Abstract 1447: Retinoic acid-induced 2 driven polymerization of c-terminal binding proteins sustains epithelial differentiation in castration-resistant prostate cancer

Abstract

Abstract Previously, we found that increased Retinoic acid-induced 2 (RAI2) protein concentration in primary tumors predicts early biochemical relapse of prostate cancer patients and described a co-repressor function of RAI2 related to androgen receptor activity (1). The presence of a non-canonical tandem ALDLS motif on RAI2 protein prompted the present investigation on a possible role of a dual interaction of RAI2 with C-terminal binding proteins (CtBPs) in relation to prostate cancer progression. We analyzed RAI2 gene expression as well as CtBP1 protein localization and constitution in circulating tumors cells (CTCs) isolated from metastatic prostate cancer patients by semi-quantitative PCR analysis and immune-fluorescence staining. We employed an integrated structural biology approach combining biophysical techniques like isothermal calorimetry, circular dichroism, size exclusion chromatography with high-resolution structural biology techniques including X-ray crystallography and single particle cryo-electron microscopy. We established RAI2-depleted VCaP cells and used this model for validation of the obtained results. RAI2 gene expression is significantly increased in CTCs isolated from patients with castration-resistant disease in comparison to CTCs isolated from patients with hormone-sensitive, aggressive or neuroendocrine variant prostate cancer. We detected CtBP1 protein foci in the nuclei of individual CTCs. We show that RAI2 induces CtBP polymerization. Using single particle cryo-electron microscopy, we obtained crucial insights into the structural organization of this filament. RAI2 inactivation in VCaP cells caused a significant reduction of both CtBP proteins, which is accompanied by the disappearance of CtBP protein aggregates in nuclei. In addition, we observed the induction of neuroendocrine features, marked by the induction of Synaptophysin and Chromogranin A, a higher nucleus to cytoplasm ratio and smaller cell size of RAI2-depleted cells. In summary, we report tandem motifs-induced polymerization of CtBP’s for the first time. We found that elevated RAI2 gene expression is a characteristic of castration-resistant prostate cancer. Because RAI2 depletion resulted in loss of CtBP aggregates and the induction of neuroendocrine traits, we conclude that RAI2-driven CtBP polymerization sustains epithelial differentiation in castration-resistant prostate cancer. These findings holds potential for the diagnosis of castration-resistant disease and inhibiting the progression from castration-resistant to neuroendocrine prostate cancer. 1. Besler K, Weglarz A, Keller L, von Amsberg G, Bednarz-Knoll N, Offermann A, et al. Expression Patterns and Corepressor Function of Retinoic Acid-induced 2 in Prostate Cancer. Clin Chem 2022;68:973-83 Citation Format: Stefan Werner, Nishit Goradia, Edukondalu Mullapudi, Lina Merkens, Sarah Greimeier, Aleksandra Węglarz, Gunhild von Amsberg, Harriet Wikman, Klaus Pantel, Matthias Wilmanns. Retinoic acid-induced 2 driven polymerization of c-terminal binding proteins sustains epithelial differentiation in castration-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 1447.