(1997). Human Cytochrome P450 Enzymes: A Status Report Summarizing Their Reactions, Substrates, Inducers, and Inhibitors. Drug Metabolism Reviews: Vol. 29, No. 1-2, pp. 413-580.

Human Cytochrome P450 Enzymes: A Status Report Summarizing Their Reactions, Substrates, Inducers, and Inhibitors

Despite widespread clinical use of antimalarial drugs such as hydroxychloroquine and chloroquine in the treatment of rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) and other inflammatory rheumatic diseases, insights into the mechanism of action of these drugs are still emerging. Hydroxychloroquine and chloroquine are weak bases and have a characteristic ‘deep’ volume of distribution and a half-life of around 50 days. These drugs interfere with lysosomal activity and autophagy, interact with membrane stability and alter signalling pathways and transcriptional activity, which can result in inhibition of cytokine production and modulation of certain co-stimulatory molecules. These modes of action, together with the drug’s chemical properties, might explain the clinical efficacy and well-known adverse effects (such as retinopathy) of these drugs. The unknown dose–response relationships of these drugs and the lack of definitions of the minimum dose needed for clinical efficacy and what doses are toxic pose challenges to clinical practice. Further challenges include patient non-adherence and possible context-dependent variations in blood drug levels. Available mechanistic data give insights into the immunomodulatory potency of hydroxychloroquine and provide the rationale to search for more potent and/or selective inhibitors. Hydroxychloroquine and chloroquine are antimalarial drugs commonly used for the treatment of rheumatic diseases. Multiple mechanisms might explain the efficacy and adverse effects of these drugs, but further investigation could lead to the development of more specific and potent drugs.

Mechanisms of action of hydroxychloroquine and chloroquine: implications for rheumatology.

Classification of orally administered drugs on the World Health Organization Model list of Essential Medicines according to the biopharmaceutics classification system.

This chapter is an update of the data on substrates, reactions, inducers, and inhibitors of human CYP enzymes published previously by Rendic and DiCarlo (1), now covering selection of the literature through 2001 in the reference section. The data are presented in a tabular form (Table 1) to provide a framework for predicting and interpreting the new P450 metabolic data. The data are formatted in an Excel format as most suitable for off-line searching and management of the Web-database. The data are presented as stated by the author(s) and in the case when several references are cited the data are presented according to the latest published information. The searchable database is available either as an Excel file (for information contact the author), or as a Web-searchable database (Human P450 Metabolism Database, www.gentest.com) enabling the readers easy and quick approach to the latest updates on human CYP metabolic reactions.

Summary of information on human cyp enzymes: human p450 metabolism data

Human P450 metabolism of warfarin.

The metabolism of clozapine by human liver has been investigated in vitro. Irreversible protein-binding and conjunction with model nucleophiles have been used as markers for bioactivation of clozapine, while stable metabolite formation has been assessed using radiometric HPLC. In all nine liver microsomal preparations investigated, clozapine was extensively metabolized to the stable products desmethylclozapine (range 19%-27.2%), N-oxide (1.5-20.5%) and three polar metabolites (0-20.8%), and was bioactivated to a protein-reactive metabolite (0.6-2.1%). The CYP2D6 genotype did not influence the capacity of the livers to form these metabolites. All metabolic pathways were inhibited by ketoconazole, indicating the involvement of the cytochrome P450 enzymes. Isozyme-selective inhibitor studies demonstrated that whereas demethylation was performed by CYP1A2, N-oxidation and chemically reactive metabolite formation were dependent upon multiple forms of P450. The N-oxide was readily reduced back to clozapine in the presence of NADPH, this conversion being inhibited by ascorbic acid. Glutathione (1 mM) decreased covalent binding by 70%. The amount of putative adduct formed in the presence of glutathione (13.4 +/- 0.9%) was much greater than the covalent binding (mean 1.1 +/- 0.2%). The bioactivation of clozapine was, like the N-oxidation of clozapine, a reversible process. In summary, our results indicate clozapine undergoes extensive metabolism by human liver to both stable and chemically reactive metabolites, the formation of which is catalyzed by the cytochrome P450 enzymes. The role of the reactive metabolite, which may be a free radical, in the pathogenesis of clozapine agranulocytosis and hepatotoxicity requires further study.

Metabolism and bioactivation of clozapine by human liver in vitro.

An investigation into the formation of stable, protein-reactive and cytotoxic metabolites from tacrine in vitro. Studies with human and rat liver microsomes.

The metabolism of drugs to chemically reactive metabolites may play a pivotal role in the pathogenesis of idiosyncratic drug toxicity. A large number of in vitro studies and a limited number of in vivo studies have demonstrated that many drugs are not toxic per se, but produce toxicity after undergoing enzyme-mediated bioactivation to chemically reactive species. Such reactive species may inflict a toxic insult on the cell either directly or indirectly by acting as a hapten and initiating an immune-mediated reaction. The enzymes responsible for bioactivation have been widely studied, both quantitatively and qualitatively, the most important being the enzymes of the cytochrome P450 (CYP) mixed function oxidase system. CYP enzymes are the most predominant drug metabolising enzymes in the liver and are also present in most other tissues of the body. The diversity of this enzyme system means that a wide range of xenobiotic substrates can be bioactivated by either a single CYP isoform or multiple isoforms of this enzyme superfamily. Other enzymes do, however, play an important role in drug bioactivation. In white blood cells, for example, myeloperoxidase has been shown to bioactivate a wide range of drugs. In other tissues low in CYP activity, prostaglandin H synthase may also be responsible for bioactivation; e.g. in the kidney paracetamol (acetaminophen) toxicity is thought to result from activation via this enzyme. The phase II or conjugation enzymes may also be important in the ultimate bioactivation of drug molecules. Whilst activation by these enzymes is, to date, apparently confined to chemicals, most drugs are also substrates for these enzymes and bioactivation by them must remain a possibility.

Idiosyncratic Drug Reactions

1. Tacrine (1,2,3,4-tetrahydro-9-aminoacridine-hydrochloride: THA) underwent metabolism in vitro by a panel (n = 12) of human liver microsomes genotyped for CYP2D6, in the presence of NADPH, to both protein-reactive and stable metabolites. 2. There was considerable variation in the extent of THA metabolism amongst human livers. Protein-reactive metabolite formation showed a 10-fold variation (0.6 +/- 0.1%-5.2 +/- 0.8% of incubated radioactivity mg-1 protein) whilst stable metabolites showed a 3-fold variation (24.3 +/- 1.7%-78.6 +/- 2.6% of incubated radioactivity). 3. Using cytochrome P450 isoform specific inhibitors CYP1A2 was identified as the major enzyme involved in all routes of THA metabolism. 4. There was a high correlation between aromatic and alicyclic hydroxylation (r = 0.92, P < 0.0001) consistent with these biotransformations being catalysed by the same enzymes. 5. Enoxacin (ENOX), cimetidine (CIM) and chloroquine (CQ) inhibited THA metabolism by a preferential decrease in the bioactivation to protein-reactive, and hence potentially toxic, species. The inhibitory potency of ENOX and CIM was increased significantly upon pre-incubation with microsomes and NADPH. 6. Covalent binding correlated with 7-OH-THA formation before (r = 0.792, P < 0.0001) and after (r = 0.73, P < 0.0001) inhibition by CIM, consistent with a two-step mechanism in the formation of protein-reactive metabolite(s) via a 7-OH intermediate. 7. The use of enzyme inhibitors may provide a useful tool for examining the relationship between the metabolism and toxicity of THA in vivo.

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S. Madden

Papers

Metabolism and bioactivation of clozapine by human liver in vitro.

An investigation into the formation of stable, protein-reactive and cytotoxic metabolites from tacrine in vitro. Studies with human and rat liver microsomes.

Idiosyncratic Drug Reactions

The effect of enzyme inhibition on the metabolism and activation of tacrine by human liver microsomes

An investigation of the formation of cytotoxic, genotoxic, protein-reactive and stable metabolites from naphthalene by human liver microsomes.