Downmodulation of Bcl-2 sensitizes metastatic LNCaP-LN3 cells to undergo apoptosis via the intrinsic pathway.
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.
Song
Renduo
R
Department of Urology, University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
Harris
Loleta D
LD
Pettaway
Curtis A
CA
D000230:Adenocarcinoma / Q000378:metabolism / Q000473:pathology; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D045304:Cytochromes c / Q000378:metabolism; D000077143:Docetaxel; D015536:Down-Regulation / Q000502:physiology*; D006801:Humans; D008297:Male; D009362:Neoplasm Metastasis; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000378:metabolism* / Q000473:pathology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D011944:Receptors, Androgen / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D043823:Taxoids / Q000494:pharmacology
D000970:Antineoplastic Agents; D019253:Proto-Oncogene Proteins c-bcl-2; D034741:RNA, Small Interfering; D011944:Receptors, Androgen; D043823:Taxoids; D000077143:Docetaxel; D045304:Cytochromes c; D011065:Poly(ADP-ribose) Polymerases; D020169:Caspases