Abstract 4815: Efficacy and toxicity of combinatorial therapy with EZH2 and androgen receptor inhibitor for castration-resistant prostate cancer
Authors: Shankar Eswar , Franco Daniel , Iqbal Omair , Moreton Stephen , Kanwal Rajnee , Gupta Sanjay 
Published: August 2018

Abstract
Enhancer of zeste homolog 2 (EZH2), the enzymatic member of the polycomb repressor complex PRC2 has emerged as a key activator of androgen receptor in castration-resistant prostate cancer (CRPC). Androgen deprivation therapy (ADT) is a main therapeutic modality for advance-stage metastatic prostate cancer targeting AR. Recent studies shed light on the fact that overabundance of EZH2 in localized cancers increases aggressiveness and their recurrence. EZH2 activates AR through methylation-mediated gene suppression. Though prostate cancers respond to ADT treatment initially they subsequently re-emerge as CRPC. In recent years, Enzalutamide (ENZU), a second generation AR antagonist exhibits survival advantage in CRPC patients, but show relapse within a year, activating AR in these tumors. Recent genomic studies reveal AR-regulated genes contribute to CRPC emergence. This apparent association between EZH2 and AR in activating target genes by cooperative recruitment may lead to the development of chemoresistance. Therefore we rationalize that combination treatment targeting EZH2 and AR could be efficacious and significantly inhibit proliferation and metastasis of CRPC cells. Human prostate cancer C4-2B and 22Rv1, representative of CRPC tumors, were treated with EZH2 inhibitor GSK126 and AR antagonist (ENZU) individually and in combination, followed by assessment of cell viability, cell cycle analysis, migration, invasion and expression of various target genes by Western blotting. ELISA was employed to assess the change in activity of histone methylation in these cells. Treatment of C4-2B and 22Rv1 cells individually with GSK126 and ENZU (2.5-80 µM) for 24 h exhibited a partial suppressive effect in cell growth. Using GSK126 and ENZU combination at 1:1, 1:5, 1:10 and 1:20 micro-molar ratio exhibited increased cell growth inhibition, where 1:10 ratio showed synergistic effect in inhibiting cell growth in both cell lines. This combination caused marked increase in G0/G1-phase cell cycle arrest followed by cell death, inhibition of migration and invasion in both cell lines. Furthermore, combination treatment led to significant reduction in the protein expression of AR, AR-V7, EZH2, SUZ12, EED, p-Akt (Ser473) in both cell lines, compared to individual treatments. Combined treatment also caused significant decrease in the levels of H3K27 methylation and its activity. Taken together, our results demonstrate that simultaneous inhibition of AR and EZH2 is more efficacious in inhibiting the growth of CRPC cells. This opens new possibilities of uncovering a novel path of treating CRPC by simultaneously targeting EZH2 and AR using a combinatorial approach.
Citation Format: Eswar Shankar, Daniel Franco, Omair Iqbal, Stephen Moreton, Rajnee Kanwal, Sanjay Gupta. Efficacy and toxicity of combinatorial therapy with EZH2 and androgen receptor inhibitor for castration-resistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4815.