While it has been demonstrated that cigarette smoke contains aromatic and heterocyclic amines that initiate carcinogenesis, the association between cigarette smoking and breast cancer remains controversial. N-acetyltransferase 2 (NAT2) catalyzes arylamine carcinogen biotransformation and NAT2 genetic polymorphisms may contribute to differential susceptibility to breast cancer. We tested whether NAT2 modified the association between cigarette smoking and breast cancer risk in a population-based study of Hispanic and non-Hispanic white women in the Southwest United States. Data were available for cigarette smoking and NAT2 polymorphisms for 717 cases (Hispanic, 251 and non-Hispanic white, 466) and 735 controls (Hispanic, 245 and non-Hispanic white, 490). NAT2 genotypes were translated into rapid, intermediate, slow, or very slow acetylator phenotypes. Odds ratios (ORs) and 95% confidence intervals (95% CIs) for the joint association of NAT2 with smoking on breast cancer risk were estimated using logistic regression. Non-Hispanic white women were more likely (p < 0.001) than Hispanic women to have a slow (41.7 vs. 33.5%) or very slow (19.0 vs. 11.1%) acetylator status and less likely to have rapid/intermediate phenotypes (39.2 vs. 54.4%). Breast cancer risk was significantly increased in non-Hispanic white women with a very slow acetylator phenotype who smoked: ever versus never (OR, 2.57; 95% CI, 1.49-4.41), never versus former (OR, 2.69; 95% CI, 1.41-5.17) or current (OR, 2.46; 95% CI, 1.07-5.65), and 16 + pack-years (OR, 2.29; 95% CI, 1.16-4.51). Results for Hispanic women were not statistically significant. These findings support smoking as a risk factor for breast cancer among non-Hispanic white women with very slow NAT2 acetylator phenotype.