Abstract 2082: Extragonadal and regional androgen biosynthesis associated with a common steroidogenic enzyme polymorphism promotes radioresistance in prostate cancer

Abstract

Abstract Purpose: Androgen deprivation therapy (ADT) is the linchpin therapeutic for locally advanced and metastatic PCa. However, resistance to ADT is common and has been associated with a gain of function mutation (A1245C) in the 3β-HSD enzyme, which catalyzes intratumoral DHT synthesis. As androgen signaling is known to upregulate the DNA damage response (DDR), we investigated whether variant HSD3B1 modulates DDR and radiosensitivity in PCa via extragonadal androgen biosynthesis. Methods: We stably knocked down HSD3B1 in LNCaP, C42 and VCaP cell lines (which carry the protein stabilizing variant allele), and overexpressed the variant HSD3B1 allele in LAPC4 (that harbors WT allele, where the protein readily undergoes degradation). We examined the proliferative and clonogenic capacity of these cells in presence and absence of substrate, the extragonadal androgen precursors DHEA, followed by treatment with IR (200,400, 600 and 800 cGy, single fraction). We studied DNA DSB formation and resolution kinetics using phospho- γH2AX labelling and neutral COMET assay. We also measured 3β-HSD dependent changes in expression of DDR response genes pre- and post-radiation Results: Control shRNA transduced prostate cancer cell lines expressing the variant HSD3B1 allele had increased cell proliferation (3.1289-fold, p<0.001) and clonogenic survival (100 x more colonies after 800 cGY single fraction, p&lt 0.001) in the presence of DHEA compared to HSD3B1 knockdown LNCaP cells. We found variant HSD3B1 cell lines were more radioresistant to 400cGY single fraction IR and exhibited more efficient γH2AX foci resolution with significantly low no. of residual foci (1 foci/nucleus, p &lt 0.05) and smaller COMET Tail moment at 24 hrs in a DHEA dependent. We observe increased basal mRNA expression of DDR genes from specific repair networks including non-homologous end joining (PRKDC, XRCC4, XRCC5) and homologous recombination (RAD51, RAD54), but not mismatch repair (MSH2, MSH6), in variant HSD3B1 cells. Transcriptional induction of DDR gene expression following radiation and in presence of DHEA, was significantly more pronounced in HSD3B1 variant cells, suggesting AR establishes a regional chromatin environment which is more permissive to DDR gene transcriptional activation. Conclusion: Increased intracellular HSD3B1, which converts extra gonadal precursors to DHT, leads to increased expression of NHEJ and HR genes, more rapid resolution of γH2AX foci, and radioresistance in prostate cancer. This work has therapeutic implications for combined radiation and androgen directed therapy in localized and low volume metastatic prostate cancer. Prospective validation of personalized treatment strategies combining blockade of extragonadal androgen biosynthesis, conventional ADT, and radiotherapy in prostate cancer are warranted. Citation Format: Shinjini Ganguly, Aysegul Balyimez, Zaeem lone, Aimalie Hardaway, Monaben Patel, Elai Davicioni, Rahul Tendulkar, Eric Klein, Nima Sharifi, Omar Mian. Extragonadal and regional androgen biosynthesis associated with a common steroidogenic enzyme polymorphism promotes radioresistance in prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2082.