The roles of ROS and P21 in apoptosis-inducing effect of cepharanthine in human colorectal cancer cells

Abstract

Cepharanthine (CEP), a natural compound isolated from Stephania cepharantha Hayata, has been shown to possess anti-cancer activity against several cancers including colorectal cancer (CRC) cells. Previously, it was found that cytotoxicity of CEP was associated with ROS generation and p21Waf1/Cip1 expression. However, information regarding to the molecular mechanisms underlying the apoptosis-inducing effects of CEP in CRC cells is still limited. Therefore, the present study aimed to evaluate the roles of ROS and p21Waf1/Cip1 in the apoptosis-inducing effect of CEP in two human colorectal cancer cells lines, p53 wild-type HCT116 cells and p53 mutant HT-29 cells. The results demonstrated that CEP significantly inhibited the growth of both HT-29 and HCT116 cells. Remarkably, CEP was more effective than L-OHP in controlling the growth of HT-29 cells. Moreover, CEP induced apoptosis was mediated through upregulation of pro-apoptotic Bak, downregulation of anti-apoptosis Bcl-xL, and induction of poly (ADP-ribose) polymerase (PARP) cleavage in both HT-29 cells and HCT116 cells. Additionally, CEP was found to induce ROS generation in HT-29 and HCT-116 cells. It should be noted that N-acetyl cysteine (NAC) could abolish the apoptosis-inducing effects of CEP in both HT-29 cells and HCT116 cells, suggesting that CEP-induced apoptosis is mediated through ROS generation. Furthermore, CEP could modulate ERK1/2 signaling pathway by inhibiting ERK1/2 phosphorylation in HT-29 cells while activating ERK1/2 phosphorylation in HCT116 cells, which was found to be associated with ROS generation. Notably, CEP markedly upregulated the expression of p21Waf1/Cip1 in p53 mutant HT-29 cells but did not alter the expression of p21Waf1/Cip1 in HCT116 cells. A p21 inhibitor, UC2288, could abolish apoptosis-inducing effect of CEP in HT-29 cells, suggesting that CEP-induced apoptosis is mediated via upregulation of p21Waf1/Cip1 in p53 mutant HT-29 cells. Taken together, the results of this study demonstrated that generation of ROS and upregulation of p21 Waf1/Cip1 play a significant role in apoptosis-inducing effect of CEP in p53 mutant CRC cells. It may be useful for treatment of colorectal cancer carrying mutant p53 which often cause resistant to current chemotherapeutic agents.