METABOLISM OF 4-(METHYLNITROSAMINO)-1-(3-PYRIDYL)-1-BUTANONE IN MOUSE LUNG MICROSOMES AND ITS INHIBITION BY ISOTHIOCYANATES

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  • 1 November 1990
    • journal article
    • research article
    • Vol. 50  (21) , 6817-6822
Abstract
The tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) induces lung tumors in rats, mice, and hamsters, and metabolic activation is required for the carcinogenicity. 2-Phenethylisothiocyanate (PEITC), whose precursor gluconasturtiin (a glucosinolate) occurs in cruciferous vegetables, has been found to inhibit carcinogenesis by NNK. The purpose of the study was to investigate the enzymes involved in the metabolism of NNK in lung microsomes and to elucidate the mechanisms of inhibition of NNK metabolism by isothiocyanates. NNK metabolism in lung microsomes (isolated from female A/J mice) resulted in the formation of formaldehyde, 4-hydroxy-1-(3-pyridyl)-1-butanone (keto alcohol), 4-oxo-4-(3-pyridyl)butyric acid (keto acid), 4-(methylnitrosamino)-1-(3-pyridyl-N-oxide)-1-butanone, and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol, displaying apparent Km values of 5.6, 5.6, 9.2, 4.7, and 2540 .mu.M, respectively. Higher Km values in the formation of formaldehyde and keto alcohol were also observed. When cytochrome P-450 inhibitors [2-(diethylamino)ethyl 2,2-diphenylpentenoate] hydrochloride (100 .mu.M), carbon monoxide (90%), and 9-hydoxyellipticine (10 .mu.M) were used, NNK metabolism was inhibited by each 70, 100, and 30%, respectively. Methimazole (1 mM), an inhibitor of the flavin-dependent monooxygenase, inhibited the formation of 4-(methylnitrosamino)-1-(3-pyridyl-N-oxide)-1-butanone and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol by 20%, but had no effect on the formation of keto alcohol. Inhibitory antibodies against cytochromes P-450IIB1 and -2, P-450IA1, and P-450IA2 inhibited the formation of keto alcohol by 25, 15, and 0%, respectively. Administration of PEITC at doses of 5 and 25 .mu.mol/mouse 2 h before sacrifice produced a 40 and 70% decrease in microsomal NNK metabolism, respectively. PEITC and 3-phenylpropyl isothiocyanate exhibited a mixed type of inhibition, and the competitive component of inhibition had apparent Ki values of 90 and 30 nM, respectively. Preincubation of PEITC in the presence of a NADPH-generating system did not result in a further decrease in the formation of NNK metabolites, indicating that the metabolism of PEITC was not required for the inhibition. When a series of isothiocyanates with varying alkyl chain length (phenylisothiocyanate, benzylisothiocyanate, PEITC, 3-phenylpropylisothiocyanate, and 4-phenylbutylisothiocyanate) were used, the potency of the inhibition increased with the increase in chain length. The results suggest that cytochrome P-450 are involved in the metabolism of NNK and that the inhibitory action of isothiocyanates is due to competitive inhibition and an inactivation of the enzyme by these compounds.

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