B ackground: HER2 (ERBB2) amplification in gastric cancer ranges from 6-23%, accordingly Trastuzumab has been incorporated into the treatment arsenal of HER2-enriched gastric cancer. We had previously demonstrated the miR-125a-3p induces overexpression of HER2 in basal-like breast cancer cells and sensitizes them to anti-HER2 therapy (AACR-NCI-EORTC 2015). We aimed to study the effect of miR-125a-3p as a potential modulator of the ERBB2/HER2 pathway in HER2-negative gastric adenocarcinoma. Methods: We generated stable KATO-III cells that overexpress miR-125a-3p and control cells that overexpress scrambled miRNA. Relative mRNA level of ERBB2 was measured by qPCR and its protein expression and localisation were examined by western blot and immunofluorescence staining. Moreover, the effect of miR-125a-3p alone or combined with anti-HER2 therapies on cellular proliferation was evaluated using EdU incorporation and XTT assays. Results: miR-125a-3p-overexpressing KATO cells showed a significant increase in the expression level of ERBB2 mRNA and protein as well as a stronger immunofluorescence staining of ERBB2 on cell membrane compared with control cells. Trastuzumab reduced cell growth and proliferation of miR-125a-3p-overexpressing KATO cells. Furthermore, this antiproliferative effect was enhanced following pre-treatment of the miR-125a-3p-overexpressing KATO cells with lapatinib, a dual ErbB1 and ErbB2 receptor tyrosine kinase inhibitor, prior tp trastuzumab administration. Conclusions: Our results indicate that miR-125a-3p is capable of inducing a shift in the expression and function of ERBB2 pathway that may convert the fate of gastric cancer cells to effectively dispose them to anti-HER2 therapies. In an era of personalized medicine, our study proposes a means to enlarge the patient population that may benefit from anti-HER2 therapies.