Background: S-1, a novel oral fluoropyrimidine, plus cisplatin (SP) regimen is one of the standard chemotherapy as first-line for advanced gastric cancer. ToGA study demonstrated that trastuzumab (T-mab) combination regimen improved the overall survival of patients with HER2-positive advanced gastric cancer. However, there was no study evaluating the efficacy and the safety of T-mab in combination with S-1 plus cisplatin (SP) regimen. Therefore, we planned this study to examine the efficacy and the safety of the SP plus T-mab. Methods: Patients confirmed to be HER2-positive by IHC and/or FISH (IHC 3+ or IHC 2+ and FISH positive) received S-1 at 80 mg/m² po, day 1-14, and cisplatin 60 mg/m² iv, day 1 plus trastuzumab 8 mg/kg iv, day 1 (6 mg/kg iv, d1 from 2nd course), repeated every 3 weeks until disease progression. Primary endpoint was response rate (RR). Secondary endpoints were progression-free survival, overall survival and safety. The threshold response rate was defined as 35%, and the expected rate was set at 50% with a 80% power and a 1-sided alpha value of 0.1 and the calculated sample size was 50 patients. Results: A total of 56 patients (median age 66) were enrolled in this study. The efficacy and the safety analyses were conducted in the full analysis set of 53 patients. (Two patients were excluded for ineligibility and one was for no treatment). The confirmed RR assessed by the independent review committee was 67.9% (95% CI: 53.7 – 80.1), and the disease control rate was 94.3%. The median PFS was 7.1 months (95% CI: 6.0 – 10.1). The median OS was not reached. (The median follow-up time: 9.2 months) The main grade 3/4 adverse events were as follows: neutropenia 34%, leucopenia 8%, anorexia 23%, diarrhea 8%, hypoalbuminemia 4%, vomiting 6%, and increased creatinine 6%. Conclusions: This tri-week regimen with SP plus T-mab showed promising results in patients with HER2-positive advanced gastric cancer. Clinical trial information: UMIN000005739.