The etiology of gastric cancer is not well-understood. Epidermal growth factor (EGF) transduces growth signals to mitogen-activated protein kinase via RAS and BRAF, and EGF/EGF receptor interaction is important for tumor growth and progression. Previous studies have reported that the EGF +61 (A/G) single nucleotide polymorphism in the 5'-untranslated region of the EGF gene is functional, and is associated with gastric cancer and various malignancy. Individuals with the EGF A/A genotype produce less EGF than individuals with G/G or G/A. We investigated a single nucleotide polymorphism at exon 1 of EGF, named rs4444903 in NCI dbSNP, in 454 Japanese subjects undergoing a health checkup and 202 patients with gastric cancer. Genotype was determined by PCR with confronting two-pair primers. Results showed that EGF polymorphism was not associated with gastric cancer but that the EGF A/A genotype showed a protective effect (odds ratios, 0.58; 95% confidence interval, 0.29-1.17 relative to G/G). Furthermore, when we divided cases into two groups, a differentiated type and an undifferentiated type, the A/A and G/A combined was found to be lower frequency in the latter type than in the former type without significance (OR, 0.81; 95% confidence interval, 0.44-1.49 relative to G/G). As is the case with any malignancy, other factors are involved, including environmental and host factors. The present results show that although EGF is necessary for cancer, it is not sufficient. We also found ethnic heterogeneity in the functional EGF polymorphism. Because the relationship between EGF polymorphism and malignancy remains inconsistent, confirmation of the role of EGF polymorphism in gastric cancer requires a much larger study.