processes that allow the regeneration of destroyed tissues. In order to skew the balance in favor of regeneration as opposed to immunopathology, we want to acquire a profound knowledge of the molecular mechanisms that mediate such biological responses. Our aim is to identify genes and regulatory RNAs and their underlying functions by which they control the activity and survival of cells that contribute to chronic inflammation or help to regenerate destroyed tissues. For the global identification of such genes and regulatory RNAs, we have established a next generation sequencing platform. This technology allows us to obtain the expression patterns of mrna and regulatory RNAs even from rare cells or from limited sample material that we obtain from patients with rheumatic diseases as well as OA. Furthermore, we want to interfere with the cellular functions by using therapeutic oligonucleotides targeting the identified mrna and regulatory RNAs in preclinical models of inflammation and osteoarthritis. So far, we have identified the microrna, mir-148a, which is upregulated in memory T helper (Th) cells isolated from inflamed tissues of patients with rheumatic joint diseases. We could show that mir-148a directly promotes the survival of repeatedly activated Th1 cells. By using a class of cholesterol-tagged inhibitory oligonucleotides against micrornas, so called antagomirs, we achieved a significant knock-down of mir-148a in repeatedly activated Th1 cells. This knockdown in turn resulted in an upregulation of the mrna coding for the proapoptotic protein Bim, which is a target of mir-148a. The consequence of Bim upregulation was enhanced apoptosis of repeatedly activated Th1 cells. Thus, mir-148a qualifies as a therapeutic target in order to deplete proinflammatory Th1 cells by systemic application of antagomirs in chronic inflammatory diseases. Our results demonstrate that therapeutic oligonucleotides are a suitable tool to reduce the expression of cell intrinsic target RNAs driving chronic inflammation and thus, to selectively modulate or ablate pathogenic cells. In the future we are going to further investigate the potential of therapeutic oligonucleotides including sirnas and antagomirs in targeting relevant genes and regulatory RNAs that are important in maintaining chronic inflammation and help to achieve remission in degenerative diseases such as OA.

70 Mashreghi, Therapeutic Gene Regulation Regenerative Rheumatology 69 Cooperation Partners Selected Publications Dr. Tobias Alexander, Klinik für Rheumatologie und klinische Immunologie, Charité-Universitätsmedizin Berlin Prof. Dr. Klemens Budde, Department of Nephrology, Charité-Universitätsmedizin Berlin Prof. Dr. Falk Hiepe, Klinik für Rheumatologie und klinische Immunologie, Charité-Universitätsmedizin Berlin Prof. Dr. Hans-Martin Jäck, Molekulare Immunologie, Universitätsklinikum Erlangen Dr. Anja Kühl, Institut für Pathologie, Charité- Universitätsmedizin Berlin Prof. Dr. Michael Lohoff, Institut für Medizinische Mikrobiologie, Philipps University Marburg Dr. Mareen Matz, Department of Nephrology, Charité-Universitätsmedizin Berlin Haftmann C, Stittrich AB, Zimmermann J, Fang Z, Hradilkova K, Bardua M, Westendorf K, Heinz GA, Riedel R, Siede J, Lehmann K, Weinberger EE, Zimmel D, Lat, Hoffmann U, Porstner M, Chen W, Grün JR, Baumgrass R, Matz M, Löhning M, Scheffold A, Wittmann J, Chang HD, Rajewsky N, Jäck HM, Radbruch A, Mashreghi MF. mir-148a is upregulated by Twist1 and T-bet and promotes Th1-cell survival by regulating the proapoptotic gene Bim. Eur J Immunol Apr;45(4): doi: /eji PubMed PMID: ; PubMed Central PM- CID: PMC Haftmann C, Riedel R, Porstner M, Wittmann J, Chang HD, Radbruch A, Mashreghi MF. Direct uptake of Antagomirs and efficient knockdown of mirna in primary B and T lymphocytes. J Immunol Methods Nov;426: doi: /j.jim PubMed PMID: ; PubMed Central PMCID: PMC Apr;100(4): doi: / TP PubMed PMID: *equal contribution Matz M, Lorkowski C, Fabritius K, Durek P, Wu K, Rudolph B, Neumayer HH, Mashreghi MF*, Budde K*. Free microrna levels in plasma distinguish T-cell mediated rejection from stable graft function after kidney transplantation. Transpl Immunol Nov;39: doi: /j. trim PubMed PMID: *equal contribution Weber JP, Fuhrmann F, Feist RK, Lahmann A, Al Baz MS, Gentz LJ, Vu Van D, Mages HW, Haftmann C, Riedel R, Grün JR, Schuh W, Kroczek RA, Radbruch A, Mashreghi MF, Hutloff A. ICOS maintains the T follicular helper cell phenotype by down-regulating Krüppel-like factor 2. J Exp Med Feb 9;212(2): doi: / jem PubMed PMID: ; Pub- Med Central PMCID: PMC Prof. Dr. Nikolaus Rajewsky, Max-Delbrück- Centrum für Molekulare Medizin und Berlin Institute for Medical Systems Biology, Berlin Prof. Dr. Kai Wucherpfennig, Dana-Farber Cancer Institute, Boston, MA, USA Matz M, Fabritius K, Lorkowski C, Dürr M, Gaedeke J, Durek P, Grün JR, Goestemeyer A, Bachmann F, Wu K, Rudolph B, Schmidt D, Weber U, Haftmann C, Unterwalder N, Lachmann N, Radbruch A, Neumayer HH, Mashreghi MF*, Budde K*. Identification of T Cell-Mediated Vascular Rejection After Kidney Transplantation by the Combined Measurement of 5 Specific Micro- RNAs in Blood. Transplantation.

71 DRFZ groups Kirsten Minden 72 Anja Strangfeld 76 Angela Zink Liaison group Denis Poddubnyy 74

72 Programme Area Epidemiology of rheumatic diseases Minden, Paediatric rheumatology and health services research...72 Poddubnyy, Spondyloarthritiden...74 Strangfeld, Pharmacoepidemiology Zink, Health services research in adult rheumatology... 78

73 Epidemiology of Rheumatic Diseases Minden, Paediatric Rheumatology and Health Services Research Optimising health care for young people with rheumatic diseases Minden, DRFZ group Prof. Dr. med. Kirsten Minden Paediatric Rheumatology and Health Services Research 72 Keywords Juvenile rheumatic diseases, outcomes, health care provision, cost of illness, transition Group Member Group leader Prof. Dr. med. Kirsten Minden Scientists Dr. rer. nat. Jens Klotsche, Dr. med Miriam Listing, Martina Niewerth (MPH), Sandra Schenck (Msc), Dr. med. Claudia Sengler, Dr. rer. medic. Florian Milatz PhD students Pascale Kurt, Paula Drechsel Data manager Nils Geisemeyer, Nadine Grösch, Ina Liedmann, Cornelia Stamme-Schäfer, Karin Weber MD students Marisa Beck, Nina Derksen, Ae-Rie Schipolowski, Julian Zink, Ewelina Türck, Svea Horn, Michaela Heinrich Students Juliane Lenz, Anneli Rieck, Katharina Teifel, Annemarie Uhlmann The focus of our research is on juvenile idiopathic arthritis (JIA), the most common chronic inflammatory rheumatic disease in childhood and adolescence. Three large prospective observational cohort studies allow us to address various research questions with regard to phenotypes, outcome and its determinants, cost and adequacy of health care provision in JIA: 1) the national paediatric rheumatologic database (NPRD), in which more than 8,000 children and adolescents with JIA from 61 paediatric rheumatology sites in Germany are recorded per year, 2) the biologic register JuMBO (Juvenile arthritis Methotrexate/Biologics long-term Observation) for young adults with JIA that follows about 1,200 patients into adulthood, 3) the Inception Cohort Of Newly diagnosed children with JIA (ICON), in which about 950 JIA patients and 480 healthy controls are observed. During the last year, our group has specifically examined risk factors, frequency and outcomes of uveitis in patients with recent onset of JIA, how JIA affects the quality of life of children and adolescents and whether current routine care reduces the burden of illness, the cost of early JIA from the societal perspective and the main cost drivers in the first two years of paediatric rheumatology care, participation in school and club sport of children and adolescents with JIA, its trend over time and associated factors by cross sectional data of the NPRD from 2000 to 2015, predictors of a successful stop of methotrexate in JIA patients who achieved remission on drug and were recorded in the JIA biologic registers BiKeR and JuMBO, the health care situation of young people in the period of transfer from child-centred to adultoriented health care services by claims data. Risk factors of uveitis, such as disease onset before the 4th birthday, the JIA category oligoarthritis, the presence of antinuclear antibodies and a high disease activity were identified. They may help in identifying a patient group that could benefit from early treatment with methotrexate, since we could demonstrate that this drug prevents uveitis. Current routine care improves the quality of life of JIA patients, approximating that of healthy peers after three years of care. During this time, most JIA patients attain inactive disease at least once. We found that the likelihood of experiencing recurrence of active disease is a function of time in inactive disease prior to drug discontinuation. Thus, patients in inactive disease should remain on treatment for at least 12 months before drug discontinuation. All projects are based on close cooperation with the paediatric rheumatology sites in Germany, the national uveitis study group, and other national and international research groups.

74 Minden, Paediatric Rheumatology and Health Services Research Epidemiology of Rheumatic Diseases 73 Cooperation Partners Dr. Ivan Foeldvari, Kinderrheumatologische Praxis am AK Eilbek, Hamburg Prof. Dr. Dirk Foell, Universität Münster Dr. Gerd Ganser, St. Joseph-Stift, Sendenhorst Prof. Dr. Johannes-Peter Haas, Deutsches Zentrum für Kinder- und Jugendrheumatologie, Garmisch Partenkirchen Prof. Dr. Gerd Horneff, Asklepios Kinderklinik Sankt Augustin Dr. Anton Hospach, Klinikum Stuttgart Prof. Dr. Hans-Iko Huppertz, Klinikum Bremen Mitte PD Dr. Tilmann Kallinich, Universitätsmedizin Charité - Berlin Prof. Dr. Jasmin Kümmerle-Deschner, Universitätsklinik Tübingen Dr. Kirsten Mönkemöller, Kinderkrankenhaus der Stadt Köln Dr. Angelika Thon, Medizinische Hochschule Hannover and 50 other pediatric rheumatologic and more than 200 adult rheumatologic sites Prof. Dr. Arnd Heiligenhaus, Augenabteilung am St. Franziskus Hospital, Münster PD Dr. Christoph Tappeiner, Universitätsklinik für Augenheilkunde, Schweiz and more than 200 opthalmologic sites Dr. Heinrike Schmeling, Alberta Children's Hospital, University of Calgary, Canada Prof. Dr. Nico Wulffraat, Wilhelmina Ziekenhuis, Utrecht, Niederlande Prof. Dr. Reinhard W. Holl, Universität Ulm / Joachim Rosenbauer, Deutsches Diabetes-Zentrum Düsseldorf - Diabetes-Patienten-Verlaufsdokumentation (DPV) Dr. Ulrich Neudorf / PD Dr. Elke Lainka, Universitätsklinik Essen / Prof. Dr. Tim Niehues, HELI- OS-Klinikum Krefeld - AID-Net Andres Luque Ramos, Falk Hoffmann, Universität Oldenburg - PROCLAIR Prof. Dr. Burkhard Tönshoff / Prof. Dr. Petra Knaup, Universitätsklinikum Heidelberg Kinderrheumatologie Online Kerndokumentation (KRhOKo) Prof. Dr. Michael Hofer, Centre Hospitalier Universitaire Vaudois, Lausanne, Schweiz - Juvenile Inflammatory Rheumatism Cohorte Karl Cattelaens, Monika Mayer, Deutsche Rheuma-Liga, Bonn Bettina Model, Deutsche Kinder-Rheumastiftung, München Selected Publications Klotsche J, Niewerth M, Haas JP, Huppertz HI, Zink A, Horneff G, Minden K. Long-term safety of etanercept and adalimumab compared to methotrexate in patients with juvenile idiopathic arthritis (JIA). Ann Rheum Dis May;75(5): Foster HE, Minden K, Clemente D, Leon L, Mc- Donagh JE, Kamphuis S, Berggren K, van Pelt P, Wouters C, Waite-Jones J, Tattersall R, Wyllie R, Stones SR, Martini A, Constantin T, Schalm S, Fidanci B, Erer B, Dermikaya E, Ozen S, Carmona L. EULAR/PReS standards and recommendations for the transitional care of young people with juvenile-onset rheumatic diseases. Ann Rheum Dis Apr;76(4): Horneff G, Klein A, Oommen PT, Hospach A, Foeldvari I, Feddersen I, Minden K. Update on malignancies in children with juvenile idiopathic arthritis in the German BIKER Registry. Clin Exp Rheumatol Nov-Dec;34(6): Klotsche J, Raab A, Niewerth M, Sengler C, Ganser G, Kallinich T, Niehues T, Hufnagel M, Thon A, Hospach T, Horneff G, Minden K. Outcome and Trends in Treatment of Systemic Juvenile Idiopathic Arthritis in the German National Pediatric Rheumatologic Database, Arthritis Rheumatol Dec;68(12): Sengler C, Klotsche J, Niewerth M, Liedmann I, Föll D, Heiligenhaus A, Ganser G, Horneff G, Haas JP, Minden K. The majority of newly diagnosed patients with juvenile idiopathic arthritis reach an inactive disease state within the first year of specialised care: data from a German inception cohort. RMD Open Dec 8;1(1):e

75 Epidemiology of Rheumatic Diseases Poddubnyy, Spondyloarthritides Early diagnosis and effective therapy of chronic inflammatory spine disease Poddubnyy, Liaison group with Prof. Dr. med. Denis Poddubnyy Spondyloarthritides 74 Keywords Spondyloarthritis, ankylosing spondylitis, diagnosis, treatment Group Member Group leader Prof. Dr. med. Denis Poddubnyy (from )/ Prof. Dr. Joachim Sieper (till ) Scientists PD Dr. Hildrun Haibel, PD Dr. Uta Syrbe, Dr. Burkhard Muche, Dr. Fabian Proft, Dr. Mikhail Protopopov, Dr. Judith Rademacher, Valeria Rios-Rodriguez, Inge Spiller, Dr. Laura Spiller, Dr. Anne-Katrin Weber MD students Agnes Hartl, Lorraine Tietz, Henning Weineck Technicians/Study nurses Annegret Langdon, Claudia Lorenz, Bianca Mandt, Anna-Lisa Meinke, Judith Reichardt, Martina Seipel, Petra Tietz In our clinical research work, we are interested in the epidemiology of spondyloarthritis as well as in improving diagnosis and therapy of the disease. In basic research work, we focus on elucidating the pathogenesis, including mechanisms of inflammation and bone formation in spondyloarthritis (SpA). In the PROCLAIR study we investigate the disease burden, standards of care and diseaserelated costs in ankylosing spondylitis in Germany. Regarding early diagnosis of SpA, we developed and tested screening parameters for identification of patients with possible SpA, which is critical for an early diagnosis of the disease. These data together with published results from other research groups resulted recently in the development of an international recommendation for the early referral of patients with a suspicion of axial spondyloarthritis by non-rheumatology specialists dealing with patients with back pain (orthopaedists, general practitioners). Further studies exploring possibilities of self-referral and developing referral strategies for other specialists (i.e., ophthalmologists, dermatologists, gastroenterologists) are ongoing. Apart from this, we initiated a prospective cohort of patients with axial spondyloarthritis, the German Early Spondyloarthritis Inception Cohort GESPIC (the project is still ongoing). In this cohort we study the natural course of the disease, predictors of a long-term outcome and of structural damage development in the sacroiliac joints and in the spine, including various biomarkers (ArthroMark project) and metabolic factors (METARTHROS project). Most recently we extended GESPIC to patients with early Crohn s disease in order to study factors (including microbiome) associated with the development of arthritis phenotype. Moreover, we performed a number of clinical trials analyzing the effect of different treatment options like glucocorticoids, TNF blockers and other biologics, such as ustekinumab, in the treatment of axial spondyloarthritis. The ESTHER study is the first study worldwide to investigate a long-term (10 years) follow-up in patients with early axial SpA treated with TNF blockers. In 2016 we initiated a large multicenter study CONSUL aimed at investigation of the effect of treatment with NSAIDs added to anti- TNF therapy versus anti-tnf therapy alone on progression of Structural damage in the spine over two years in patients with ankylosing spondylitis. In the ongoing NEUROIMPA project we investigate intraarticular treatment options in patients with arthritis of the knee joint.

76 Poddubnyy, Spondyloarthritides Epidemiology of Rheumatic Diseases 75 Cooperation Partners Selected Publications Prof. D. Baeten, Amsterdam University, Netherlands Prof. D. van der Heijde, Leiden University, Netherlands Dr. A. van Tubergen, University of Maastricht, Netherlands Prof. R. Landewe, Amsterdam University, Netherlands Prof. R. Lories, Universität Leuven, Belgium Prof. M. Ostergaard, Dr. S. Pedersen, University of Copenhagen, Denmark Germany Prof. J. Braun, Rheumazentrum Ruhrgebiet, Herne Prof. G.-R. Burmester, Med. Klinik m.s. Rheumatologie und Klinische Immunologie, Charité - Campus Mitte, Berlin PD Dr. K.-G. Hermann, Radiologie, Charité - Campus Mitte, Berlin Prof. U. Müller-Ladner, PD Dr. E. Neumann, Justus-Liebig-Universität Gießen Prof. A. Thiel, Immunologie im BCRT, Charité - Cambus Virchow Klinikum, Berlin Prof. G. Schett, Universität Erlangen Poddubnyy D, Fedorova A, Listing J, Haibel H, Baraliakos X, Braun J, et al. Physical Function and Spinal Mobility Remain Stable Despite Radiographic Spinal Progression in Patients with Ankylosing Spondylitis Treated with TNF-alpha Inhibitors for Up to 10 Years. J Rheumatol Dec; 43(12): Poddubnyy D, Protopopov M, Haibel H, Braun J, Rudwaleit M, Sieper J. High disease activity according to the Ankylosing Spondylitis Disease Activity Score is associated with accelerated radiographic spinal progression in patients with early axial spondyloarthritis: results from the GErman SPondyloarthritis Inception Cohort. Ann Rheum Dis Dec; 75(12): Sieper J, Listing J, Poddubnyy D, Song IH, Hermann KG, Callhoff J, et al. Effect of continuous versus on-demand treatment of ankylosing spondylitis with diclofenac over 2 years on radiographic progression of the spine: results from a randomised multicentre trial (ENRADAS). Ann Rheum Dis Aug; 75(8): Poddubnyy D, van Tubergen A, Landewe R, Sieper J, van der Heijde D, Assessment of SpondyloArthritis international S. Development of an ASASendorsed recommendation for the early referral of patients with a suspicion of axial spondyloarthritis. Ann Rheum Dis Aug; 74(8): Poddubnyy D, Listing J, Sieper J. Brief Report: Course of Active Inflammatory and Fatty Lesions in Patients With Early Axial Spondyloarthritis Treated With Infliximab Plus Naproxen as Compared to Naproxen Alone: Results From the Infliximab As First Line Therapy in Patients with Early Active Axial Spondyloarthritis Trial. Arthritis Rheumatol Aug; 68(8):

77 Epidemiology of Rheumatic Diseases Strangfeld, Pharmacoepidemiology Safety and effectiveness of drug treatment in daily rheumatologic care Strangfeld, DRFZ group Dr. med. Anja Strangfeld Pharmacoepidemiology 76 Keywords Pharmacovigilance, Drug safety, Real world effectiveness, Observational long-term cohort, New therapies Group Member Group leader Dr. med Anja Strangfeld Deputy Group leader: Anne Regierer MD Scientists Lisa Baganz MSc, Dörthe Kühl-Habich MSc, Joachim Listing PhD, Dagmar Pattloch MSc, Adrian Richter PhD, Anja Weiß MSc PhD students Yvette Meissner MD students Madlen Spilka Medical data managers Christina Bungartz, Tanja Ellmann, Mandy Grope, Ulrike Kamenz, Lisa Lindner, Bettina Marsmann, Steffen Meixner, Juliane Reinke, Carina Wittkopp, Birgit Wuthe, Susanna Zernicke, Steffi Zühlke Database administrators Claudia Fritz, Dina Schreckeis Students Juliana Hoffmann, Ina Kammermeier, Lea Karnstedt, Ronald Müller, Steve Mvogo Ondoua, Linda Ruppert, Jasmin Sandor, Amy Schmiedeskamp, Julia Seidel The main interest of our group is to investigate longterm outcomes of rheumatic diseases including the effects of drug treatment in daily routine care. In 2001 we established the longitudinal observational cohort study RABBIT (Rheumatoid Arthritis: observation of BIologics Therapy) to closely monitor patients with rheumatoid arthritis (RA) treated with newly approved drugs and to compare the outcomes with an internal control group of patients under conventional synthetic treatment. Until December 2016, more than patients starting a treatment were enrolled by about 300 rheumatologists all over Germany. Patients are observed for at least 5 and up to 10 years, regardless of any treatment changes. Every 6 months details about treatment course and adverse events are collected. We are continuously expanding the observed therapies by newly approved drugs. In 2015 and 2016, the first three biosimilars were incorporated in the observation. Serious infections are common adverse events in patients with RA. A great concern is the development of sepsis where case fatality reach rates up to 50%. With data of the RABBIT cohort, we could confirm that older patients and those with a chronic renal disease are more likely to develop sepsis after a serious infection. The highlight of the analysis was a significantly lower risk for sepsis in patients treated with a biologic drug at the time of the serious infection compared to patients on conventional synthetic DMARD treatment. This result, for the first time and with striking effect size, confirmed observations from animal studies with epidemiologic data. This had not been possible in humans before. With Rhekiss, started in 2015, we run a prospective cohort study dedicated to investigate the safety of therapies in pregnancy and lactation. We aim to add substantial evidence to the limited knowledge about the course and outcome of pregnancies in patients with various inflammatory rheumatic diseases and the influence of drug treatment. Women wishing for a child or at the beginning of pregnancy can be enrolled. The course of pregnancy including treatments and complications, the outcome and the first two years of child development are reported from rheumatologists and patients. One hundred rheumatologic units enrolled more than 400 pregnant patients until December The first results were reported at EULAR and ACR. In contrast to RABBIT, Rhekiss is a web-based system. A new project currently under development in our research group is RABBIT-SpA, a web-based observational cohort study to observe patients with axial spondyloarthritis and psoriatic arthritis. The results of our projects are highly relevant for clinicians and are directly transferred into daily clinical practice. The identification of prognostic factors or better treatment strategies for specific groups of patients, but as well the detection of risk factors for specific adverse events help clinicians to individualize treatments and to find the most suitable therapy for each patient. One example is the widely used calculator for the risk of serious infection (RABBIT Risk Score Calculator) which is available on our website.

78 Strangfeld, Pharmacoepidemiology Epidemiology of Rheumatic Diseases 77 Cooperation Partners Selected Publications 300 rheumatologists in Germany PD Dr. Rebecca Fischer-Betz, Universität Düsseldorf (Rhekiss project) PD Dr. med. Jutta Richter, Universität Düsseldorf (REBRA satellite project, Rhekiss) Prof. Dr. med. Matthias Schneider (REBRA satellite project, Rhekiss) Members of the scientific advisory boards: Prof. Dr. Jürgen Braun, Herne; Prof. Dr. Bernhard Manger, Erlangen; Prof. Dr. Jörn Kekow, Vogelsang-Gommern; Prof. Dr. Peter Herzer, München; Prof. Dr. Matthias Schneider, PD Dr. Jutta Richter, PD Dr. Rebecca Fischer-Betz, Düsseldorf; Dr. Susanna Späthling-Mestekemper, München; Prof. em. Dr. Jochen Sieper, Berlin; Prof. Dr. Christoph Specker, Essen; International registers: AIR (Montpellier, F), ARTIS (Stockholm, S), ATTRA (Prag, CZ), BIOBADASER (Madrid, E), BSRBR (Manchester, GB), DANBIO (Kopenhagen, DK), GISEA (Bari, I), ORA (Straßburg, F), RATIO (Paris, F), REGATE (Straßburg, F), SCQM (Genf, S), Rheuma.pt (Lissabon, P) Research Networks: EULAR Working Group RODS (Register and Observational Drugs Studies), ENCePP (European Network of Centers of Pharmacovigilance and Pharmacoepidemiology), EuNeP (European Network of Pregnancy Registers) Strangfeld A, Richter A, Siegmund B, Herzer P, Rockwitz K, Demary W, Aringer M, Meißner Y, Zink A, Listing J. Risk for lower intestinal perforations in patients with rheumatoid arthritis treated with tocilizumab in comparison to treatment with other biologic or conventional synthetic DMARDs. Ann Rheum Dis Mar; 76(3): doi: /annrheumdis [Online first: July 2016] Richter A, Listing J, Schneider M, Klopsch T, Kapelle A, Kaufmann J, Zink A, Strangfeld A. Impact of treatment with biologic DMARDs on the risk of sepsis or mortality after serious infection in patients with rheumatoid arthritis. Ann Rheum Dis Sep; 75(9): doi: /annrheumdis Pattloch D, Richter A, Manger B, Dockhorn R, Meier L, Tony HP, Zink A, Strangfeld A. [The first biologic for rheumatoid arthritis: factors influencing the therapeutic decision]. Z Rheumatol Aug 12. [Epub ahead of print] German. Meissner Y, Zink A, Kekow J, Rockwitz K, Liebhaber A, Zinke S, Gerhold K, Richter A, Listing J, Strangfeld A. Impact of disease activity and treatment of comorbidities on the risk of myocardial infarction in rheumatoid arthritis. Arthritis Res Ther 2016 Aug 5; 18(1):183. doi: /s z. Listing J, Kekow J, Manger B, Burmester G-R, Pattloch D, Zink A, Strangfeld A. Mortality in rheumatoid arthritis: the impact of disease activity, treatment with glucocorticoids, TNFα inhibitors and rituximab. Ann Rheum Dis 2015 Feb; 74(2):

79 Epidemiology of Rheumatic Diseases Zink, Health Services Research in Adult Improving health care provision by continuous outcome monitoring Zink, DRFZ group Prof. Dr. rer. pol. Angela Zink Health Services Research in Adult Rheumatology 78 Keywords Health services research, Trends in treatment, Long-term observation, Cost of illness, Predictors of outcome Group Member Group leader Prof. Dr. rer. pol. Angela Zink Deputy Group leader: Dr. Katinka Albrecht Scientists Dr. Johanna Callhoff, Dr. Dörte Huscher Imke Redeker Medical documentalists Katja Thiele, Sascha Bischoff, Gregor Förster Technician Sabine Achtelstetter Students Mariya Chubrieva Our research focuses on the systematic and longterm evaluation of the quality and outcomes of health care for patients with rheumatic and musculoskeletal diseases in Germany. We investigate changes in health care provision, in clinical and patient-reported outcomes and deficits in health care within rheumatology as well as on the population level. We regularly give feedback to the treating rheumatologists and inform decision makers on the current state and gaps in health care. Our data are used to plan future health care provision and to improve the quality of care. The National Database of the German Collaborative Arthritis Centres which is now in its 25th year, with 17,000 patients observed annually, is a tool not only used for our research but also for the work of several stakeholders such as rheumatologists, patient representatives, politicians and health authorities. It describes changes in health care, patient profiles and outcomes as well as their economic consequences. Over the years, the data show continuous improvements in clinical and patient-reported outcomes, increasing costs of treatment and decreasing societal costs due to work disability, and the importance of social and psychological factors. One of our most recent observations is a continuously increasing age at onset as well as increasing current age in all patient groups, reflecting longer life expectancy as well as changes in risk factors. The group is involved in two nation-wide research networks funded by the Federal Minister of Health. In the METARTHROS network (Metabolic impact on joint and bone disease) we investigate the impact of obesity, diabetes and metabolic syndrome on juvenile idiopathic arthritis and rheumatoid arthritis, using all cohorts available in the Unit, supporting the idea of obesity as contributor to the development of inflammatory arthritis. In the PROCLAIR network (Combining Patient-Reported Outcomes with CLAims data) we found significant regional differences in health care, a strong influence of social status on disease burden and undersupply of specialized care in particular in old, disabled persons with rheumatoid arthritis. Our next studies in PROCLAIR will focus on spondyloarthritides and osteoarthritis. Data from the early inception cohort Course And Prognosis of Early Arthritis (CAPEA) are currently used for the collaborative investigation of poor prognostic markers in rheumatoid arthritis and for the investigation of biomarkers in early RA in cooperation with the universities of Duesseldorf and Granada.

80 Zink, Health Services Research in Adult Epidemiology of Rheumatic Diseases 79 Cooperation Partners Selected Publications T Alexander, Charité Universitätsmedizin Berlin; R Alten, Schlossparkklinik Berlin; M Aringer, Universitätsklinik Dresden; H Burckhardt, Universitätsklinikum Frankfurt; F Buttgereit, Charité Universitätsmedizin Berlin; R de la Camp, Erlangen; T Dörner Charité Universitätsmedizin Berlin; E Edelmann, Rheumapraxis Bad Aibling, T Eidner, Rheumazentrum Jena; K Fischer, Greifswald; E Gromnica-Ihle, Berlin; KP Günther, Universitätsklinikum Dresden; J Henes, Universitätsklinikum Tübingen; U von Hinüber und W Demary, Hildesheim; G Hoese, Stadthagen; F Hoffmann, Universität Oldenburg; A Krause, Immanuel-Krankenhaus Berlin; J Martin, Granada/Spain, W Ochs, Bayreuth; G Riemekasten, Charité Universitätsmedizin Berlin; J Richter und M Schneider, Universitäts-klinikum Düsseldorf; G Schett, Universitätsklinikum Erlangen; A Seifert, Berlin; S Späthling-Mestekemper, München; S Wassenberg und R Weier, Ratingen; Javier Martin, Granada, Spain Albrecht K, Richter A, Callhoff J, Huscher D, Schett G, Strangfeld A, Zink A. Body mass index distribution in rheumatoid arthritis: a collaborative analysis from three large German rheumatoid arthritis databases. Arthritis Res Ther Jun 23;18:149. Albrecht K, Callhoff J, Westhoff G, Dietrich T, Dörner T, Zink A. The Prevalence of Dental Implants and Related Factors in Patients with Sjögren Syndrome: Results from a Cohort Study. J Rheumatol Jul;43(7): Albrecht K, Callhoff J, Buttgereit F, Straub RH, Westhoff G, Zink A. The association between the use of oral contraceptives and patient-reported outcomes in an early arthritis cohort. Arthritis Care Res (Hoboken) Mar;68(3): Huscher D, Mittendorf T, von Hinüber U, Kötter I, Hoese G, Pfäfflin A, Bischoff S, Zink A; German Collaborative Arthritis Centres. Evolution of cost structures in rheumatoid arthritis over the past decade. Ann Rheum Dis Apr;74(4): Huscher D, Thiele K, Rudwaleit M, Albrecht KC, Bischoff S, Krause A, Karberg K, Wassenberg S, Zink A. Trends in treatment and outcomes of ankylosing spondylitis in outpatient rheumatological care in Germany between 2000 and RMD Open May 25;1(1):e K Karberg, Berlin; S Kleinert, Erlangen; I Kötter, Asklepios Klinik Hamburg Altona;

81 80

82 Core Facilities and Technical Units Central Laboratory for Microscopy...82 Central Laboratory Flow Cytometry & Cell Sorting (FCCF) Immune Monitoring Mass cytometry Regine von Ramin-Laboratory for Molecular Rheumatology Animal Facility...94 Embryo Technology Laboratory...95

83 Core Facilities and Technical Units Central Laboratory for Microscopy Cinima - Core facility for Innovative Imaging and Microscopy Approaches 82 Prof. Dr. Anja E. Hauser Dr. Raluca Niesner Group Member Group leader Prof. Dr. Anja E. Hauser Dr. Raluca Niesner Bioinformatics Ralf Köhler Cooperation Partners Max Delbrück Centrum for Molecular Medicine, Berlin, Germany MPI for Biophysical Chemistry (Bionanophotonic group), Göttingen, Germany Due to the successful combination of intrinsic molecular specificity with high spatial resolution, fluorescence microscopy is a versatile technique largely employed in both routine and high-end research applications of biosciences and biomedicine. Currently, these applications reach from the investigation of fixed biopsies, living cell cultures, vital organ models or even living organisms on a molecular basis and down to electron-microscope-like spatial resolution, i.e. supra-resolution techniques (like STED microscopy, PALM, STORM or SSIM/SPEM). The expertise at the Core facility for inovative imaging und analysis (CINIMA) at the DRFZ covers standard fluorescence techniques like wide-field and confocal fluorescence microscopy to the newest developments in vital organ explants and intravital multi-photon microscopy. Confocal laser-scanning microscopy and wide-field fluorescence microscopy For fast screening of histology samples of different organs, the DRFZ research groups employ the fully automated wide-field fluorescence microscope (Keyence). In order to achieve three-dimensional images at high resolution the Zeiss LSM710 confocal microscope is used. Both whole field images reaching to a few mm² (when tile scans are made) and images depicting cellular interaction/co-localization at sub-cellular level using the 63x oil-immersion lens (NA = 1.3, i.e. resolution at 488 nm excitation wavelength of 229 nm laterally and 577 nm axially) can be acquired. A central feature of the confocal microscope at the DRFZ is the possibility of live cell visualization and monitoring in a temperature- and CO2-regulated incubation chamber. With this system, spectral resolution of up up to five colors can be acquired. In order to achieve multiplexing in histology, we a Toponome imaging cycler is used. This system allows to detect more than 100 markers in one histological section, by sequentially staining and bleaching the sample, a process termed Multieptiope Ligand Cartography (MELC). We are developing customized algorithms to analyze the data generated using this system, in the sense of multiplexed histocytometry. Multi-photon laser-scanning microscopy for deep-tissue and intravital microscopy While the standard wide-field and confocal microscopy based on single-photon excitation intrinsically allows for high resolution due to UV/visible illumination, they do not allow for high imaging depths in intact, living organs. Due to the benefits of ultra-short pulsed near infrared (NIR) and infrared (IR) illumination, multiphoton microscopy counteracts exactly this shortcoming of standard fluorescence microscopy. The intrinsic 3D resolution and the excitation within the tissue optical window, i.e. at wavelengths where neither water nor hemoglobin absorb radiation, makes multiphoton microscopy an ideal tool for the visualization of cellular motility and communication deep in vital organ models or in organs within living organisms at low endogenous fluorescence and sub-cellular resolution. The DRFZ and its partners at the Charité already have a wide expertise concerning multi-photon imaging within explanted organs, e.g. hippocampal brain slices (Dr. Jan Leo Rinnenthal, Dr. Helena Radbruch, Prof. Frank Heppner, Institute for Neuropathology, Charité) and spleen slices (Prof. A.E. Hauser, Dr. R. Niesner), and even more, within living organisms in intestine

84 Central Laboratory for Microscopy Core Facilities and Technical Units 83 (group of Prof. A.E. Hauser), lymph node (group of Prof. A.E. Hauser), bone-marrow (group of Prof. A.E. Hauser) and organs of the central nervous system, i.e. brain stem (Dr. Helena Radbruch), brain cortex (Dr. Jan Leo Rinnenthal, Prof. Frank Heppner, Simon Bayerl, Prof. Vajcozy) and neuronal retina (Dr. R. Niesner, Prof. Paul). Currently, we focus our attention in extending this expertise to intravital spleen and kidney imaging, to longitudinal bone marrow endoscopy and to the use of optical coherence tomography in the mouse eye. While extending the expertise in the field of deep-tissue and intravital multiphoton imaging with applications especially in immunology plays a central part in the work at the DRFZ imaging core-facility CINIMA, tight collaborations with other imaging facilities in Berlin as well as with systems biology groups ensure the access of DRFZ scientists to a broad palette of fluorescence imaging techniques and data evaluation. In 2011 JIMI (Joint network for Intravital MIcroscopy) was founded in order to bundle the expertise of Multi-Photon intravital imaging present at the DRFZ, Berlin (Dr. R. Niesner und Prof. Dr. A. Hauser) with the expert knowledge of microscopy at the MDC (Dr. A. Sporbert) and image analysis present at the HKI in Jena (Prof. M.T. Figge). The goal of JIMI is to make intravital microscopy accessible for researchers from different fields in life science. We provide help in planning and performing intravital microscopy experiments as well as in the analysis of the data. This is completed by consulting and assistance through regular seminars of the JIMI members and of invited experts of the field of optical imaging in biosciences and biomedicine. Between 2012 and 2016, JIMI has been granted funding by the DFG. In this time period, JIMI supported 38 projects out of 8 national and international institutions and published 27 scientific articles (www. jimi-network.de). Furthermore, JIMI is majorly involved in leading national and international imaging networks like CTLS (headed by Prof. Spencer Shorte, Imagopole, Institute Pasteur, Paris) or German BioImaging (led by Prof. Elisa May, University of Konstanz) and closely cooperates with leading microscopy core-faclities as well as microscopy companies in Germany, France and UK. To further increase the access of the JIMI users to information and to enhance scientific exchange, we organized yearly work-shops dedicated to live and intravital microscopy: the Imaging cytometry meetings at the annual meeting of the DGfZ (German Society for Cytometry), in October 2015 and October 2016, both in Berlin. Technical equipment 1 Lavision TrimScope1 (equipped with Ti:sa Laser) 1 Lavision TrimScope 2 (equipped with Ti:sa Laser and optoparametric oscillator. Detection with PMTs or time-correlated single photon counting device) 1 Double header Lavision TrimScope 2 (equipped with Ti:sa Laser and optoparametric oscillator. Detection with PMTs or time-correlated single photon counting device). 1 Zeiss LSM710 confocal microscope 1 Keyence Biorevo fluorescent microscope 1 Zeiss Axiovert fluorescent microscope 1 Leica fluorescent microscope for multiplex imaging 1 Amnis Imaging flow cytometer

85 Core Facilities and Technical Units Central Laboratory Service facility for experimental research Central Laboratory Heidi Schliemann Tuula Geske Heidi Hecker-Kia Anette Peddinghaus Dörner, 84 Group Members Dr. rer. nat. Heidi Hecker-Kia Tuula Geske, Dipl.-Biol. Heidi Schliemann, Dipl.-Chem. Anette Peddinghaus Cooperation partners Max Planck Institut für Infektionsbiologie Charité Universitätsmedizin Berlin, Campus Mitte und Campus Benjamin Franklin The central laboratory facility of the DRFZ was founded in the year The lab managers provide services for all research groups, the main focus being on the supply with antibodies and their conjugates. Apart from this central task, the lab managers are also responsible for infrastructural aspects concerning the DRFZ, such as: the estimation and acquisition of financial contributions of the liaison groups to the DRFZ infrastructure, the ordering system regarding the supply with cell culture and general materials, management of general scientific equipment, including acquisition, calculation and service, in collaboration with the administration of the DRFZ and the supply of groups with basic materials, chemicals and chemical solutions for working with cell cultures. Antibodies Specific antibodies and their fluorochrome derivatives are of central importance for doing experimental cell-biological research. Therefore, the team of lab managers working in the central laboratory particularly focuses on the production and supply of more than substances. More than 250 antibody-producing hybridomas are cultivated. Specific antibodies are isolated from cell culture supernatants, purified and, if necessary, conjugated with various fluorochromes. Thus, the lab managers provide the basic supply with all relevant biological tools necessary for doing FACS analysis, cell sorting, histology, ELISA, and other immunological techniques. (Fig.1) The huge variety of antibodies and conjugates is accessible to scientists working at the DRFZ, the Max Planck Institute for Infection Biology, the Charite- Universitätsmedizin Berlin, and in other collaborating groups. The intranet folder OnlineAntibodiesManagement offers information on the current quantity of samples available in the stock. Users are also provided with background information such as, for instance, the name of clone, concentration, and titer, or with other data regarding the samples. Service On request, the lab managers also offer their advice and assistance in case of technical problems, for instance concerning the isolation and purification of proteins, derivatisation, fusion or fragmentation of antibodies and other proteins, production of specific affinity matrices for chromatographic purposes and for the establishment of ELISAs. All preparations go hand in hand with specific quality controls. (Fig.2+3) The field of activity of the central laboratory also includes the introduction and establishment of new methods and techniques, in particular, regarding new fluorochromatic markers. Perspectives The acquisition of further hybridomas for new scientific questions is essential.the fight against mycoplasma or endotoxin contamination has to be adapted continuously to state-of-the- art technology. The increasing use of multicolor techniques in FACS requires a continuous increase in the number of available fluorochromes, to provide enough substances with sufficient brightness, photo stability and narrow emission spectra in different combinations. Beyond that the introduction of the CyTOF technology in the

86 Central Laboratory Core Facilities and Technical Units Hybridoma Microscopy Mycoplasma testing Pos. Clone from Conc (mg/ml) lay on 1 2.4G2 1/6/2016 3,31 1,51µl BOLT MiniGel Novex (Life) ph3-10 Antibody Subcloning Fab F(ab) 2 Conjugation with fluorochromes, or other hapten, peptides, proteins, etc. ELISA SDS PAGE IEF Mycoplasma testing Endotoxin testing HPLC SDS PAGE FPLC Titration 2 2.4G2 1/8/2016 5,95 0,84µl 3 R4.6A2 1/6/2016 4,49 1,12µl 4 R4.6A2 1/5/2016 3,81 1,32µl 5 R4.6A2 1/5/2016 1,25 4µl date: Marker 5µl 7 4G11 1/5/2016 1,36 3,68µl run by: HS 8 MP6-XT22 1/6/2016 5,48 0,92µl Fig 1: General overview of antibody production and quality control 9 MP6-XT22 1/6/2016 1,36 3,68µl 10 XMG1.2 1/6/2016 3,7 1,35µl 11 XMG1.2 1/6/2016 1,91 2,62µl DRFZ implies new fields of responsibility, e.g. the establishment of antibody conjugation with heavy metals. The growing number of groups and scientists working at the DRFZ makes the adaption of infrastructural aspects of the central laboratory necessary and represents another great challenge for the lab management. The main aim is to guarantee the availability of necessary equipment and facilities at all times. Fig 2: Quality control of antibodies: isoelectric focusing Fig 3: Titration protocol: amb220 (RA3.6B2) - Alexa 594

87 Core Facilities and Technical Units Flow Cytometry & Cell Sorting (FCCF) State-of-the-art cell analysis and cell sorting through technical innovation Flow Cytometry & Cell Sorting (FCCF) 86 Group Member Scientific Head of the FCCF Dr. rer. nat. Hyun-Dong Chang Technical Head of the FCCF Dipl. Ing. Toralf Kaiser Service operator and advisor for biological issues Jenny Kirsch Bioinformatics Dr. Kristen Feher Technical Assistence Ana-Catalina Teichmüller patent Kaiser T, Kirsch J, Grützkau A, WO/2012/048906: Principle component analysis (PCA) - based analysis of discontinuous emission spectra in multichromatic flow cytometry Kaiser T, Kolbe T, Kneissl M, US B2: Flow cytometer disinfection module The Flow Cytometry Core Facility (FCCF) provides a comprehensive range of services and proprietary technological innovations to ensure state-of-the-art analysis and sorting of cells. The Central Laboratory, operated by the DRFZ, was established in 2000 and is equipped with a broad spectrum of machines. It is used by more than 300 scientists and clinicians from the DRFZ, the Charité, the Max Planck Institute for Infection Biology and other research facilities. Flow cytometry is an analytical method used for the quantitative measurement of physical, biochemical/cell biological and immunogenetic parameters of single cells by optoelectronical means. On the basis of these parameters various cell features, such as viability, quantification of antigens, phenotype, function, and cell cycle can be determined. In a flow cytometer, up to cells per second can be analyzed for more than 15 parameters simultaneously and quantitatively using light scattering and combinations of antibodies labelled with different fluorescent dyes. The high sensitivity of the cell analyzer allows the detection of as few as 50 molecules per cell. This technology allow us to monitor patients with rheumatic diseases before and during therapy on a cellular level, aiding us not only in the identification of cellular signature of different rheumatic diseases, prediction of therapy response, and monitoring of therapy response, but may also help to understand the cellular processes underlying rheumatic diseases. By cell sorting cells of interest can be isolated from complex mixtures and collected for further biochemical or functional analyses. For this, cells Technical equipment 4 Analyser MACSQuant (3-lasers) 1 Analyser FACS Canto (3-lasers) 1 Analyser Symphony (5-lasers) 1 Analyser LSR Fortessa (4-lasers) 1 Analyser Attune NxT (4 Laser) 2 Cell Sorter FACS Aria (3-lasers) 1 Cell Sorter FACS Aria II (4-lasers) 1 Cell Sorter Influx (5-lasers) are measured (as in the cell analyzer) and subsequently packaged into individual droplets. The droplets are then given an electrical charge according to their staining pattern and then deflected by charged electrodes into waiting sample tubes. In combination with suitable preenrichment methods, such as magnetic cell sorting, also rare cells, such as antigen-specific T cells can be analysed and isolated with high purity for subsequent molecular analyses. For the isolation of sensitive cells, such as plasma cells or stroma cells from the bone marrow, the FCCF utilizes a BD Influx sorter with adapted high-speed sorting protocols that ensure high viability and purity of the cells. To improve the sensitivity of multiparameter flow cytometry we are, in cooperation with the company APE, working on the hardware development for a new multispectral Flow Cytometer. A spectral flow cytometer detects the entire spectra of an individual fluorescence labelled cell, instead of only detecting a specific peak of a certain fluorochrome which has passed

88 Flow Cytometry & Cell Sorting (FCCF) Core Facilities and Technical Units Hyun-Dong Chang, Florian Oswald, Toralf Kaiser, Kristen Feher, Jenny Kirsch 87 through an optical band-pass filters, and thus collects more light increasing the detection sensitivity (Feher et al., 2016). In cooperation with APE we have also developed a LED based calibration tool (quantiflash ) which enables us to perform quality control of our machines with respect to the dynamic detection range as well as the signal-to-noise ratio (Giesecke et.al., 2017). It also allows us to calibrate the measured light intensities of a flow cytometer to absolute values. By this we are now for the first time able to quantify absolute numbers of proteins expressed by a cell. To train our users, we offer a two hour basic course on the proper use of the techniques used in flow cytometry every month. The course is open for all who are interested. Together with various groups working at the DRFZ, the FCCF offered workshops for users from around the world. Every year the FCCF opens its doors for the Long Night of Sciences for making cell sorting and cellular immunology more perceptible for public visitors, such as rheumatic patients, students or pupils. Our service offers for scientists are accessible via internet. After the registration, the users get access to the online-scheduler located on: There, the use of equipment, the documentation of experiments, and billing can be managed. Users also have access to the central data server in order to view their flow cytometry data. Cooperation partners Max Planck Institute for Infection Biology, Berlin Charité - Universitätsmedizin Berlin (Campus Mitte, Campus Steglitz, Campus Virchow) Technische Universität Berlin Angewandte Physik und Elektronik GmbH (A P E), Berlin European development fond EFRE Selected Publications C. Giesecke, K. Feher, K.v. Volkmann, J. Kirsch, A. Radbruch, T. Kaiser, Determination of background, signal-to-noise and dynamic range of a flow cytometer - a novel practical method for instrument characterization and standardization, (under review) K. Feher, K. von Volkmann, J. Kirsch, A. Radbruch, J. Popien, and T. Kaiser, Multispectral flow cytometry: The consequences of increased light collection, Cytometry A, Jun Comparison study of various flow cytometers using a novel ultra stable calibration light source. von Volkmann, K., Feher, K., Popien, J., Kaiser, T. Poster at DGfZ meeting, Berlin, Classification of Flow Cytometry Samples with a Dimension Reduction and Binning Approach. Feher, K., Radbruch, A., Kaiser, T. Poster at Cyto conference, Glasgow, UK, 2015 Modification of BD FACS Calibur for multispectral flow cytometry (photograph:t.kaiser)

89 Core Facilities and Technical Units Immune Monitoring Immune Monitoring Dr. rer. nat. Andreas Grützkau 88 Keywords Biomarker Personalized medicine Type I interferon signatures Mass cytometry Immunophenotyping Group Member Gruppenleitung Dr. rer. nat. Andreas Grützkau Scientists Dr. rer. nat. Sabine Baumgart Dr. rer. nat. Axel Schulz Technische Assistenz Heike Hirseland Silke Stanislawiak Anette Peddinghaus PhD and MD students Ursula Schulte-Wrede Patrizia Batel Hannah Brandt Marie Urbicht cooperation Partners Dr. med. Thomas Häupl Till Sörensen Bruno Stuhlmöller Dr. rer. nat. Biljana Smiljanovic Dr. med. Robert Biesen Dr. med. Thomas Rose Dr. Ute Syrbe Prof. Falk Hiepe Prof. Jochen Sieper Prof. Dennis Poddubnyy Prof. Stefan Schreiber Prof. Andreas Thiel Dr. med. Phillip Enghard Dr. rer. nat Sven Olek (Epiontis) Dr. rer. nat. Udo Baron (Epiontis) Dr. med. Guido Heine Prof. Margitta Worm Immune monitoring includes the combination of various procedures of diagnosis, which provide information about the immune state of a patient. Depending on the clinical discipline, humoral factors, like cytokines, antibody titres or complement factors, the cellular composition of peripheral blood, functional-cellular parameters or rather a combination of them all, can be determined. Immune monitoring programmes already assist in the diagnosis and treatment of many complex clinical manifestations. For example, physicians use immune monitoring in transplantation medicine, in cases of sepsis, in the course of immune modulatory therapies, and in vaccine strategies to evaluate specific immune responses. The immune monitoring core facility aims to identify immunophenotypic signatures as biomarkers for monitoring disease activity and prediction of treatment responses. Siglec-1 a new biomarker in lupus diagnostics Siglec-1 is an adhesion molecule specifically expressed in peripheral monocytes that was originally identified by cell-specific transcriptomic studies as a type I interferon surrogate marker. Interestingly, in cooperation with our clinical liaison partners it could be shown that an increased expression of Siglec-1 correlates with disease activity in patients suffering in SLE and Sjögren's syndrome (Biesen et al., 2008; Rose et al., 2016). Based on these promising results, Siglec-1 has been extensively investigated as a new biomarker in recent years and has been validated in various clinical studies (Rose et al. 2013; Rose et al. 2016; Rose et al. 2017), so that it could be shown to have a higher diagnostic potential than other gold standards in lupus diagnostics, e.g. autoantibody titres or the consumption of complement factors. Meanwhile, this biomarker has been successfully translated from bench to routine diagnostics and the quantitative measurement of this parameter is offered in the portfolio of the diagnostic laboratory of the Charité ("Labor Berlin"). Therefore, this is a paradigm of how modern high-throughput technologies can be used to identify promising, new molecular biomarkers, which will be successfully validated in welldesigned clinical trials. This is all the more remarkable because similar to the development of drugs, only very few biomarker candidates overcome hurdles of further validation studies. In the context of Siglec-1 as type I interferon surrogate marker, it was also investigated in detail why up to now no other blood-based biomarkers, such as the interferon signature, i.e. a specific pattern of genes regulated by IFN-α/β in blood cells, has been reached diagnostic routine in rheumatology. Here, we could show by a comprehensive study comparing own gene expression data and those published by other groups, that interferon signatures are strongly influenced by the cellular composition of white blood cells. Each type of immune cell type responds with another set of interferonassociated genes and it could be concluded that only interferon signatures generated in a cellspecific manner are useful for diagnostic purposes (Strauss et al., 2017). NK cells as predictors for a successful anti-tnf-a therapy response Unfortunately, at this stage it is not possible to predict whether a patient will respond to a specific therapy or not before starting of therapy. However, this is exactly the goal of the so-called personalized or individualized medicine aiming

90 Immune Monitoring Core Facilities and Technical Units Sabine Baumgart, Heike Hirseland, Andreas Grützkau, Marie Urbicht 89 to match the best possible therapy to the individual patient. This minimizes the risk of adverse side effects and can save enormous health care costs, since on average only 50%-60% of rheumatoid patients treated with expensive biologics benefit from a noticeable improvement in their disease symptoms. In collaboration with Prof. Jochen Sieper, Prof. Dennis Poddubnyy and Dr. Uta Syrbe at the Charité campus Benjamin Franklin, we conducted an extensive immune monitoring study on ankylosing spondylitis patients treated with TNF-α blocking drugs. It could be shown that in blood of these patients, the frequency of certain natural killer cell subpopulations allows a prediction of the treatment success with TNF-α blockers even before starting therapy. These results have just been submitted for publication in "Scientific Reports". Bioinformatics Multidimensional data sets generated especially by mass cytometry requires new algorithms which automatically recognize and quantify relevant cell populations. For this purpose, a software ("immunoclust", Sörensen et al., 2015) has been developed by bioinformaticians and mathematicians of the DRFZ and the department for medical bioinformatics of the rheumatology at the Campus Charité Mitte. Perspectives The complex immune monitoring approach described is used to accompany therapy studies to identify predictors of therapy responses. Furthermore, attempts are currently being made to develop algorithms which allow automation of the primary data analysis. Ultimately, it is necessary to look for ways to adjust and optimize immune monitoring of larger sample quantities with regard to a highthroughput method. Selected Publications Strauß R, Rose T, Flint SM, Klotsche J, Häupl T, Peck-Radosavljevic M, Yoshida T, Kyogoku C, Flechsig A, Becker AM, Dao KH, Radbruch A, Burmester GR, Lyons PA, Davis LS, Hiepe F, Grützkau A, Biesen R. Type I interferon as a biomarker in autoimmunity and viral infection: a leukocyte subset-specific analysis unveils hidden diagnostic options. J Mol Med (Berl) Mar 29. Rose T, Grützkau A, Klotsche J, Enghard P, Flechsig A, Keller J, Riemekasten G, Radbruch A, Burmester G-R, Dörner T, Hiepe F and Biesen R. Are interferon-related biomarkers advantageous for monitoring disease activity in SLE? A longitudinal benchmark study. Rheumatol (in press) Rose T, Szelinski F, Lisney A, Reiter K, Fleischer SJ, Burmester GR, Radbruch A, Hiepe F, Grützkau A, Biesen R, Dörner T. SIGLEC1 is a biomarker of disease activity and indicates extraglandular manifestation in primary Sjögren's syndrome. RMD Open Dec 30;2(2):e Sörensen T, Baumgart S, Durek P, Grützkau A, Häupl T. immunoclust--an automated analysis pipeline for the identification of immunophenotypic signatures in high-dimensional cytometric datasets. Cytometry A Jul;87(7): Rsk2 controls synovial fibroblast hyperplasia and the course of arthritis.derer A, Böhm C, Grötsch B, Grün JR, Grützkau A, Stock M, Böhm S, Sehnert B, Gaipl U, Schett G, Hueber AJ, David JP. Ann Rheum Dis Feb;75(2): doi: /annrheumdis PMID: [PubMed - indexed for MEDLINE]

91 Core Facilities and Technical Units Mass Cytometry Cellular diversity and biomarkers in rheumatic diseases Dr. rer. nat. Henrik Mei Mass Cytometry 90 Keywords High dimensional single-cell profiling Deep immune phenotyping Cytomics Biosensors, biomarkers and biosignatures Cellular diversity Group Member Group leader Dr. rer. nat. Henrik Mei Dr. rer. nat. Andreas Grützkau Scientists Dr. Sabine Baumgart Dr. Axel Ronald Schulz Bachelor/Diploma/ Master students Sarah Gillert Christina Schäfer Technician Silke Stanislawiak Mass cytometry permits deep cellular phenotyping in clinical immunology and basic research, with more than 50 parameters that can be measured simultaneously in a single assay. This is achieved by combining principles of flow cytometry and inductively coupled plasma mass spectrometry (ICP-MS), a technology that has been developed for trace metal analysis. Using mass cytometry, we perform biomarker discovery studies in several cohorts of patients with different chronic inflammatory diseases such as rheumatoid arthritis, SLE, and inflammatory bowel diseases. We follow the idea that the blood cytome contains important yet covered information about the patient s condition and response to future therapy. We pursue cytomics to achieve precision medicine in the treatment of chronic inflammation and aim at developing a diagnostic that can be used to select only effective treatments - individually for each patient. In basic research, we use mass cytometry to capture the diversity of different cell types implicated in chronic inflammatory processes such as T and B lymphocytes, plasma cells and myeloid immune cells. Such diversity relates to e.g. states of cell differentiation and activation, migration and adhesion, apoptosis and survival. Understanding the complexity of cellular systems and provides new insight into the homeostasis and dynamics of immunity, which are of particular interest in cases of specialized lymphocytes bearing protective vs. autoreactive immune memory, and of immune cells that fuel acute inflammation such as monocytes. To serve these aims at maximum quality, we strive for improving mass cytometry endeavours by developing and improving cell sample barcoding, expanding the measurement capacity by generating new probes and by developing and validating accessory techniques to minimize experimental variations such as cell preservation and antibody cocktail lyophilisation. Implementing these features helps to improve the accuracy of measurements and the interpretation of highly complex multi-dimensional datasets by downstream computational analyses. The DRFZ mass cytometry unit serves a variety of internal and external collaborations and is an active part of the worldwide mass cytometry community. The DRFZ spearheads the German Mass Cytometry Network that will be established in 2017.

92 Mass Cytometry Core Facilities and Technical Units 91 Cooperation Partners Selected Publications Prof. Holden Maecker, HIMC/ Microbiology & Immunology, Stanford, USA Dr. Norbert Jakubowski, Dr. Jochen Vogl, BAM, Berlin, Deutschland Prof. Margitta Worm and PD Dr. Guido Heine, Charité Berlin Prof. Andreas Krause, Dr. Julia Patermann, Immanual Hospital Berlin-Wannsee PD Dr. Philipp Engelhard, PD Dr. Adrian Schreiber, Charité Berlin Prof. Susann Müller, Leipzig, Helmholtz- Zentrum für Umweltforschung Dr. Uta Kossatz, Univ. Tübingen, Deutschland Prof. Stefan Schreiber, Univ. Klinik Schleswig- Holstein, Kiel, Deutschland PD Dr. Thomas Häupl, Dr. Till Sörensen, Dr. Melanie Conrad, PD Dr. Adrian Schreiber, Dr. Angela Ariza de Schellenberger, Charité Berlin Dr. Desiree Kunkel, BCRT, Berlin, Deutschland Dr. Christian Dose, Dr. Susanne Krauthäuser, Miltenyi Biotec, Bergisch-Gladbach, Deutschland Dr. Larissa Müller, Thermo Fisher, Breme Ben Pacheco, PhD, Cytodiagnostics, Burlington, ON, Canada Fluidigm Sciences, South San Francisco, CA, USA Spherotech Inc., Lake Forest, IL, USA Baumgart, S., Peddinghaus, A., Schulte- Wrede, U., Mei, H. E. & Grutzkau, A. OMIP-034: Comprehensive immune phenotyping of human peripheral leukocytes by mass cytometry for monitoring immunomodulatory therapies. Cytometry A 91, 34-38, doi: / cyto.a (2017). Schulz, A. R., Stanislawiak, S., Baumgart, S., Grutzkau, A. & Mei, H. E. Silver nanoparticles for the detection of cell surface antigens in mass cytometry. Cytometry A 91, 25-33, doi: /cyto.a (2017). Mei, H. E., Leipold, M. D. & Maecker, H. T. Platinum-conjugated antibodies for application in mass cytometry. Cytometry A, doi: /cyto.a (2015). Mei, H. E., Leipold, M. D., Schulz, A. R., Chester, C. & Maecker, H. T. Barcoding of live human peripheral blood mononuclear cells for multiplexed mass cytometry. J Immunol 194, , doi: / jimmunol (2015). CD25 expression Sorensen, T., Baumgart, S., Durek, P., Grutzkau, A. & Haupl, T. immunoclust--an automated analysis pipeline for the identification of immunophenotypic signatures in high-dimensional cytometric datasets. Cytometry A 87, , doi: /cyto.a (2015). RA patient before TSO therapy RA patient after 13 weeks of TSO therapy Blood immune cells of a rheumatoid arthritis (RA) patient analyzed by 43-parameter mass cytometry before and after induction of an experimental helminth therapy (TSO). Data were analyzed by automated SPADE clustering (Qiu et al., Nat. Biotech. 2011). Each circle represents a cell cluster. Regulatory T cells (arrow) were expanded after induction of TSO therapy.

93 Core Facilities and Technical Units Regine von Ramin-Laboratory for Molecular Rheumatology Regine von Ramin-Laboratory for Molecular Rheumatology Regine von Ramin-Laboratory for Molecular Rheumatology 92 Dr. rer. nat. Andreas Grützkau PD Dr. med. Thomas Häupl Group Member Group leader Dr. rer. nat. Andreas Grützkau PD Dr. med. Thomas Häupl Technische Assistenz Heidi Schliemann Heike Hirseland Gudrun Steinhauser Through the generous bequest from Mrs. Regine von Ramin and further financing by the Berlin senate-administration WiFoku, the Regine-von- Ramin Laboratory for Molecular Rheumatology (RvR-Laboratory) was founded in December of This facility will be used jointly by research groups of the DRFZ and the affiliated groups of the Charité for the purposes of genome-wide gene expression analysis using Affymetrix- Technology. In addition to global transcription analysis, it will also be possible to carry out microarray-based mirna expression analysis. Technical equipment belonging to the Regine von Ramin Laboratory The RvR-Laboratory contains an Affymetrixstation, which is equipped with the the Scanner G and the Fluidics FS450. For the purposes of RNA-sample quality control, a Bioanalyzer (Agilent) and Nanodrop ND-1000 spectral photometer are available in addition to standard instruments. Additionally, the ahybtm- Hybridisation station (Milenyi Biotec) allows for processing of array formats that are spotted on slides, and thereby processes like a mirna expression profiling using the mirxplore Array (Miltenyi Biotec). An ideal supplement to this facility is a Laser Capture Microdissection machine (LCM) from Fa. Arcturus, which allows a cutting out of morphologically and immune-histologically characterized tissue samples for molecular analysis without enzymatically digesting them. Both systems are primarily used for molecular analysis of cells at the level of their DNA, RNA and proteins. Through the integration of LCMtechnology in the RvR-Laboratory, it is possible to perform global gene expression analysis on tissue samples that are morphologically and immuno-histologically identifiable (Figure 1). Bioinformatic Analysis A close cooperation with the Department of Bioinformatics (Joachim Grün, DRFZ; Thomas Häupl, Charité) and the Charité spin-off company BioRetis,, allowed the establishment of a comprehensive mouse- and disease-relevant human transcriptome database. The datawarehousing of these data by the BioRetisanalysis platform allows for comfortable and precise group comparative analysis that can be implemented worldwide by existing national and international research organizations within the framework of a defined rights allocation system. An overview of the basic analysis strategies can be found in the following chapter on Bioinformatics by Joachin Grün. Generating disease-, cell- and cytokinespecific gene expression profiles One research focus at the DRFZ is the analysis of disease- and cell-specific global gene expression profiles. It has been shown that peripheral monocytes isolated from RA, SLE and Bechterew s disease-patients show disease-specific gene signatures that couldbe successfully used for disease-classification. The large number of differentially expressed genes reflects the complexity of chronic-inflammatory rheumatic diseases. The central pathophysiological role of pro-inflammatory cytokines, such as TNF-α or INF-α/γ, in the inflammatory-processes could be shown clearly with the clinical success of cytokine-specific biologicals. However, there is currently no reliable molecular biomarker that would allow for a targeted and thereby individualized therapy. Therefore, cytokine-

94 Regine von Ramin-Laboratory for Molecular Rheumatology Core Facilities and Technical Units Figure1: process of global gene expression analysis beginning with cell isolation using FACS or Laser-Microdissection to the generation and analysis of the transcriptome using microarray-technology. These methods allow for the identification of so-called gene-signatures that shed light on the cell-specific pathophysiology and can also be used for diagnostic purposes. 93 specific gene signatures have been generated and analyzed ex-vivo in two EU-supported research-projects (Autocure and IMI JU BTCure). Through a comparison of these expression profiles with the diseasespecific profile, Biljana Smiljanovic was able to show, in the course of the research conducted for her dissertation, that peripheral monocytes are highlysensitive biomarkers that express disease-dependent, qualitative and quantitative variability in their cytokine signature. While monocytes from Lupus-patients are characterized primarily by an IFN-α/γ-induced immune-response, RA- patients were characterized by a dominance of TNF-α in this process. This knowledge will be used in further investigations to clarify if these molecular cytokine-response signatures can be used to predict a positive response to therapy. Cooperation Partners Dr. rer. nat. Joachim Grün, DRFZ Dr. rer. nat. S Fillatreau, Dr. rer. nat. Chiara Romagnani, Dr. rer. nat. A Hutloff, Dr. rer. nat. K Tokoyoda, DRFZ PD Dr. med. K Pels, PD. Dr. med. E. Schott, Dr. rer. nat. C Daridon, Prof. Dr. med. T Dörner, Dr. med. S Mercan,Dr. rer. nat. K Kappert, Charité - Universitätsmedizin Berlin Prof. Dr. rer. nat. A Thiel, BCRT - Charité Prof. Dr. rer. nat. F Melchers, Max-Planck- Institut für Infektionsbiologie, Berlin PD Dr. rer. nat. U Höpken, Max Delbrück Zentrum, Berlin-Buch PD Dr. rer. med. H Eibel, Universitätsklinikum, Freiburg Prof. Dr. med. B Bröker, Immunologie, Greifswald Dr. F Apparailly, Dr. H Yssel, INSERM, Montpellier, Frankreich Prof. Dr. W Maslinski, Institute of Rheumatology, Warschau, Polen Dr. rer. nat S. Wild, Miltenyi Biotech, Bergisch- Gladbach References Thiel A.Effects of aging on human leukocytes (part II): immunophenotyping of adaptive immune B and T cell subsets. Age (Dordr) Oct;37(5):93. Stervbo U, Meier S, Mälzer JN, Baron U, Bozzetti C, Jürchott K, Nienen M, Olek S, Rachwalik D, Schulz AR, Waldner JM, Neumann A, Babel N, Grützkau A, Thiel A. Effects of aging on human leukocytes (part I): immunophenotyping of innate immune cells. Age (Dordr) Oct;37(5):92 Jürchott K, Schulz AR, Bozzetti C, Pohlmann D, Stervbo U, Warth S, Mälzer JN, Waldner J, Schweiger B, Olek S, Grützkau A, Babel N, Thiel A, Neumann AU. Highly Predictive Model for a Protective Immune Response to the A(H1N1) pdm2009 Influenza Strain after Seasonal Vaccination. PLoS One Mar 8;11(3): e Alissafi T, Hatzioannou A, Ioannou M, Sparwasser T, Grün JR, Grützkau A, Verginis P. De novo-induced self-antigen-specific Foxp3+ regulatory T cells impair the accumulation of inflammatory dendritic cells in draining lymph nodes. J Immunol Jun 15;194(12): Derer A, Böhm C, Grötsch B, Grün JR, Grützkau A, Stock M, Böhm S, Sehnert B, Gaipl U, Schett G, Hueber AJ, David JP. Rsk2 controls synovial fibroblast hyperplasia and the course of arthritis. Ann Rheum Dis Feb;75(2): Stervbo U, Bozzetti C, Baron U, Jürchott K, Meier S, Mälzer JN, Nienen M, Olek S, Rachwalik D, Schulz AR, Neumann A, Babel N, Grützkau A, Duroux-Richard I, Roubert C, Ammari M, Présumey J, Grün JR, Häupl T, Grützkau A, Lecellier CH, Boitez V, Codogno P, Escoubet J, Pers YM, Jorgensen C, Apparailly F. ir-125b controls monocyte adaptation to inflammation through mitochondrial metabolism and dynamics. Blood Dec 29;128(26):

95 Core Facilities and Technical Units Animal Facility Animal Facility Animal Facility 94 Dr. med. vet. Anja Schulz, Dr. med. vet. Elena Noe, Astrid Puppe Group Member Facility management Dr. med. vet. Anja Schulz Animal welfare officer Astrid Puppe (Elternzeit) Dr. med. vet. Elena Noe (ab 04/2015) Administration and Technical Staff Sabine Gruczek Maria Hoffmann Christina Kasprzak (ab 06/2015) Manuela Ohde Laura Prüfer (bis 05/2015) Helmut Schäfer (bis 06/2015) Paul Streckert (ab 09/2015) Vivien Theißig Patrick Thiemann Trainee Claudia Arndt (bis 09/2015) Julien Frey Although the list of methods replacing animals in experimental research is constantly growing, for some scientific questions the use of animal models is still necessary. Especially for the understanding of multifactorial diseases like autoimmune disorders, in which different organs and cell types are involved, the complexity of a living organism is required. Therefore, the DRFZ runs a state of the art animal facility. This facility is divided into an experimental area in Berlin-Mitte and a separate breeding area in Berlin-Marienfelde, where numerous mouse strains are kept. Trained animal caretakers and veterinarians ensure best husbandry conditions and a seamless monitoring of the animals in favor of animal welfare. All mice are bred and held under so called SPF (specific pathogen free) conditions to ensure that animals are free from pathogens that could interfere with animal experimental studies. For this purpose, the facility is equipped with personnel airlocks, and all material is thoroughly autoclaved before entering the facility. Moreover, mice undergo a microbiological check-up on a regular basis in order to detect potentially pathogenic microorganisms. As a part of the Leibniz-Gemeinschaft the DRFZ intends to communicate openly about animal experiments. Scientists working with mice at the DRFZ put the principles of the 3Rs (Replacement, Reduction and Refinement) into practice to minimize suffering and improve animal welfare. The DRFZ provides a training program on animal experimental work according to EU and German animal welfare legislation, consisting of a theoretical and practical part. Successful participation is officially certified. Cooperation Partners Charité - Universitätsmedizin Berlin, Campus Mitte and Campus Benjamin Franklin Max Planck Institute for Infection Biology, Berlin

96 Embryo Technology Laboratory Core Facilities and Technical Units Embryo Technology Laboratory Embryo Technology Laboratory 95 Dr. Andreas Hutloff, Paolo Rosellini Tognetti The Embryo Technology Laboratory is a joint core facility together with the Robert Koch Institute. It is located within the animal facility in Berlin-Marienfelde and provides state of the art technologies for modern mouse colony management like in vitro fertilization (IVF) and cryopreservation. The following services are offered (Fig. 1): 1) Embryotransfer. To guarantee a defined health status in the SPF breeding area, new transgenic mouse lines have to be imported via embryotransfer. Two cell embryos (generated by IVF or classical mating) are washed extensively to Fig. 1: Workflow for services offered by the ET-Lab. remove any pathogens (Fig. 2) and implanted into pseudopregnant foster mice. 2) Cryopreservation of mouse sperm and embryos. Mouse strains which are temporarily not needed can be stored as 2-cell embryos or sperm in liquid nitrogen. Frozen material is also an option for world-wide shipping of transgenic mouse lines. 3) In vitro fertilization. IVF is used to recover mouse lines from frozen sperm and is generally a very efficient method to generate mouse embryos. Scientist Scientific facility management: Dr. Andreas Hutloff Technical staff: Paolo Rosellini Tognetti Additional assistance: Janni Breinl Cooperation Partners Dr. Uwe Klemm, Experimental Animals, Max Planck Institute for Infection Biology, Berlin Dr. Geert Michel, Transgene Facility, FEM, Charité University Medicine Berlin, Germany Ronald Naumann, Transgenic Core Facility, Max Planck Institute of Molecular Cell Biology and Genetics, Dresden Frank Zimmermann, Biotechnology Laboratory, University Heidelberg Fig. 2: Washing of embryos. The fertilized oocyte is protected by the zona pellucida which is impermeable for pathogens. Residual sperm is still sticking to the zona. Fig. 3: Developmental stages of mouse embryos. a) Sperm and oocytes (hidden in the cumulus cells) 5 min after IVF. b) 45 min after IVF. The sperm almost completely dissolved the cumulus cells. c) 2-cell embryos. d) 4- and 8-cell embryos. e) Morula stage. f) Blastocysts.

97 96

98 Appendix Third party funded projects - DRFZ Groups Third party funding projects - Liaison Groups Third party funds Qualifications Sitemap Imprint...108

99 Appendix Third party funded projects - DRFZ Groups Third party funded projects - DRFZ Groups Project Title/ Acronym (sub)-project Funding Source Scientist(s) in Charge Start End IMMUNOBONE SFB 650 Cellular approaches to the suppression of unwanted immune reactions - from bench to bedside Mesenchymal stroma cells as organizers of immunological memory in the bone marrow An integrated high-resolution immunophenotyping and computational biomarker discovery approach for monitoring clinical trials Engineering resting B cells for the suppression of unwanted immunity DFG Chang, Radbruch 07/ /2016 DFG Grützkau 01/ /2016 DFG Fillatreau 01/ /2016 Induction of stable Foxp3+ regulatory T cells DFG Hamann 01/ /2016 Redirecting the antigen-specificity of polyclonal regulatory T cells to optimize their therapeutic efficacy and safety DFG Fillatreau 01/ / Targeting of pathogenic T helper cells in chronic rheumatic inflammation Exploiting NK cells for suppression of unwanted immune responses in transplantation DFG Radbruch, Chang 01/ /2016 DFG Romagnani 01/ /2016 JIMI JIMI - Joint Intravital Microscopy and Imaging Network (JIMI) DFG Niesner 08/ /2016 DFG FOR 2165 SFB 633 Induction and Modulation of T-cell- Mediated Immune Reactions in the Gastrointestinal Tract Longitudinal intravital imaging of dynamics of osteoimmunological interactions during bone healing Role of pro-inflammatory T cells in chronic mucosal inflammation Analysis of tolerance induction of interleukin-17-producing T helper cells in the small intestine Role and potential therapeutic use of interleukin-22+ Natural Killer cells in colitis Induction of specific homing receptors and of regulatory properties in mucosal T cells DFG Niesner 09/ /2017 DFG Radbruch 07/ /2015 DFG Hauser, Esplugues 07/ /2015 DFG Romagnani 07/ /2015 DFG Hamann 07/ /2015 Dynamics of memory plasma cell niches DFG Radbruch 11/ /2017 SFB TRR 130 B cells: Immunity and Autoimmunity BIS DFG SPP 1937: Innate Lymphoid Cells Deciphering the antibody-independent functions of antibodyproducing cells in autoimmune diseases Dynamic multi-photon microscopy and microendoscopy quantifying communication and function of B lymphocytes in vivo Characterization of IL-10-producing B cells and their suppressive activities during primary and secondary Salmonella typhimurium infection Molecular characterization of survival niches for memory T helper cells in bone marrow Angeborene lymphoide Zellen (ILCs) des humanen Darms: Signaturen und Polarisierungssignale Rolle angeborener lymphoider Zellen und des Arylhydrocarbonrezeptors bei pulmonalen bakteriellen Infektionen Molecular regulation of T follicular helper cell maintenance by Bach2 and RankL DFG Fillatreau 10/ /2017 DFG Niesner 10/ /2017 DFG Fillatreau 10/ /2017 DFG Tokoyoda 05/ /2019 DFG Romagnani 03/ /2019 DFG Romagnani 11/ /2019 DFG Hutloff 11/ /2019 NEU² IL SysInflame - sysinflame - Netzwerk für Systemmedizin chronischentzündlicher Erkrankungen Identification and functional analysis of new small molecules inducing anti-inflammatory IL-27 and IL-35 High dimensional immunophenotyping to study therapymodulated immune cell dynamics BMBF Fillatreau, Hamann 09/ /2017 BMBF Grützkau, Radbruch 12/ /2016

100 Third party funded projects - DRFZ Groups Appendix EPILYZE Project Title/ Acronym OA TREAT Technology Transfer NEUROIMPA - Neuroimmunologie und Schmerz DEEP Deutsches Epigenom Programm e:bio T-Sys Funding (sub)-project Source DNA Methylierungs-Signaturen als innivative Biomarker für die quantitative und qualitative Analyse von Immunzellen, Teilprojekt E Prospektive randomisierte, doppel-blinde und plazebokontrollierte klinische Studie mit Hydroxychloroquin (HCQ) bei Patienten mit einer entzündlichen Osteoarthritis (OA) der Hände Verstetigung der Verwertungskonzepte in den lebens- und umweltwissenschaftlichen Instituten der Leibniz-Gemeinschaft - Deutsches Rheuma-Forschungszentrum Berlin Scientist(s) in Charge Start End BMBF Grützkau, Radbruch 12/ /2016 BMBF Buttgereit 07/ /2018 BMBF Moser 08/ /2016 Neuromodulation in entzündungsfördernden T Zellen BMBF Chang, Radbruch 02/ /2019 Epigenetik von inflammatorischen T-Zellen BMBF Hamann 09/ /2017 Modulation of transcriptional regulation of Th cells BMBF Baumgrass 01/ /2016 MicroRNAs and genes regulating the transition of protective into pathogenic memory T helper cells in chronic inflammatory diseases BMBF Radbruch, Mashreghi 01/ / OA TREAT Klinische Stidue OA Treat BMBF Listing/Zink 10/ /2018 METARTHROS metabolic impact on joint and bone diseases ICON - Frühkohorte Auswirkungen von Diabetes mellitus und Übergewicht auf den Verlauf der rheumatoiden Arthritis Frühkohorte juvenile idiopathische Arthritis-Arbeitspaket Studienprotokoll BMBF Zink 02/ /2018 BMBF Zink, Minden 09/ /2015 PROCLAIR ZIM-RETI-IM ZIM_ibi C3ID Gesundheitsversorgung und Krankheitslast bei Personen mit rheumatoider Arthritis (RA) Retina-Bildgebungsverfahren zur Frühdiagnose neurodegenerativer und neuroimmunologischer Erkrankungen (RETI-IM); Entwicklung der Gesamtintegration eines murinen Retina-Mikroskopiesystems mit der Erarbeitung von Anwendungsszenarien Entwicklung einer RNA-Sequenzierungs-Pipeline für Dienstleistungen im Bereich der Forschung, Klinik und Industrie; Optimierung und Validierung der speziellen RNA-Aufreinigung für RNA-Seq aus histo-pathologischen Proben Leibnitz-Wissenschaftscampus "Comprehensive Center for Chronic Inflammatory Diseases" BMBF Zink, Minden 02/ /2017 BMWi Niesner 05/ /2016 BMWi Baumgrass 05/ /2017 Leibniz Association Luger, Radbruch 07/ /2020 LRA Healthy Aging Leibniz Research Alliance "Healthy Aging" Leibniz Association Radbruch, Luger 05/ /2017 Immuno Memory International Leibniz Research Cluster (ILRC) - ImmunoMemory Leibniz Association Tokoyoda 04/ /2015 LGRh Leibniz Graduate School for Rheumatology Leibniz Association Radbruch, Moser 06/ /2018 ITN EUTRAIN European Translational training for Autoimmunity & Immune manipulation Network (EUTRAIN) European Commission Radbruch 01/ /2015 ITN OSTEOIMMUNE Unraveling the interactions between the immune system and bone (OSTEOIMMUNE) European Commission Chang, Radbruch 06/ /2016 IMMEMO Protective and pathogenic immunological memory and its organisation by stroma cells (ERC Grant) European Research Council Radbruch 08/ /2016 IEF-CR NKG2D-Immuno European Commission Romagnani 05/ /2016 IMI BTCure BeTheCure (IMI Inflammation - Translational Research and Adaptive Immunity) EC, EFPIA Radbruch, Chang, Grützkau 04/ /2017 Infect-ERA ABIR Identifying DC subsets that induce protective effector and memory T cells during Listeria and Salmonella infection EC,BMBF Fillatreau 10/ /2017

101 Appendix Third party funded projects - DRFZ Groups Project Title/ Acronym (sub)-project Funding Source Scientist(s) in Charge Start End ProFit MultiSpecFlowCytometry EC,BMWi Radbruch, Kaiser 04/ /2017 Irc-p B-Cells RABBIT Investigating the role of cytokine-producing B cells in the pahtogenesis of autoimmune diseases of the central nervous system Langzeitbeobachtung der Therapie mit Biologika bei rheumatoider Arthritis Foundation Fillatreau 08/ /2015 Industry Zink/Strangfeld 04/ /2030 RABBIT-SpA Rabbit-Spondyloarthritis Register Industry Zink 07/ /2017 Kerndoc Kerndokumentation Industry Zink 11/ /2018 Biologic register JuMBO Juvenile arthritis Methotrexate/Biologics long-term Observation Industry Minden, Zink 05/ / Irc-p B-Cells Investigating the role of cytokine-producing B cells in the pahtogenesis of autoimmune diseases of the central nervous system Foundation Fillatreau 08/ /2015 Humboldt Fellowship Humboldt Research Fellowship for Postdoctoral Researchers Foundation - Alexander von Humboldt- Stiftung Shintaro 04/ /2017 NPRD National paediatric rheumatologic database Foundation - Deutsche Kinderheumastiftung Minden EINSTEIN The role of dendritic cells in pesenting tumor and autoimmune epitopes produced by proteasomal peptide splicing Foundation - Einstein Stiftung Berlin Chang, Radbruch 07/ /2016 The Indicilble T cell Co-Stimulator ICOS as target for elimination of tissue-resident follicular helper-like T cells Foundation - Fritz Thyssen Stiftung Hutloff 09/ /2017 Ideenwettbewerb "Ist Rheuma heilbar" Molekulare Adaption pathogener Gedächtnis T Helferzellen bei rheumatischen Entzündungen Foundation - Reumastiftung Chang 01/ /2018 Pitzer- Stiftungsprofessur "Arthrose" Pitzer-Stiftungsprofessur "Arthrose" Foundation - Willy Robert Pitzer Stiftung Löhning 09/ /2020 Kinderkerndokumentation Kinderkerndokumentation für Kinder und Jugendliche mit entzündlich-rheumatischen Erkrankungen Others Minden 01/ /2017 VPSS Mundgesundheitliche Probleme und Implantat-Versorgung bei Patienten mit primäem Sjörgren-Syndrom Others Zink 12/2011 Univerity Chiba Outsourcing the operation of Chiba University's Germany office Others - Chiba University Tokoyoda 04/ /2016 RITUXIMAB in RA DGRh Start-Up 2014 ZMV Transition UOS Combined cellular and serological biomarkers for the prediction of responsiveness to B cell depleting therapy with rituximab in patients with RA Functional stabilization of clinically suitable Treg populations by RNA mediated "epigenetic editing". Transition Empowering young people with rheumatic diseases for transition to adult care Uveitis Outcome Study Others - Deutsche Gesellschaft für Rheumatologie e.v. (DGRh) Others - Deutsche Gesellschaft für Rheumatologie e.v. (DGRh) Others - Deutsche Rheuma-Liga Bundesverband e.v. Others - Hamburger Elterninitiative rheumakranker Kinder e.v. Mei 01/ /2016 Polansky 01/ /2016 Minden 01/ /2017 Minden 06/ /2016 as of December, 31, 2016

102 Third party funding projects - Liaison Groups Appendix Third party funding projects - Liaison Groups Project Title/ Acronym (sub)-project Funding Source Scientist(s) in Charge Start End DFG FOR 2165 DFG SPP 1937: Innate Lymphoid Cells IMMUNOBONE Longitudinal intravital imaging of dynamics of osteoimmunological interactions during bone healing Analysing the heterogeneity of innate lymphoid cells and relationship with their microenvironments in situ and in vivo Delineation of human plasma cell subsets and their bone marrow niche DFG Hauser 10/ /2017 DFG Hauser 01/ /2019 DFG Dörner 06/2016 JIMI JIMI - Joint Intravital Microscopy and Imaging Network (JIMI) DFG Hauser 08/ SFB 633 Induction and Modulation of T-cell- Mediated Immune Reactions in the Gastrointestinal Tract Characterization of human peripheral and intestinal T cell responses after mucosal antigen exposition: Induction of tolerance vs. immunity Analysis of the presentation of orally applied antigens and the specific T-cell reaction: tolerance versus immunity DFG Dörner 06/2015 DFG Scheffold 01/ / New generation anti-b cell therapy DFG Dörner 12/2016 SFB 650 Cellular approaches to the suppression of unwanted immune reactions - from bench to bedside Induction of tolerance by immunoablation followed by autologous stem cell transplantation in refractory autoimmune diseases Targeting of pathogenic T helper cells in chronic rheumatic inflammation Redirecting the antigen-specificity of polyclonal regulatory T cells to optimize their therapeutic efficacy and safety Redirecting the antigen specificity of polyclonal regulatory T cells to optimise their therapeutic efficacy and safety DFG DFG Hiepe, Arnold, Alexander Hiepe, Radbruch, Burmester 01/ / / /2016 DFG Löhning 01/ /2016 DFG Scheffold 01/ /2016 Immune modulation of allergic diseases by nuclear hormone receptor ligands DFG Worm /2016 B cell dynamics in chronic neuroinflammation DFG Hauser 10/ /2017 SFB TRR 130 B cells: Immunity and Autoimmunity Selective plasma cell targeting DFG Hiepe 10/ /2017 Regulation of B cell differentiation by vitamin A and D DFG Worm, Heine /2017 Characteristics and regulation of human, antigen-specific B cell memory DFG Dörner 05/2018 Transkriptionelle Programmierung und Umprogrammierung entzündungsverstärkender T-Helfer-1 Zellen Nutzen des Placeboeffektes bei atopischer Dermatitis Steigerung der pharmakologischen Wirkung bei Juckreiz durch Konditionierungs- und Erwartungsprozesse Cyclooxygenasen als Modulatoren allergischer Reaktionen, mechanistische Hintergründe und therapeutisches Potential DFG Löhning 12/ /2016 DFG Worm 2013 ongoing DFG Worm 2015 ongoing Cellular and molecular mechanisms of TNF (tumor necrosis factor-alpha) mediated protection in atopic dermatitis DFG Worm 2015 ongoing Cluster of Excellence NeuroCure Immune Dynamics in the central nervous system DFG Hauser 10/ /2017

103 Appendix Third party funding projects - Liaison Groups 3DBoMo Project Title/ Acronym ArthroMark CONSUL e:bio T-Sys (sub)-project Entwicklung eines 3D-Modells zur Simulation der initialen Frakturheilungsphase in vitro - Effektive Reduzierung von Tierversuchen in der Frakturheilungsforschung In vitro und in silico Modellierung der Immunpathogenese von Arthritiden zur effektiven Reduzierung der Zahl von Versuchstieren im Bereich der Therapieforschung Biomarker und Bildgebung zur Diagnose und Stratifizierung der Rheumatoiden Arthritis und Spondyloarthritis COmparison of the effect of treatment with NSAIDs added to anti-tnf therapy versus anti-tnf therapy alone on progression of StrUctural damage in the spine over two years in patients with ankylosing spondylitis: a randomized controlled multicentre trial Interplay of epigenetic and transcriptional imprinting of T helper cell fate Funding Source Scientist(s) in Charge Start End BMBF Buttgereit 04/ /2017 BMBF Buttgereit 04/ /2019 BMBF Poddubnyy BMBF Poddubnyy 2016 ongoing BMBF Löhning 01/ / InfectControl ART4Fun InfectControl ART4Fun BMBF Scheffold 09/2016 ongoing InfectControl Transsektorale Forschungsplattform METARTHROS metabolic impact on joint and bone diseases NEUROIMPA - Neuroimmunologie und Schmerz PROCLAIR T-Zellen als sensitive diagnostische Sensoren zur Identifizierung und Differenzierung A. fumigatus-assozierter (Immun-) Pathologien Metabolic Impact on Joint and Bone Diseases, TP 6 (Soluble markers of energy and glucose homeostasis in arthritis Intraartikuläre Applikation von Opioiden bei chronischer Arthritis Linking Patient-Reported Outcomes with CLAims data for health services research In Rheumatology BMBF Scheffold 03/2015 ongoing BMBF Poddubnyy 02/ /2018 BMBF Poddubnyy BMBF Poddubnyy GESPIC German SPondyloarthritis Inception Cohort BMBF, BIH, Industry Poddubnyy 2000 ongoing GLORIA Comparing the effectiveness and safety of additional low-dose glucocorticoid in treatment strategies for elderly patients with rheumatoid arthritis European Commission Buttgereit 09/ /2020 QuanTI, ITN Quantitative T Cell Immunology Cytokine memory in T cells European Commission Löhning 05/ /2017 RAPID Rheumatoid Arthritis and Periodontal Inflammatory Disease, Marie Curie Initial Training Network European Commission Buttgereit 11/ /2015 IMI PRECISESADS ESTHER HORIZON Project PRECISESADS Molecular reclassification to find clinically useful biomarkers for systemic autoimmune diseases Effects of etanercept versus sulfasalazine in early axial spondyloarthritis on active inflammatory lesions as detected by whole-body MRI: a randomised controlled trial Effects of glucocorticoids on the expression of Thioredoxin in human CD4+ T cells under hypoxic conditions Effects of glucocorticoids on membrane bound glucocorticoid receptor (mgcr) expression in human monocytes under hypoxic conditions EC, EFPIA Hiepe 02/ /2019 Industry Poddubnyy 2005 ongoing Industry Buttgereit 07/ /2018 Industry Buttgereit 07/ /2018 PreTheraX PreTheraX Industry Buttgereit 02/ /2018 RhGIOP Glucocorticoid-induced Osteoporosis in Patients With Chronic Inflammatory Rheumatic Diseases or Psoriasis Industry Buttgereit 10/ /2018 RLX GR Interaktion RLX GR Interaktion Industry Buttgereit 01/ /2016 Compliance und Lebensqualität (CELL) bei PSS-Patienten - Entwicklung eines strukturierten Schulungsprogramms Industry Worm 2016 ongoing Einfluss von LC-PUFA auf die Ausprägung der allergischen Immunantwort im Mausmodell Industry Worm ongoing

104 Third party funds Appendix Project Title/ Acronym ArthroMo SNF: Sinergia BIH, SPARK RefineMOMo (sub)-project Pathogen-specific T helper cells as diagnostic sensors in CF patients Etablierung und Validierung eines humanen 3D in vitro Arthrose- Modells The alarmin interleukin-33 in infection, immunity and autoimmunity Affinity matrix for the depletion of plasma cells secreting pathogenic autoantibodies Optimierung des Schmerzmanagements im Maus-Osteotomie- Modell Integration von Refinement-Untersuchungen in einer grundlagenwissenschaftlichen Studie Funding Source Foundation - Cystic Fibrosis Foundation (USA) Foundation - Wolfgang- Schulze-Stiftung Others - Schweizerischer Nationalfonds Others - Berlin Institute of Health Others - BfR Bundesinstitut für Risikobewertung Scientist(s) in Charge Start End Scheffold 07/2015 ongoing Buttgereit 10/ /2017 Löhning 01/ /2018 Hiepe 07/ /2018 Buttgereit 01/ /2017 EULAR Task Force Project on Glucocorticoids Others - EULAR Buttgereit 05/ /2017 Antigen-spezifische T-Zellen als diagnostische Sensoren von latenter vs. aktiver Tuberkulose Others - Labor Berlin Charité Vivantes Services GmbH Scheffold 04/2015 ongoing 103 as of December,31, 2016 Third party funds DRFZ Groups DFG-SFB/TR Further DFG projects Government (BMBF BMWI) EC/ FP-7/ Horizon 2020/ERC Industry Leibniz Competition Foundations Others Total Liaison Groups (Charité) DFG-SFB/TR ,4 Further DFG projects ,7 Government (BMBF BMWI) ,6 EC/ FP-7/ Horizon ,1 Industry Leibniz Competition - - Foundations ,4 Others 0,0 144,9 Total as of December,31, 2016 in T Euro as of March,22, 2017 in T Euro

105 Appendix Qualifications 104 Qualifications 2015 Bachelor theses Sema Basaran, Der Einfluss von potentiell frakturheilungsfördernden Substanzen auf die zelluläre Immunantwort im Maus-Osteotomie- Modell, TU-Berlin Elzbieta L. Gralinska, Systemic data integration for identification of crucial transcription factors for Th2 cell fate decision, TU-Berlin Ruth Leben, Asymmetrische Zwei-Photon-Anregung zur Erweiterung der simultan detektierbaren Chromophoren intravitalmikroskopie, Beuth Hochschule für Technik Berlin Franziska I. Szelinski, Assessment of SIGLEC-a (CD169) as possible biomarker for IFN-a in patients with primary Sjögren s syndrome, TU- Berlin Florian Padberg, The role of the Transcription factor Cax2 in Th17 Cell Development and Function, Technische Universität Braunschweig, Fakultät Machinenbau Roman Rauch, Einfluss von superparamagnetischen Nanopartikeln (SPIONs) auf die Differenzierung von humanen Monozyten zu Makrophagen, TU-Berlin Kerstin Schönbeck, Expression und Funktion von Zytotoxischem T- Lymphozyten Antigen-4 (CTLA-4) in humanen mesenchymalen Stromazellen (hmsc), TU-Berlin Erik Schröter, Impact of the fracture hematoma on MSC activation, Beuth Hochschule für Technik Berlin Mario Simonetti, Tolerogene Wirksamkeit PEGylierter Peptide, TU- Berlin Caroline Spee-Mayer, Role of Treg and IL-2 in the pathogenesis and treatment of SLE, HU Berlin Meike Storms, Krankheitskosten der frühen juvenilen idiopathischen Arthritis, Charité Aderajew Waka, Vergleichende Untersuchungen zur diagnostischen Relevanz von verschiedenen Immunoassay-Testsystemen zum Nachweis von Anti-dsDNA- Autoantikörpern beim Systemischen Lupus Erythematodes (SLE) des Menschen, Charité Katharina Wylon, Immunologische Wirkung einer Einmalgabe von I.E. Cholecalciferol (Vitamin D), HU Berlin PhD theses Janine Bleil, Histopathologische Charakterisierung des Entzündungsinduzierten Gelenkumbaus bei Patienten mit Ankylosierender Spondylitis, TU-Berlin Qingyu Cheng, Autoantibodies from long-lived memory plasma cells of NZB/W mice drive immune complex nephritis, Charité Master theses/diploma theses Stefanie Gryzik, Funktionelle Charakterisierung Autoimmunerkrankungen-assoziierter SNPs in nicht-kodierenden Regionen des humanen STAT4-Gens, Uni Potsdam Nicole Affinass, Generation of regulatory T cell library for detection and functional characterization of SLEassociated human antigen-specific regulatory T cells, FU Berlin Julia Bluhm, The Role of Epigenetics in Foxp1 Gene Regulation in CD4+ T Lymphocytes, Biologisch- Pharmazeutischen Fakultät der Friedrich-Schiller-Universität Jena Lisa Deichgräber, Untersuchungen zur HIF-Expression in T-Helferzellsubpopulationen, Beuth Hochschule für Technik Berlin Maxim Nosenko, Engineering of artificial lymph nodes, Lomonosov Moscow University Fabian Kriegel, Validierung von intravitalmikroskopieschen Daten mittels in vitro und in silico Analysen im Mausmodell der akuten Darmentzündung, FU Berlin Katja Börnert, The influence of Interleukin-6 Receptor antibodies on breast cancer metastases in bone, Charité MD theses Friedericke Breitenfeld, Untersuchungen zum Zusammenhang einer systemischen Glucocorticoidtherapie mit dem Infektionsrisiko sowie dem Auftreten unerwünschter Hautveränderungen bei Patienten mit rheumatoider Arthritis, Charité Benedikt Paßmann, Validierung des Anaphylaxie-Registers, HU Berlin Franziska Rauhut, Untersuchung zur Relevanz von Interleukin-17 bei Patienten mit systemischem Lupus erythematodes, Charité Kristina Conrad, Quantifizierung löslicher und zellulärer Biomarker bei Patienten mit Spondyloarthritiden, HU Berlin Duc van Dang, Novel molecular mechanisms involved in the suppressive function in B cells, HU Berlin Alexander Daum, Einfluss der CD22-Ligation mittels Epratuzumab auf B-Zell-Effektorfunktionen über die Aktivierung von TLR-Rezeptoren bei SLE, FU Berlin Sarah Fleischer, Dysbalanced BCR signaling in B cells of patients with SLE, HU Berlin Claudia Giesicke, Molecular and phenotypic studies of human antigenspecific effector- and memory B cells, HU Berlin Vandana Kumari, Mechanisms underlying the regulatory function of tumor necrosis factor-α in skin inflammation, HU Berlin

106 Qualifications Appendix Patrick Maschmeyer, Hepatic stellate cells induce gut tropism of CD8+ T lymphocytes, TU-Berlin Maria Nassiri, Herz-Kreislauf- Medikamente als Kofaktoren der Anaphylaxie, HU Berlin Anna Okhrimenko, Human bone marrow hosts resting polyfunctional long-term memory T cells, HU Berlin René Riedel, Localization and Characterization of Murine Memory B Lymphocytes, FU Berlin Stefanie Ries, New insights into pathways controlling B cell fate and IL-6 expression, TU-Berlin Angelika Rose, Mechanisms of IL-2 therapy in murine models of systemic lupus erythematosus (SLE), TU-Berlin Caroline Winsauer, The contribution of TNF from distinct cellular sources to autoimmune colitis., FU Berlin Ina Wirries, Phänotypische und funktionelle Charakterisierung von CD19- Plasmazellen im humanen Knochenmark, HU Berlin Jakob Zimmermann, The Role of Th1, Th17, and Th17+1 Cells for the Initiation and Perpetuation of Inflammatory Bowel Disease, HU Berlin Habilitation Hildrun Haibel, Therapie der axialen Spondyloarthritis, Charité Bimba Hoyer, Die Rolle von Plasmazellen bei Autoimmunerkrankugen, Charité 2016 Bachelor theses Alina Liebheit, Role of plasma cells in chronic neuroinflammation, HU Berlin Janet Pubanz, Funktionelle Analysen einer potentiellen Enhancer- Region im murinen Irf8 Genlocus, Uni Potsdam Veronica Raba, Vergleich der Photoaktivierung von PA-GFP mittels Zwei-Photonen-Mikroskopie in Milz und Lymphknoten der Maus, Hochschule Niederrhein Julia Schubert, Classification of Human GM-CSF Producing T Helper Cell Subpopulations, Uni Potsdam Dipl. Ing. Peggy Kunath, Bedeutung von HIF in der Regulation von VEGFR-1 und VEGFR-2, TU-Berlin Master theses/diploma theses Merce Conill Cortes, Characterization of the GM-CSFproducing human T helper cell subpopulation, Charité - Universitätsmedizin Berlin Melanie de Almeida, Epigenetic Regulation of FOXP1 during CD4+ Memory T cell Differentiation, University of Copenhagen Almut Sophia Eisele, Molecular Signatures of NKG2D+ CD4+ T helper 1 Cells, Charité - Universitätsmedizin Berlin Ralf Köhler, SIBILLA algorithm - improving lateral resolution in stripedillumination intravital microscopy, Beuth Hochschule für Technik Berlin Rick Nolte, Die Auswirkung von Vitamin-D-Mangel auf die Brustkrebs Metastasierung, TU-Berlin Sabrina Schüngel, Identifizierung u Validierung immunmodulatorischer Eigenschaften von Substanzen, Hochschule f Technik und Wirtschaft Christopher Skopnik, Structure and function relationship of the IgM Fc receptor (FcµR), TU-Berlin Olga Trupp, Tropheryma whipplei as potential source for novel immunoregulatory agents, Technische Universität Braunschweig Siska Wilantri, Immunomodulation by inhibitory cytokines coupled to antigenic peptide and carrier peptide, Charite MD theses Agata Mossakowski, Funktionelle Echtzeitanalyse der zellulären und molekularen Mechanismen der neuronalen Dysfunktion in chronischer Neuroinflammation, Charité Universitätsmedizin Berlin Pascal Klaus, Modulation of the humoral immune response by antithymocyte globulin (ATG), Charité - Universitätsmedizin Berlin" Marie Lettau, Etablierung immunohistochemischer Multifluoreszenzfärbungen für Gedächtnis-B-Zell- Lokalisationsstudien in humanen lymphatischen Geweben, Charité - Universitätsmedizin Berlin Agata Mossakowski, Oxidative stress memory in chronic neuroinflammation by NAD(P) H-FLIM Julia Sieber, Zytokinprofil von TLR9-aktivierten B-Lymphozyten bei Systemischen Lupus Erythematodes, Charité - Universitätsmedizin Berlin Magdalena Kraft, Lebensdauer der im Rahmen einer mukosalen Immunantowrt generierten Plasmazellen, Charité Universitätsmedizin Berlin Dr. rer. med. Adrian Richter, Vergleich von Therapien der rheumatoiden Arthritis in Langzeitbeobachtungsstudien unter besonderer Berücksichtigung fehlender Daten, Charité Universitätsmedizin Berlin 105

107 Appendix Qualifications 106 PhD theses Jannike Bayat Sarmadi, In vivo analysis of CD3-specific antibody treatment-induced cellular changes in the small intestine, FU Berlin Alexander Beller, Regulation der mukosalen Immunität durch eosinophile Granulozyten und die intestinale Mikroflora, Freie Universität Berlin Christian Gabriel, Analysis of T Cell Receptor-Induced Signaling Modules by Mass Spectrometry: NFAT Interactions and the CBM-Complex, FU Berlin Manja Jargosch, Identifikation von wichtigen Transkriptionsfaktoren bei der Differenzierung von T Helferzellen mittels integrativer Netzwerkanalysen, FU Berlin Elisabeth Kenngott, Analysis and characterization of the 13C carrier peptide, Humboldt-Universität Maybritt Mardahl, Molecular regulation of P-selectin ligand in Th1 and Th2 cells in vitro, Freie Universität Berlin Anna Pascual-Reguant, Deciphering tissue-specific signals and molecular mechanisms in control of TH17 cells, FU Berlin Jennifer Pfeil, Induction of antigen-specific tolerance bx carrierconjugated peptides, Humboldt- Universität Karolin Pollok, Charakterisierung von Plasmazellen und deren Einfluß in der experimentellen autoimmunen Enzephalomyelitis, TU Berlin Laura Tech, Analysing the effectors contributing to germinal center dynamics: proliferation, migration and differentiation of B cells in vivo, FU Berlin Habilitation Paula Hoff, Osteoimmunologische Prozesse in der initialen inflammatorischen Phase der Frakturheilung, Charité - Universitätsmedizin Berlin

108 Sitemap Appendix Sitemap Sitemap 107 Approach with local trafic approx. 10 min walking distance from Berlin Central Station "Hauptbahnhof " (yellow) Bus TXL, Stop "Charité - Campus Mitte Bus 147, Stop Schumannstraße from Airport Berlin-Schönefeld: Train RE7 or RB14 to S+U Hauptbahnhof (approx. 30 min.)

109 Appendix Imprint Annual Report Title: Intestinal microbiota has crucial functions in chronic inflammatory diseases. One of the immune-mediated mechanisms is the production of immunoglobulin A (IgA) at mucosal surfaces. Here we show the localization of IgA producing B cells in the small intestine. Picture taken by Andrey Kruglov and Sandra Prepens 108 Imprint Imprint Deutsches Rheuma-Forschungszentrum Berlin (DRFZ) A Leibniz Institute Charitéplatz 1, Berlin, Germany Andreas Radbruch, Scientific Director Petra Starke, Administrative Director Phone +49 (0) Fax +49 (0) Contact info@drfz.de Coordination Graphics Photographs Translation other help Print Jacqueline Hirscher, Katrin Moser, Eva Kreiß Jacqueline Hirscher Jacqueline Hirscher, if not noted otherwise Christine Raulfs, Eva Kreiß, Katrin Moser, Randall Lindquist Tanja Durez Laserline, Berlin This annual report is available for free: ISSN