Downmodulation of Bcl-2 sensitizes metastatic LNCaP-LN3 cells to undergo apoptosis via the intrinsic pathway

• Published in: The Prostate Vol. 70; no. 6; pp. 571 - 583
• Main Authors: Song, Renduo, Harris, Loleta D., Pettaway, Curtis A.
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 01.05.2010
• Subjects: Adenocarcinoma - metabolism; Adenocarcinoma - pathology; androgen resistance; anti-Bcl-2 therapy; Antineoplastic Agents - pharmacology; apoptosis; Apoptosis - physiology; Bcl-2; Caspases - metabolism; Cell Line, Tumor; Cell Proliferation; Cytochromes c - metabolism; Down-Regulation - physiology; Humans; Male; Neoplasm Metastasis; Poly(ADP-ribose) Polymerases - metabolism; prostate cancer; Prostatic Neoplasms - metabolism; Prostatic Neoplasms - pathology; Proto-Oncogene Proteins c-bcl-2 - metabolism; Receptors, Androgen - metabolism; RNA, Small Interfering - pharmacology; Signal Transduction - physiology; Taxoids - pharmacology
• ISSN: 0270-4137; 1097-0045
• DOI: 10.1002/pros.21091

BACKGROUND We explored the mechanisms of apoptosis after Bcl‐2 protein downmodulation in metastatic LNCaP‐LN3 cells (LN3). METHODS LNCaP, LNCaP‐Pro5 (Pro5) and LN3 cells were cultured in 5% charcoal‐stripped serum (CSS) or in R1881 (synthetic androgen) and bicalutamide (synthetic anti‐androgen) and growth inhibition was assessed. Expression levels of androgen receptor (AR) and Bcl‐2 were determined. LN3 cells were transfected with small interfering RNA Bcl‐2 (siRNA Bcl‐2) or control siRNA oligonucleotides. Rates of apoptosis and proliferation were obtained. Cytochrome c localization in treated and control cells was assessed ± cyclosporine A (CsA). Caspases 9, 3, and poly (ADP‐ribose) polymerase cleavage (PARP) were measured upon downmodulation of Bcl‐2; and cell growth inhibition in vitro after Bcl‐2 modulation combined with docetaxel chemotherapy was determined. RESULTS LN3 cells maintained growth under castrate conditions in vitro. AR protein amplification did not explain castrate‐resistant LN3 cell growth. Bcl‐2 protein levels in LN3 cells were significantly higher than in Pro5 cells, and were effectively downmodulated by siRNA Bcl‐2. Subsequently increased apoptosis and decreased proliferation mediated by cytochrome c was noted and this was reversed by CsA. siRNA Bcl‐2‐transfected LN3 cells exhibited elevated levels of caspases 9, 3, and PARP cleavage. Exposure of LN3 cells to docetaxel led to increased apoptosis, and simultaneous downmodulation of Bcl‐2 substantially enhanced this effect. CONCLUSIONS Downmodulation of Bcl‐2 in metastatic castrate‐resistant LNCaP‐LN3 cells led to apoptosis via a cytochrome c‐dependent pathway that was enhanced with docetaxel treatment. Prostate 70: 571–583, 2010. © 2009 Wiley‐Liss, Inc.