CYP1A1 is one of the main cytochrome P450 enzymes, examined extensively for its capacity to activate compounds with carcinogenic properties. Continuous exposure to inhalation chemicals and environmental carcinogens is thought to increase the level of CYP1A1 expression in extrahepatic tissues, through the aryl hydrocarbon receptor (AhR). Although the latter has long been recognized as a ligand-induced transcription factor, which is responsible for the xenobiotic activating pathway of several phase I and phase II metabolizing enzymes, recent evidence suggests that the AhR is involved in various cell signaling pathways critical to cell cycle regulation and normal homeostasis. Disregulation of these pathways is implicated in tumor progression. In addition, it is becoming increasingly evident that CYP1A1 plays an important role in the detoxication of environmental carcinogens, as well as in the metabolic activation of dietary compounds with cancer preventative activity. Ultimately the contribution of CYP1A1 to cancer progression or prevention may depend on the balance of procarcinogen activation/detoxication and dietary natural product extrahepatic metabolism.

Introduction: The natural product eupatorin has been reported to have antiproliferative activity in tumour cell lines, but the exact mechanism is unclear. The cytochromes P450 CYP1B1, CYP1A1, and CYP1A2 have been shown to participate in the activation of various xenobiotics, compounds derived from the diet as well as chemotherapeutic drugs. CYP1B1 and CYP1A1 have also been proposed as targets for cancer chemotherapy for their differential and selective overexpression in tumour cells. In this study, we aimed to identify a possible mechanism of action for the antiproliferative effect of eupatorin, which can be attributed to CYP1 family-mediated metabolism. Methods: The study focuses on the antiproliferative action of eupatorin on the human breast carcinoma cell line MDA-MB-468 and on a cell line derived from normal mammary tissue, MCF-10A. The cytotoxicity of the flavone, its effect on the cell cycle of the abovementioned cell lines, and its metabolism by CYP1 family enzymes were examined. Results: Eupatorin showed a dose-dependent inhibitory effect of cell growth on MDA-MB-468 cells with a submicromolar median inhibition concentration (IC₅₀) whereas the IC₅₀ of this compound in MCF-10A cells was considerably higher. The antiproliferative effect, as measured by EROD (ethoxyresorufin-O-deethylase) assay and Western immunoblotting, was attributed mainly to CYP1A1 expression in MDA-MB-468 cells but not in MCF-10A cells. Moreover, CYP1 family enzymes were shown to metabolise eupatorin in vitro to the flavone cirsiliol and two other unidentified metabolites. Metabolism of eupatorin was also detected in MDA-MB-468 cell cultures, whereas metabolism by MCF-10A cells was negligible. Eupatorin was further shown to arrest the cell cycle of the CYP1-expressing cell line MDA-MB-468 in G₂/M phase, whereas no effect was observed in MCF-10A cells, which do not express CYP1 enzymes. The effect of eupatorin on the MDA-MB-468 cell cycle could be reversed by co-application of the CYP1 inhibitor acacetin. Conclusion: The flavone eupatorin is selectively activated in breast cancer cells, but not in normal breast cells, due to CYP1 family metabolism. This provides a basis for selectivity which is desired against breast tumour cells. In this sense, eupatorin is shown by this study to be a very promising chemopreventative candidate that should be examined further in an in vivo study.