Inhibition of mTOR signaling by oleanolic acid contributes to its anti-tumor activity in osteosarcoma cells.

Oleanolic acid (OA), a pentacyclic triterpenoid exhibits potent anti-tumor activity against many tumor cell lines. But the mechanisms through which OA inhibits osteosarcoma cells are not known. The mammalian target of rapamycin (mTOR) serves as a central regulator of cell growth, proliferation, survival, and metabolism by integrating intracellular and extracellular signals. In this study, we examined effects of OA on proliferation, cell cycle progression, apoptosis in osteosarcoma cells, and involvement of mTOR signaling in this process. OA inhibited cell proliferation and colony formation, induced G1 arrest in osteosarcoma MG63 and Saos-2 cells dose and time dependently. The protein level of cyclin D1, which plays critical role in G1 to S phase transition and servers as a downstream target of mTOR complex 1 (mTORC1) was down-regulated by OA. Phosphorylation of p70 ribosomal S6 kinase 1 (p70 S6K1) (T389) and S6 (S235/236), mediators of mTORC1 signaling in controlling protein translation and cell growth, was also inhibited by OA. Furthermore, OA inhibited phosphorylation of Akt, a pro-survival factor and substrate for mTORC2. Inactivation of Akt correlated with pro-apoptotic role of OA in osteosarcoma cells, as manifested by an increase in annexin V-FITC binding, cleavage of poly (ADP-ribose) polymerase (PARP) and activation of caspases 3. Our results suggest that OA is a promising agent for treatment of osteosarcoma and mTOR signaling may contribute to its anti-tumor effects on osteosarcoma cells.