Reevaluation of the Microsomal Metabolism of Montelukast: Major Contribution by CYP2C8 at Clinically Relevant Concentrations

Abstract

ABSTRACT:According to published in vitro studies, cytochrome P450 3A4 catalyzes montelukast 21-hydroxylation (M5 formation), whereas CYP2C9 catalyzes 36-hydroxylation (M6), the primary step in the main metabolic pathway of montelukast. However, montelukast is a selective competitive CYP2C8 inhibitor, and our recent in vivo studies suggest that CYP2C8 is involved in its metabolism. We therefore reevaluated the contributions of different cytochrome P450 (P450) enzymes, particularly that of CYP2C8, to the hepatic microsomal metabolism of montelukast using clinically relevant substrate concentrations in vitro. The effects of P450 isoform inhibitors on montelukast metabolism were examined using pooled human liver microsomes, and montelukast oxidations by human recombinant CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4, and CYP3A5 were investigated. The results verified the central role of CYP3A4 in M5 formation. The CYP2C8 inhibitors gemfibrozil 1-O-␤ glucuronide and trimethoprim inhibited the depletion of 0.02 M montelukast and formation of M6 from 0.05 M montelukast more potently than did the CYP2C9 inhibitor sulfaphenazole. Likewise, recombinant CYP2C8 catalyzed montelukast depletion and M6 formation at a 6 times higher intrinsic clearance than did CYP2C9, whereas other P450 isoforms produced no M6. On the basis of depletion of 0.02 M montelukast, CYP2C8 was estimated to account for 72% of the oxidative metabolism of montelukast in vivo, with a 16% contribution for CYP3A4 and 12% for CYP2C9. Moreover, CYP2C8 catalyzed the further metabolism of M6 more actively than did any other P450. In conclusion, CYP2C8 plays a major role in the main metabolic pathway of montelukast at clinically relevant montelukast concentrations in vitro.

In Vitro Metabolism of Montelukast by Cytochrome P450s and UDP-Glucuronosyltransferases

“…The sensitivity of the LC-MS/MS was too low to monitor the metabolite a-hydroxy montelukast. In agreement with a previous study (Filppula et al, 2011), the montelukast sulfoxide was present as contamination in the montelukast synthetic standard. Therefore, it was not possible to obtain an accurate quantification of montelukast sulfoxide produced during the metabolism of montelukast; consequently, it was not possible to identify precisely the P450s responsible for its formation.…”

“…We found that 36-hydroxylation accounts for over 83% approximately of the total oxidative in vitro Cl int , whereas 21-and 25-hydroxylation represent minor metabolic routes, which is consistent with previous in vitro Filppula et al, 2011;VandenBrink Kinetic parameters for the formation of 1,2 diol, 21(R)-OH montelukast, 21(S)-OH montelukast, and 25-OH montelukast from montelukast in expressed in CYP2C8, CYP2C9, and CYP3A4…”

“…In an earlier in vitro study and on the product label CYP2C9 and CYP3A4 are reported as the principal P450s in montelukast metabolism; CYP2C9 was considered selective toward montelukast 36-hydroxylation . In contrast, recent in vitro studies implicate CYP2C8 in montelukast 36-hydroxylation (Filppula et al, 2011;VandenBrink et al, 2011), although close examination of the in vitro studies indicate that P450s other than CYP2C8 may also be involved. The potential role of CYP2C8 is further supported by the ability of montelukast to tightly bind to the CYP2C8 active site (Schoch et al, 2008) and to potently and competitively inhibit this enzyme in vitro (Walsky et al, 2005).…”

“…In recent years, montelukast has been recommended as a selective probe of CYP2C8 activity in vitro (Filppula et al, 2011;VandenBrink et al, 2011) and in vivo (Karonen et al, 2010(Karonen et al, , 2012 primarily on the basis of two assumptions: 36-hydroxylation to 1,2-diol and then to montelukast dicarboxylic acid (M4) is the main clearance mechanism of montelukast Karonen et al, 2010Karonen et al, , 2012; and this pathway is selectively catalyzed by CYP2C8 (Filppula et al, 2011;VandenBrink et al, 2011).…”

Dose-Dependent Interaction between Gemfibrozil and Repaglinide in Humans: Strong Inhibition of CYP2C8 with Subtherapeutic Gemfibrozil Doses

“…The importance of CYP2C8-mediated drug interactions is increasing continuously, because the list of CYP2C8 substrates, and therefore the list of potential victim drugs of CYP2C8-mediated interactions, is increasing. To date, for example, paclitaxel, cerivastatin, loperamide, rosiglitazone, repaglinide, amiodarone, amodiaquine, and montelukast have been recognized as CYP2C8 substrates (Rahman et al, 1994;Ohyama et al, 2000;Backman et al, 2002;Wang et al, 2002;Niemi et al, 2003a;Kim et al, 2004;Jaakkola et al, 2005;Kajosaari et al, 2005a;Totah and Rettie, 2005;Niemi et al, 2006;Lai et al, 2009;Karonen et al, 2010;Filppula et al, 2011). When developing new therapeutic agents, it is important, among other things, to assess whether their metabolism is dependent on CYP2C8 (Huang et al, 2007(Huang et al, , 2008.…”