The effect of epigenetic therapies on Glioblastoma Multiforme cancer stem cells

Glioblastoma Multiforme (GBM) is one of the most aggressive types of brain tumour; it is hard to treat with conventional therapy and often shows tumour regrowth after surgical removal. Recent studies have indicated the presence of cancer stem cells in GBM tumours, which are more resistant to therapy and driver of tumour recurrence after treatment. Here, we investigate the effect of several epigenetic inhibitors on two GBM stem cell lines, G7 and E2. Cells were not sensitive to the BET inhibitor OTX015, but the EZH2 inhibitors EPZ6438, UNC1999 and GSK343 all significantly reduced colony formation. The three EZH2 inhibitors induced expression of senescence-associated β-galactosidase, sensitised both cell lines to the alkylating agent temozolomide, and sensitised the E2 cell line to radiation. RNA-seq analysis following treatment with EZH2 inhibitors revealed a significant induction of genes related to neurogenesis and neuronal function, particular in the G7 cell line. ChIP-seq analysis of the epigenetic changes following EZH2 treatment was used to identify genes that were both upregulated and showed a decrease in histone H3 lysine 27 tri-methylation, which is the epigenetic mark deposited by EZH2. Several regulatory factors were identified that are potential candidates for driving the neuronal differentiation of the cancer stem cells. One of these candidates is the transcription factor PAX2, which is upregulated in both G7 and E2 cells. PAX2 is essential for the correct development of the inner ear and the eye, and is known to regulate expression of the neural transcription factors NEUROG1, NEUROD1 and ATOH1. Based on these data, it is hypothesise that the EZH2 inhibitors reduce GBM stem cell proliferation by inducing neuronal differentiation of the cancer stem cells, in addition to causing therapy induced senescence. Further work is required to investigate whether the candidate regulatory genes that were identified, including PAX2, do indeed play a role in driving this differentiation. Sensitisation of the GBM stem cells to temozolomide and radiation is a promising line of enquiry for further investigation into improving treatments for GBM