Cervical cancer as one of the major malignant tumors seriously threatens women's health. More than 270,000 women die of cervical cancer each year. Warangalone is an isoflavone compound isolated from Cudrania tricuspidata with excellent antitumor activity. In this research, we investigated the molecular mechanism of warangalone-induced apoptosis in HeLa cells. The results show that warangalone can selectively and effectively inhibit HeLa cells proliferation. Warangalone can effectively inhibit the invasion and migration of HeLa cells. Furthermore, warangalone was confirmed to activate p53 and mitogen-activated protein kinase (MAPK) family signaling pathways to cause apoptosis. In this case, the expression of the B-cell lymphoma-2 (Bcl-2) family is regulated, and caspase-3 is eventually cleaved, finally triggering the mitochondrial apoptosis. In conclusion, warangalone can induce HeLa cells apoptosis via a mitochondria-mediated endogenous pathway, which represented the potential therapeutic effect of warangalone on cervical cancer.