Hydroxylation of chlorzoxazone as a specific probe for human liver cytochrome P-450IIE1

Abstract

Human cytochrome P-450IIE1 has been implicated in the oxidation of a number of substrates, including protoxins and -carcinogens. To date, no drugs have been identified that are exclusive substrates for the protein and are applicable for use as noninvasive probes of the in vivo function of the enzyme in humans. Chlorzoxazone was found to be oxidized only to 6-hydroxychlorzoxazone in human liver microsomes. Results of steady-state kinetics are consistent with the view that only a single enzyme catalyzes the reaction. The microsomal reaction was strongly inhibited by rabbit anti-P-450IIE1 and, in a competitive manner, by known P-450IIE1 substrates. Rates of chlorzoxazone 6-hydroxylation in different human liver microsomal preparations were well correlated with levels of immunochemically measured P-450IIE1 and rates of (CH3)2NNO oxidation. Chlorzoxazone 6-hydroxylation was also found to be catalyzed by purified human liver P-450IIE1. These results provide strong evidence that P-450IIE1 is the primary catalyst of chlorzoxazone 6-hydroxylation in human liver. Rates of chlorzoxazone 6-hydroxylation vary considerably among human liver samples, and chlorzoxazone 6-hydroxylation may have potential use as a noninvasive probe in estimating the in vivo expression of human P-450IIE1 and its significance as a risk factor in the toxicity and carcinogenicity of a number of solvents, nitrosamines, and drugs.

Differential induction of ethanol-metabolizing CYP2E1 and nicotine-metabolizing CYP2B1/2 in rat liver by chronic nicotine treatment and voluntary ethanol intake

“…The induction of hepatic CYP2E1 by nicotine not only increases ethanol consumption and ethanol-mediated liver damage, but can also increase the risk for a number of non-ethanol associated liver diseases. Elevated CYP2E1 levels could also alter the metabolism of clinically relevant substrates of CYP2E1 such as acetaminophen (Patten et al, 1993), chlorzoxazone (Peter et al, 1990), isoniazid (Ryan et al, 1986), tamoxifen (Styles et al, 1994), and halothane (Spracklin et al, 1997). The altered metabolism of these drugs could alter the therapeutic efficacy for some drugs and even cause toxicity in the case of drugs with a narrow therapeutic index.…”

Hepatic cytochrome P450 2E1 activity in nondiabetic patients with nonalcoholic steatohepatitis

“…CHZ, a centrally acting muscle relaxant, is primarily hydroxylated by CYP2E1 to 6-hydroxy CHZ and therefore has been extensively used as a selective probe for measuring hepatic CYP2E1 activity in humans. [17][18][19][20][21] Subjects were asked to avoid certain foods (grapefruit, vegetables from the mustard green family, and beverages containing xanthine and alcohol) for at least 72 hours, and, when possible, nonessential medications were discontinued one week before this test to reduce the chances of drug interference. Blood and urine samples were collected for 8 hours after administering a 500-mg dose of chlorzoxazone orally, followed immediately by 240 mL of water.…”

Structures of Human Cytochrome P-450 2E1

“…These substrates include industrial solvents such as benzene, toluene, aniline, and halogenated solvents (18); alcohols such as ethanol (19), glycerol, phenol, and p-nitrophenol (20); and bicyclic heterocycles such as caffeine (21) and the muscle relaxant chlorzoxazone (22). However, CYP2E1 is also known to metabolize endogenous fatty acids, including lipids associated with signaling mechanisms such as arachidonic acid (23) and epoxyeicosatrienoic acids (24).…”