<p>Background: The Human Epidermal growth factor Receptor-3 (HER-3) is a member of ErbB receptor family and has deficient kinase activity. HER-3 should heterodimerize with other members of ErbB receptor family, especially with HER-2, to transduce downstream signaling pathways. HER-3 co-expresses with other ErbB receptors in different cancers and overexpresses while the oncogenic signaling pathways such as Jak/Stat, MAPK, and PI3K/Akt are activated and promoted. Here, the expression level of HER-3 was evaluated in Iranian gastric adenocarcinoma&#39;s patients and the effects of HER-3 knocking down was investigated on cell cycle and cell viability of human gastric adenocarcinoma cell line of MKN45.<br /> Methods: In this study, 38 paraffin-embedded surgical adenocarcinoma specimens and their marginal non-tumor tissue samples were collected. Total RNAs were extracted and cDNAs were synthesized. Finally, the expression level of HER-3 was evaluated by real time PCR approach. Moreover, the human adenocarcinoma cell line of MKN45 was transfected with siRNA against HER-3 and the effects of its down-regulation were evaluated using MTT assay and cell-cycle analysis.<br /> Results: The data obtained from this study revealed HER-3 is significantly overexpressed in gastric tumors rather than non-tumor marginal tissues. Also, it was found that the expression level of HER-3 is elevated with tumor depth of invasion. Moreover, HER-3 knocking down promotes cell accumulation in G2/M phase of cell cycle and decreases cell viability in MKN45 cells which suggests a potential role for HER-3 in gastric adenocarcinoma tumorigenesis.<br /> Conclusion: Taken together, these results emphasize the importance of HER-3 receptor in diagnosis and prognosis of gastric adenocarcinoma.</p>