The prognosis of metastasis gastric cancer patients remains poor and the identification of novel molecular markers will improve the management of gastric cancer patients. The present study aimed to investigate the clinical significance and functional role of miR-466 in gastric cancer peritoneal metastasis. miR-466 expression was confirmed by RT-qPCR. The biological functions were examined by MTT assay, Transwell migration, and invasion assays. The Kaplan-Meier survival curve and multivariate Cox regression analysis were used to investigate the clinical role of miR-466. The logistic regression analysis was performed to reveal the risk factors associated with peritoneal metastasis. miR-466 expression was downregulated in gastric cancer cell lines, tumor tissues, and peritoneal metastasis tissues compared with respective controls. Increased miR-466 expression inhibited proliferation, migration, and invasion of gastric cancer cells. Besides, the lower expression of miR-466 in gastric cancer patients was associated with peritoneal dissemination. Furthermore, multivariate Cox proportional hazard regression analyses demonstrated miR-466 expression level as an independent predictor of prognosis of gastric cancer. The present study provides novel evidence for the clinical and biological significance of miR-466 expression as a possible biomarker for the prognosis and identifying patients with peritoneal metastasis, as well as a potential therapeutic target in patients with gastric cancer.