Abstract 1445: TBX2 acts as a molecular switch to downregulate androgen receptor and upregulate glucocorticoid receptor signaling in castrate resistant prostate cancer

Abstract

Abstract Background: A major obstacle in the treatment of metastatic castrate resistant prostate cancer (mCRPC) is acquired resistance to androgen deprivation therapy (ADT). It is now recognized that ADT, particularly the 2nd generation androgen receptor (AR) antagonists, such as enzalutamide, orchestrate plasticity changes/molecular alterations leading to therapy resistance. It is hypothesized in the field that the emergence of castrate resistance to 2nd generation ADT is driven by a switch/bypass from AR signaling to the glucocorticoid receptor (GR) signaling. Therefore, identifying the molecular mechanisms that concurrently drive the loss of AR and gain of GR signaling may be crucial in devising novel and effective therapeutic modalities against mCRPC. We have previously reported that TBX2, a T-box transcription factor (TF) with both repressor and activator functions, is over-expressed in CRPC and that TBX2 drives PCa bone metastatic progression. In agreement with our findings, a recent report showed that TBX2 is a key TF that drives plasticity associated with CRPC. In this study, we investigated the molecular mechanisms by which TBX2 drives plasticity associated with enzalutamide resistance. Methods: We genetically modulated TBX2 using the dominant negative (DN), sh-RNA, and overexpression (OE) approaches. RNA-seq was performed, and qRT-PCR, Western blot and immunohistochemical (IHC) analyses were used for validation. Further, we performed chromatin immunoprecipitation (ChIP) and site directed mutagenesis (SDM). Results: Using publicly available databases, we observed a negative correlation between TBX2 and AR; and a positive correlation between TBX2 and GR. Strikingly, blocking TBX2 expression in wildtype PC3 human PCa cells that do not express AR mRNA or protein led to a marked elevation of AR signature as assessed by RNA-seq analysis. These results were confirmed/validated at the protein level using additional human PCa cell lines. Further, these results were validated using all three approaches to genetically modulate TBX2, i.e. DN, shRNA, and OE. Further, ChIP and SDM analyses confirmed that TBX2 directly binds and transcriptionally represses AR. Conversely, genetic modulation of TBX2 was positively associated with GR expression, and TBX2 was found to directly bind to the GR promoter. Further, we found that knock-down of GR in LNCaP cells overexpressing TBX2 (LNCaPTBX2OE) reversed enzalutamide resistance that is associated with elevated TBX2 expression. Conclusions: Our studies suggest that TBX2 acts as the molecular switch that on one hand represses AR and on the other hand activates GR. Further our study paves the way for potential therapeutic strategies against the AR/GR switch/bypass in CRPC wherein AR and GR signaling could be co-inhibited through targeting TBX2. Citation Format: Sayanika Dutta, Girijesh Patel, Hamed Khedmatgozar, Daniel Latour, Manisha Tripathi, Srinivas Nandana. TBX2 acts as a molecular switch to downregulate androgen receptor and upregulate glucocorticoid receptor signaling in castrate resistant prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1445.