1. 2009.11.21 成大婦產部學術研討會 黃體素受體基因之多型性與習慣性流產 的關聯性 蘇美慈 醫師 / 郭保麟 教授

2. Genetic background Common disease are caused by combination of genetic and environmental factors Genetic studies linkage studies: broad chromosome region association studies: specific genetic variants and disease – candidate gene approach

4. Genetic markers Common variants arose from single historical events, and are associated with nearby variants

5. HapMap Project Goal determine common patterns of DNA sequence variants in human genome Public resource To increase power and efficiency of genetic association studies to medical traits High-density SNP genotyping across genome

7. Tag SNPs

8. Genetic association study for the role of progesterone receptor in recurrent pregnancy loss (RPL) 黃體素受體基因之多型性與習慣性 流產的關聯性

9. Introduction Recurrent pregnancy loss (RPL) repeated abortion before 20 weeks of gestation 1-5% of women who conceive multi-factorial disorder insufficient progesterone in the luteal phase and early pregnancy

10. Progesterone oocyte maturation, embryo implantation and placenta maintenance progesterone receptor knockout (PRKO) female mice significant defects in all reproductive tissues Mifepristone (RU486, block PR) Termination success rates before 7 weeks of gestation is >90 %

11. Progesterone receptor gene hPR (human PR) Size: 92Kb (8 exons) chromosome 11q22-23 2 alternative promoters and translated into two different zinc-finger proteins: PR-A, PR-B

12. Two functional SNPs +331G/A (rs10895068) polymorphism creates a unique transcription start site ↑ transcription of PGR, favoring the B isoform associated with endometrial cancer PROGINS

13. Definitions (3 polymorphisms) Alu element in intron 7 missense SNP in exon 4 (rs1042838) silence SNP in exon 5 (rs1042839) in complete linkage disequilibrium with the Alu insertion Association studies associated with breast cancer, ovarian cancer and endometrial cancer

14. Rationale Progesterone and its receptor is critical for female reproduction, esp for luteal phase support, embryo implantation & early placentation

15. Hypothesis Women who carries risk allele of PR may have more tendency for recurrent pregnancy loss, or vice versa

16. Materials and Methods Inclusion: Total analyzed 121 RPL patients normal controls: 179 Exclusion: Patients who could be identified obvious causal factors (27 patients) chromosome, coagulation, autoimmune, endocrine, uterine anomalies …

17. HapMap exploration 9 tag SNPs (MAF ≧.05, r2 ≧ 0.8) 2 functional SNPs PROGINS (rs1042838), +331G/A (rs10895068) Genotyping analysis ABI: assay-on-demand or assay-by-design (Taqman, VIC & FAM) Read by ABI 370

18. Results

20. Table 1: Significant allele and genotype frequency difference in 2 SNPs

21. Haplotype analysis

22. Conclusion In this study, two SNPs (rs9282823 and rs1042838) and several haplotypes of PR gene were associated with RPL and may increase the risk of recurrent pregnancy loss

23. Discussion

24. PROGINS 306 base pair insertion allele (Alu) higher mRNA stability and is transcribed to a more stable and transcriptionally active mutated protein (PR-B) --2004 JCEM Aqoulnik et al. increased in-cis transcription of PROGINS allele but reduces the stability of the transcript --2007 JME Romano et al. segregate with progesterone-dependent neoplasms, such as breast cancer, ovarian cancer and endometrial cancer

25. V660L polymorphism (rs1042838) the V660L allele may loss of cis-acting, DNA binding SF2-type slicing enhancer site, and definding the boundary of intron/exon  result in abnormal RNA splicing and exon skipping steric interference between progesterone-binding domain and the DNA-binding domain  affecting hormone-receptor binding & downstream transcription PR phosphorylation and degradation in the two PR variants upon ligand binding --2007 JME Romano et al.

26. Other PR polymorphism association studies in RPL 2004, Schweikert A. et al 3 SNPs of PR genes were linked with PROGINS (Alu-Insertion) and may be associated with an increased risk of RPL exon 1(G1031CSer344Thr), exon 4 (G1978TLeu660Val) and exon 5(C2310THis770His) 2001, Kurz el al did not supported the association between PROGINS Alu-Insertion and idiopathic RPL

27. Thank you for your attention!