Gastric cancer (GC) is one of the most common malignant tumors of the digestive system. A deeper understanding of the mechanism of proliferation and metastasis is needed to improve patient survival. T cell lymphoma invasion and metastasis 1 (TIAM1) has been proven to play an essential role in the proliferation and metastasis of GC. The aim of this study was to explore the relevant upstream regulatory mechanism of TIAM1. Bioinformatic analysis, RT-qPCR, and dual luciferase reporter assays were used to predict and validate microRNAs that target the TIAM1 gene. Among eleven predicted microRNAs, eight (miR-10b-5p, miR-589-3p, miR-651-3p, miR-335-3p, miR-653-5p, miR-373-3p, miR-372-3p, and miR-205-3p) affected TIAM1 expression; and only miR-10b-5p regulated TIAM1 expression by directly binding to the 3'-UTR of TIAM1 mRNA. miR-10b-5p levels were determined in both normal and cancerous tissues retrieved from GC patients. We observed that by targeting TIAM1 expression, miR10b-5p inhibited the proliferation, migration, and invasion of GC cells. To verify our observations, we evaluated the participation of runt-related transcription factor 3 (RUNX3), a known regulator of microRNA expression and tumor suppressor. Tumor-suppressor RUNX3 combined with core-binding factor subunit beta (CBF beta) upregulated miR-10b-5p and suppressed GC. In conclusion, we identified a CBF beta/RUNX3-miR10b-TIAM1 molecular axis that inhibits GC progression and metastasis and may provide suitable treatment targets for GC.