Abstract A20: Disruption and restoration of pioneer transcription factor function in bladder cancer

Abstract

Abstract Urothelial bladder cancer (BLCA) is characterized by the failure to complete epithelial-differentiation. The mechanisms underlying this failure are unknown. To address this knowledge gap, we used unbiased analyzes to identify 1,000 genes significantly suppressed in BLCA vs normal bladder (p<0.001, FDR<0.01) (n=426, TCGA RNA-seq). Remarkably, these genes are located in compacted chromatin in embryonic stem cells (ESC) (decreased H3K4me3, increased CpG methylation), while 1,000 genes upregulated in BLCA vs normal bladder are located in open chromatin at this same ultimate baseline. This striking pattern in unbiased analyses suggested pioneer transcription factor (TF) loss of function in BLCA: pioneer TF access compacted chromatin to initiate the remodeling necessary for gene activation. The pioneer TF hub containing FOXA1, GATA4 is highly expressed in normal bladder, and these pioneer TFs are frequently haploinsufficient in BLCA, 32% and 56%, respectively. We pulled down endogenous FOXA1, the most expressed TF, from UM-UC-6 BLCA cells. Proteomic analyzes by mass spectrometry revealed enrichment for corepressors (DNMT1) over coactivators (ARID1A) in this pioneer TF hub. Genetic analyses showed frequent mutation/deletion of the coactivators: 43% bi-allelic ARID1A inactivation, while corepressors, e.g., CHD4 and DNMT1, were amplified or unaltered. Restoration of ARID1A into UM-UC-6 upregulated epithelial genes, downregulated MYC, upregulated CDKN1B, and terminated proliferation. To translate into therapy, we confirmed using immunoprecipitation LCMS/MS that DNMT1-depletion using noncytotoxic concentrations of the clinical drug decitabine (0.2-0.5 μM) shifted hub composition to coactivators, to thereby induce terminal epithelial-differentiation of BLCA cells. This was also seen in vivo in UM-UC-6 xenograft: very low dose non-cytotoxic decitabine (0.1 mg/kg) combined with an inhibitor of its degradation, THU, decreased tumor size more than standard high-dose cytotoxic gemcitabine/cisplatinum. Thus, BLCA suppresses hundreds of epithelial-differentiation genes via corepressor/coactivator imbalance in the bladder pioneer TF hub. Sensitivity of the pioneer TF hub to corepressor/coactivator stoichiometry, however, also enables pharmacologic reduction of corepressors to compensate for genetic reduction of coactivators, to thereby release BLCA cells to destined terminal epithelial fates. Citation Format: Caroline Schuerger, Xiaorong Gu, Yvonne Parker, Daniel Lindner, Babal Jha. Disruption and restoration of pioneer transcription factor function in bladder cancer [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2019 May 18-21; Denver, CO. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(15_Suppl):Abstract nr A20.