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.

Inflammation is a major regulator of drug metabolizing enzymes and transporters: Consequences for the personalization of drug treatment

“…For example, because studies investigating the impact of diabetes on various animal models were recently reviewed ( Wu & Lin, 2019 ) , this table does not include the works discussed in that review, except those which were published after its publication. b According to Yao et al (2019) , human CYP2C9 and CYP2C19 corresponds to CYP2C11 and CYP2C22 in rats. …”

“… b According to Yao et al (2019) , human CYP2C9 and CYP2C19 corresponds to CYP2C11 and CYP2C22 in rats. …”

Study on the Interaction between Deuterium Clopidogrel and Omeprazole

“…The derived plasma was collected and stored at −80°C till further use. The contents of active metabolites in the plasma were determined by HPLC-MS/MS as described previously [14] on the Agilent 1100 high-performance liquid chromatography, AB SCIEX Q-trap 5500 platform.…”

“…The BCA protein quantitative kit was used for quantitative analysis of liver microsomal proteins, and the cocktail method was used to detect the activities of CYP450 enzymes, including CY-P1A2 (rats: CYP1A2), CYP2B6 (rats: CYP2B1/2/3), CYP2C8 (rats: CYP2C6), CYP2C9 (rats: CYP2C7), CYP2C19 (rats: CYP2C11), CYP2D6 (rats: CYP2D1/4/6/26), and CYP3A4 (rats: CYP3A1). The specific experimental steps from a previously published study were followed [14].…”

Antibiotics-Induced Disruption of Gut Microbiota Increases Systemic Exposure of Clopidogrel Active Metabolite in Type 2 Diabetic Rats

“…The enzymatic activities of CES and all studied CYPs in rat liver microsomes were measured as described previously (Yao et al, 2019).…”