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Background: We aimed to investigate the effect of PI3K/Akt signaling pathway on PRAS40Thr246 phosphorylation in gastric cancer cells. Methods: The study was conducted from April 2017 to January 2018 in Zhongshan Hospital, Xiamen University, Xiamen, China. Gastric cancer cells were divided into three groups: gastric cancer cell group, LY294002 group and MK-2206 group. Specific tests were conducted accordingly.   Results: Inhibition of PI3K/Akt signaling pathway activation and PRAS40Thr246 phosphorylation could inhibit proliferation and invasion and promote apoptosis of gastric cancer cells, and PRAS40Thr246 phosphorylation could activate PI3K/Akt signaling pathway. Conclusion: The levels of PI3K/Akt signaling pathway related proteins and p-PRAS40Thr246 were significantly increased in gastric cancer cells. p-PRAS40-Thr246 was able to reflect the activation of the PI3K/Akt signaling pathway, reflecting the sensitivity of the PI3K/AKT signaling pathway to inhibitors.