O-甲基鸟嘌呤-DNA 甲基转移酶（MGMT）mRNA 表达预测恶性神经胶质瘤的预后，与 MGMT 启动子甲基化无关。

BACKGROUND

We analyzed prospectively whether MGMT (O(6)-methylguanine-DNA methyltransferase) mRNA expression gains prognostic/predictive impact independent of MGMT promoter methylation in malignant glioma patients undergoing radiotherapy with concomitant and adjuvant temozolomide or temozolomide alone. As DNA-methyltransferases (DNMTs) are the enzymes responsible for setting up and maintaining DNA methylation patterns in eukaryotic cells, we analyzed further, whether MGMT promoter methylation is associated with upregulation of DNMT expression.

METHODOLOGY/PRINCIPAL FINDINGS: ADULT PATIENTS WITH A HISTOLOGICALLY PROVEN MALIGNANT ASTROCYTOMA (GLIOBLASTOMA: N = 53, anaplastic astrocytoma: N = 10) were included. MGMT promoter methylation was determined by methylation-specific PCR (MSP) and sequencing analysis. Expression of MGMT and DNMTs mRNA were analysed by real-time qPCR. Prognostic factors were obtained from proportional hazards models. Correlation between MGMT mRNA expression and MGMT methylation status was validated using data from the Cancer Genome Atlas (TCGA) database (N = 229 glioblastomas). Low MGMT mRNA expression was strongly predictive for prolonged time to progression, treatment response, and length of survival in univariate and multivariate models (p<0.0001); the degree of MGMT mRNA expression was highly correlated with the MGMT promoter methylation status (p<0.0001); however, discordant findings were seen in 12 glioblastoma patients: Patients with methylated tumors with high MGMT mRNA expression (N = 6) did significantly worse than those with low transcriptional activity (p<0.01). Conversely, unmethylated tumors with low MGMT mRNA expression (N = 6) did better than their counterparts. A nearly identical frequency of concordant and discordant findings was obtained by analyzing the TCGA database (p<0.0001). Expression of DNMT1 and DNMT3b was strongly upregulated in tumor tissue, but not correlated with MGMT promoter methylation and MGMT mRNA expression.

CONCLUSIONS/SIGNIFICANCE: MGMT mRNA expression plays a direct role for mediating tumor sensitivity to alkylating agents. Discordant findings indicate methylation-independent pathways of MGMT expression regulation. DNMT1 and DNMT3b are likely to be involved in CGI methylation. However, their exact role yet has to be defined.

背景

我们前瞻性地分析了在接受放疗联合辅助替莫唑胺或单独替莫唑胺治疗的恶性胶质瘤患者中，MGMT（O(6)-甲基鸟嘌呤-DNA 甲基转移酶）mRNA 表达是否获得了独立于 MGMT 启动子甲基化的预后/预测影响。由于 DNA-甲基转移酶（DNMTs）是负责建立和维持真核细胞中 DNA 甲基化模式的酶，我们进一步分析了 MGMT 启动子甲基化是否与 DNMT 表达上调有关。

方法/主要发现：纳入了 53 例组织学证实的恶性星形细胞瘤（胶质母细胞瘤：N=53，间变性星形细胞瘤：N=10）的成年患者。通过甲基化特异性 PCR（MSP）和测序分析确定 MGMT 启动子甲基化。通过实时 qPCR 分析 MGMT 和 DNMTs mRNA 的表达。预后因素来自比例风险模型。使用癌症基因组图谱（TCGA）数据库（N=229 例胶质母细胞瘤）的数据验证了 MGMT mRNA 表达与 MGMT 甲基化状态之间的相关性。在单变量和多变量模型中，低 MGMT mRNA 表达强烈预测进展时间延长、治疗反应和生存时间延长（p<0.0001）；MGMT mRNA 表达的程度与 MGMT 启动子甲基化状态高度相关（p<0.0001）；然而，在 12 例胶质母细胞瘤患者中发现了不一致的结果：6 例具有高 MGMT mRNA 表达的甲基化肿瘤患者的预后明显差于低转录活性患者（p<0.01）。相反，低 MGMT mRNA 表达的未甲基化肿瘤患者的预后好于其对应患者。通过分析 TCGA 数据库，获得了几乎相同频率的一致和不一致的发现（p<0.0001）。DNMT1 和 DNMT3b 在肿瘤组织中表达强烈上调，但与 MGMT 启动子甲基化和 MGMT mRNA 表达无关。

结论/意义：MGMT mRNA 表达直接介导肿瘤对烷化剂的敏感性。不一致的发现表明 MGMT 表达调控存在甲基化非依赖性途径。DNMT1 和 DNMT3b 可能参与 CGI 甲基化。然而，它们的确切作用尚待确定。