Targeting Epigenetic Processes in Glioblastoma Mulitforme

Among tumors of the central nervous system, gliomas are the most prevalent and are associated with the poorest prognosis. Overall survival rates are low, and combined radioand chemo-therapy provides only marginal benefit; thus it is essential to develop novel methods to treat patients with these aggressive tumors. One promising new area of research focuses upon epigenetics, the regulation of genetic expression without modifying the DNA base-pair sequence. Epigenetic mechanisms, such as histone acetylation, methylation, and cytosine methylation, have been shown to modulate tumor-suppressor genes in nontransformed cells. We hypothesized that epigenetic mechanisms contribute to the transformed phenotype of gliomas by targeting critical tumor suppressor genes. Our preliminary research showed that low doses (1 - 3 mM) of the histone deacetylase inhibitor HDAC-42 led glioma cell lines to undergo cell cycle arrest and apoptosis in a dose dependent fashion. Similar concentrations of HDAC-42 sensitized glioma cell lines to apoptosis induced by standard chemotherapeutic drugs (cisplatin, temozolomide). The diminished apoptotic threshold induced by HDAC-42 correlated with decreased sphingosine kinase-1 (SK1) activity, an important enzyme promoting survival in GBM tumors. Despite this promising result, the apoptotic effect was modest in several glioma cell lines, which led us to examine the importance of other enzymes that contribute towards tumor suppressor gene silencing. Recent work has demonstrated that HDAC enzymes associate with repressive SWI/SNF complexes containing other co-repressive elements to silence tumor suppressor genes. Among the epigenetic enzymes implicated in tumor-suppressor silencing is the protein arginine methyl transferase-5 (PRMT5), an enzyme that methylates histones H3 (H3R4) and H4 (H4R8) that localize to promoter regions of specific genes. Chromatin immunoprecipitation (ChIP) experiments have shown that PRMT5 directly targets the ST7 promoter, a known tumor suppressor gene. Western blot analysis of protein lysates isolated from 6 glioma cell lines has shown that PRMT5 is overexpressed, a finding that we have found to be critical for cellular transformation. Western blots have shown that ST7 not expressed in glioma lines but is abundantly expressed in non-transformed cells (B cells and fibroblasts). Through sequencespecific RNA interference, we are presently targeting PRMT5 expression and examining ST7 expression. These experiments will allow us to determine if re-expression of ST7 correlates with restoration of features associated with control of cell growth and survival. Experimental therapeutic strategies to target multiple co-repressor enzyme activities are discussed