Increased TGF-β1-mediated suppression of growth and motility in castrate-resistant prostate cancer cells is consistent with Smad2/3 signaling

BACKGROUND Elevated TGF-beta levels are associated with prostate cancer progression. Although TGF-beta is a tumor suppressor for normal epithelial and early-stage cancer cells, it may act paradoxically as a tumor promoter in more advanced cancers, although its effects are largely cell and context dependent. This study analyzed prostate cancer responses to TGF-beta signaling in an isogenic model of androgen-sensitive and castration-resistant prostate cancer cells. METHODS Phosphorylation and nuclear translocation of Smad2 and Smad3 were analyzed using immunoblotting. Proliferation and cell cycle responses to TGF-beta 1 (5?ng/ml) were assessed using growth assays and flow cytometry for DNA content, as well as Western blot and immunoprecipitation of cell cycle proteins. RESULTS both androgen-sensitive (LNCaP) and castration-resistant (C4-2 and C4-2b) prostate cancer cell lines demonstrated TGF-beta 1-induced phosphorylation and nuclear translocation of Smad2/3 that was robust in metastatic lines. Smad phosphorylation was completely abrogated with inhibition of ALK-5 kinase activity using the kinase inhibitor, Sb-431542. Increased sensitivity to TGF-beta 1-mediated growth inhibition was observed in C4-2 and C4-2b cells, as compared to LNCaP cells. This was paralleled with downregulation of Cyclin D and increased association of p15(Ink4b) or p27(Kip) with CDK's. Additionally, TGF-beta 1 inhibited motility and invasion of metastatic cell lines. CONCLUSIONS TGF-beta-mediated suppression of growth and motility is enhanced in metastatic, castration-resistant prostate cancer cells. Enhanced TGF-beta 1-induced Smad2 and -3 signaling in prostate cancer cells may correlate with tumor suppressive activity. Therefore, the direct effects of TGF-beta 1 on prostate cancer cells post-castration may be anti-tumorigenic and growth-suppressive. Prostate 72:13391350, 2012. (c) 2012 Wiley Periodicals, Inc.