To explore the mechanism of action of bufalin in the treatment of gastric cancer based on network pharmacology,molecular docking and cellular experiments in the study.The PubChem and PharmMapper databases were used to retrieve the drug targets of bufalin,and the related targets of gastric cancer were collected in the GeneCards database,and the PPI network diagram for common targets of drugs and diseases was constructed by Venny software and STRING database,and GO enrichment and KEGG pathway analysis were performed through the DAVID database.Cytoscape3.9.0 software was used to visualize and screen the core targets,and the relationship between the core targets and gastric cancer and bufalin was analyzed by GIEPA database,Kaplan-Meier method,AutoDock and PyMOL software,respectively.Finally,the targets and pathways predicted by network pharmacology were experimentally verified by cellular experiments.The results of network pharmacology showed that 285 drug targets and 909 gastric cancer targets were screened out.The number of common targets between bufalin and gastric cancer were 91,HSP90AA1 and SRC were core targets based on the ranking of PPI network value,the expression level in gastric cancer patients,and the correlation of survival and prognosis.KEGG enrichment analysis showed that bufalin in the treatment of gastric cancer may be related to PI3K-Akt,FoxO,Ras and MAPK signaling pathways.The results of survival analysis showed that the expression of HSP90AA1 and SRC mRNA in gastric cancer tissues was significantly up-regulated,and the overall survival of patients with high expression was significantly lower than that of the low expression group.The molecular docking results showed that the core targets had relatively strong binding activities with bufalin.The results of cellular experiments showed that compared with the control group,the proliferation and migration of gastric cancer cells MGC-803 were decreased,and the number of apoptosis was increased.The expression levels of HSP90AA1,SRC,Bcl-2,p-AKT and FoxO1 were significantly down-regulated,and the expression of Bax protein was significantly up-regulated.In summary,bufalin may play a role in the treatment of gastric cancer by regulating the core protein of HSP90AA1 and SRC and the Akt/FoxO1 signaling pathway.