The aim of the present study was to evaluate the association between the polymorphisms of the Ku80 DNA repair gene, which plays an important role in maintaining genome stability, and the risk of bladder cancer in Central Taiwan. Materials and Methods: In this hospital-based case-control study, the association of Ku80 G-1401T rs828907, Ku80 C-319T rs11685387 and Ku80 intron 19 rs9288518 polymorphisms with bladder cancer risk in a central Taiwanese population was investigated. In total, 288 patients with bladder cancer and 288 age- and gender-matched healthy controls recruited from the China Medical Hospital in central Taiwan were genotyped. Results: A significantly different distribution was found in the frequency of the Ku80 G-1401T polymorphism genotypes, but not the Ku80 C-319T or intron 19 polymorphism genotypes, between the bladder cancer and control groups. The T allele of Ku80 G-1401T conferred a significant (p=0.0055) increased risk of bladder cancer. Gene-environment interactions with smoking, but not with alcohol consumption, were significant for the Ku80 G-1401T polymorphism. The Ku80 G-1401T GT and TT genotypes, in association with smoking, conferred an increased risk of 2.053-fold (95% confidence interval=1.232-3.419) for bladder cancer. Conclusion: The first evidence that the T allele of the Ku80 G-1401T may be associated with the development of bladder cancer and may be a novel useful marker for primary prevention and anticancer intervention is provided.