The hMSH2 (human MutS homolog 2) gene plays a central role in DNA mismatch repair. Structural variations in the gene may lead to protein instability and deficient mismatch repair. However, the role of polymorphic variants of the hMSH2 gene have not been defined in head and neck cancer. In this study, the roles of three polymorphic variants in the functional domains of the gene were investigated in 166 patients with head and neck cancer by allele-specific PCR, electronical array addressing, and PCR/RFLP (restriction fragment length polymorphism). This is the first study to investigate the gIVS12-6T -> C polymorphism in head and neck cancer. A significant association between the CC genotype and reduced risk of disease suggests that the gIVS12-6T -> C substitution at the splice-acceptor site may affect the risk of head and neck cancer. We did not observe an association between the Asn127Ser and Gly322Asp polymorphisms and cancer risk. A possible role of the gIVS12-6T -> C substitution warrants further validation in larger cohorts because of low allele frequency.