The hepatic cytochrome P4502A6 (CYP2A6) enzyme mediates the oxidative metabolism of several procarcinogens that have liver as their primary target. Mouse models indicate that liver tumors invariably overexpress CYP2A forms, and that inflammation and cirrhosis may regulate the CYP2A expression pattern. In this study, the distribution of the CYP2A6 protein was investigated in a series of 24 human hepatocellular carcinoma (HCC) samples by immunohistochemical analysis. A polyclonal antibody was raised in chicken against CYP2A5, the mouse orthologue of CYP2A6. The antibody was characterized and found to be specific for CYP2A members. In DBA/2 mouse liver, a strong increase of CYP2A5 protein amount, localized in the perivenous region, occurred in response to treatment with pyrazole. In human HCC samples, overexpression of CYP2A6 protein was associated with the presence of chronic inflammation and cirrhosis. CYP2A6 protein was observed in 9 of 16 (56%) of samples with non-neoplastic hepatocytes and in 10 of 24 (42%) HCC samples. The staining for CYP2A6 protein was very heterogeneous in tumor cells, suggesting that increased expression of CYP2A6 occurred in a distinct subpopulation of neoplastic cells. In Kaplan-Meyer survival analysis, there was a tendency toward a more favorable prognosis in patients with CYP2A6-positive tumors in comparison with patients with CYP2A6-negative tumors. These data suggest that, in human HCC, in contrast to mouse liver tumors, CYP2A6 overexpression is not an invariable phenotype.