2-PHENYLPHENANTHRIDINONE AND RELATED-COMPOUNDS - ARYL-HYDROCARBON RECEPTOR AGONISTS AND SUICIDE INACTIVATORS OF P4501A1

The effects of several a-substituted phenanthridinones (2-nitro-, 2-t- butyl-, 2-bromo-, 2-phenyl-, 2-ethyl-, 2-methoxy-, 2-iodo-, 2-n-butyl- , 2-chloro-, 2-trifluoromethyl-, 2-fluoro-, 2-isopropyl-, and 2-methyl ) and phenanthridinone as ligands for the rat liver cytosolic aryl hyd rocarbon (Ah) receptor were determined using a competitive binding ass ay and 2,3,7,8-[H-3]tetrachlorodibenzo-p-dioxin (TCDD) as the radiolig and. The competitive binding IC50 values varied from 317 (2-trifluorom ethyl-) to 5870 nM (2-methoxyphenanthridinone); the relative low Ah re ceptor binding affinities for these compounds were paralleled by their weak activity as inducers of ethoxyresorufin O-deethylase (EROD) acti vity in rat hepatoma H4IIE cells; however, there was not a correlation between their structure-binding and structure-induction relationship. In cells cotreated with 1 nM TCDD plus different concentrations (0.01 -10 mu M) of the 2-substituted phenanthridinones, several of these com pounds inhibited induction of EROD activity by TCDD; 2-t-butyl- and 2- phenylphenanthridinone (2-PP) were the most active compounds, causing a >80% reduction in the induced response. 2-PP was selected as a model to further investigate the mechanism of this inhibitory response. The results of interactive studies in rat hepatoma H4IIE cells cotreated with 2-PP plus TCDD or [H-3]TCDD showed that 2-PP did not inhibit form ation of the nuclear Ah receptor complex or induction of CYP1A1 mRNA l evels or CYP1A1 protein. In contrast, incubation of 2-PP with either r at hepatoma H4IIE cells treated with TCDD or hepatic microsomes from T CDD-treated rats resulted in a rapid loss of EROD activity. In paralle l experiments, [H-3]2-PP was incubated with hepatic microsomes from TC DD-treated rats and analysis by denaturing electrophoresis showed that [H-3]2-PP formed a covalent adduct with a 50- to 55-kDa protein and t hus acted as a suicide inactivator of CYP1A1. (C) 1994 Academic Press, Inc.