MGMT Promoter Methylation is a Poor Predictor of Gene Expression in Low Grade Gliomas

Abstract

Radiation therapy with adjuvant temozolomide (TMZ) is the standard of care for glioblastoma multiforme (GBM). However, management of adult low grade gliomas (LGG) remains more complex. Expression of O6-methylyguanine-DNA-methyltransferase (MGMT) has been demonstrated to confer treatment resistance, while hypermethylation of the promoter is commonly used as a prognostic biomarker in GBM. In contrast, MGMT promoter methylation does not appear to improve progression-free survival in LGG. Here, we report an unexpected discordant relationship between MGMT promoter methylation and mRNA expression in LGG. This finding indicates that assays for MGMT promoter methylation may not properly identify treatment sensitive LGG. Grade II and III gliomas cases with clinical, methylation (450k infinium methylation beadchip) and gene expression (RNAseq) data were obtained from the TCGA-LGG dataset. Gene expression cutoffs were stratified by Youden index. The gplots R package was used to perform non-hierarchical cluster analysis using MGMT CpG sites for stratification. Differential methylation between samples was determined using t-tests with Bonferroni correction. In addition, differential methylation by two common MGMT promoter assays, DMR1 and MS-MLPA, was estimated using annotated CpG sites. Clinical variables were compared using t-tests for continuous variables and Fisher’s exact for categorical variables. Of the 516 TCGA-LGG cases eligible for analysis, 454 had complete clinical, methylation and sequencing data. Thirty-eight (8.4%) had high gene expression of MGMT. Non-hierarchical clustering analysis identified two distinct groups; 53% had hypermethylation of the MGMT promoter (Δ20% in methylation, q < 0.01), and by DMR1 (Δ13%, q = 0.02) and MS-MLPA sites (Δ31%, q < 0.01). There were no significant differences in average methylation across the entire MGMT gene (137 sites, Δ04%, q = 0.61) or the whole genome (38808 sites, Δ03%, q = 0.99). LGG cases with high MGMT expression and promoter methylation were four times more likely to have an IDH mutation (P < 0.01). Our analysis reveals an unexpected discordant relationship between MGMT promoter methylation and gene expression. Direct MGMT mRNA quantification may provide a better method for evaluating sensitivity to radiation and adjuvant TMZ in LGG.