Autophagy contributes to the antitumor activity of Ziyu II in CRC cells
We next intend to evaluate whether autophagy was involved in the anti-CRC effect of Ziyu II. First, CRC cells were treated with Ziyu II combined with CQ or 3-MA. As shown in Fig. 6A and Fig. S4A, combinational use of CQ or 3-MA with Ziyu II partially restored Ziyu II-induced growth inhibition in CRC cells. In addition, LDH release assay also revealed that CQ or 3-MA counteracted Ziyu II-induced cytotoxicity (Fig. 6B and Fig. S4B). A similar increase in cell proliferation was also observed in Ziyu II-treated CRC cells in combination with CQ or 3-MA, as evidenced by colony formation analysis (Fig. 6C, D and Fig. S4C, D). Consistently, similar results were obtained by inhibition of autophagy by transfecting a constitutively active form of Akt (Fig. 6E-H and Fig. S4E, F). Collectively, these results demonstrate that autophagy is involved in Ziyu II-induced CRC growth suppression.
We further investigated whether Ziyu II enhanced the chemosensitivity of CRC cells to 5-fluorouracil (5-FU), the first-line chemotherapeutic drug for CRC treatment. As indicated, combinational treatment of Ziyu II with 5-FU markedly decreased the cell growth (Fig.S5A) and proliferation rate (Fig.S5B and C) compared with monotherapy in CRC cells, suggesting that Ziyu II effectively sensitizes CRC cells to the treatment of 5-FU.