Abstract 4619: Prolonged inhibition of androgen receptor signaling induces expression of Nuclear ErbB3 which renders prostate cancer cells susceptible to targeted inhibitors

Abstract Objective: Nuclear expression of the receptor tyrosine kinase (RTK) ErbB3/HER3 increases in highly aggressive prostate cancer cells, but its nuclear transport mechanism is currently unknown. Prostate tumors rely on the androgen receptor (AR), whose activation transcriptionally controls ErbB3 expression, but its role in ErbB3 nuclear localization had not previously been reported. Here, we investigated the mechanism by which subcellular localization of ErbB3 was altered in prostate cancer. Methods: ErbB3 localization was investigated in the human prostate cancer tumor progression model LNCaP, C4, C4-2 and C4-2B. ErbB3 was stimulated with heregulin-1β (HRG). Nuclear translocation was tested with a nucleocytoplasmic transport inhibitor panel (chlorpromazine, filipin III, amiloride and Leptomycin B) LNCaP and C4 cells were continuously cultured with the AR activation inhibitor abiraterone acetate (AbiAc) or ethanol (VEH) or treated with the AR inhibitors enzalutamide, darolutamide and apalutamide. ErbB3 activity and subcellular localization were analyzed using confocal microscopy/subcellular fractionation/immunoblot. Cell viability was determined by MTT assay. Proliferation and apoptosis were determined by flow cytometry. Invasive potential was investigated by crystal violet staining of cell colonies. ErbB3 expression was reduced by siRNA technology. Results: The ratio of nuclear to cytoplasmic ErbB3 increased in untreated cells from LNCaP<C4-2<C4<C4-2B. In all four lines, nuclear localization of ErbB3 peaked at 30 minutes after HRG treatment, with a rapid return to the cytoplasm in LNCaP cells (at 1 hour), slower in C4 cells (at 4 hours), and continued accumulation in the nucleus in C4-2 and C4-2B 8 hours following HRG treatment. Treatment with the transport inhibitor panel showed that nuclear accumulation of ErbB3 was prevented by the clathrin-dependent endocytosis inhibitor Chlorpromazine (CPZ) but not by the others. In LN-VEH and C4-VEH cells, HRG induced nuclear translocation of ErbB3 but not in LN-AbiAc or C4-AbiAc cells. Accordingly, HRG increased ErbB3 phosphorylation at Y1328 and nuclear Akt phosphorylation at S473 in VEH but not AbiAc cells. C4 cells, which had high baseline nuclear ErbB3, were more sensitive to the AR inhibitors and showed decreased viability and invasive potential compared to LNCaP cells. Conclusions: These results indicate that (1) ErbB3 nuclear localization required clathrin-dependent endocytosis (2) ligand binding of ErbB3 as well as the presence of an active AR is necessary for ErbB3 nuclear localization and (3) the presence of nuclear ErbB3 increases sensitivity to AR inhibitors. Citation Format: Maitreyee K. Jathal, Maria M. Mudryj, Paramita M. Ghosh. Prolonged inhibition of androgen receptor signaling induces expression of Nuclear ErbB3 which renders prostate cancer cells susceptible to targeted inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4619.