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Extensive genomic and gene expression studies have been performed in gliomas, but the epigenetic alterations that characterize different subtypes of gliomas remain largely unknown. Here, we analyzed the methylation patterns of 807 genes (1536 CpGs) in a series of 33 low-grade gliomas (LGGs), 36 glioblastomas (GBMs), 8 paired initial and recurrent gliomas, and 9 controls. This analysis was performed with Illumina's Golden Gate Bead methylation arrays and was correlated with clinical, histological, genomic, gene expression, and genotyping data, including IDH1 mutations. Unsupervised hierarchical clustering resulted in 2 groups of gliomas: a group corresponding to de novo GBMs and a group consisting of LGGs, recurrent anaplastic gliomas, and secondary GBMs. When compared with de novo GBMs and controls, this latter group was characterized by a very high frequency of IDH1 mutations and by a hypermethylated profile similar to the recently described glioma CpG island methylator phenotype. MGMT methylation was more frequent in this group. Among the LGG cluster, 1p19q codeleted LGG displayed a distinct methylation profile. A study of paired initial and recurrent gliomas demonstrated that methylation profiles were remarkably stable across glioma evolution, even during anaplastic transformation, suggesting that epigenetic alterations occur early during gliomagenesis. Using the Cancer Genome Atlas data set, we demonstrated that GBM samples that had an LGG-like hypermethylated profile had a high rate of IDH1 mutations and a better outcome. Finally, we identified several hypermethylated and downregulated genes that may be associated with LGG and GBM oncogenesis, LGG oncogenesis, 1p19q codeleted LGG oncogenesis, and GBM oncogenesis.

The loss of chromosomes 1p-19q is the only prognostic molecular alteration identified in low-grade gliomas (LGGs) to date. Search for loss of heterozygosity (LOH) on chromosomes 1p, 9p, 10q, and 19q was performed in a series of 231 LGGs. Loss of chromosomes 1p-19q was strongly correlated with prolonged progression-free survival (PFS) and overall survival (OS) in univariate and multivariate analyses. LOH on 9p and 10q were associated with shortened PFS (P = .01 and .03, respectively) on univariate analysis. On multivariate analysis, LOH on 9p remained significant for PFS (P = .05), whereas LOH on 10q had a significant effect on OS (P = .02). Search for LOH 9p and 10q appears to be a useful complement to analysis of chromosomes 1p-19q in LGGs.

The O(6)-methylguanine-DNA methyltransferase gene (MGMT) is methylated in several cancers, including gliomas. However, the functional role of cysteine-phosphate-guanine (CpG) island (CGI) methylation in MGMT silencing is still controversial. The aim of this study was to investigate whether MGMT CGI methylation correlates inversely with RNA expression of MGMT in glioblastomas and to determine the CpG region whose methylation best reflects the level of expression. The methylation level of CpG sites that are potentially related to expression was investigated in 54 glioblastomas by pyrosequencing, a highly quantitative method, and analyzed with respect to their MGMT mRNA expression status. Three groups of patients were identified according to the methylation pattern of all 52 analyzed CpG sites. Overall, an 85% rate of concordance was observed between methylation and expression (p < 0.0001). When analyzing each CpG separately, six CpG sites were highly correlated with expression (p < 0.0001), and two CpG regions could be used as surrogate markers for RNA expression in 81.5% of the patients. This study indicates that there is good statistical agreement between MGMT methylation and expression, and that some CpG regions better reflect MGMT expression than do others. However, if transcriptional repression is the key mechanism in explaining the higher chemosensitivity of MGMT-methylated tumors, a substantial rate of discordance should lead clinicians to be cautious when deciding on a therapeutic strategy based on MGMT methylation status alone.

Various gene amplifications have been observed in gliomas. Prognostic-genomic correlations testing simultaneously all these amplified genes have never been conducted in anaplastic oligodendrogliomas. A set of 38 genes that have been reported to be amplified in gliomas and investigated as the main targets of amplicons were studied in a series of 52 anaplastic oligodendrogliomas using bacterial artificial chromosome-array based comparative genomic hybridization and quantitative polymerase chain reaction. Among the 38 target genes, 15 were found to be amplified in at least one tumor. Overall, 27% of anaplastic oligodendrogliomas exhibited at least one gene amplification. The most frequently amplified genes were epidermal growth factor receptor (EGFR) and cyclin-dependent kinase 4/sarcoma amplified sequence (CDK4/SAS) in 17% and 8% of anaplastic oligodendrogliomas, respectively. Gene amplification and codeletion of chromosome arms 1p/19q were perfectly exclusive (p = 0.005). In uni- and multivariate analyses, gene amplification was a negative prognostic factor for progression-free survival and overall survival in anaplastic oligodendrogliomas, providing complementary information to the classic prognostic factors identified in anaplastic oligodendrogliomas (extent of surgery, KPS, and chromosome arms 1p/19q status).