Abstract B033: Characterizing the global function of NCOR2 in prostate cancer cells and its contribution to PCa progression
Authors: Mark D. Long, Prashant K. Singh, Gerard Llimos, Spencer Rosario, Dominic J. Smiraglia, Moray J. Campbell
Journal: Cancer Research (2018)

Abstract The nuclear receptor corepressors (NCORs) serve as critical mediators between transcription factors and epigenetic regulators, and thus their functions are critical in the maintenance of epigenetic and transcriptional states. As such, NCORs have important developmental and homeostatic roles, and disruptions of their functions are implicated in various cancers. In hormonally driven cancers, such as breast and prostate cancer (PCa), NCORs are implicated in de novo and acquired changes to steroid hormone signaling and therapeutic resistance. Supporting the importance of NCOR function in the prostate, inactivating genomic aberrations to NCORs have been identified in androgen deprivation therapy (ADT) resistant disease. However, to date, the global functions of NCORs in prostate cancer cells and their overall contributions to PCa progression remain enigmatic. To characterize the global functions of NCOR2/SMRT in prostate cancer we have undertaken efforts to map the NCOR2 dependent transcriptome (RNA-seq), miRnome (miRNA-seq), methylome (EPIC methylation array) and cistrome (ChIP-seq) in both androgen-sensitive (LNCaP) and ADT-resistant (LNCaP-C42) prostate cancer cells upon shRNA targeted knockdown. NCOR2 loss in LNCaP-C42 cells revealed a striking degree of global hypermethylation (87,078 CpGs &amp;gt; 10% gain, adj.pval &amp;lt; 0.01), while relatively little hypomethylation was observed (1,742 CpGs &amp;gt; 10% loss). Hypermethylation was largely exclusive of TSS loci and CpG islands, occurring at distal regions. Conversely, the NCOR2 associated transcriptome was more divergent in its expression changes by comparable criteria (1,491 upregulated, 1,195 downregulated). Similar patterns were observed in LNCaP cells, albeit to a lesser extent, suggesting broader regulatory roles in ADT resistant cells. NCOR2 knockdown deregulated a number of miRNA including let-7e and miR-200a with target binding motifs significantly enriched among the NCOR2 regulated transcriptome, and several of these miRNA stratified survival in PCa patient data. Furthermore, NCOR2 binding at candidate loci distinguished normal from malignant prostate cells, and global binding profiles revealed significant overlap with androgen receptor and pioneering factor FOXA1. To examine the effect of NCOR2 loss in the context of androgen signaling, similar experimental conditions were performed in the presence of dihydrotestosterone (DHT). NCOR2 control of androgen sensitivity was complex, resulting in amplified signaling of some classic AR target genes (KLK3, HERC3) while dampening others (TMPRSS2, TIPARP). Ongoing data integration efforts will dissect novel NCOR2 methylome-cistrome-transcriptome relationships within and across models to define target gene networks that are epigenetically governed through NCOR2 function. To determine whether NCOR2 loss alters PCa progression we have utilized stable introduction of NCOR2 targeting shRNA in the CWR22 xenograft model that simulates androgen-dependent primary growth, regression upon ADT, and subsequent recurrence at established proportions. NCOR2 loss had no effect on primary tumor growth rate or size of tumor at ADT. However, shNCOR2 expressing tumors had significantly reduced regression in response to ADT, and furthermore at 180 days post injection, a significantly higher proportion of shNCOR2 expressing tumors (17/49) had reached recurrence compared to shCTL tumors (7/46). In total, these findings place NCOR2 as an important epigenetic regulator in prostate cancer cells, the dysfunction of which contributes to PCa progression. Citation Format: Mark D. Long, Prashant K. Singh, Gerard Llimos, Spencer Rosario, Dominic J. Smiraglia, Moray J. Campbell. Characterizing the global function of NCOR2 in prostate cancer cells and its contribution to PCa progression [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr B033.