Abstract 5182: Chromatin reader machinery as target for overcoming resistance to DNA-demethylating epi-drug decitabine

Abstract

Based on the current understanding of the epigenetic landscape, cancer methylome is highly disrupted, which makes DNA methylation an excellent target for anti-cancer cures. To date, azacytidine and its congener, decitabine (DAC) are the most successful DNA demethylating epigenetic drugs. However, scientists are yet to find the reversal of clinical resistance to these drugs. Yet another challenge remains to be the decreased efficacy of these promising therapies in the cure of solid tumors. We used a soild tumor, HCT116 colorectal cancer cell line and developed resistance against DAC. DAC-resistant HCT116 cells were used to study the epigenetic cross-talk between DNA methylation and chromatin modifications. The screening of parental and DAC-resistant HCT116 cells against inhibitors of epigenetic writers-erasers-readers unveiled increased sensitivity of resistant cells to BET inhibitor (inhibitor of reader enzymes), (+)-JQ1. BET inhibitor further mediated augmented response on cell cycle phases of resistant cells, increased anti-proliferative effects in xenograft models of resistant cells, and synergystic effects in combination with DAC in HCT116 parental cells, both in vitro and in vivo . We then sequenced the transcriptome of DAC-sensitive and -resistant HCT116 cells, and (+)-JQ1 treated-(DAC-resistant) cells, using RNA-seq. The RNA-seq data revealed the overexpression of critical oncogenes, and their binding inactivation of key tumor suppressor genes (TSGs) in resistant cells. The most significant and biologically relevant transcriptional changes were further validated by qRT-PCR, and in addition their methylation status was determined by bisulphite sequencing. We discovered that the expressions of down-regulated TSGs were driven by promoter methylation, exposing these tumor-suppressive signatures as biomarkers which might differentiate between DAC-resistance and sensitivity, whereas, overexpression of oncogenes was independent of promoter methylation. Interestingly, the expressions of up-regulated oncogenes which define cell identity, mainly those involved in signaling of inflammatory pathways (including some bromodomain-specific genes) were reversed on treatment with (+)-JQ1. Further, siRNA-mediated genetic inhibition of bromodomains in resistant cells phenocopied therapeutic inhibition by (+)-JQ1. These data unveil the chromatin “reader proteins”, as regulators of dysregulated oncogenic expressions in DAC-resistant cells. The present study provides novel insights into the epigenomic landscape of DAC-resistant colorectal cancer cells, and put forward, the alternative therapeutic regimen for DAC-resistant patients. This study was supported by Czech Ministry of Education, Youth and Sports (LO1304, LM2015091, LM2015064), Technology Agency of the Czech Republic (TE01020028) and Internal Grant Agency of Palacky University (IGA_LF_2016_019). Citation Format: Khushboo Agrawal, Petr Vojta, Rastislav Slavkosky, Ivo Frydrych, Petr Dzubak, Marian Hajduch. Chromatin reader machinery as target for overcoming resistance to DNA-demethylating epi-drug decitabine [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5182.