Epigenetic adaptation to therapy in glioblastoma: popping the hood on therapeutic resistance

Abstract

Glioblastoma (GBM), the most common and aggressive primary brain tumor in adults, remains one of most intractable diseases. Median survival remains at only 15 months, despite aggressive combination therapy involving surgical resection, radiation, and alkylating chemotherapy. While initially responsive to these therapies, GBM tumors quickly adapt and generate treatment-resistant recurrent growths. We and others have previously shown that the stress induced by standard of care therapies itself activates a remarkable plasticity in GBM cells, converting previously differentiated cells to a therapy-resistant glioma stem cell (GSC) state. The exact processes governing this conversion, however, remain to be fully elucidated. Preliminary investigation revealed that polycomb group protein EZH2, a well-established epigenetic regulator, plays a crucial role in this process. To determine how EZH2 responds to therapy, we performed Genome-wide chromatin immunoprecipitation (ChIP) in parallel with DNA sequencing analyses (ChIP-seq), which identified 1449 distinct regions with elevated EZH2 binding, including critical genes PTPRT, CDK5R2, and Siglec6. Microarray analysis demonstrated subsequent attenuation in their gene expression, leading to heightened activity of STAT3, a critical regulator for promotion of the GSC phenotype. To better understand the epigenetic adaptions occurring during therapeutic stress, we performed ChIP seq analysis for histone 3 lysine 27 acetylation (H3K27ac), a marker for active chromatin. Chemotherapy induces H3K27ac enrichment in 452 unique loci, while radiation caused H3K27ac enrichment at 1029 sites. Comparison of these sites to canonical H3K27me3 sites revealed specific de novo binding in the homeobox transcription factor binding motif (p=0.025). Combination of our epigenomic data from patient-derived xenograft models and GBM patient data with H3K27me3 enrichment profile allowed us to pinpoint several novel homeobox transcription factors potentially linked to GBM plasticity during therapeutic stress. These results provide critical perspective on the global epigenetic changes driving this plasticity occurring during anti-glioma therapy and provide novel avenues for targeting this adaptation therapeutically.