Discovery of epigenetically masked tumor suppressor genes in gyneoobgical cancer

点击翻译按钮获取中文摘要

Realization that many tumor suppressor genes are silenced by epigenetic mechanisms has stimulated discovery of novel tumor suppressor genes. We used a variety of research tools to search for genes that are epigenetically silenced in human endometrial cancers. Changes in global gene expression of the endometrial cancer cell line Ishikawa was analyzed after treatment with the demethylating agent 5-aza-2'-deoxycytidine (5-Aza-CdR) combined with the histone deacetylase inhibitor, suberoyl anilide bishydroxamide (SAHA). By screening over 22,000 genes, candidate tumor suppressor genes were identified. Additional microarray analysis and real-time RTPCR of normal and cancerous endometrial samples, and search for CpG islands further refined the list. Tigl and C/ebpa were chosen for further study. Expression of both genes was low in endometrial cancer cell lines and clinical samples, but high in normal endometrial tissues. Bisulfite sequencing and restriction analysis revealed aberrant methylation of the CpG island in the Tigl gene of all 6 endometrial cancer cell lines examined, whereas the C/ebpa promoter remained unmethylated in endometrial cancers Chromatin immunoprecipitation showed increased acetylated histones H3 bound to both Tigl and C/ebpa genes after treatment with 5-Aza-CdR and/or SAHA. Forced expression of either TIG1 or C/EBPa led to significant growth reduction of Ishikawa cells. Immunoprecipitation and reporter gene assays demonstrated that C/EBPa bound to and suppressed transcriptional activity of E2F1, a key cell cycle regulator. Our data suggest that C/ebpa and Tigl function as tumor suppressor proteins in endometrial cancers and that their reexpression may be a therapeutic target.