Cervical cancer is the fourth most common female cancer worldwide, and drug targeted therapy plays a crucial role in delaying the progression of cervical carcinoma. Chelerythrine hydrochloride (CHE) is a natural alkaloid, which is a focal point in anti-tumor research. In this study, we investigated the effect of CHE on HeLa cells by using MTT assay, RTCA, and colony formation assay. In addition, the flow cytometric analysis, immunofluorescence staining assay and western blot analysis were performed to study the mechanism of CHE. The results showed that CHE exhibited a significant inhibitory effect in HeLa cells, and it could suppress the expression of PI3K subunits in a dose-dependent manner. Moreover, we found that the treatment of CHE further restrained the downstream AKT/mTOR and PKCα signaling. In addition, CHE induced mitochondrial apoptosis of HeLa cells by regulating the BCL-2 family member's expression. Immunofluorescence staining assay indicated that AIF and Cytochrome C were translocated from mitochondria to cytoplasm or nucleus, and notable changes in mitochondrial morphology of HeLa cells were also observed. Finally, the aberrant expression of CHE led to the mitochondrial apoptosis by upregulating the expression of APAF1, Cleaved-Caspase9, Cleaved-Caspase3, and Cleaved-PARP. In summary, CHE suppresses the proliferation of HeLa cells by trigging the mitochondrial apoptosis through the PI3K/BAD signaling pathway.