The adult liver performs many metabolic functions for maintaining homeostasis. There are several sex differences in liver function and disease pathogenesis. One important function of the liver is drug metabolism, where cytochrome p450s (CYPs) in hepatocytes are the main enzymes involved. The toxicity of various drugs and chemicals differs with sex due to differences in hepatocytic CYP expression. However, the molecular mechanism regulating sex-related differences in drug metabolism remains unknown. In this study, we identified transcriptional regulator B-cell lymphoma 6 (Bcl6) as an important factor in sex-biased differential CYP expression. Microarray analysis of livers derived from liver-specific Bcl6-knockout mice showed that Bcl6 is required for sex-biased CYP expression patterns in the liver. Additionally, quantitative PCR analysis revealed that hepatocytic expression of male-biased genes, such as Cyp2d9, Cyp2u1, Cyp4a12a/12b, and Cyp7b1, in liver-specific Bcl6-knockout male mice significantly decreased to levels similar to those observed in wild-type female mice. Conversely, hepatocytic expression of female-biased genes, such as Cyp2a4/2a5, Cyp2b9, Cyp3a41, and Cyp17a1, significantly increased in liver-specific Bcl6-knockout male mice. Deletion of Bcl6 caused female-like expression of CYPs in male livers. These results suggest that Bcl6 is a key regulator of sex-related differential regulation of drug metabolism. Moreover, serum sex hormone levels and fertility did not change in liver-specific, Bcl6-knockout mice. Hepatocytic Bcl6 regulates sex-related differential CYP expression in the liver without changing the sex of the whole body. Thus, this mouse model is useful for analyzing liver-specific sex-dependent regulation of drug metabolism and pathogenesis.