Involvement of Cytochrome P450 3A4 Enzyme in the N-Demethylation of Methadone in Human Liver Microsomes

Abstract

Methadone has become one of the most widely used drugs for opiate dependency treatment. This drug is extensively metabolized by the cytochrome P450 hepatic enzyme family in man, yielding an N-demethylated metabolite that cyclizes spontaneously into 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine. The specific forms of cytochrome P450 involved in this oxidative N-demethylation were examined in a panel of 20 human liver microsomal preparations previously characterized with respect to their P450 enzyme contents. Methadone was demethylated with an apparent Km of 545 +/- 258 microM (n = 3). The metabolic rates were 745 +/- 574 pmol/(min.mg of protein). This metabolic pathway was strongly correlated with estradiol 2-hydroxylation, testosterone 6 beta-hydroxylation, nifedipine oxidation, erythromycin N-demethylation, and toremifene N-demethylation, all of these monooxygenase activities being supported by P450 3A4. Furthermore, the total P450 3A content of liver microsomal samples, determined by immuno-quantification using a monoclonal anti-human P450 3A4 antibody, was correlated with methadone demethylation (r = 0.72; p < 0.003). Methadone metabolism was 60-72% inhibited either by three mechanism-based inhibitors of P450 3A4 (gestodene, TAO, and erythralosamine) or by four reversible inhibitors of P450 3A (ketoconazole, dihydroergotamine, quercetin, and diazepam with an apparent Ki of 50 microM) and by two nonspecific inhibitors (metyrapone and SKF-525A). Conversely, quinidine (inhibitor of P450 2D6), 7,8-benzoflavone (inhibitor of P450 1A), or sulfaphenazole (inhibitor of P450 2C) did not significantly inhibit, and may even have activated, methadone metabolism. Four heterologously expressed P450 proteins were able to catalyze the N-demethylation of methadone, namely, P450 2C8, P450 2C18, P450 2D6, and P450 3A4. However, referring to their relative liver content, it can be asserted that P450 3A4 is the major enzyme involved in the N-demethylation of methadone on average. Accordingly, caution should be advised in the clinical use of methadone when other drugs are also administered that induce or inhibit P450 3A4, such as rifampicin or diazepam, respectively.

Differential in vitro inhibition of M3G and M6G formation from morphine by (R)‐ and (S)‐methadone and structurally related opioids

“…Methadone is primarily metabolized by the cytochrome P450 enzyme system, specifically CYP3A4, to 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) [25][26][27]. Although not a substrate for glucuronidation itself, methadone inhibits the glucuronidation of 3 ′ -azido-3 ′ -deoxythmidine (AZT) in vitro [28].…”

“…In this investigation, it was also observed that methadone-maintained subjects had higher plasma morphine concentrations than non-opioid-dependent controls when given the same i.v. dose, suggesting decreased morphine clearance in this population.Methadone is primarily metabolized by the cytochrome P450 enzyme system, specifically CYP3A4, to 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) [25][26][27]. Although not a substrate for glucuronidation itself, methadone inhibits the glucuronidation of 3 ′ -azido-3 ′ -deoxythmidine (AZT) in vitro [28].…”

Inhibitory Effects of Juices Prepared from Individual Vegetables on CYP3A4 Activity in Recombinant CYP3A4 and LS180 Cells

“…9) In addition, quercetin is known to be a reversible inhibitor of CYP3A4 activity. 15) Therefore, quercetin contained in onion may be considered to be a major inhibitory component in the mixture of orange vegetables juices.…”

A Review of the Toxicity of HIV Medications II: Interactions with Drugs and Complementary and Alternative Medicine Products

“…Methadone is a racemic synthetic opioid that acts by binding the μ-opioid receptor, with the affinity for the R-enantiomer being 10 times higher than that for the S-enantiomer [74]. Methadone is a substrate of CYP2B6 and 2C19 and also of pgp, while the role of CYP3A4 remains debated [75,76]. When methadone maintenance patients were started on NNRT Is such as efavirenz and nevirapine, methadone AUCs were observed to decrease to 55 and 63 %, respectively, precipitating withdrawal in nine out of 10 participants [77,78].…”