Laboratory of Brain Tumor Biology, Department of Clinical Neurosciences, Centre Hospitalier Universitaire Vaudois and University of Lausanne, Lausanne, Switzerland.
Clin Cancer Res. 2011 Jan 15;17(2):255-66. doi: 10.1158/1078-0432.CCR-10-1931. Epub 2010 Nov 19.
Quantitative methylation-specific tests suggest that not all cells in a glioblastoma with detectable promoter methylation of the O6-methylguanine DNA methyltransferase (MGMT) gene carry a methylated MGMT allele. This observation may indicate cell subpopulations with distinct MGMT status, raising the question of the clinically relevant cutoff of MGMT methylation therapy. Epigenetic silencing of the MGMT gene by promoter methylation blunts repair of O6-methyl guanine and has been shown to be a predictive factor for benefit from alkylating agent therapy in glioblastoma.
Ten paired samples of glioblastoma and respective glioblastoma-derived spheres (GS), cultured under stem cell conditions, were analyzed for the degree and pattern of MGMT promoter methylation by methylation-specific clone sequencing, MGMT gene dosage, chromatin status, and respective effects on MGMT expression and MGMT activity.
In glioblastoma, MGMT-methylated alleles ranged from 10% to 90%. In contrast, methylated alleles were highly enriched (100% of clones) in respective GS, even when 2 MGMT alleles were present, with 1 exception (<50%). The CpG methylation patterns were characteristic for each glioblastoma exhibiting 25% to 90% methylated CpGs of 28 sites interrogated. Furthermore, MGMT promoter methylation was associated with a nonpermissive chromatin status in accordance with very low MGMT transcript levels and undetectable MGMT activity.
In MGMT-methylated glioblastoma, MGMT promoter methylation is highly enriched in GS that supposedly comprise glioma-initiating cells. Thus, even a low percentage of MGMT methylation measured in a glioblastoma sample may be relevant and predict benefit from an alkylating agent therapy.
定量甲基化特异性检测提示，在 O6-甲基鸟嘌呤 DNA 甲基转移酶（MGMT）基因启动子甲基化可检测到的胶质母细胞瘤中，并非所有细胞均携带甲基化的 MGMT 等位基因。这一观察结果可能表明存在具有不同 MGMT 状态的细胞亚群，从而引发有关 MGMT 甲基化治疗的临床相关截止值的问题。MGMT 基因启动子甲基化导致 MGMT 基因的表观遗传沉默，削弱了 O6-甲基鸟嘌呤的修复，已被证明是胶质母细胞瘤中烷化剂治疗获益的预测因素。
对 10 对胶质母细胞瘤及其相应的在干细胞条件下培养的胶质母细胞瘤球体（GS）样本进行分析，通过甲基化特异性克隆测序、MGMT 基因剂量、染色质状态以及对 MGMT 表达和 MGMT 活性的各自影响，评估 MGMT 启动子甲基化的程度和模式。
在胶质母细胞瘤中，MGMT 甲基化等位基因的范围为 10%至 90%。相比之下，在各自的 GS 中，甲基化等位基因高度富集（100%的克隆），即使存在 2 个 MGMT 等位基因，也只有 1 个例外（<50%）。每个胶质母细胞瘤的 CpG 甲基化模式具有特征性，在 28 个被检测的位点中，有 25%至 90%的 CpG 被甲基化。此外，MGMT 启动子甲基化与非许可染色质状态相关，相应地，MGMT 转录物水平非常低，MGMT 活性无法检测到。
在 MGMT 甲基化的胶质母细胞瘤中，MGMT 启动子甲基化在 GS 中高度富集，GS 可能包含神经胶质瘤起始细胞。因此，即使在胶质母细胞瘤样本中测量到的 MGMT 甲基化百分比很低，也可能具有相关性并预测烷化剂治疗的获益。
