A powerful model of endometrial carcinogenesis! molecular analysis and rational design of immunological intervention approaches
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1 A powerful model of endometrial carcinogenesis! molecular analysis and rational design of immunological intervention approaches Afrouz Behboudi PhD, Associate Prof. School of Life Sciences, University of Skövde Bild 1
2 Outlines!! Uterine tumors!! Endometrial cancer (EC)!! A relevant tumor model for EC!! The BDII rat tumor model!! Recent Molecular data!! Development of this model for Immunological intervention approaches Bild 2
3 Uterine Tumors Malignant Endometrial cancer (EC) Benign!! Fibroids!! Endometriosis!! Hyperplasia Bild 3
4 Endometrial cancer!! The most common pelvic gynecologic malignancy - 2% to 3% lifetime risk!! develops in over women worldwide!! an estimated women die!! The incidence is increasing (" 4% per year)!! About 75% of tumors are localized!! " 20% recur after primary surgery, showing limited response to systemic therapy Bild 4
5 Endometrial cancer!! Hereditary!! Sporadic!! Type I!! Type II Bild 5
6 Type I Type II Histology Hormonedependency Microsatellite instability PTEN mutation K-RAS mutation ERBB2 overexpression Tp53 mutation P16 inactivation CDH1 downregulation endometrioid Yes 20 45% 50 80% 10 30% 10 30% 10 20% 10% 10-20% papillary serous or clear cell No 0 5% 10% 0 5% 45 80% 90% 40% 80-90% Bild 6
7 The BDII inbred rat strain! a relevant tumor model for endometrial carcinogenesis Bild 7
8 Project I Molecular classification of spontaneous endometrial adenocarcinomas in BDII rats Bild 8
9 Type I Type II BDII rat ECs Histology Hormonedependency Microsatellite instability PTEN mutation K-RAS mutation ERBB2 overexpression Tp53 mutation P16 inactivation CDH1 downregulation endometrioid 20 45% 50 80% 10 30% 10 30% 10 20% 10% papillary serous or clear cell 0 5% 10% 0 5% 45 80% 90% 40% 80-90% endometrioid Yes No Yes 10-20% 0-5% no mutation, but significant down-regulation no mutation No 67% 57% deletion and significant down-regulation significant down-regulation Bild 9 Samuelson et al. Endocr Relat Cancer. 2009
10 Conclusion!! Endometrial carcinogenesis in BDII rat females is related to higher-grade human type I tumors Bild 10
11 Project II Identification of novel tumor suppressor genes in this model Bild 11
12 Cytogenetic analysis of tumors Bild 12
13 Experimental design!! Cytogenetic: G-banding, CGH, SKY!! Gene mappings: FISH, RH mapping!! Detailed study of chromosome aberration: microsatellite allelotyping, array-cgh!! Molecular and functional analysis: mutation sequencing, expression analysis, epigenetic silencing, functional assays, Bild 13
14 IsTp53 the target? At least seven tumors with AI/ deletions in the region, but no Tp53 mutation Tp53 mutation analysis Tp53 Bild 14
15 AI/deletions, unassociated with Tp53 mutation? Bild 15
16 An independent TSG close to Tp53? Similar observations in several human malignancies (breast, lung, hepatocellular carcinomas, neuronal tumors, ) 1-6 Mb TP53 Bild 16
17 Similar works? Bild 17
18 Ongoing work!! Three candidates: Hic1, Skip and Myosin!! Mutation screening (SNPs)!! analysis of epigenetic silencing!! Functional assays!! IHC in human EC!! expression analysis Bild 18
19 Project III Development of the rat model towards tumor intervention approaches Bild 19
20 Background!! The mucosal immune system protects body against pathogens, and tumors!! Research on the development of mucosal vaccine adjuvants:!! Mucosal immunization generates protective immunity to viral infection in mice Bild 20
21 Question Would induced immunity in rat genital tract interfere with EC development? Bild 21
22 Toll-like receptors: new generation of vaccine adjuvants! all about copying nature! Bild 22
23 Adjuvant TLR activity and cancer!! Negative regulator of cancer!! TLR stimulation drives tumorigenesis!! "! to avoid counterproductive stimulation of TLRs: select adjuvants which will be recognized by immune cell, but not cancer cell TLRs Bild 23
24 Experimental design 1.! BDII rats in estrus and diestrus phases!! vagina and endometrium 2.! BN rats in estrus and diestrus phases 3.! Tumors developed in BDII rats Expression analysis for: Cd180, Tlr2, Tlr3, Tlr4, Tlr5, Tlr6, Tlr7, Tlr9, Myd88, Nfkb1, Ppia, Sigirr, Esr1, Esr2, Pgr Bild 24
25 BDII estrus vs BDII diestrus Endometrium Vagina Bild 25
26 BDII estrus vs BN estrus Endometrium Vagina Bild 26
27 BDII endometrium vs EC tumors Bild 27
28 Candidate receptor targets!! Tlr9 -- bacterial DNA, synthetic ODN with unmethylated CpG (CpG ODN)!! Cd180 (RP105) -- modulator of Tlr4 signaling!! Tlr7 -- imidazoquinoline, ss RNA Bild 28
29 Future directions!! Assess immune response to Tlr stimulations in rat vaginal tract!! Evaluate influence of Tlr stimulation prior to or at the time of EC on the natural courses of this malignancy!! High throughput molecular analysis!! new sets of rat tumors!! a panel of human ECs Bild 29
30 Developed tumor screening protocol! MRI! Ultrasound Bild 30
31 Developed tumor screening protocol Bild 31
32 University of Gothenburg Carola Hedberg, PhD Emma Samuelson, PhD Diego Garcia Dios Anna Linder Ali Harandi, PhD Katarina Ejeskär, PhD Charlotte Örndal, MD, PhD Karin Sundfeldt, MD, PhD University of Skövde Prof. Karin Klinga-Levan Björn Olsson, PhD Chalmers University of Technology Staffan Nilsson, PhD Hannover Medical School Dirk Wedekind, PhD University of Québec at Trois-Riviéres Prof. Eric Asselin Bild 32
33 !! Gunvor och Ivan Svenssons stiftelse!! Åke Wibergs Stiftelse!! Kungl. Vetenskapsakademien (KVA)!! ALF-Göteborg!! The Royal Physiographic Society in Lund (Nilsson-Ehle Foundation)!! Assar-Gabrielsson Foundation Bild 33
34 A powerful model of endometrial carcinogenesis! molecular analysis and rational design of immunological intervention approaches Afrouz Behboudi PhD, Associate Prof. School of Life Sciences, University of Skövde Bild 34
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