Abstract 275: Apalutamide inhibits the expression of regulatory genes for prostate cancer cell invasion and migration in LNCaP prostate cancer cells and Hi-Myc mouse prostate

Abstract The next generation antiandrogen, apalutamide (APA), improves both overall survival and metastasis-free survival in men with castration-resistant prostate cancer (CRPC). To investigate mechanisms of APA action in CRPC, in vitro and in vivo studies were performed to characterize the effects of APA on prostate cancer cell proliferation, invasion, and migration, and the expression of genes that regulate these processes. Exposure to 40 or 80 nM concentrations of APA for 24 hr inhibited the proliferation of LNCaP cells in both the presence and absence of dihydrotestosterone (DHT). APA also inhibited LNCaP cell migration/invasion in the Boyden chamber assay. At the mRNA level (RT-PCR), APA down-regulated the expression of androgen receptor (AR), c-Myc, MMP-2, MMP-9, DANCR, and incRNA, and up-regulated TIMP-2 expression. Similar data were obtained for protein expression (western blot): APA down-regulated expression of AR, c-Myc, PCNA, MMP-2 and MMP-9, and up-regulated expression of TIMP-2. In the in vivo study, heterozygous male Hi-Myc mice received daily oral (gavage) administration of APA at doses of 0 (vehicle), 2, or 15 mg/kg/day beginning at age 8 weeks. Cohorts of mice were euthanized for prostate collection after 2 months, 3.5 months, or 5 months of APA administration. Daily oral administration of both dose levels of APA reduced accessory sex gland weights by over 50% at all three time points. Confirming and extending the results of in vitro studies in LNCaP cells, RT-PCR demonstrated that APA inhibited the expression of AR, c-Myc, lncRNA, DANCR, MMP-2, and MMP-9 in the prostate of Hi-Myc mice, and upregulated the expression of TIMP-2. The maximum efficacy of APA was seen at 3.5 months of exposure. Because resistance to antiandrogen therapy commonly develops over time in clinical populations, a subline of LNCaP cells that is resistant to APA was derived and used to repeat in vitro cell proliferation, migration, invasion, and gene expression assays. The APA-resistant subline no longer expressed AR, but did express the AR variant, AR-V7. To determine whether cells can be resensitized to APA by inhibiting AR-V7 expression, AR-V7 was knocked down by siRNA. Comparisons of the effects of APA on gene expression in APA-resistant LNCaP cells and APA-resistant LNCaP cells in which AR-V7 had been knocked down demonstrated that knockdown of AR-V7 in APA-resistant LNCaP cells resensitizes those cells to APA. Although APA is an effective inhibitor of prostate cancer growth, development, and invasion in both preclinical models and clinical populations, strategies to overcome the apparent AR-V7-mediated resistance to antiandrogens are necessary to optimize its efficacy when administered over extended periods. Citation Format: Gina Qualter, Genoveva Murillo, Xinjian Peng, David McCormick. Apalutamide inhibits the expression of regulatory genes for prostate cancer cell invasion and migration in LNCaP prostate cancer cells and Hi-Myc mouse prostate [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 275.