Krüppel-like factor 9 and histone deacetylase inhibitors synergistically induce cell death in glioblastoma stem-like cells

Abstract

Background: The dismal prognosis of patients with glioblastoma (GBM) is attributed to a rare subset of cancer stem cells that display characteristics of tumor initiation, growth, and resistance to aggressive treatment involving chemotherapy and concomitant radiation. Recent research on the substantial role of epigenetic mechanisms in the pathogenesis of cancers has prompted the investigation of the enzymatic modifications of histone proteins for therapeutic drug targeting. In this work, we have examined the function of Krüppel-like factor 9 (KLF9), a transcription factor, in chemotherapy sensitization to histone deacetylase inhibitors (HDAC inhibitors). Methods: Since GBM neurosphere cultures from patient-derived gliomas are enriched for GBM stem-like cells (GSCs) and form highly invasive and proliferative xenografts that recapitulate the features demonstrated in human patients diagnosed with GBM, we established inducible KLF9 expression systems in these GBM neurosphere cells and investigated cell death in the presence of epigenetic modulators such as histone deacetylase (HDAC) inhibitors. Results: We demonstrated that KLF9 expression combined with HDAC inhibitor panobinostat (LBH589) dramatically induced glioma stem cell death via both apoptosis and necroptosis in a synergistic manner. The combination of KLF9 expression and LBH589 treatment affected cell cycle by substantially decreasing the percentage of cells at S-phase. This phenomenon is further corroborated by the upregulation of cell cycle inhibitors p21 and p27. Further, we determined that KLF9 and LBH589 regulated the expression of pro- and anti- apoptotic proteins, suggesting a mechanism that involves the caspase-dependent apoptotic pathway. In addition, we demonstrated that apoptosis and necrosis inhibitors conferred minimal protective effects against cell death, while inhibitors of the necroptosis pathway significantly blocked cell death. Conclusions: Our findings suggest a detailed understanding of how KLF9 expression in cancer cells with epigenetic modulators like HDAC inhibitors may promote synergistic cell death through a mechanism involving both apoptosis and necroptosis that will benefit novel combinatory antitumor strategies to treat malignant brain tumors.