Abstract

Desloratadine (Clarinex^®), the major active metabolite of loratadine (Claritin^®), is a non-sedating long-lasting antihistamine widely used for the treatment of allergic rhinitis and chronic idiopathic urticaria. For over 20 years, it has remained a mystery as to which enzymes are responsible for the formation of 3-hydroxydesloratadine, the major active human metabolite, largely due to the inability of any in vitro system tested thus far to generate this metabolite. In this study, we demonstrated that cryopreserved human hepatocytes (CHH) form 3-hydroxydesloratadine and its corresponding O-glucuronide. CHHs catalyzed the formation of 3-hydroxydesloratadine with a K_m of 1.6 μM and V_max of 1.3 pmol/min/million cells. Chemical inhibition of cytochrome P450 (CYP) enzymes in CHH demonstrated that gemfibrozil glucuronide (CYP2C8 inhibitor) and 1-aminobenzotriazole (general P450 inhibitor) inhibited 3-hydroxydesloratadine formation by 91% and 98%, respectively. Other inhibitors of CYP2C8 (gemfibrozil, montelukast, clopidogrel glucuronide, repaglinide and cerivastatin) also caused extensive inhibition of 3-hydroxydesloratadine formation (73-100%). Assessment of desloratadine, amodiaquine and paclitaxel metabolism by a panel of individual CHHs demonstrated that CYP2C8 marker activity robustly correlated with 3-hydroxydesloratadine formation (r² of 0.70-0.90). Detailed mechanistic studies with sonicated or saponin-treated CHHs, human liver microsomes and S9 fractions showed that both NADPH and UDP-glucuronic acid are both required for 3-hydroxydesloratadine formation, and studies with recombinant UGT and CYP enzymes implicated the specific involvement of UGT2B10 in addition to CYP2C8. Overall, our results demonstrate for the first time that desloratadine glucuronidation by UGT2B10, followed by CYP2C8 oxidation and a de-conjugation event are responsible for the formation of 3-hydroxydesloratadine.