〔Abstract〕 Objective To investigate the effect and mechanism of cordycepin(CP) on autophagy in oral squamous cell carcinoma SCC9 cell line. Methods The SCC9 blank group, CP(5 μg/ml) group, CP(10 μg/ml) group and CP(20 μg/ml) group were set in the experiment. Cell proliferation was detected by 5-ethynyl-2′-deoxyuridine(EdU) staining. Apoptosis was detected by Hoechst staining. Western blot was used to detect the expressions of Ki67, PCNA, Caspase-3, Caspase-9, Beclin 1, p62, LC3 A/B, PI3 K, p-PI3 K, AKT, p-AKT, mTOR and p-mTOR. The autophagy marker molecule LC3 Ⅱ was detected by immunofluorescence. After adding PI3 K inhibitor LY294002, cell proliferation was detected by EdU staining, apoptosis was detected by Hoechst staining, and autophagy marker molecule LC3 Ⅱ was detected by immunofluorescence. Results Compared with the SCC9 blank group, the cells in each CP treatment group showed significant inhibition of proliferation, accompanied by apoptosis, down-regulation of proliferation-related molecules, up-regulation of apoptosis-related molecules, and increased concomitant with increasing concentration of CP(P<0.01); The expression of autophagy-promoting related molecules(Beclin 1 and LC3 Ⅱ) was up-regulated, the expression of autophagy-inhibiting molecules(p62) was down-regulated, and the activity of the upstream signal molecules(PI3 K/AKT/mTOR) decreased, and the difference increased with the increase of CP concentration(P<0.01). Compared with CP treatment, LY294002 combined with CP treatment inhibited cell proliferation and promoted apoptosis and autophagy. Conclusion CP can increase the level of autophagy in oral squamous cell carcinoma, inhibit cell proliferation and promote apoptosis. The main reason may be related to the down-regulation of PI3 K/AKT/mTOR pathway.