In Vitro Oxidative Metabolism of 6-Mercaptopurine in Human Liver: Insights into the Role of the Molybdoflavoenzymes Aldehyde Oxidase, Xanthine Oxidase, and Xanthine Dehydrogenase

Abstract

Anticancer agent 6-mercaptopurine (6MP) has been in use since 1953 for the treatment of childhood acute lymphoblastic leukemia (ALL) and inflammatory bowel disease. Despite being available for 60 years, several aspects of 6MP drug metabolism and pharmacokinetics in humans are unknown. Molybdoflavoenzymes such as aldehyde oxidase (AO) and xanthine oxidase (XO) have previously been implicated in the metabolism of this drug. In this study, we investigated the in vitro metabolism of 6MP to 6-thiouric acid (6TUA) in pooled human liver cytosol. We discovered that 6MP is metabolized to 6TUA through sequential metabolism via the 6-thioxanthine (6TX) intermediate. The role of human AO and XO in the metabolism of 6MP was established using the specific inhibitors raloxifene and febuxostat.Both AO and XO were involved in the metabolism of the 6TX intermediate, whereas only XO was responsible for the conversion of 6TX to 6TUA. These findings were further confirmed using purified human AO and Escherichia coli lysate containing expressed recombinant human XO. Xanthine dehydrogenase (XDH), which belongs to the family of xanthine oxidoreductases and preferentially reduces nicotinamide adenine dinucleotide (NAD + ), was shown to contribute to the overall production of the 6TX intermediate as well as the final product 6TUA in the presence of NAD + in human liver cytosol. In conclusion, we present evidence that three enzymes, AO, XO, and XDH, contribute to the production of 6TX intermediate, whereas only XO and XDH are involved in the conversion of 6TX to 6TUA in pooled HLC.

Review article: recent advances in pharmacogenetics and pharmacokinetics for safe and effective thiopurine therapy in inflammatory bowel disease

“…Both AOX and XO were involved in the metabolism of the 6TX intermediate, whereas only XO was responsible for the conversion of 6TX to 6TUA. Xanthine dehydrogenase (XDH), which belongs to the family of xanthine oxidoreductases and preferentially reduces nicotinamide adenine dinucleotide (NAD + ), was shown to contribute to the overall production of the 6TX intermediate as well as the final product 6TUA in the presence of NAD + in human liver cytosol …”

Determination of Urinary Caffeine Metabolites as Biomarkers for Drug Metabolic Enzyme Activities

“…Xanthine oxidase is important in gout patients, because XO produces uric acid, which is a crucial factor in gout. In addition, xanthine oxidase is involved in the catabolism of xenobiotics; for example, it converts a prodrug (mercaptopurine) into the active form 6-thioinosine-5′triphosphate [30]. About 20 genetic variants are reported, and each XO variant may differ in its enzymatic activity [20]; however, decreased enzyme activity is shown only in 4% or fewer volunteers [31].…”

Role of Molybdenum-Containing Enzymes in the Biotransformation of the Novel Ghrelin Receptor Inverse Agonist PF-5190457: A Reverse Translational Bed-to-Bench Approach

“…An increasing number of compounds in the past few decades have been identified to be substrates of these enzymes, especially involving AO, which also has broader substrate specificity in comparison with XO (O'Connor et al, 2006;Pryde et al, 2010;Sanoh et al, 2015;Battelli et al, 2016). There are only a few substrates that show biotransformation by both enzymes, such as 6-deoxyclovir, 6-thioxanthine, and recently VU0409106, a lead compound for childhood developmental disorders (Krenitsky et al, 1984;Morrison et al, 2012;Choughule et al, 2014). Although some overlap exists between the substrates, there have been chemical inhibitors identified specifically for AO and XO including raloxifene (Obach, 2004), hydralazine (Strelevitz et al, 2012), allopurinol (Panoutsopoulos et al, 2004), and febuxostat (Weidert et al, 2014).…”