Interaction of cotinine with rat hepatic microsomal P-450. Comparison with metyrapone and immunomodulation of cotinine and metyrapone binding by monoclonal anti-cotinine antibodies

RJ Bjercke, DK Hammond, HW Strobel and JJ Langone

Department of Medicine, Baylor College of Medicine, Houston 77030.

The ability of the major nicotine metabolite, cotinine, to interact with rat liver microsomal cytochrome P-450 and the immunomodulatory effects of anti-cotinine antibodies were studied. Cotinine induced type II spectral changes with both microsomes from phenobarbital (PB)- induced rats and purified P-450 with apparent Ks values of 97 and 750 microM, respectively. In contrast, the Ks value was 0.3 microM for metyrapone and 5 microM for nicotine with both the microsomes and purified enzyme. The apparent Ki value for cotinine inhibition of 7- pentoxyresorufin O-dealkylase activity with the microsomes (87 microM) was approximately 87- and 870-fold higher than for nicotine and metyrapone, respectively. Monoclonal antibodies produced against cotinine cross-reacted equally well with metyrapone. They specifically blocked enzyme binding of both drugs based on dose-dependent inhibition of spectral changes, and reversed the metyrapone-induced inhibition of microsomal O-dealkylase activity. In contrast, antibodies to nicotine did not cross-react with cotinine or metyrapone and had no effect on their activity, although they did block the action of nicotine. These results demonstrate that cotinine binding to P-450 from PB-induced rats and inhibition of functional activity in vitro are qualitatively like the effects of metyrapone and nicotine, and that monoclonal anti- cotinine antibodies are useful molecular probes of the interactions between cotinine and metyrapone with the enzyme.

Volume 18, Issue 5, pp. 759-764, 09/01/1990
Copyright © 1990 by American Society for Pharmacology and Experimental Therapeutics