Abstract 4316: Disruption and restoration of pioneer transcription factor hub function in bladder cancer
Authors: Schuerger Caroline , Saunthararajah Yogen 
Published: August 2018

Urothelial bladder cancer is characterized by the failure to complete transitional epithelial-differentiation. The mechanisms underlying this failure are unknown. To address this gap in knowledge, we analyzed 2000 genes differentially expressed between normal bladder vs. bladder cancer (BLCA) (n=426, TCGA RNA sequencing). Remarkably, genes suppressed in BLCA are found in embryonic stem cells (ESC) in compacted chromatin (decreased H3K4me3, increased CpG methylation), while upregulated genes in BLCA are found in ESCs in open chromatin (H1 ESC Encode).This striking property of genes suppressed in BLCA suggested loss-of-function of pioneer transcription factor (TF) function in BLCA, as pioneer TFs access compacted chromatin to initiate the necessary remodeling for subsequent gene activation. The pioneer master TF hub containing FOXA1, GATA4/6 is highly expressed in normal bladder, and these master TFs were frequently haploinsufficient in BLCA, 32%, 56%, and 24% respectively. To better understand why target genes of FOXA1, the most expressed bladder master TF, are unremodeled and repressed, we pulled down endogenous FOXA1 from UM-UC-6 and -3 BLCA cells and used mass spectrometry and Western blot to analyze interacting coregulators, which execute the biochemical work of chromatin remodeling. These analyses revealed enrichment for corepressors, e.g., DNMT1, in the FOXA1 interactome over coactivators, e.g., ARID1A. Genetic analysis of BLCA show frequent mutation/deletion of coactivators: 43% bi-allelic ARID1A inactivation, while corepressors SMARCA5 and DNMT1 were amplified or unaltered to remain functional. Restoration of ARID1A into ARID1A haploinsufficient BLCA (UC-6) upregulated epithelial-differentiation genes and induced terminal epithelial-differentiation, downregulation of MYC and upregulation of p27 protein. To translate into therapy, we restored coactivator/corepressor balance by pharmacologic corepressor depletion: we used the clinical drug decitabine to deplete DNMT1, to compensate for genetic coactivator inactivation. Decitabine recapitulated the effects of ARID1A introduction, inducing terminal epithelial-differentiation of BLCA cells. This was also seen in vivo in a xenograft model of BLCA (UC-6). Decitabine combined with an inhibitor of its in vivo degradation, tetrahydrouridine, decreased tumor size significantly with vehicle tumor size 917.8 mm3 to treated tumor size 321.0 mm3 (p&lt;0.01). In sum, genetic alterations in BLCA alter composition of a pioneer master TF hub to remove remodeling needed for transitional epithelial-differentiation. Dynamic coactivator/corepressor opposition in gene regulation, however, enables pharmacologic reduction of corepressors to compensate for genetic reduction of coactivators.

Citation Format: Caroline Schuerger, Yogen Saunthararajah. Disruption and restoration of pioneer transcription factor hub function in bladder 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 4316.