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Epigenetic Activation of TUSC3 Sensitizes Glioblastoma to Temozolomide Independent of MGMT Promoter Methylation Status

ABSTRACT: 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.

ORGANISM(S): Homo sapiens

PROVIDER: GSE282886 | GEO | 2024/11/26

REPOSITORIES: GEO

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Project description:Glioblastoma (GBM) is the most common and most aggressive primary brain tumor in adults. The existence of a small population of stem-like tumor cells that efficiently propagate tumors and resist cytotoxic therapy is one proposed mechanism leading to the resilient behavior of tumor cells and poor prognosis. In this study, we performed an in-depth analysis of the DNA methylation landscape in GBM-derived cancer stem cells (GSCs). Parallel comparisons of primary tumors and GSC lines derived from these tumors with normal controls (a neural stem cell (NSC) line and normal brain tissue) identified groups of hyper- and hypomethylated genes that display a trend of either increasing or decreasing methylation levels in the order of controls, primary GBMs, and their counterpart GSC lines, respectively. Interestingly, concurrent promoter hypermethylation and gene body hypomethylation were observed in a subset of genes including MGMT, AJAP1 and PTPRN2. These unique DNA methylation signatures were also found in primary GBM-derived xenograft tumors indicating that they are not tissue culture-related epigenetic changes. Integration of GSC-specific epigenetic signatures with gene expression analysis further identified candidate tumor suppressor genes that are frequently down regulated in GBMs such as SPINT2, NEFM and PENK. Forced re-expression of SPINT2 reduced glioma cell proliferative capacity, anchorage independent growth, cell motility, and tumor sphere formation in vitro. The results from this study demonstrate that GSCs possess unique epigenetic signatures that may play important roles in the pathogenesis of GBM. The reduced representation bisulfite sequencing (RRBS) approach (Meissner et al., 2008) was used to generate genome-wide single-base resolution CpG methylation profiles of three primary GBMs (1063T, 1133T, and 1142T) and three GSC lines (1063S, 1133S, 1142S) derived from these primary GBM tumors. The NSC line and the normal brain (NB) tissue sample were used as controls for comparison purposes. In addition, we analyzed three GBM xenograft tumor tissue samples (Mayo22, Mayo39, Mayo59) developed by Dr. Jann N. Sarkaria of Mayo Clinic.

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Epigenetic Activation of TUSC3 Sensitizes Glioblastoma to Temozolomide Independent of MGMT Promoter Methylation Status

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