Vol. 296, Issue 2, 329-337, February 2001

Assessment of the Effects of Metabolism on the Estrogenic Activity of Xenoestrogens: A Two-Stage Approach Coupling Human Liver Microsomes and a Yeast Estrogenicity Assay

Robert Elsby, James L. Maggs, John Ashby, David Paton, John P. Sumpter and B. Kevin Park

Department of Pharmacology and Therapeutics, University of Liverpool, Liverpool, United Kingdom (R.E., J.L.M., B.K.P.); AstraZeneca Central Toxicology Laboratory, Alderley Park, Macclesfield, Cheshire, United Kingdom (J.A., D.P.); and Department of Biological Sciences, Brunel University, Uxbridge, Middlesex, United Kingdom (J.P.S.)

Concern that the reproductive health of humans is being affected by exposure to xenoestrogens has led to the development of various in vitro and in vivo screening assays for the identification of suspected xenoestrogens. However, the estrogenic activity of a chemical determined in vitro may not necessarily predict its activity in vivo if the chemical is metabolized during the assay and/or in vivo. Therefore, to investigate the role of metabolism in modulating the estrogenic activity of suspected xenoestrogens, we have devised a two-stage approach coupling incubations with either human or rat hepatic microsomes with a yeast estrogenicity (transcription) assay. We have assessed the activity of the proestrogenic pesticide 99.5% methoxychlor [1,1,1-trichloro-2,2-bis-(4-methoxyphenyl)ethane, MXC] (EC₅₀ = 4.45 ± 1.9 µM, n = 6) and a structural analog, methoxybisphenol A [2,2-bis-(4-methoxyphenyl) propane, MBPA], in the yeast estrogenicity assay and also established that yeast (Saccharomyces cerevisiae), unlike human liver microsomes, are not able to demethylate MXC or MBPA to estrogenic metabolites. This indicates that the proestrogen MXC has weak intrinsic estrogenic activity. Using 99.5% MXC and 17-estradiol as paradigms, we have demonstrated how metabolism can enhance or suppress, respectively, estrogenic activity. The effect of metabolism on the activities of the weak xenoestrogens 3,17-bisdesoxyestradiol [1,3,5(10)-estratriene] and 6-hydroxytetralin (5,6,7,8-tetrahydro-2-naphthol) was also assessed. This two-stage approach can distinguish the estrogenic activity of a suspect chemical from the activity due to its more, or less, active metabolites and will aid in the evaluation of novel xenoestrogens and, more importantly, proestrogens.