Hepatocellular carcinoma (HCC) is one of the most common malignant tumors and the prognosis of patients was poor. Cantharidin is an effective anti-tumor component of Chinese traditional medicine. N-methylcantharidinimidum is a novel cantharidin derivative with better curative effect and less side effect. In this study, the effects of N-methylcantharidinimidum on proliferation, apoptosis and cell cycle of human hepatocellular carcinoma cell line HepG2 were examined. The results demonstrated that N-methylcantharidinimidum significantly induced apoptotic cell death in a dose and time-dependent manner and arrested cell cycle in HepG2 cells. N-methylcantharidinimidum also suppressed cyclin D1. This study suggests that N-methylcantharidinimidum may serves as a potential chemopreventive agent for live cancer.