With HeLa cells stably expressing a fusion protein FOXO3A-GFP and several other cancer cell lines, we have discovered that PL treatment induces dephosphorylation and nuclear translocation of FOXO3A and promotes its binding to the BIM gene promoter, leading to increased regulation of BIM in cancer cell lines. The nuclear translocation induced by the piperolongumine of the transcription factor FOXO3A triggers BIM-mediated apoptosis in cancer cells. In summary, our results show that PL activates the apoptotic axis of FOXO3A-BIM promoting dephosphorylation and nuclear translocation of FOXO3A by inhibiting Akt signaling. It should be noted that the siRNA-mediated FOXO3A knockdown saved cells from PL-induced cell death. However, the mechanism of PL-induced cancer cell death is not fully understood. These results reveal a critical mechanism underlying the effects of PL on cancer cells. In vivo treatment with PL significantly inhibited the tumor growth of xenograft, and this inhibition was accompanied by activation of the FOXO3A-BIM axis. In addition, PL promoted the dephosphorylation of FOXO3A by inhibiting the phosphorylation and activation of Akt, a kinase that phosphorylates FOXO3A. As a result, PL inhibited cell viability and caused intrinsic apoptosis in a FOXO3A-dependent manner. Before undertaking any natural, integrative or conventional treatment, it is recommended to consult a licensed physician.