Clopidogrel pharmacotherapy is associated with substantial interindividual variability in clinical response, which can translate into an increased risk of adverse outcomes. Clopidogrel, a recognized substrate of hepatic carboxylesterase 1 (CES1), undergoes extensive hydrolytic metabolism in the liver. Significant interindividual variability in the expression and activity of CES1 exists, which is attributed to both genetic and environmental factors. We determined whether CES1 inhibition and CES1 genetic polymorphisms would significantly influence the biotransformation of clopidogrel and alter the formation of the active metabolite. Coincubation of clopidogrel with the CES1 inhibitor bis(4-nitrophenyl) phosphate in human liver s9 fractions significantly increased the concentrations of clopidogrel, 2-oxo-clopidogrel, and clopidogrel active metabolite, while the concentrations of all formed carboxylate metabolites were significantly decreased. As anticipated, clopidogrel and 2-oxo-clopidogrel were efficiently hydrolyzed by the cell s9 fractions prepared from wild-type CES1 transfected cells. The enzymatic activity of the CES1 variants G143E and D260fs were completely impaired in terms of catalyzing the hydrolysis of clopidogrel and 2-oxo-clopidogrel. However, the natural variants G18V, S82L, and A269S failed to produce any significant effect on CES1-mediated hydrolysis of clopidogrel or 2-oxo-clopidogrel. In summary, deficient CES1 catalytic activity resulting from CES1 inhibition or CES1 genetic variation may be associated with higher plasma concentrations of clopidogrel-active metabolite, and hence may enhance antiplatelet activity. Additionally, CES1 genetic variants have the potential to serve as a biomarker to predict clopidogrel response and individualize clopidogrel dosing regimens in clinical practice.
Title: Activation of the antiviral prodrug oseltamivir is impaired by two newly identified carboxylesterase 1 variants Zhu HJ, Markowitz JS Ref: Drug Metabolism & Disposition: The Biological Fate of Chemicals, 37:264, 2009 : PubMed
Oseltamivir phosphate is an ethyl ester prodrug widely used in the treatment and prevention of both Influenzavirus A and B infections. The conversion of oseltamivir to its active metabolite oseltamivir carboxylate is dependent on ester hydrolysis mediated by carboxylesterase 1 (CES1). We recently identified two functional CES1 variants p.Gly143Glu and p.Asp260fs in a research subject who displayed significant impairment in his ability to metabolize the selective CES1 substrate, methylphenidate. In vitro functional studies demonstrated that the presence of either of the two mutations can result in severe reductions in the catalytic efficiency of CES1 toward methylphenidate, which is required for hydrolysis and pharmacological deactivation. The aim of the present study was to investigate the function of these mutations on activating (hydrolyzing) oseltamivir to oseltamivir carboxylate using the cell lines expressing wild type (WT) and each mutant CES1. In vitro incubation studies demonstrated that the S9 fractions prepared from the cells transfected with WT CES1 and human liver tissues rapidly convert oseltamivir to oseltamivir carboxylate. However, the catalytic activity of the mutant hydrolases was dramatically hindered. The V(max) value of p.Gly143Glu was approximately 25% of that of WT enzyme, whereas the catalytic activity of p.Asp260fs was negligible. These results suggest that the therapeutic efficacy of oseltamivir could be compromised in treated patients expressing either functional CES1 mutation. Furthermore, the potential for increased adverse effects or toxicity as a result of exposure to high concentrations of the nonhydrolyzed prodrug should be considered.
The human carboxylesterase 1 (CES1) gene encodes for the enzyme carboxylesterase 1, a serine esterase governing both metabolic deactivation and activation of numerous therapeutic agents. During the course of a study of the pharmacokinetics of the methyl ester racemic psychostimulant methylphenidate, profoundly elevated methylphenidate plasma concentrations, unprecedented distortions in isomer disposition, and increases in hemodynamic measures were observed in a subject of European descent. These observations led to a focused study of the subject's CES1 gene. DNA sequencing detected two coding region single-nucleotide mutations located in exons 4 and 6. The mutation in exon 4 is located in codon 143 and leads to a nonconservative substitution, p.Gly143Glu. A deletion in exon 6 at codon 260 results in a frameshift mutation, p.Asp260fs, altering residues 260-299 before truncating at a premature stop codon. The minor allele frequency of p.Gly143Glu was determined to be 3.7%, 4.3%, 2.0%, and 0% in white, black, Hispanic, and Asian populations, respectively. Of 925 individual DNA samples examined, none carried the p.Asp260fs, indicating it is an extremely rare mutation. In vitro functional studies demonstrated the catalytic functions of both p.Gly143Glu and p.Asp260fs are substantially impaired, resulting in a complete loss of hydrolytic activity toward methylphenidate. When a more sensitive esterase substrate, p-nitrophenyl acetate was utilized, only 21.4% and 0.6% catalytic efficiency (V(max)/K(m)) were determined in p.Gly143Glu and p.Asp260fs, respectively, compared to the wild-type enzyme. These findings indicate that specific CES1 gene variants can lead to clinically significant alterations in pharmacokinetics and drug response of carboxylesterase 1 substrates.
