Metabolism of Nicotine and Cotinine by Human Cytochrome P450 2a13

Abstract

ABSTRACT:Nicotine, a major constituent of tobacco, plays a critical role in smoking addiction. In humans, nicotine is primarily metabolized to cotinine, which is further metabolized to trans-3-hydroxycotinine. Recently, we have demonstrated that heterologously expressed human CYP2A13 is highly active in the metabolism of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a nicotine-derived carcinogen. In the present study, CYP2A13-catalyzed NNK metabolism was found to be inhibited competitively by nicotine and Nnitrosonornicotine (NNN), suggesting that both nicotine and NNN are also substrates of CYP2A13. We have further demonstrated that human CYP2A13 is indeed an efficient enzyme in catalyzing C-oxidation of nicotine to form cotinine, with the apparent K m and V max values of 20.2 M and 8.7 pmol/min/pmol, respectively. CYP2A13 also catalyzes the 3-hydroxylation of cotinine to form trans-3-hydroxycotinine, with the apparent K m and V max values of 45.2 M and 0.7 pmol/min/pmol, respectively. The importance of CYP2A13-catalyzed nicotine and cotinine metabolism in vivo remains to be determined.Nicotine is a major constituent of tobacco, playing a critical role in establishing and maintaining tobacco dependence (Henningfield et al., 1985). The major pathway of nicotine metabolism in humans is the C-oxidation to form cotinine, which is a two-step process. The first step is catalyzed by the cytochrome P450 (P450) system to produce the intermediate nicotine-⌬ Ϫ1Ј(5Ј) -iminium ion, which is further oxidized to cotinine by cytosolic aldehyde oxidase (Gorrod and Hibberd, 1982;Peterson et al., 1987;Williams et al., 1990). Other major metabolites of nicotine identified in human urine so far include trans-3Ј-hydroxycotinine and nicotine NЈ-oxide (Jacob et al., 1988). The conversion of cotinine to 3Ј-hydroxycotinine is highly stereospecific in vivo, with the trans isomer as a predominating product (Jacob et al., 1990). trans-3Ј-Hydroxycotinine is excreted in the urine to a much greater extent than cotinine itself. Cotinine may also be metabolized to 5Ј-hydroxycotinine, norcotinine, and cotinine N-oxide (Murphy et al., 1999).In humans, liver is the primary site of nicotine metabolism. Nicotine is also metabolized to some extent in the lung and kidney (Benowitz et al., 1987). Previous studies demonstrated that human CYP2A6 is a key enzyme for the metabolism of nicotine to cotinine, and cotinine to trans-3Ј-hydroxycotinine (Flammang et al., 1992;McCracken et al., 1992; Nakajima et al., 1996a,b;Shimada et al., 1996;Messina et al., 1997). CYP2A6 is the best characterized member in the CYP2A subfamily (Pelkonen and Raunio, 1995; FernandezSalguero and Gonzalez, 1995;Pelkonen et al., 2000). The other two members are CYP2A7 and CYP2A13, which share a high degree of sequence identity with CYP2A6 (Fernandez-Salguero and Gonzalez, 1995). Although the transcripts of these three genes were all found in the liver, CYP2A6 was more abundant than CYP2A7 and CYP2A13 (Koskela et al., 1999). The highest level of CYP2A13 mRNA was found in huma...

Nicotine and 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone Binding and Access Channel in Human Cytochrome P450 2A6 and 2A13 Enzymes

“…The human CYP2A enzymes 2A13 and 2A6 are 94% identical at the amino acid sequence level, but there are substantial differences in their metabolism of the key pharmacologically important agents in tobacco, primarily the addictive agent nicotine and a key procarcinogen, NNK. Although the primary nicotine metabolite for both enzymes results from 5Ј-oxidation, CYP2A13 demonstrates 5-23-fold higher in vitro enzymatic efficiency (11,31). Nicotine also binds to CYP2A13 with a binding affinity 8.7-fold greater than CYP2A6.…”

“…Activation of NNK in the lung results in DNA adducts and lung tumors in mice (10) and is thought to be a potent determinant for lung adenocarcinoma in smokers. The catalytic efficiency of CYP2A13 with nicotine is also 22-fold higher than CYP2A6 (11), although rapid nicotine distribution suggests that hepatic CYP2A6 is likely primarily responsible for most phase I metabolism of nicotine in vivo (3). Although the CYP2A6 and CYP2A13 enzymes are 94% identical, the bases for differential binding and metabolism of nicotine and NNK are unknown.…”

CYP2A13 Metabolizes the Substrates of Human CYP1A2, Phenacetin, and Theophylline

“…CYP2A6 is involved in the metabolism of coumarin and nicotine and the metabolic activation of tobacco-specific nitrosamines such as 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (Yamano et al, 1990;Tiano et al, 1993;Nakajima et al, 1996). CYP2A13 is also active toward these CYP2A6 substrates von Weymarn and Murphy, 2003;Bao et al, 2005). Although CYP2A13 is less active for coumarin 7-hydroxylation than CYP2A6, it is much more active for nicotine, cotinine, and especially 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone metabolisms He et al, 2004;Bao et al, 2005).…”

Selective Expression of CYP2A13 in Human Pancreatic α-Islet Cells

“…CYP2A13 can also metabolically activate 4-aminobiphenyl, another carcinogen in cigarette smoke (Nakajima et al, 2006). In addition, CYP2A13 is highly efficient in metabolizing nicotine (Bao et al, 2005) as well as the metabolic activation of aflatoxin B 1 , naphthalene, and styrene (Fukami et al, 2008).…”