Involvement of p21 and FasL in Induction of Cell Cycle Arrest and Apoptosis by Neochamaejasmin A in Human Prostate LNCaP Cancer Cells

Neochamaejasmin A (1), a biflavonoid isolated from the roots of a traditional Chinese medicine, Stellera chamaejasme L., was shown to inhibit cellular 3H-thymidine incorporation (IC50 12.5 µg/mL) and subsequent proliferation of human prostate cancer LNCaP cells. Treatment of LNCaP cells with low doses of 1 (≤6.25 µg/mL) suppressed DNA-binding activities of the transcription factors NFκB and AP-1 to the promoter of cyclin D and also inhibited expression of the cell cycle regulatory proteins cyclin D, proliferating cell nuclear antigen, and nucleolin, thus arresting cells in G1 phase of the cell cycle. A lengthy exposure with higher doses of 1 (≥12.5 µg/mL) revealed the production of reactive oxygen species, dissipation of the mitochondrial membrane potential, up-regulation of cyclin-dependent kinase inhibitor p21, and induction of cell apoptosis. An aggregation of Fas−procaspase 8−procaspase 3 and p21−procaspase 3 proteins by coimmunoprecipitation, immunoblotting analysis, and MALDI-mass spectrometry indicated the involvement of Fas and p21 in 1-mediated cytotoxicity, and pretreatment of cells with antisense FasL oligonucleotides partially abolished apoptosis. Thus, 1 blocked cell cycle progression at the G1 phase by activating the p21 protein and ultimately promoting the Fas−caspase 8−caspase 3 apoptotic machinery.