Study on the Interaction between Deuterium Clopidogrel and Omeprazole.

INTRODUCTION: Dual antiplatelet therapy with aspirin in combination with a P2Y12 receptor antagonist is a cornerstone for treating patients with acute coronary syndrome and in percutaneous coronary intervention. However, as this combination of antiplatelet therapy increases the risk of bleeding, proton pump inhibitors (PPIs) are currently recommended to prevent gastrointestinal ulcers and bleeding. The cytochrome P450 (CYP450) isoenzyme system metabolizes both clopidogrel (CLP) and PPIs. Unfortunately, omeprazole (OM) reduce the antiplatelet activity of CLP and increases the probability of recurrence of cardiovascular events by competitively inhibiting the CYP450 isoenzyme CYP2C19. METHODS: To address these abovementioned problems, we designed and synthesized deuterium CLP (D-CL) using selective deuterium technology. Our previous research results showed that D-CL had better pharmacokinetic and pharmacodynamic properties. Thus, the HPLC-MS/MS method, cocktail method, Born method, electro-stimulated thrombus generation, and thrombus elastography were used to detect the production of thiol active metabolites (AM), CYP450 enzyme activities, platelet aggregation, time and length of thrombus formation, and the maximum clot strength after combination therapy. We investigated the pharmacokinetics and pharmacodynamics properties of D-CL combined with OM. RESULTS: As compared to CLP, D-CL was less affected when combined with OM, which was reflected in lower inhibitory effects of CYP450 enzyme activities, a greater area under the curve of AM, and better antiplatelet and antithrombotic effects. CONCLUSION: D-CL may reduce drug-drug interactions and address the clinical disadvantages of CLP.

Complex Cytochrome P450 Kinetics Due to Multisubstrate Binding and Sequential Metabolism. Part 2. Modeling of Experimental Data.

Three CYP3A4 substrates, midazolam, ticlopidine, and diazepam, display non-Michaelis-Menten kinetics, form multiple primary metabolites, and are sequentially metabolized to secondary metabolites. We generated saturation curves for these compounds and analyzed the resulting datasets using a number of single-substrate and multisubstrate binding models. These models were parameterized using rate equations and numerical solutions of the ordinary differential equations. Multisubstrate binding models provided results superior to single-substrate models, and simultaneous modeling of multiple metabolites provided better results than fitting the individual datasets independently. Although midazolam datasets could be represented using standard two-substrate models, more complex models that include explicit enzyme-product complexes were needed to model the datasets for ticlopidine and diazepam. In vivo clearance predictions improved markedly with the use of in vitro parameters from the complex models versus the Michaelis-Menten equation. The results highlight the need to use sufficiently complex kinetic schemes instead of the Michaelis-Menten equation to generate accurate kinetic parameters. SIGNIFICANCE STATEMENT: The metabolism of midazolam, ticlopidine, and diazepam by CYP3A4 results in multiple metabolites and sequential metabolism. This study evaluates the use of rate equations and numerical methods to characterize the in vitro enzyme kinetics. Use of complex cytochrome P450 kinetic models is necessary to obtain accurate parameter estimates for predicting in vivo disposition.

A New Workflow for Drug Metabolite Profiling by Utilizing Advanced Tribrid Mass Spectrometry and Data-Processing Techniques.

Drug metabolite profiling utilizes liquid chromatography with tandem mass spectrometry (LC/MS/MS) to acquire ample information for metabolite identification and structural elucidation. However, there are still challenges in detecting and characterizing all potential metabolites that can be masked by a high biological background, especially the unknown and uncommon ones. In this work, a novel metabolite profiling workflow was established on a platform using a state-of-the-art tribrid high-resolution mass spectrometry (HRMS) system. Primarily, an instrumental method was developed based on the novel design of the tribrid system that facilitates in-depth MSn scans with two fragmentation devices. Additionally, different advanced data acquisition techniques were assessed and compared, and automatic background exclusion and deep-scan approaches were adopted to promote assay efficiency and metabolite coverage. Finally, different data-analysis techniques were explored to fully extract metabolite data from the information-rich MS/MS data sets. Overall, a workflow combining tribrid mass spectrometry and advanced acquisition methodology has been developed for metabolite characterization in drug discovery and development. It maximizes the tribrid HRMS platform's utility and enhances the coverage, efficiency, quality, and speed of metabolite profiling assays.

Impact of selected genetic factors on clopidogrel inactive metabolite level and antiplatelet response in patients after percutaneous coronary intervention.

BACKGROUND AND OBJECTIVE: Clopidogrel is frequently used as part of optimal dual antiplatelet therapy in high-bleeding risk patients with the acute coronary syndrome. The concentration of the inactive carboxylic acid metabolite of clopidogrel might be useful to evaluate the response to clopidogrel therapy. Therefore, we sought to correlate the inhibition of platelet aggregation with the plasma level of the inactive metabolite of clopidogrel in patients after percutaneous coronary interventions (PCI) and their associations with the most frequently studied genetic polymorphisms. For this purpose, the fast and simple HPLC method for determining the concentration of the inactive metabolite was developed. METHODS: The effect of CYP2C19, CYP3A4/5, ABCB1 and PON1 genes on the plasma inactive metabolite concentration of clopidogrel and the platelet aggregation was investigated in 155 patients before and after PCI. RESULTS: The concentration of the inactive metabolite of clopidogrel was not significantly different in the intermediate metabolizers (IM) of CYP2C19 compared with extensive metabolizers (EM) both before and after PCI, while inhibition of platelet aggregation was found to be significantly better in EM than in IM. The presence of the A allele at position 2677 in the ABCB1 gene was associated with a significantly lower concentration of inactive metabolite of clopidogrel before PCI. CONCLUSION: The CYP2C19*2 allele was associated with decreased platelet reactivity during clopidogrel therapy before and after PCI. Simultaneous determination of platelet aggregation and concentration of the inactive clopidogrel metabolite may be useful in clinical practice to find the cause of adverse effects or insufficient treatment effect in patients chronically treated with clopidogrel.

Implementation of a Reference-Scaled Average Bioequivalence Approach for Highly Variable Acetylsalicylic Acid in Fixed-Dose Combination with Clopidogrel Versus Enteric Aspirin in Chinese Subjects Under Fasting Conditions: A Phase 1, Open-Label, Randomized, Crossover Study.

INTRODUCTION: Dual antiplatelet therapy, aspirin and a P2Y12 inhibitor, is recommended to prevent thrombotic complications of acute coronary syndrome. Clopidogrel plus acetylsalicylic acid combination is the most commonly used dual antiplatelet therapy recommended by international guidelines and in Chinese clinical practice. Poor adherence to dual antiplatelet therapy or premature interruption of dual antiplatelet therapy is an important contributor to cardiovascular mortality and lethal cardiovascular events. Clopidogrel + acetylsalicylic acid fixed-dose combination enhances adherence to dual antiplatelet therapy. Herein, we aimed to evaluate bioequivalence of acetylsalicylic acid and clopidogrel in fixed-dose combination compared with simultaneous administration of their individual formulations in healthy Chinese subjects under fasting conditions. METHODS: This was a randomized, single-center, open-label, three-sequence, three-period, two-treatment, crossover study with a washout period of 10 days conducted in healthy Chinese volunteers. Subjects were randomized to receive Co-Plavix® (test formulation- fixed-dose combination of 100 mg acetylsalicylic acid and 75 mg clopidogrel) once and reference formulations (coadministration of individual formulations of 100 mg acetylsalicylic acid and 75 mg clopidogrel) twice during the study period. Pharmacokinetic parameters were analyzed for acetylsalicylic acid, its metabolite salicylic acid, clopidogrel, and its metabolite SR26334. As acetylsalicylic acid shows high intrasubject variability, the reference-scaled average bioequivalence (RSABE) approach was implemented for acetylsalicylic acid analysis, while bioequivalence of clopidogrel was assessed using the average bioequivalence method. Point ratios and confidence intervals (CIs) for AUC, AUClast, and Cmax for acetylsalicylic acid and clopidogrel were calculated. RESULTS: In total, 171 healthy subjects were enrolled in this study. Subjects were randomized and 170 subjects were treated with test or reference formulation; 164 subjects completed the study. Regarding acetylsalicylic acid exposure, as reference within-subject standard deviation (SDW) was at least 0.294 for acetylsalicylic acid Cmax, AUClast, and AUC, the RSABE analysis method was used to assess bioequivalence for all three parameters. The point estimates were within the 0.80-1.25 range (1.19, 1.09, and 1.04, respectively), and upper one-sided 95% CIs of scaled average bioequivalence metric were at most 0 (- 0.30, - 0.14, and - 0.10, respectively). Thus, bioequivalence was demonstrated with acetylsalicylic acid. Bioequivalence was also achieved with clopidogrel as the 90% CIs for geometric mean ratios of clopidogrel Cmax, AUClast, and AUC were within the bioequivalence range (0.80-1.25). CONCLUSION: Application of the reference-scaled average bioequivalence approach to evaluate bioequivalence of acetylsalicylic acid in Chinese male and female healthy volunteers under fasting conditions demonstrated bioequivalence of test and reference formulations. TRIAL REGISTRATION: CTR20181695.

Semimechanistic Population Pharmacokinetic Model to Predict the Drug-Drug Interaction Between S-ketamine and Ticlopidine in Healthy Human Volunteers.

Low-dose oral S-ketamine is increasingly used in chronic pain therapy, but extensive cytochrome P450 (CYP) mediated metabolism makes it prone to pharmacokinetic drug-drug interactions (DDIs). In our study, concentration-time data from five studies were used to develop a semimechanistic model that describes the ticlopidine-mediated inhibition of S-ketamine biotransformation. A mechanistic model was implemented to account for reversible and time-dependent hepatic CYP2B6 inactivation by ticlopidine, which causes elevated S-ketamine exposure in vivo. A pharmacokinetic model was developed with gut wall and hepatic clearances for S-ketamine, its primary metabolite norketamine, and ticlopidine. Nonlinear mixed effects modeling approach was used (NONMEM version 7.3.0), and the final model was evaluated with visual predictive checks and the sampling-importance-resampling procedure. Our final model produces biologically plausible output and demonstrates that ticlopidine is a strong inhibitor of CYP2B6 mediated S-ketamine metabolism. Simulations from our model may be used to evaluate chronic pain therapy with S-ketamine.

Effects of Danshen capsules on the pharmacokinetics and pharmacodynamics of clopidogrel in healthy volunteers.

Nowadays, the Herb-drug combination is becoming increasingly popular in China. However, the possible interaction induced by their combination was examined rarely. The aim of this study was to investigate the effect of multi-dose administration of Danshen capsules on clopidogrel pharmacokinetics and pharmacodynamics in healthy volunteers. A sequential, open-label, and two-period pharmacokinetic drug interaction study was designed to compare clopidogrel pharmacokinetic parameters before and after 7 days of administration of Danshen capsules in twenty healthy male volunteers. Co-administration of multiple doses of Danshen capsules caused increases in apparent oral clearance of clopidogrel and its metabolite by 96.5% and 73.7% and apparent volume of distribution by 94.2% and 75.1%, corresponding declines in Cmax by 41.7% and 32.9%, AUC0-t by 50.3% and 41.8%, and AUC0-∞ by 49.3% and 41.5% in human volunteers, respectively. Corresponding pharmacokinetic findings, co-administration of Danshen capsules with clopidogrel decreased the antiplatelet activity compared with individual agents. The results suggested that multiple dose administration of Danshen capsules could induce cytochrome P450 (CYP) isoenzymes, thereby increasing the clearance of clopidogrel. Therefore, caution should be taken when Danshen products containing lipophilic components are used in combination with therapeutic drugs metabolized by CYP3A4.

Quantitative in vitro phenotyping and prediction of drug interaction potential of CYP2B6 substrates as victims.

1. Determination of fm, CYP for a compound is critical to assess the potential risk of a drug candidate as a victim of DDI. Several compounds are identified as CYP2B6 substrates, but the fm, CYP2B6 values are not determined quantitatively. 2. Two methods of reaction phenotyping, the chemical inhibition method and metabolism in rCYP enzymes, were used to determine the relative contributions of the enzymes. Chemical inhibition method was also conducted in the presence of BSA (0.5% w/v). 3. The results confirm with the earlier studies concerning the identity of the CYP2B6 enzyme. The fm, CYP2B6 values for artemisinin, bupropion, clopidogrel, ketamine, selegiline, sertraline and ticlopidine were 0.24, 0.28, 0.15, 0.45, 0.46, 0.42 and 0.54, respectively, in HLM determined by chemical inhibition method. The fm, CYP2B6 values for artemisinin, bupropion, clopidogrel, ketamine, selegiline, sertraline and ticlopidine were 0.46, 0.17, 0.15, 0.60, 0.51, 0.66 and 0.77, respectively, in HLM determined by chemical inhibition method in the presence of BSA (0.5% w/v). 4. Bupropion metabolism is majorly mediated by CYP2C19 (0.41) with a minor contribution from CYP2B6 (0.16) in the presence of BSA. Ticlopidine is a time-dependent inhibitor of both CYP2B6 and CYP2C19 that can inhibit the bupropion metabolism by 50-60%.

Effects of Genetic Variants on Carboxylesterase 1 Gene Expression, and Clopidogrel Pharmacokinetics and Antiplatelet Effects.

Several single nucleotide variations (SNVs) affect carboxylesterase 1 (CES1) activity, but the effects of genetic variants on CES1 gene expression have not been systematically investigated. Therefore, our aim was to investigate effects of genetic variants on CES1 gene expression in two independent whole blood sample cohorts of 192 (discovery) and 88 (replication) healthy volunteers and in a liver sample cohort of 177 patients. Furthermore, we investigated possible effects of the found variants on clopidogrel pharmacokinetics (n = 106) and pharmacodynamics (n = 46) in healthy volunteers, who had ingested a single 300 mg or 600 mg dose of clopidogrel. Using massively parallel sequencing, we discovered two CES1 SNVs, rs12443580 and rs8192935, to be strongly and independently associated with a 39% (p = 4.0 × 10-13 ) and 31% (p = 2.5 × 10-8 ) reduction in CES1 whole blood expression per copy of the minor allele. These findings were replicated in the replication cohort. However, these SNVs did not affect CES1 liver expression, or clopidogrel pharmacokinetics or pharmacodynamics. Conversely, the CES1 c.428G>A missense SNV (rs71647871) impaired the hydrolysis of clopidogrel, increased exposure to clopidogrel active metabolite and enhanced its antiplatelet effects. In conclusion, the rs12443580 and rs8192935 variants reduce CES1 expression in whole blood but not in the liver. These tissue-specific effects may result in substrate-dependent effects of the two SNVs on CES1-mediated drug metabolism.

Development and validation of a sensitive and rapid UHPLC-MS/MS method for the simultaneous quantification of the common active and inactive metabolites of vicagrel and clopidogrel in human plasma.

Vicagrel, an analog of clopidogrel, is a novel thienopyridine P2Y12 antagonist in clinical development in China for the treatment of acute coronary syndromes. Vicagrel and clopidogrel are prodrugs requiring metabolic activation to produce a pharmacologically active thiol metabolite (H4) in vivo. The formation of H4 from vicagrel in humans is promising more efficient and consistent than that from clopidogrel. After oral dosing of vicagrel or clopidogrel to humans, the active thiol metabolite H4 and two inactive metabolites closely related to H4 formation (the thiol metabolite H3 and the S-methylated metabolite SM3) were observed in plasma; SM3 was the most abundant drug-related component of vicagrel in circulation. In this study, a sensitive, rapid, and reliable UHPLC-MS/MS method was developed for the simultaneous determination of derivatized H3 (MP-H3), derivatized H4 (MP-H4), and SM3 in human plasma to investigate the pharmacokinetics of vicagrel in healthy subjects compared with clopidogrel. After extracted from 50µL of plasma by simple protein precipitation, the analytes and stable isotope-labeled internal standards were separated on an Acquity UPLC BEH C18 column (100mm×2.1mm, 1.7µm). The mobile phase was acetonitrile-water-formic acid (45:55:0.0275, v/v/v) delivered at a flow rate of 0.45mL/min under isocratic elution. The total chromatographic run time was 6min. MP-H3, MP-H4, and SM3 were separated from their corresponding isomers under the chromatographic conditions. Mass spectrometric detection was conducted in multiple reaction monitoring mode on an AB Sciex Qtrap 5500 System using a positive electrospray ionization interface. The calibration curves were linear in the following ranges: 0.100-200ng/mL for MP-H3 and MP-H4 and 1.00-1000ng/mL for SM3. The intra- and inter-day accuracy and precision for each analyte at all concentrations were within acceptable limits (±15%). The validated method was successfully applied to compare the pharmacokinetics of vicagrel and clopidogrel after a single oral administration to healthy subjects in the first-in-human study of vicagrel.

Impaired liver cytochrome P450 2C11 activity after dual antiplatelet therapy with aspirin and clopidogrel in rats.

1. Aspirin (ASA) and clopidogrel (CLP) are used in combination as dual antiplatelet therapy (DAPT) for acute coronary syndrome based on their complementary mechanisms for platelet aggregation inhibition. However, the pharmacokinetics of such drug combination usage has not been thoroughly investigated. 2. In the current study, an LC-MS/MS method was developed to simultaneously determine the plasma concentrations of ASA and its metabolite salicylic acid (SA) with CLP and its metabolites, clopidogrel carboxylic acid (CLPM) and clopidogrel active metabolite derivative (CAMD). The pharmacokinetics of ASA, SA, CLP, CLPM and CAMD in rats receiving two-week DAPT with ASA and CLP were then determined. 3. After two-week DAPT with ASA and CLP in rats, the activities of aspirin esterase and rCyp2c11, enzymes mediating rat metabolism of ASA and CLP, respectively, in prepared rat liver microsomes were measured followed by further determination of rCyp2c11 mRNA expressions. The results demonstrated that DAPT led to minimal impact on aspirin esterase activity but significant decrease in rCyp2c11 activity and mRNA expression. 4. In conclusion, our findings on impairment in rCyp2C11 activity and mRNA expression by DAPT in rats could provide guidance on its safe clinical use with other CYP 2C19 substrates.

Long-term pharmacodynamic effects of Ticagrelor versus Clopidogrel in fibrinolytic-treated STEMI patients undergoing early PCI.

The long-term pharmacodynamic effects of Ticagrelor versus Clopidogrel in patients undergoing early percutaneous coronary intervention (PCI) after fibrinolytic therapy is unknown. From May 2014 to August 2016, 212 patients undergoing PCI within 24 h of Tenecteplase (TNK), Aspirin, and Clopidogrel for ST-elevated myocardial infarction (STEMI) were randomized at four Canadian sites to receive additional Clopidogrel or Ticagrelor initiated prior to PCI. The platelet reactivity units (PRU) were measured with the VerifyNow Assay before study drug administration (baseline), at 4 and 24 h post PCI, and follow-up appointment. A mixed-model analysis with time as the repeated measure and drug as the between-subjects factor was calculated using 2 separate 1 × 4 ANOVAs, with students t-tests used to compare drugs within each time point. Complete clinical follow-up data (median 115.0 days; IQR 80.3-168.8) was available in 50 patients (23.6%) randomized to either Clopidogrel (n = 23) or Ticagrelor (n = 27). Analyses revealed significant decreases in PRU from baseline to 4 h (261.4 vs. 71.7; Mdiff = - 189.7; p < 0.001) to 24 h (71.7 vs. 27.7; Mdiff = - 44.0; p < 0.001) to end of follow-up (27.7 vs.17.9; Mdiff = - 9.9. p = 0.016) for those randomized to Ticagrelor and significant decreases in PRU only from baseline to 4 h (271.3 vs. 200.8; Mdiff = - 70.5, p = < 0.001) in patients receiving Clopidogrel, and a significantly greater proportion of patients with adequate platelet inhibition (PRU < 208) on long-term follow-up (Clopidogrel, 82.6% vs. Ticagrelor, 100.0%; p = 0.038). Our results demonstrate that in patients undergoing PCI within 24 h of fibrinolysis for STEMI, Ticagrelor provides prolonged platelet inhibition compared with Clopidogrel.

Effect of cytochrome P450 2C19 polymorphism on adverse cardiovascular events after drug-eluting stent implantation in a large Hakka population with acute coronary syndrome receiving clopidogrel in southern China.

BACKGROUND AND OBJECTIVES: The objective of this study is to evaluate the effects of cytochrome P450 2C19 (CYP2C19) polymorphism on adverse cardiovascular events (MACE) in Hakka patients with acute coronary syndrome (ACS) receiving clopidogrel who had undergone coronary drug-eluting stent placement after percutaneous coronary intervention (PCI) in southern China. METHODS: Genotyping of CYP2C19 and MACE of 934 ACS patients with PCI on clopidogrel maintenance therapy were analyzed. Patients who carried loss-of-function CYP2C19 were treated with a 150-mg maintenance dose of clopidogrel or 90 mg of ticagrelor antiplatelet therapy, and patients who were non-carriers received clopidogrel therapy daily at a maintenance dose of 75 mg and the patients were followed-up for at least 12 months. The primary efficacy endpoint was a composite of cardiovascular death, myocardial infarction, and target vessel revascularization and stroke. RESULTS: The allelic frequency of CYP2C19*2 and CYP2C19*3 of Hakka patients in the current study was 31.64 and 5.19%, respectively. The CYP2C19 wild-type homozygotes (*1/*1) were the most predominant among the patients (40.36%), followed by the CYP2C19*2 heterozygotes (*1/*2) (40.26%). The distribution of CYP2C19 phenotypes was divided into extensive metabolizers (EM; 40.36%), intermediate metabolizers (IM; 45.61%), and poor metabolizers (PM; 14.03%). Based on the genotype-guided antiplatelet therapy, there was no significant association between the carrier status and the clinical outcome at 1, 6, and 12 months. In addition, no significant difference in the rates of bleeding was found among the three groups. After logistic regression analysis, hypertension was the only independent predictor of cardiovascular events (relative risk, 1.501; 95% CI, 1.011 to 2.229; P = 0.044). CONCLUSIONS: Our results shed new light on the important benefit of testing CYP2C19 polymorphisms before prescribing clopidogrel in patients treated with drug-eluting stent implantation after PCI. The testing may help to optimize pharmacotherapy effectiveness by providing individualized treatment to the Chinese population. Our findings mandate further studies aimed at initiating genome-based personalized antiplatelet therapy in a Hakka population in southern China.

Bioequivalence and pharmacokinetic/pharmacodynamic correlation of clopidogrel in healthy Thai subjects
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Clopidogrel is an antiplatelet drug, selectively binding to the platelet P2Y12 receptor of adenosine diphosphate. Clopidogrel is a prodrug modified through active metabolite in the liver by two steps of CYP enzyme. The active metabolite is responsible for inhibiting platelet aggregation. OBJECTIVE: The study aimed to assess the bioequivalence of clopidogrel 75 mg generic and reference drugs and to investigate the correlation between pharmacokinetics of active metabolites and its antiplatelet activities. MATERIALS AND METHODS: Determination of clopidogrel, carboxylic acid form, and active metabolite were done by liquid chromatography tandem mass spectrometry, and evaluation of platelet function was also investigated by light transmission aggregometer. 20 subjects were randomized and assigned in a crossover design to take a single 75-mg oral dose of clopidogrel generic and reference drugs in two periods with washout. Pharmacokinetic parameters Cmax, AUC0-t, and AUC0-inf of clopidogrel, carboxylic acid form, and active metabolite were analyzed. RESULTS: Bioequivalence could be shown when testing parameters with ANOVA, as 90% confidence intervals were found to be within the acceptance range of 80 - 125%. For the maximum of platelet aggregation after administration of both products, no significant differences were found. Significant correlation of Cmax of clopidogrel active metabolite and maximum platelet aggregation was found after receiving 0 - 6 hours of both formulations. CONCLUSION: The study found bioequivalence of clopidogrel generic and reference drugs. There were also significant correlations between Cmax of clopidogrel active metabolite and maximum platelet aggregation.
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Review of aspirin and clopidogrel resistance in peripheral arterial disease.

OBJECTIVE: Aspirin resistance (AR) and clopidogrel resistance (CR) are terms used to describe a reduction in the medication's efficacy in inhibiting platelet aggregation despite regular dosing. This review gives context to the clinical role and implications of antiplatelet resistance in peripheral arterial disease (PAD). METHODS: A review of English-language literature on AR and CR in PAD involving human subjects using PubMed and MEDLINE databases was performed in April 2017. A total of 2075 patients in 22 relevant studies were identified. To give this issue context, a review of the larger, more established literature on antiplatelet resistance in coronary disease was undertaken, identifying significant research associating resistance to major adverse cardiovascular events (MACEs). RESULTS: Studies in the coronary arterial disease literature have strongly associated antiplatelet resistance with increased MACE. Prevalence of AR or CR in coronary disease appears to be >55% for each in some studies. Meta-analyses of >50 studies revealed that AR and CR are significantly associated with MACE (relative risk of 2.09 and 2.8, respectively). This adds further weight to the literature reporting antiplatelet resistance as an independent predictor of and a threefold risk factor for major adverse cardiovascular events. The prevalence of resistance in PAD in this review was comparable to that in the coronary disease literature, with AR and CR prevalence up to 60% and 65%, respectively. There is evidence that the adverse effects of antiplatelet resistance are significant in PAD. In fact, research directly studying stent thrombosis populations with either coronary arterial disease or PAD revealed more significantly impaired platelet responsiveness to clopidogrel and aspirin in PAD compared with similar individuals with coronary disease. AR in PAD was found in studies to be a significant risk factor for iliofemoral stent reocclusion (P = .0093) and stroke in patients with symptomatic carotid disease (P = .018). CR was found to be a significant, independent risk factor in predicting ischemic outcomes in several recent PAD studies (P < .0001). Loss-of-function carriers of enzyme CYP2C19, important in clopidogrel metabolism, have a 30% greater risk of ischemic events (P < .001). Importantly, less antiplatelet drug resistance may be encountered with newer antiplatelet agents, including ticagrelor and prasugrel, because of reduced enzymatic polymorphisms. CONCLUSIONS: The limited research addressing AR and CR in PAD suggests that further research is required to clarify the role of platelet assays and potential for individualized antiplatelet therapy.

Simultaneous confidence regions for multivariate bioequivalence.

Demonstrating bioequivalence of several pharmacokinetic (PK) parameters, such as AUC and Cmax , that are calculated from the same biological sample measurements is in fact a multivariate problem, even though this is neglected by most practitioners and regulatory bodies, who typically settle for separate univariate analyses. We believe, however, that a truly multivariate evaluation of all PK measures simultaneously is clearly more adequate. In this paper, we review methods to construct joint confidence regions around multivariate normal means and investigate their usefulness in simultaneous bioequivalence problems via simulation. Some of them work well for idealised scenarios but break down when faced with real-data challenges such as unknown variance and correlation among the PK parameters. We study the shapes of the confidence regions resulting from different methods, discuss how marginal simultaneous confidence intervals for the individual PK measures can be derived, and illustrate the application to data from a trial on ticlopidine hydrochloride. An R package is available.

Influence of genetic co-factors on the population pharmacokinetic model for clopidogrel and its active thiol metabolite.

PURPOSE: A high interindividual variability is observed in the pharmacokinetics of clopidogrel, a widely used antiplatelet drug. In the present study, a joint parent-metabolite population pharmacokinetic model was developed to adequately describe observed concentrations of clopidogrel and its active thiol metabolite (H4). METHODS: The study included 63 patients undergoing elective coronarography or percutaneous coronary intervention. The population pharmacokinetic model was developed in the NONMEM 7.3 software, and first-order conditional estimation method with interaction was applied. Also, the influence of covariates was evaluated (age, weight, body mass index (BMI), obesity defined as BMI ≥ 30 kg/m2, sex, diabetes mellitus, co-administration of PPI or statins, presence of CYP2C19*2, CYP2C19*17, CYP3A4*1G alleles, and ABCB1 3435 TT genotype). RESULTS: It was found that the only significant covariate was the presence of CYP2C19*2 allele, which had an impact on lower conversion of clopidogrel to H4. As a result, predicted area under the time-concentration curve values was lower in carriers of this allele, with median 5.94 ng h/ml (interquartile range 3.92-12.51 [ng∙h/ml]) vs. 12.70 ng h/ml in non-carriers (interquartile range, 7.00-19.39 [ng∙h/ml]), respectively (p = 0.004). CONCLUSIONS: Developed model predicts that the only significant covariate influencing the observed concentrations and therefore the exposure to the active H4 metabolite is the presence of CYP2C19*2 allele.

Effects of statin therapy on platelet reactivity after percutaneous coronary revascularization in patients with acute coronary syndrome.

Statin use is associated with enhanced pharmacodynamic response to clopidogrel in patients with stable coronary artery disease undergoing percutaneous coronary intervention (PCI). However, the impact of statin therapy on clopidogrel response profiles in patients with acute coronary syndrome (ACS) undergoing PCI has not been established and represents the objective of this investigation. On-treatment P2Y12 platelet reactivity was measured using the vasodilator stimulated phosphoprotein (VASP) phosphorylation assay before PCI, at hospital discharge, and at 1 month after PCI in ACS patients enrolled in the multicenter, prospective GEne polymorphisms, Platelet Reactivity, and Syntax Score (GEPRESS) study (n = 962). High platelet reactivity (HPR) was defined as platelet reactivity index ≥50%. Statins were prescribed at hospital discharge in 87% (n = 835) of patients. All patients were followed for 1 year. The 1-month HPR rate was lower in statin than in non-statin treated patients (39.6 vs 52%, respectively, p = 0.009). This finding was confirmed also among statin-treated patients with high Syntax score (≥15). After adjustment for differences in baseline characteristics, statin use at discharge was independently associated with 1-month HPR rate (odds ratio, 0.58, 95% confidence interval, 0.38-0.89; p = 0.015). In ACS patients undergoing PCI treated with clopidogrel the use of statins at discharge was associated with significantly lower 1-month HPR rates compared with patients not treated with statins.

Time course of the antiplatelet effect after switching to clopidogrel from initial prasugrel therapy in patients with acute coronary syndrome.

Prasugrel is often replaced with clopidogrel after a certain period of time following coronary stenting. However, the time course of platelet aggregation during this replacement is unknown. We performed a prospective, single-arm study to monitor platelet reactivity before and after the replacement. Forty-five patients (mean age 62.6 ± 13 years, 40 male) who received coronary stenting for acute coronary syndrome were initially treated with the loading dose (20 mg) of prasugrel followed by the maintenance dose (3.75 mg/day) for 7 days, then switched to 75 mg/day of clopidogrel. The P2Y12 reaction unit (PRU) level was measured at baseline and selected time points. Prasugrel effectively suppressed PRU from 248 ± 59 at baseline to 145 ± 65 on day 1 (P < 0.001). The PRU value on the final day of prasugrel treatment (day 7) was 156 ± 68 (P < 0.001 vs. baseline). After switching to clopidogrel, PRU was consistently suppressed [146 ± 60, 139 ± 54, and 135 ± 60 on days 9, 11, and 13, respectively (P < 0.001, each point vs. baseline)]. Switching from the initial prasugrel therapy to clopidogrel using the maintenance dose does not cause a drug efficacy gap and stays effective for preventing stent thrombosis.