C5 extract induces apoptosis in B16F10 murine melanoma cells through extrinsic and intrinsic apoptotic pathways and Sub-G1 phase arrest

Abstract

Purpose: To investigate the anti-cancer activities of C5 extract (C5E), a new herbal preparation from Korea, on B16F10 cells. Methods: The anti-proliferative effects of C5E were assessed by culturing B16F10 cells in the presence or absence of C5E. Cell cycle progression was analyzed by PI staining using flow cytometry. The quantities of apoptosis-inducing proteins were measured by Western blot. Results: C5E inhibited the proliferation of B16F10 cells but not human keratinocytes. C5E induced S phase arrest by interfering with cell regulatory factors such as cycling B1, D1, D3, and E, and cycling-dependent kinas 2, in B16F10 cells. Furthermore, immunoblot analysis confirmed that treatment with C5E induced apoptosis and cleaved caspase-3, poly (ADP-ribose) polymerase, via extrinsic pathway, whereas Bcl-2 expression was down-regulated. In addition, the suppression of cell proliferation by C5E is through down-regulation of p-Act, up-regulation of phosphates and tensing homolog protein expression via phosphoinositol 3 kinas survival signaling pathways in B16F10 cells. The combined cytotoxic effects of C5E and vinblastine generated 10% increase in activity in contrast to the sum of the inhibitory effects of the individual agents. Conclusion: C5E shows promising anti-cancer activity and can be a useful adjuvant with vinblastine in combination therapeutic treatment of skin cancer.