The NAT1 gene exhibits polymorphisms in the non-coding polyadenylation region with a number of alleles. Of these alleles, NAT1*10 is responsible for increased NAT1 enzyme levels and is reported to be associated with increased risk for colorectal and bladder cancers. In view of the possible role of the NAT1 gene product in the metabolism of a number of cigarette smoke carcinogens, we tested the possibility that genetic variation in the NAT1 gene might also be associated with increased risk for lung cancer. Allelic variances of the NAT1 gene were analyzed in 45 lung cancer patients and 47 controls who were matched with respect to age, race and gender using restriction fragment length polymorphism (RFLP) analysis and allele-specific (AS)-PCR. Our results indicate that individuals who inherited the NAT1*10 allele had a 3.7-fold increased relative risk for lung cancer (95% CL = 1.2-16.0, p < 0.02). There was a 6.8-fold increase in relative risk for lung cancer associated with the inheritance of the NAT1*10 allele in younger individuals (< 60 years of age) compared to 2.2-fold increase in older individuals (> 60 years old) (OR = 6.8; 95% CL = 1.1-40.7, p < 0.01 and OR = 2.2; 95% CL = 0.5-11.1, p = 0.2, respectively). We have also applied the sensitive fluorescence in situ hybridization (FISH) tandem probe assay to elucidate the frequency of chromosome breakage among a subgroup of the studied individuals harboring the NAT1*10 allele (17 lung cancer patients, 17 smoking controls and 7 non-smoking controls). Our results indicate a significant increase (p < 0.001) in the frequency of chromosome breaks in lung cancer patients (mean +/- SE per 100 cells = 1.45 +/- 0.11) and in smoking controls (1.30 +/- 0.13) compared to non-smoking controls (0.47 +/- 0.07). Regression analysis indicated a highly significant positive correlation between the duration of smoking in years and the frequency of chromosome breaks in lung cancer patients (r = 0.62, p = 0.008), but not in smoking controls (r = 0.02; p = 0.91). These findings suggest that NAT1 polymorphism may be an important genetic determinant of lung cancer risk. In addition, these data provide a mechanistic link between the inheritance of the NAT1*10 allele and smoking-induced lung cancer. Given that the NAT1 enzyme can mediate activation and detoxication pathways for numerous carcinogens and given that this polymorphism is prevalent in the general population (20-50% frequency), it may play a significant role in influencing the outcome of a variety of environmental cancers.