肝内和肝外胆管癌肿瘤抑制基因启动子甲基化分析

Promoter Methylation Profiles of Tumor Suppressor Genes in lntrahepatic and Extrahepatic Cholangiocarcinoma

目的 探讨胆管癌的表遗传学改变。方法 用甲基化特异PCR(MSP)法 ,检测了 12个候选肿瘤抑制基因 (APGE cad herin/CDH1,MGMT ,RASSF1A ,GSTP ,RAR β ,p14 ARF,p15 INK4b,p16INK4a,p73 ,hMLH1,DAPK)启动子在 72例胆管癌中的甲基化情况 ,其中肝内和肝外胆管癌各 3 6例 ,10例良性胆管上皮作为对照。结果 85 %的胆管癌至少有一个肿瘤抑制基因的甲基化 ,在胆管癌中 ,肿瘤抑制基因的甲基化顺序是 :RASSF1A(65 %) ,p15 INK4b(5 0 %) ,p16INK4a(5 0 %) ,APC(4 6%) ,E cadherin/CDH1(4 3 %) ,p14 ARF(3 8%) ,p73 (3 6%) ,MGMT(3 3 %) ,hMHL1(2 5 %) ,GSTP(14 %) ,RAR β(14 %)和DAPK(3 %)。虽然单个肿瘤抑制基因的甲基化可见于良性胆管上皮 ,但是多个肿瘤抑制基因的甲基化只见于胆管癌。约 70 %(5 0 /72 )的胆管癌有 3个或 3个以上的肿瘤抑制基因的甲基化 ,5 2 %(3 8/72 )有 4个或 4个以上肿瘤抑制基因的甲基化。多个肿瘤抑制基因的协同甲基化 ,和RASSF1A ,p15 INK4b,p16INK4a和 /或hMHL1密切相关。RASSF1A的甲基化在肝外胆管癌 (83 %)较肝内胆管癌更常见 (4 7%) (P =0 . 0 0 3 ) ,而GSTP更多见于肝内胆管癌(肝内 3 1%,肝外 6%,P =0 . 0 12 ) ,本研究提示肿瘤抑制基因启动子CpG岛的甲基化在胆管癌中是一?

Objective To further understand epigenetic alterations in cholangiocarcinoma.Methods We have studied the meth3lation profiles of 12 candidate tumor suppressor genes (APC. E-cadberin/CDHl, MGMT, RASSF1A, GSTP, RAR-β pl4 ARF, p15 INK4b. p16 INK4a, p73, hMLHI annDAPK) in 72 cases ofcholangiocarcinoma, including equal munber cases of intrahepatic cholangiocarcinoma and extrahepatic cholangiocarcinoma. Ten cases of benign biliary epithelia were included as controls. The methylation status of tumor suppressor genes was analyzed using methylation-specific PCR. Results We found that 85% of all cholangiocarcinomas had methylation of at least one tumor suppressor gene. The fi'equency of tumor suppressor gene methylation in cholangiocarcinoma was: RASSF1A (65%), p15 INK4b (50%). p16 INK4a (50%), APC (46%), E-cadherin/CDHl (43%), p14 ARF(38%), p73 (36%), MGMT (33%), hMHLI (25%), GSTP (14%),RAR-β(14%) and DAPK (3%). Although single tumor suppressor gene methylation can be seen in benign biliary epithelium, methylation of multiple tumor suppressor genes is only seen in cholangiocarcinorna. About 70% (50/72) of the cholangiocarcinomas had three or more tumor suppressor genes methylated and 52% (38/72) of cases had lbur or more tumor suppressor genes methylated. Concerted methylafion of multiple tumor suppressor genes was closely associated with methylation of RASSF1,4, pi6 and/or hMHLI. Methylation of RASSF1A was more common in extrahepatic eholangiocarcinoma than intrahepatic cholangiocarcinoma (83% vs 47%, P=0.003) while GSTP was more frequently seen in intrahepatic compared to extrahepatic cholangiocarcinoma (31% vs 6%, p 0.012). Our study indicates that methylation of promoter CpG islands of tumor suppressor genes is a common epigenetic event in cholangiocarcinoma. Based on distinct methylation profiles, intrahepatic cholangiocarcinoma and extrahepatic cholangiocarcinoma are two closely related but biologically unique neoplastic processes.Conclusions Taking advantage of the unique concurrent methylation profile of multiple genes in cholangiocarcinoma may facilitate the distinction ofcholangiocarcinoma fi'om benign biliary epithelium in clinical settings.