Abstract 5429: Inhibition of LSD1 and HDACs causes synergistic cell death in glioblastoma cells

Abstract

Abstract Glioblastoma multiforme is a particularly aggressive brain tumor and remains a clinically devastating disease. More than 90% of children diagnosed with glioblastoma die within 2 years making it among the leading causes of cancer related morbidity and mortality. Despite innovative therapies for the treatment of glioblastoma, there has not been a significant increase in patient survival over the past decade. Enzymes that control epigenetic alterations have become popular targets for cancer therapy due to their ability to control cellular processes that lead to oncogenesis. Several inhibitors of histone deacetylases (HDACi) have been developed and are in clinical use for cancer therapy. Histone methylation can be modulated by HDACi and here we propose to examine the effect of dually inhibiting deacetylases and demethylases. Specifically, we focused on LSD1, lysine specific demethylase 1, a demethylase known to be in complex with HDAC1/2 and implicated in several high risk cancers. We evaluated apoptosis induction upon knockdown of LSD1 in combination with HDACi in glioblastoma cell lines. Our results demonstrate a greater than two-fold increase in apoptosis in LSD1 knockdown cells treated with increasing doses (1-5 μM) of HDACi for 48 hours as compared to control as measured by DNA fragmentation by propidium iodide staining and analysis by flow cytometry. Furthermore, pretreatment of LSD1 knockdown cells with the general caspase inhibitor zVAD-fmk prior to treatment with vorinostat only partially protects against DNA fragmentation suggesting that cell death proceeds through both caspase-dependent and -independent mechanisms. Similar results were obtained by pharmacologically inhibiting LSD1 with the monoamine oxidase inhibitor tranylcypromine. Treatment of LN-18 cells with combinations of tranylcypromine (100-1000 μM) and vorinostat (1-5 μM) for 72 hours demonstrate that these two agents synergize to promote cell death with 1000 μM tranylcypromine and 5 μM vorinostat showing the most synergistic effect (CI 0.313). Surprisingly, the increase in apoptosis observed upon LSD1 inhibition, by knockdown and pharmacological inhibitors, and HDACi also correlates with an increase in intracellular superoxide as measured by hydroethidum staining and analysis by flow cytometry. These data suggest that reactive oxygen species may play a role in the apoptosis induced by the combination of LSD1 and HDAC inhibition. Further studies aimed at understanding the mechanism by which LSD1 and HDAC inhibition leads to enhanced tumor cell death will substantially increase our knowledge of how epigenetic modifiers interact to control cell survival and may suggest novel options for cancer therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5429.