MKK4 (mitogen-activated protein kinase kinase 4, NM_003010.2), which belongs to the mitogen-activated protein kinase pathways, possesses functions in tumorigenesis. We hypothesized the genetic variants in MKK4 gene may alter its functions and thus cancer risk. In current hospital-based case-control study of 706 patients with sporadic colorectal cancer (CRC) and 723 sex-age-frequency-matched control subjects in a southern Chinese population, we genotyped two polymorphisms of MKK4 promoter (i.e., -1304T>G, rs3826392 and -1044A>T, rs3809728) and assessed their associations with the risk of sporadic CRC. Compared with -1304TT genotypes, -1304TG had a significantly decreased risk of CRC (adjusted odds ratios [OR] = 0.56; 95% CI = 0.44-0.72; p = 3.53 x 10(-6)), the -1304GG carriers had a further decreased risk of CRC (OR = 0.40; 95% CI = 0.23-0.70; p = 1.32 x 10(-3)), and there was a significant trend for an allele dose effect on risk of CRC (p(trend) = 2.64 x 10(-7)). The decreased risk associated with -1304G variant genotypes (i.e., TG+GG) was more pronounced in the subjects older than 60 years (adjusted OR = 0.41; 95% CI = 0.29-0.57; p = 2.25 x 10(-7)), in ever drinkers (adjusted OR = 0.41; 95% CI = 0.28-0.59; p = 2.42 x 10(-6)). The age and alcohol drinking status interacted with -1304G variant genotypes on reducing cancer risk (p values for interaction were 0.015 and 0.043, respectively). Western blotting analysis showed that the levels of Mkk4 protein in sporadic CRC neoplastic tissues were significantly higher in the carriers of -1304G variant genotypes than that in those with -1304TT genotypes. However, no significant association was observed between -1044A>T polymorphism and risk of CRC. To the best of our knowledge, this is the first study of genetic variants in MKK4 and cancer susceptibility. Larger studies are needed to validate our findings.