N-Glucuronidation of Carbamazepine in Human Tissues Is Mediated by UGT2B7

Abstract

Carbamazepine (CBZ) is one of the most widely prescribed anticonvulsants despite a high incidence of idiosyncratic side effects. Metabolism of CBZ is complex, and of the more than 30 metabolites identified, one of the most abundant is CBZ Nglucuronide. To date the uridine diphosphate glucuronosyltransferase (UGT) isoform responsible for the N-glucuronidation of CBZ has not been identified. We have developed a sensitive liquid chromatography/mass spectrometry assay to quantify CBZ glucuronidation, and we report that CBZ is specifically glucuronidated by human UGT2B7. Kinetics of CBZ glucuronidation in human liver, kidney, and intestine microsomes were consistent with those of recombinant UGT2B7, which displayed a K m value of 214 M and V max value of 0.79 pmol/mg/min. In addition to revealing the isoform responsible for CBZ glucuronidation, this is the first example of primary amine glucuronidation by UGT2B7.Carbamazepine (5H-dibenzo[b,f]azepine-5-carboxamide) is one of the most widely prescribed anticonvulsants and is used to treat a variety of conditions from epilepsy to muscle spasm and trigeminal neuralgia. However its use is associated with a number of idiosyncratic adverse side effects, including skin rash, blood disorders, and hepatitis in onethird to one-half of patients (Ju and Uetrecht 1999). These adverse side effects have been associated with the formation of CBZ metabolites (Shear and Spielberg 1988;Riley et al., 1989); therefore, the study of all CBZ metabolites has important clinical implications.The metabolism of CBZ is complex and has been widely studied in human and in animal models (Madden et al., 1996;Maggs et al., 1997) with over 30 metabolites (Lertratanangkoon and Horning, 1982) identified. The major metabolites are the 10,11-epoxide and its hydrolytic trans-dihydrodiol product. Glucuronidation is also an important detoxification pathway because the CBZ N-glucuronide and glucuronides of the hydroxylated metabolites are significant urinary metabolites; however, to date no glucuronide metabolite has been implicated in the incidence of side effects. Epoxide hydrolase has been the focus of most attention as a potential source of toxic metabolites; however, the available data do not support a major role for this enzyme in causing side effects (Pirmohamed et al., 1992;Green et al., 1995b). One minor metabolite, 2-hydroxy-CBZ, formed through loss of the carboxamide, could be metabolized to an iminoquinone, which due to its potential chemical reactivity might be the metabolite responsible for the idiosyncratic reactions, although this has not been shown to date (Ju and Uetrecht, 1999). In addition CBZ is also a well known enzyme inducer up-regulating cytochrome P450 (Luo et al., 2002) and UGT activity/expression (Tanaka, 1999) Carbamazepine is metabolized to an N-glucuronide (Bauer et al., 1976); in addition, glucuronide metabolites have been demonstrated for all 13 of the hydroxylated metabolites of CBZ (Maggs et al., 1997). Formation of N-glucuronides has been principally studied in nonro...

Characterization of 1′-Hydroxymidazolam Glucuronidation in Human Liver Microsomes

“…UGT2B7 is a very important human UGT isoform in that it appears to be expressed in many tissues besides liver, and it catalyzes the glucuronidation of a wide range of xenobiotics, including polycyclic aromatic hydrocarbons, phenols, opioids, aliphatic alcohols, carboxylic acids, and tetrazoles (King et al, 2000). Recently, UGT2B7 was reported to catalyze the N-glucuronidation of an amide and a primary amine (Staines et al, 2004;Zhang et al, 2004). In contrast, UGT2B4 catalyzes only a limited number of substrates, and the data so far are not consistent among different laboratories.…”

Human UDP-Glucuronosyltransferase (UGT) 2B10 in DrugN-Glucuronidation: Substrate Screening and Comparison with UGT1A3 and UGT1A4

“…Desipramine and nortriptyline carry secondary amines, and were previously reported not to be glucuronidated by UGT1A4 (Green and Tephly, 1996). Carbamazepine, a substrate of UGT2B7, contains a primary amine (Staines et al, 2004). Afloqualone, diphenhydramine, tamoxifen, ketoconazole, and midazolam were also examined in the present study for glucuronidation by UGT2B10 because these drugs have amine structures and were reported to be substrates for UGT1A4 (Green and Tephly, 1996;Kaku et al, 2004;Kaji and Kume, 2005;Klieber et al, 2008;Bourcier et al, 2010).…”

Regio- and Stereospecific N-Glucuronidation of Medetomidine: The Differences between UDP Glucuronosyltransferase (UGT) 1A4 and UGT2B10 Account for the Complex Kinetics of Human Liver Microsomes

“…Among the human UGTs, UGT1A4 was largely considered as the enzyme "specializing" in N-glucuronidation because it is capable of conjugating different types of amines: primary aromatic amines, secondary and tertiary aliphatic amines, and secondary and tertiary aromatic N-heterocycles (Green and Tephly, 1998;Kaivosaari et al, 2002;Zenser et al, 2002;Kuehl and Murphy, 2003;Rowland et al, 2006). Several other human UGTs, including 1A1, 1A3, 1A7, 1A9, and 2B7, were documented to catalyze different N-glucuronidation reactions (Green and Tephly, 1998;Kaivosaari et al, 2002;Zenser et al, 2002;Staines et al, 2004;Girard et al, 2005;Kaji and Kume, 2005;Borlak et al, 2006;Rowland et al, 2006;Omura et al, 2007). Nevertheless, for all the latter enzymes, N-glucuronidation seems to be a minor reaction, whereas for UGT1A4, it is probably the major type of activity.…”