MGMT基因启动子甲基化检测在脑胶质瘤化疗中的意义

背景与目的：如何预测和克服肿瘤细胞对化疗药物的耐药性，实施个体化治疗是肿瘤化疗急需解决的问题。与基因启动子甲基化密切相关的DNA损伤修复基因O^6-甲基鸟嘌呤-DNA甲基转移酶（O^6-methylguanine-DNA methyhransferase，MGMT）表观沉默与肿瘤对烷化剂药物化疗敏感性密切相关。本研究探讨检测MGMT基因启动子CpG岛甲基化在判断脑胶质瘤患者预后及预测肿瘤对烷化剂药物耐药性中的意义。方法：甲基化特异性PCR（MSP）法检测脑胶质瘤组织及肿瘤细胞株MGMT基因启动子甲基化状态，蛋白印迹和免疫组化法测定蛋白表达。MTF法检测肿瘤细胞株对烷化剂药物敏感性，将患者随访资料针对MGMT甲基化状态绘制Kaplan-Meier生存曲线，并进行log—rank检验分析。结果：39例脑胶质瘤患者组织MGMT基因启动子甲基化发生率为46．2％，蛋白表达阳性率为61．5％，且肿瘤组织中MGMT基因甲基化状态与蛋白表达显著相关（P〈0．05）：6例正常组织均未检测出基因甲基化。MGMT基因过甲基化的脑胶质瘤SHG44细胞株用5-Aza-CdR处理后完全脱甲基化．MGMT蛋白恢复了表达，同时细胞株对烷化剂药物敏感性也发生逆转．由敏感转变为耐受。在采用手术、放疗和烷化剂尼莫司汀化疗等综合治疗的39例脑胶质瘤患者中，MGMT基因甲基化的患者生存率显著高于MGMT基因未甲基化患者（P〈0．05）。结论：MGMT基因甲基化状态与蛋白表达及肿瘤细胞对烷化剂药物敏感性密切相关，有可能替代MGMT蛋白检测成为判断脑胶质瘤患者预后和预测肿瘤对烷化剂化疗耐药性的标志分子。

Detection of O6-methylguanine-DNA methyltransferase promoter methylation in chemotherapy for glioma

Background and Objective： Epigenetic silencing of the DNA repair gene, O^6-methylguanine-DNA methyltransferase （MGMT）, is associated with the therapeutic response to methylating agents. This study was to assess the value of detecting the promoter methylation of MGMT gene in chemotherapy for glioma. Methods. Methylation-specific PCR （MSP） was employed to detect MGMT promoter CpG isrand methylation in 39 samples of glioma taken from surgery. Western blot and immunohistochemistry were used to detect protein expression. MTT were employed to detect the sensitivity of two glioma cell lines to alkylating agents, ACNU and TMZ. The Kaplan-Meier curve was adopted to estimate the overall survival according to the methylation status of the MGMT promoter. Results. Methylation of MGMT promoter CpG island was detectable in 46.2% of glioma tissues, but not in any normal tissues. The expression rate of MGMT protein was 61,5%. The status of MGMT methylation status was association with the protein level of MGMT （P〈0.05）. The MGMT gene was demethylated in glioma cell line SHG-44 following 5-Aza-CdR treatment; the expression of MGMT protein was restored and the resistance of SHG44 cells to alkylating agents was reversed. The overall survival was higher in patients with methylated MGMT promoter than in those with unmethylated MGMT promoter （P〈0.05）. Conclusions. The status of MGMT promoter CpG island methylation is closely correlated to MGMT protein expression and sensitivity of cells to alkylating agents in glioma. Detection of the methylated sequences of MGMT may be used as a predictive factor for the treatment of giloma.