Cytochrome P450 1B1 (CYP1B1) and catechol-O-methyltransferase (COMT) are important estrogen-metabolizing enzymes that may affect breast cancer risk. Few studies have directly measured the expression of CYP1B1 and COMT genes in breast tissue samples. The subjects in this study were a subgroup of participants of the Shanghai Breast Cancer Study including 64 patients diagnosed with breast cancer and 68 patients diagnosed with benign breast diseases (BBD) who provided samples of tumor tissue and adjacent nontumor tissue to the study. We compared CYP1B1 and COMT mRNA expression in tumor tissue and adjacent nontumor tissue in both breast cancer patients and BBD patients. High levels of CYP1B1 expression and low levels of COMT expression in adjacent nontumor tissue were associated with a significantly increased breast cancer risk in a nonlinear manner. Odds ratios and 95% confidence intervals (in parentheses) for the midpoints of the first, second, fourth, and fifth quintiles of gene expression levels compared with the overall median levels in BBD subjects were 0.21 (0.07-0.67), 0.81 (0.69-0.95), 1.20 (1.05-1.38), and 1.55 (1.12-2.15) for CYP1B1 and 1.72 (1.17-2.55), 1.19 (1.05-1.35), 0.83 (0.73-0.95), and 0.78 (0.65-0.93) for COMT, respectively. These results support the hypothesis that the formation and accumulation of catechol estrogens in breast tissue through increased CYP1B1 expression and reduced COMT expression may play a significant role in breast cancer risk.