Enhanced Platelet Response to Clopidogrel in Zucker Diabetic Fatty Rats due to Impaired Clopidogrel Inactivation by Carboxylesterase 1 and Increased Exposure to Active Metabolite.

Abstract

Clopidogrel (Clop), a thienopyridine antiplatelet prodrug, is metabolized by cytochrome P450s (CYPs) to an active metabolite, Clop-AM, and hydrolyzed by carboxylesterase (CES)1 to the inactive Clop-acid. Patients with type 2 diabetes (T2DM) tend to have a poor response to Clop due to reduced generation of Clop-AM. Whether a similar response occurs in the Zucker diabetic fatty (ZDF) rat, a commonly used animal model of T2DM, has not been explored. In this work, we compared ZDF and control rats for hepatic CES1- and CYP-mediated Clop metabolism; pharmacokinetics of Clop, Clop-AM, and Clop-acid; and the antiplatelet efficacy of Clop. In contrast to clinical findings, Clop-treated ZDF rats displayed significantly less (50%) maximum platelet aggregation at 4 hours than control rats; the enhanced efficacy was accompanied by higher formation of Clop-AM and lower formation of Clop-acid. In vitro studies showed that hepatic levels of CES1 protein and activity and Ces1e mRNA were significantly lower in ZDF than in control rats, as were the mRNA levels of CYP2B1/2, CYP2C11, and CYP3A2, and levels of CYP2B6-, CYP2C19-, and CYP3A4-related proteins and enzymatic activities in liver microsomes of ZDF rats. Interestingly, liver microsomes of ZDF rats produced higher levels of Clop-AM than that of control rats despite their lower CYP levels, although the addition of fluoride ion, an esterase inhibitor, enhanced Clop-AM formation in control rats more than in ZDF rats. These results suggest that the reduction in CES1-based Clop inactivation indirectly enhances Clop efficacy in ZDF rats by making more Clop available for CYP-mediated Clop-AM formation.