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ABSORPTION, DISTRIBUTION, METABOLISM, AND EXCRETION

Inactivation of CYP2A6 and CYP2A13 during Nicotine Metabolism

Department of Biochemistry, Molecular Biology, and Biophysics and The Cancer Center University of Minnesota, Minneapolis, Minnesota

Nicotine is the major addictive agent in tobacco. The primary catalyst of nicotine metabolism in humans is CYP2A6. However, the closely related enzyme CYP2A13 is a somewhat better catalyst. CYP2A13 is an extrahepatic enzyme that is an excellent catalyst of the metabolic activation of the tobacco-specific carcinogen 4-(methylnitrosamine)-1-(3-pyridyl)-1-butanone (NNK). Here we report that both CYP2A6 and CYP2A13 were inactivated during nicotine metabolism. Inactivation of both enzymes was dependent on NADPH and increased with time and concentration. Alternate substrates for CYP2A6 and CYP2A13 protected these enzymes from inactivation. Inactivation of CYP2A13 was irreversible upon extensive dialysis and seems to be mechanism-based. The K_I of CYP2A13 inactivation by nicotine was 17 µM, the rate of inactivation, k_inact, was 0.1 min^-1, and the t_1/2 was 7 min. However, the loss in enzyme activity occurred after nicotine metabolism was complete, suggesting that a secondary or possible tertiary metabolite of nicotine may be responsible. [5-³H]Nicotine metabolism by CYP2A13 was monitored by radioflow high-pressure liquid chromatography during the course of enzyme inactivation; the major product was the ^1'(5')iminium ion. However, cotinine was a significant metabolite even at short reaction times. The metabolism of the nicotine ^1'(5')iminium ion to cotinine did not require the addition of aldehyde oxidase. CYP2A13 catalyzed this reaction as well as further metabolism of cotinine to 5'-hydroxycotinine, trans-3'-hydroxycotinine, and N-(hydroxymethyl)-norcotinine as enzyme inactivation occurred. Studies are on-going to identify the metabolite responsible for nicotine-mediated inactivation of CYP2A13.

Address correspondence to: Dr. Sharon E. Murphy, The Cancer Center, University of Minnesota, Mayo Mail Code 806, 420 Delaware Street, S.E., Minneapolis, MN 55455. E-mail: murph062{at}umn.edu

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				P. J. Brown, L. L. Bedard, K. R. Reid, D. Petsikas, and T. E. Massey Analysis of CYP2A Contributions to Metabolism of 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone in Human Peripheral Lung Microsomes Drug Metab. Dispos., November 1, 2007; 35(11): 2086 - 2094. [Abstract] [Full Text] [PDF]