BACKGROUND: Although substantial evidence has shown that the mammalian target of rapamycin (mTOR) pathway is an important therapeutic target in gastric cancer, the overall response rates in patients to mTOR inhibitor everolimus have been less than initially expected. We hypothesized that the limited efficacy of everolimus in gastric cancer is due to the activation of extracellular signal-regulated kinase (ERK). METHODS: ERK activation was investigated using western blot. The effects of dual inhibition of ERK and mTOR via genetic and pharmacological approaches were determined using cellular assays and xenograft mouse model. RESULTS: We observed the decreased phosphorylation of mTOR, rS6, and 4EBP1 and increased phosphorylation of ERK and p90RSK in gastric cancer cells exposed to everolimus at clinically relevant concentration. Using both in vitro cell culture assays and in vivo xenograft mouse model, we found that trametinib overcame everolimus resistance by either effectively targeting resistant cells or further enhancing everolimus' efficacy in sensitive cells. Mechanism studies confirmed that trametinib overcame everolimus resistance via specifically inhibiting ERK and regulating ERK-mediated Bcl-2 family proteins in gastric cancer cells. CONCLUSIONS: Inhibition of mTOR pathway can induce "paradoxical" activation of ERK in gastric cancer, and this activation can be reversed by trametinib. Since both drugs are clinically available, our findings might accelerate the initiation of clinical trials on gastric cancer using everolimus and trametinib combination.