Background: Molecular targeted therapy and immunotherapy have shown promise in the management of gastric adenocarcinoma. The amplified expression of Human epidermal growth factor receptor-2 (HER-2) and microsatellite stable (MSI) status serve as indicators of response to targeted therapy and immunotherapy, respectively. Aims: This study was done to assess the frequency of MSI status and HER-2 expression in a pretreated sample of Iranian patients with gastric adenocarcinoma. Methods and results: We conducted HER-2/neu expression and mismatch repair (MMR) system analyses on specimens from patients with gastric and gastroesophageal adenocarcinoma in the Cancer Institute of Iran. Archival tissues from 135 mainly pre-treated surgical specimens of gastric adenocarcinoma cases were used for HER-2 analysis, and 66 specimens were used for MSI analysis. All specimens were tested for HER-2 amplification, revealing that 11 patients (8.1%) had HER-2 amplification, and three out of 66 examined patients exhibited MSI-H. Patients with HER-2 overexpressed specimens had a shorter median overall survival compared to HER-2 negative cases (21 months (CI95%: 1.4-40.6) vs. 31 months (CI95%: 22.9-39), P=0.184). The median disease-free survival (DFS) for HER-2 positive and negative patients was 15 and 28 months, respectively (P= 0.249). The estimated median overall and disease-free survival for patients with negative MSI was 39 and 36 months, respectively. None of the patients with MSI-positive status experienced recurrence, metastases, or death within the follow-up period. Conclusion: HER-2 expression, while less common in our Iranian population compared to North American or Western European countries, is associated with poor outcomes in patients with gastric adenocarcinoma. MSI-H status is highly infrequent in our population, suggesting that immunotherapy may not be a beneficial treatment for a significant fraction of Iranian patients with gastric adenocarcinoma. However, a minority may still benefit from it. Therefore, this hypothesis warrants further investigation in clinical trials.