CKLF-like MARVEL transmembrane domain containing 3 (CMTM3) is considered to be a tumor suppressor gene in multiple types of malignancies. Previous studies have indicated that CMTM3 suppresses metastasis and epithelial-mesenchymal transition (EMT) in gastric cancer. However, its role in gastric cancer cell proliferation has rarely been discussed. Moreover, the regulatory mechanisms of CMTM3 in gastric cancer remain unclear. In this study, RT‑qPCR and IHC were used to assess the expression of CMTM3 and miR‑135b‑5p in gastric cancer tissues and cell lines. We found that the expression of miR‑135b‑5p was negatively associated with CMTM3 in gastric cancer tissues, and we verified that miR‑135b‑5p directly targeted CMTM3 in gastric cancer cells by dual-luciferase reporter assay. CCK8 assay, Transwell assay and flow cytometric analysis were conducted to examine the functions of CMTM3 and miR‑135b‑5p in vitro. Our results demonstrated that the overexpression of CMTM3 or the suppression of miR‑135b‑5p using an inhibitor suppressed SGC‑7901 gastric cancer cell proliferation, invasion and cell cycle progression, and promoted SGC‑7901 cell apoptosis. Furthermore, a BALB/c nude mouse subcutaneous xenograft model was used to verify the function of miR‑135b‑5p and CMTM3. Our results revealed that miR‑135b‑5p inhibitor significantly suppressed SGC‑7901 cell tumorigenesis in vivo. In addition, IHC revealed that CMTM3 expression was markedly increased in tumors infected with miR‑135b‑5p inhibitor lentivirus. On the whole, the findings of the present study suggest that the overexpression of miR‑135b‑5p inhibits CMTM3 expression, and promotes gastric cancer progression and metastasis. Our findings provide a novel therapeutic target for gastric cancer.