AIM: To explore the role of miR-195-5p in mediating trastuzumab resistance in gastric cancer and to validate its potential as a therapeutic target along with its target gene GPRC5A. METHODS: Trastuzumab-resistant gastric cancer cell lines (NCI-N87 and MKN45) were established. Cell viability under trastuzumab treatment was assessed using CCK-8 assays. Expression levels of miR-195-5p were determined by RT-qPCR. Transfection with miR-195-5p mimics was performed to evaluate changes in trastuzumab sensitivity and proliferation. GPRC5A expression was also measured by RT-qPCR, and the targeting relationship between miR-195-5p and GPRC5A was confirmed using a dual-luciferase reporter assay. RESULTS: Parental cells showed higher sensitivity to trastuzumab than resistant cells, with miR-195-5p expression significantly lower in the latter. Overexpression of miR-195-5p in resistant cells enhanced trastuzumab sensitivity and reduced proliferation. GPRC5A was found to be upregulated in resistant cells, and miR-195-5p directly targeted GPRC5A, affecting cell proliferation under trastuzumab treatment. CONCLUSION: miR-195-5p may regulate trastuzumab sensitivity in gastric cancer by targeting GPRC5A, suggesting potential as a molecular marker for trastuzumab therapy guidance.