Dataset Information

Extent of MGMT promoter methylation correlates with outcome in glioblastomas given temozolomide and radiotherapy.

ABSTRACT:

Background

Epigenetic silencing of O(6)-methylguanine-DNA-methyltransferase (MGMT) by promoter methylation is associated with improved survival in glioblastomas treated with alkylating agents. In this study, we investigated MGMT promoter methylation in glioblastomas treated with temozolomide and radiotherapy in a single UK treatment centre.

Methods

Quantitative methylation data at individual CpG sites were obtained by pyrosequencing for 109 glioblastomas.

Results

Median overall survival (OS) was 12.4 months with 2-year survival of 17.9%. Pyrosequencing data were reproducible with archival samples yielding data for all glioblastomas. Variation in methylation patterns of discrete CpG sites and intratumoral methylation heterogeneity were observed. A total of 58 out of 109 glioblastomas showed average methylation >non-neoplastic brain in at least one clinical sample; 86% had homogeneous methylation status in multiple samples. Methylation was an independent prognostic factor associated with prolonged progression-free survival (PFS) and OS. Cases with methylation more than 35% had the longest survival (median PFS 19.2; OS 26.2 months, 2-year survival of 59.7%). Significant differences in PFS were seen between those with intermediate or high methylation and unmethylated cases, whereas cases with low, intermediate or high methylation all showed significantly different OS.

Conclusions

These data indicate that MGMT methylation is prognostically significant in glioblastomas given chemoradiotherapy in the routine clinic; furthermore, the extent of methylation may be used to provide additional prognostic stratification.

SUBMITTER: Dunn J

PROVIDER: S-EPMC2713697 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Glioblastoma (GBM) is the most malignant glioma, with a median overall survival (OS) of 14-16 months. Temozolomide (TMZ) is the first-line chemotherapy drug for glioma, but whether TMZ should be withheld from patients with GBMs that lack O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation is still under debate. DNA methylation profiling holds great promise for further stratifying the responses of MGMT promoter unmethylated GBMs to TMZ. In this study, we studied 147 TMZ-treated MGMT promoter unmethylated GBM, whose methylation information was obtained from the HumanMethylation27 (HM-27K) BeadChips (n = 107) and the HumanMethylation450 (HM-450K) BeadChips (n = 40) for training and validation, respectively. In the training set, we performed univariate Cox regression and identified that 3,565 CpGs were significantly associated with the OS of the TMZ-treated MGMT promoter unmethylated GBMs. Functional analysis indicated that the genes corresponding to these CpGs were enriched in the biological processes or pathways of mitochondrial translation, cell cycle, and DNA repair. Based on these CpGs, we developed a 31-CpGs methylation signature utilizing the least absolute shrinkage and selection operator (LASSO) Cox regression algorithm. In both training and validation datasets, the signature identified the TMZ-sensitive GBMs in the MGMT promoter unmethylated GBMs, and only the patients in the low-risk group appear to benefit from the TMZ treatment. Furthermore, these identified TMZ-sensitive MGMT promoter unmethylated GBMs have a similar OS when compared with the MGMT promoter methylated GBMs after TMZ treatment in both two datasets. Multivariate Cox regression demonstrated the independent prognostic value of the signature in TMZ-treated MGMT promoter unmethylated GBMs. Moreover, we also noticed that the hallmark of epithelial-mesenchymal transition, ECM related biological processes and pathways were highly enriched in the MGMT unmethylated GBMs with the high-risk score, indicating that enhanced ECM activities could be involved in the TMZ-resistance of GBM. In conclusion, our findings promote our understanding of the roles of DNA methylation in MGMT umethylated GBMs and offer a very promising TMZ-sensitivity predictive signature for these GBMs that could be tested prospectively.

Project description:Temozolomide (TMZ) is an important first-line treatment for glioblastoma (GBM), but there are limitations to TMZ response in terms of durability and dependence on the promoter methylation status of the DNA repair gene O6-methylguanine DNA methyltransferase (MGMT). MGMT-promoter-hypermethylated (MGMT-M) GBMs are more sensitive to TMZ than MGMT-promoter-hypomethylated (MGMT-UM) GBMs. Moreover, TMZ resistance is inevitable even in TMZ-sensitive MGMT-M GBMs. Hence, epigenetic reprogramming strategies are desperately needed in order to enhance TMZ response in both MGMT-M and MGMT-UM GBMs. In this study, we present novel evidence that the epigenetic reactivation of Tumor Suppressor Candidate 3 (TUSC3) can reprogram sensitivity of GBM stem cells (GSCs) to TMZ irrespective of MGMT promoter methylation status. Interrogation of TCGA patient GBM datasets confirmed TUSC3 promoter regulation of TUSC3 expression and also revealed a strong positive correlation between TUSC3 expression and GBM patient survival. Using a combination of loss-of-function, gain-of-function and rescue studies, we demonstrate that TUSC3 reactivation is associated with enhanced TMZ response in both MGMT-M and MGMT-UM GSCs. Further, we provide novel evidence that the demethylating agent 5-Azacitidine (5-Aza) reactivates TUSC3 expression in MGMT-M GSCs, whereas the combination of 5-Aza and MGMT inhibitor Lomeguatrib is necessary for TUSC3 reactivation in MGMT-UM GSCs. Lastly, we propose a pharmacological epigenetic reactivation strategy involving TUSC3 that leads to significantly prolonged survival in MGMT-M and MGMT-UM orthotopic GSCs models. Collectively, our findings provide a framework and rationale to further explore TUSC3-mediated epigenetic reprogramming strategies that could enhance TMZ sensitivity and outcomes in GBM. Mechanistic and translational evidence gained from such studies could contribute towards optimal design of impactful trials for MGMT-UM GBMs that currently do not have good treatment options.