Induction of apoptosis by a novel intestinal metabolite of ginseng saponin via cytochrome c-mediated activation of caspase-3 protease

Abstract

Ginseng saponins exert various important pharmacological effects with regard to the control of many diseases including cancer. The novel intestinal bacterial metabolites of ginseng protopanaxadiol saponins have recently been found and isolated after the oral administration of ginseng extract in human and rats. 20- O-(β- d-Glucopyranosyl)-20( S)-protopanaxadiol (IH-901) formed from ginsenosides Rb1, Rb2, and Rc is of particular interest in cancer chemoprevention and treatment. We investigated the effects of IH-901 on the human myeloid leukemia cell line HL-60 in terms of inhibition of proliferation and induction of apoptosis. IH-901 showed a significant cytotoxic activity in HL-60 cells ( ic 50 = 24.3 μM) following a 96-hr incubation. Treatment of HL-60 cells with IH-901 resulted in the formation of internucleosomal DNA fragments. The dose- and time-dependent induction of apoptosis by IH-901 was demonstrated in sandwich enzyme immunoassay and the results were confirmed by flow cytometric analysis. Morphological examination of IH-901-treated samples showed cells with chromatin condensation, cell shrinkage, and nuclear fragmentation, all typical characteristics of apoptotic cells. The treatment of HL-60 cells with IH-901 caused activation of caspase-3 protease and subsequent proteolytic cleavage of poly(ADP-ribose) polymerase. IH-901 did not affect the expression of antiapoptotic protein Bcl-2 but did cause a release of mitochondrial cytochrome c into cytosol. In conclusion, our results demonstrate that IH-901 dramatically suppresses HL-60 cell growth by inducing programed cell death through activation of caspase-3 protease, which occurs via mitochondrial cytochrome c release independently of Bcl-2 modulation. These results may provide a pivotal mechanism for the use of IH-901 in the prevention and treatment of leukemia.