Bufadienolides from Venenum Bufonis Inhibit mTOR-Mediated Cyclin D1 and Retinoblastoma Protein Leading to Arrest of Cell Cycle in Cancer Cells

OBJECTIVE: Bufadienolides, the main components in Venenum Bufonis secreted from toads, have been proved to be with significant anticancer activity aside from the positive inotropic action as cardenolides. Here an underlying anticancer mechanism was further elucidated for an injection made from Venenum Bufonis containing nine bufadienolides. METHODS: One solution reagent and cell cycle analyses were for determining effect of bufadienolides on cancer cells. Western blotting was used for protein expression. RESULTS: Bufadienolides inhibit cell proliferation and arrest cells in G1 phase. Bufadienolides also inhibit the mammalian target of rapamycin (mTOR) signaling pathway, which is evidenced by the data that bufadienolides inhibit type I insulin-like growth factor- (IGF-1-) activated phosphorylation of mTOR by a concentration- and time-dependent way, as well as phosphorylation of p70 S6 kinase 1 (S6K1) and eukaryotic initiation factor 4E (eIF4E) binding protein 1 (4E-BP1). Subsequent results indicated that cyclin D1 expression and phosphorylation of retinoblastoma protein (Rb)—two characterized regulators in cell cycle of G1—are also inhibited and the process is dependent on mTOR pathway. CONCLUSION: Bufadienolides inhibit proliferation partially due to arresting cell cycle in G1 phase, which is mediated by inhibiting mTOR-cyclin D1/Rb signal pathway.