Targeting Histone Deacetylase‐7 as a Novel Therapeutic Strategy for Glioblastoma Multiforme

Abstract

BackgroundGlioblastoma multiforme (GBM) is the most aggressive primary brain tumor. GBM has an ominous prognosis with a survival rate of 14‐15 months after diagnosis and despite worldwide initiatives to optimize therapeutic approaches, GBM is still among the most challenging diseases to treat and the fastest to relapse in clinical oncology. The inevitable treatment resistance and relapse are mainly attributed to the presence of glioma stem cells (GSCs), which have the ability to self‐renew, indefinitely proliferate and to differentiate into different cell types. We hypothesize that HDAC7, as an epigenetic regulator, is playing a crucial mechanistic role in GBM and constitutes a novel druggable target for this universally fatal disease. By uncovering the HDAC7 interactome in the GSC nucleus, we provide mechanistic insights into the role of HDAC7 in regulating cancer stemness and possible unprecedented targeting strategies.MethodologyWe quantified the HDAC7 expression in tissue samples from 80 GBM patients admitted at Rhode Island Hospital using Custom Nanostring Panels and correlated the HDAC7 expression to overall patient survival. Also, we compared HDAC7 mRNA expression in GBM to the non‐tumor samples from TCGA data using GlioVis webtool. Next, we performed siRNA knock‐down (KD) of HDAC7 in GSCs followed by RNA‐seq and transcriptomic, gene enrichment and stemness enrichment analysis. We used mass spectrometry (Mod Spec) in si‐HDAC7 KD versus Control to quantify the potential enzymatic de‐acetylation properties of HDAC7 on the global post‐translational modifications of histones. Finally, to unravel the protein partners for HDAC7 in the GSC nucleus, we performed Rapid immunoprecipitation mass spectrometry of endogenous protein (RIME).ResultsSurvival analysis of GBM patients showed significant decrease in survival for patients with high HDAC7 mRNA expression. In addition, GlioVis webtool revealed that HDAC7 expression is significantly higher in GBM tumors relative to non‐tumor samples. Differential gene expression analysis of RNASeq performed on HDAC7 siRNA KD GSCs versus control, revealed 4963 differentially expressed genes with the Gene Ontology enrichment showing significant enrichment for cell cycle and cell division suppression in HDAC7 KD GSCs. Additionally, 653 genes that were downregulated in HDAC7 KD GSCS were found to play significant role in stemness processes of embryonic stem cells and embryonal carcinoma stem cells. Mass spectrometry on GSCs following HDAC7 siRNA KD, revealed non‐significant de‐acetylation changes on global histone marks suggesting that HDAC7 has minimal or no histone deacetylation activity. Finally, RIME revealed novel protein partners in the proximity of 2.5 A° to HDAC7 in the GSCs nucleus. The newly discovered protein complex of HDAC7 suggests that HDAC7 regulates heterochromatin formation and spreading, which mechanistically control transcriptional programs of GSCs.ConclusionOur data suggest that HDAC7 plays a pivotal role in GBM. It regulates expression of transcripts that define cancer stemness, while inhibition of HDAC7 results in global increase in heterochromatin and transcriptional repression in GSCs. This study supports HDAC7 as a novel druggable target for GBM in a preclinical setting.