Gastric cancer (GC) is a fatal form of tumor that has no effective therapeutic agents. In this study, an anticancer tracking method was applied to ioslate Gelsemium elegans alkaloids (GEAs) in vitro. GEA-9 (11-methoxy-N-1-demethoxygelsemamide) exhibited the highest antiproliferative activity in MKN-45 cells and induced mitochondrion-mediated apoptosis, It decreased the mitochondrial potential and increased the Bax/Bcl-2 and p53/MDM2 expression ratio. It also induced cell cycle arrest in MKN-45 cells by regulating the cyclin expression. It Inhibited the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway and the growth of MKN-45 cell xenograft tumors, which were associated with light chain 3-II activation and mTOR inhibition. In summary, we first demonstrated the antineoplastic effect of GEA-9 in vitro and in vivo. We also elucidated that the underlying antineoplastic mechanisms of GEA-9 involved the crosstalk between apoptosis and autophagy by inhibiting of the PI3K/Akt/mTOR pathway. This study might provide a rationale for future clinical applications of GEAs as a chemotherapeutic agents for GC.