Cytochrome P450 2E1 is the primary enzyme responsible for low-dose carbon tetrachloride metabolism in human liver microsomes

• Published in: Chemico-biological interactions 2000-03, Vol.125 (3), p.233-243
• Main Authors: Zangar, Richard C, Benson, Janet M, Burnett, Vicki L, Springer, David L
• Format: Article
• Language: English
• Subjects: Acetonitriles - pharmacology; Animals; Antibodies - pharmacology; Aryl Hydrocarbon Hydroxylases; Carbon tetrachloride; Carbon Tetrachloride - metabolism; CYP2E1; Cytochrome P-450 CYP2B6; Cytochrome P-450 CYP2E1 - immunology; Cytochrome P-450 CYP2E1 - metabolism; Cytochrome P-450 CYP2E1 Inhibitors; Cytochrome P-450 Enzyme Inhibitors; Cytochrome P-450 Enzyme System - immunology; Cytochrome P-450 Enzyme System - metabolism; Enzyme Inhibitors - pharmacology; Human; Humans; In Vitro Techniques; Liver; Microsomes, Liver - drug effects; Microsomes, Liver - enzymology; Microsomes, Liver - metabolism; Oxidoreductases, N-Demethylating - antagonists & inhibitors; Oxidoreductases, N-Demethylating - immunology; Oxidoreductases, N-Demethylating - metabolism; Rodentia; Trichloromethyl radical
• ISSN: 0009-2797; 1872-7786
• DOI: 10.1016/S0009-2797(00)00149-6

We examined which human CYP450 forms contribute to carbon tetrachloride (CCl 4) bioactivation using hepatic microsomes, heterologously expressed enzymes, inhibitory antibodies and selective chemical inhibitors. CCl 4 metabolism was determined by measuring chloroform formation under anaerobic conditions. Pooled human microsomes metabolized CCl 4 with a K m of 57 μM and a V max of 2.3 nmol CHCl 3/min/mg protein. Expressed CYP2E1 metabolized CCl 4 with a K m of 1.9 μM and a V max of 8.9 nmol CHCl 3/min/nmol CYP2E1. At 17 μM CCl 4, a monoclonal CYP2E1 antibody inhibited 64, 74 and 83% of the total CCl 4 metabolism in three separate human microsomal samples, indicating that at low CCl 4 concentrations, CYP2E1 was the primary enzyme responsible for CCl 4 metabolism. At 530 μM CCl 4, anti-CYP2E1 inhibited 36, 51 and 75% of the total CCl 4 metabolism, suggesting that other CYP450s may have a significant role in CCl 4 metabolism at this concentration. Tests with expressed CYP2B6 and inhibitory CYP2B6 antibodies suggested that this form did not contribute significantly to CCl 4 metabolism. Effects of the CYP450 inhibitors α-naphthoflavone (CYP1A), sulfaphenazole (CYP2C9) and clotrimazole (CYP3A) were examined in the liver microsome sample that was inhibited only 36% by anti-CYP2E1 at 530 μM CCl 4. Clotrimazole inhibited CCl 4 metabolism by 23% but the other chemical inhibitors were without significant effect. Overall, these data suggest that CYP2E1 is the major human enzyme responsible for CCl 4 bioactivation at lower, environmentally relevant levels. At higher CCl 4 levels, CYP3A and possibly other CYP450 forms may contribute to CCl 4 metabolism.