Activity of benzo[a]pyrene and its hydroxylated metabolites in an estrogen receptor-α reporter gene assay

• Published in: Toxicological sciences Vol. 55; no. 2; pp. 320 - 326
• Main Authors: CHARLES, G. D, BARTELS, M. J, ZACHAREWSKI, T. R, GOLLAPUDI, B. B, FRESHOUR, N. L, CARNEY, E. W
• Format: Journal Article
• Language: English
• Published: Cary, NC Oxford University Press 01.06.2000
• Subjects: Benzo(a)pyrene - metabolism; Benzo(a)pyrene - pharmacology; Benzoflavones - pharmacology; beta-Galactosidase - metabolism; Biological and medical sciences; Breast Neoplasms - enzymology; Breast Neoplasms - genetics; Chemical and industrial products toxicology. Toxic occupational diseases; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Estradiol - analogs & derivatives; Estradiol - pharmacology; Estrogen Antagonists - pharmacology; Estrogen Receptor alpha; estrogen receptors; Female; Fulvestrant; Gene Expression - drug effects; Genes, Reporter - drug effects; Genetic Techniques; Humans; Hydroxylation; Luciferases - metabolism; Medical sciences; Receptors, Estrogen - agonists; Receptors, Estrogen - genetics; Receptors, Estrogen - metabolism; Toxicology; Transfection; Tumor Cells, Cultured; Various organic compounds; Human; Mimetic hormone; Hydrocarbon; Metabolite; Toxicity; Estrogen; Endocrine disruptor; Polycyclic aromatic compound; In vitro; Dose activity relation; Ah receptor; Cell line; Phenols; Mammary gland; Mechanism of action; Hormonal receptor
• ISSN: 1096-6080; 1096-0929; 1096-0929
• DOI: 10.1093/toxsci/55.2.320

A human breast cancer cell line, MCF-7, transiently transfected with a chimeric estrogen receptor (Gal4-HEG0) and a luciferase reporter plasmid (17m5-G-Luc), was used to investigate the estrogenic activity of benzo[a]pyrene (B[a]P), a prototypical polyaromatic hydrocarbon (PAH). B[a]P at concentrations > or = 1 microM produced responses comparable to that of 0.1 nM 17beta-estradiol (E2). The ER antagonist ICI 182,780 (ICI) completely inhibited the response to both E2 and B[a]P, indicating that the responses were ER-mediated. However, 2 microM alpha-napthoflavone (alpha-NF), an Ah receptor antagonist and P450 inhibitor, also decreased the response to B[a]P but not to E2. Analysis of the profile of B[a]P metabolites in the transfected MCF-7 cultures indicated that alpha-NF inhibited the production of the 3- and 9-hydroxy (3-OH and 9-OH), as well as the 7, 8- and 9,10-dihydroxy (7,8-OH and 9,10-OH) B[a]P species. In the ER-alpha reporter assay, the 3-OH and 9-OH metabolites produced maximal responses comparable to E2, with EC50 values of 1.2 microM and 0.7 microM, respectively. The 9,10-OH metabolite exhibited minimal activity in the assay. These responses were inhibited by ICI for both the 3-OH and the 9-OH species; however, alpha-NF inhibited only the response to the 9-OH metabolite. The 7,8-OH metabolite did not exhibit significant estrogenic activity. Furthermore, 7,8-OH B[a]P displayed observable cytotoxicity at concentrations > or = 10(-7) M. This cytotoxic response was completely inhibited by alpha-NF, suggesting that 7,8-OH B[a]P was being further metabolized to one or more cytotoxic metabolites.