DNA Methylation Profiling of Glioblastoma: Impact on Gene Expression and Clinical Outcome (Agilent Expression Study)

Genome-scale measurements of DNA methylation levels are necessary to decipher the epigenetic events involved in glioblastoma aggressive phenotype, and to guide new therapeutic strategies. In that purpose, we performed a whole genome integrative analysis of the methylation and expression profiles for 40 newly diagnosed glioblastoma patients. We have also screened for associations between CpG sites methylation levels and overall survival in a cohort of 50 patients uniformly treated with radiotherapy and chemotherapy with concomitant and adjuvant temozolomide (STUPP protocol). The methylation analysis identified 616 CpG sites differentially methylated between glioblastoma and control brain, a quarter being differentially expressed in a concordant way. Among these concordant CpG sites, 13 genes displayed, within our glioblastoma cohort, an inverse correlation between promoter methylation and expression levels: B3GNT5, FABP7, ZNF217, BST2, OAS1, SLC13A5, GSTM5, ME1, UBXD3, TSPYL5, FAAH, C7orf13, and C3orf14. The expression of these genes may be tightly regulated by epigenetic mechanisms. The survival analysis identified six CpG sites associated with overall survival. The SOX10 promoter methylation status (two CpG sites) stratifies the patients in a way similar to MGMT with improved performance based on Area Under the Curve criteria (0.78 vs. 0.71, p-value < 5.10-4). The methylation status of FNDC3B, TBX3, DGKI, and FSD1 promoters identify patients with MGMT methylated tumors non-responding to STUPP treatment (p-value < 1.10-4). These markers have a potential impact on therapeutic decision. 40 glioblastoma samples and 6 control brain samples were analysed. 2 distinct series of hybridizations were carried out, each containing GBMs and control brains.

全基因组水平的DNA甲基化水平检测，对于解析与胶质母细胞瘤侵袭性表型相关的表观遗传事件、指导新型治疗策略至关重要。为此，我们对40例新确诊胶质母细胞瘤患者的甲基化与表达谱开展了全基因组整合分析。我们还在50例接受同步联合辅助替莫唑胺放化疗的STUPP方案标准化治疗队列中，筛选了CpG位点（CpG site）甲基化水平与总生存期之间的关联。甲基化分析共鉴定出616个在胶质母细胞瘤与正常对照脑组织间存在差异甲基化的CpG位点，其中四分之一的位点同时存在差异表达且变化趋势一致。在这些趋势一致的CpG位点中，我们的胶质母细胞瘤队列中共发现13个基因的启动子甲基化水平与基因表达水平呈负相关：B3GNT5、FABP7、ZNF217、BST2、OAS1、SLC13A5、GSTM5、ME1、UBXD3、TSPYL5、FAAH、C7orf13以及C3orf14。上述基因的表达可能受到表观遗传机制的精准调控。生存分析则鉴定出6个与总生存期相关的CpG位点。SOX10启动子甲基化状态（涉及2个CpG位点）对患者的分层效果与O6-甲基鸟嘌呤-DNA甲基转移酶（MGMT）相似，且基于曲线下面积（Area Under the Curve）的评估显示其性能更优（0.78 vs. 0.71，P值<5×10^-4）。FNDC3B、TBX3、DGKI以及FSD1的启动子甲基化状态，可识别出MGMT甲基化肿瘤却对STUPP方案治疗无应答的患者（P值<1×10^-4）。上述标志物或可对临床治疗决策产生潜在影响。本研究共分析了40例胶质母细胞瘤样本与6例正常对照脑组织样本。实验共开展了2组独立的杂交实验，每组均包含胶质母细胞瘤样本与对照脑组织样本。