Pharmacokinetics and Bioequivalence of Clopidogrel Hydrogen Sulfate Tablets in Fed and Fasted Conditions: An Open-Label, Randomized, Semireplicated Crossover Study in Healthy Chinese Volunteers.

Clopidogrel is an antiplatelet drug with high intraindividual variability in systemic exposure and efficacy. It has been used for treating atherosclerosis and acute coronary syndrome and in preventing stent restenosis and thrombotic complications after stent implantation in clinical practice for nearly 20 years. In this study we aimed to evaluate the bioequivalence of 2 clopidogrel hydrogen sulfate formulations (75-mg tablets) under fed (n = 66) and fasted (n = 66) conditions by using the reference-scaled average bioequivalence method. An open-label, randomized, 3-sequence and 3-period crossover (3×3), semireplicated study was designed and conducted. Clopidogrel concentration of plasma samples was measured by high-precision liquid chromatography and tandem mass spectrometry. The pharmacokinetic parameters were derived by a noncompartmental model. In the fed condition the geometric least-squares mean ratios of peak concentration (Cmax ) and area under the concentration-time curve (AUC0-t ) were, respectively, 103.38% and 98.97%, and the corresponding 90%CIs were 95.68% to 111.70% and 94.67% to 103.47%. In the fasted condition the geometric least squares mean ratios of Cmax and AUC0-t were, respectively, 106.53% and 105.77%, and the corresponding 90%CIs were 97.62% to 116.25% and 96.96% to 115.38%. According to the criteria for bioequivalence (80.00% to 125.00%), the test formulations of clopidogrel and Plavix were determined to be bioequivalent.

Safety and Pharmacokinetics of DS-1040 Drug-Drug Interactions With Aspirin, Clopidogrel, and Enoxaparin.

DS-1040, a novel low-molecular-weight inhibitor of activated thrombin-activatable fibrinolysis inhibitor, is under development for the treatment of thromboembolic diseases including venous thromboembolism and acute ischemic stroke. Here we describe the results of 3 studies that evaluated the safety and tolerability of DS-1040 along with the effect on DS-1040 pharmacokinetic (PK) parameters, when dosed alone or when coadministered with aspirin (NCT02071004), clopidogrel (NCT02560688), or enoxaparin in healthy subjects. Concomitant administration of single-dose DS-1040 with multiple-dose aspirin, multiple-dose clopidogrel, or single-dose enoxaparin, consistent with clinically relevant dose regimens, was safe and well tolerated with no serious treatment-emergent adverse events (TEAEs), TEAEs leading to discontinuation, bleeding-related TEAEs, and no significant changes in coagulation parameters. DS-1040 did not prolong bleeding time when administered concomitantly with aspirin or clopidogrel. In the aspirin study, DS-1040 PK was evaluated following the concomitant administration with multiple-dose aspirin, where the plasma DS-1040 exposure (peak plasma concentration [Cmax ] and area under the concentration-time curve [AUCinf ]) was to be similar to the data previously published in the first-in-human study of DS-1040 in healthy subjects. The PK parameters of DS-1040 coadministered with clopidogrel were similar to those of DS-1040 alone, with small increases in geometric means for Cmax (7%) and AUClast (9%). When coadministered with enoxaparin, the PK parameters of DS-1040 were not affected (1.1% and 1.5% decreases in geometric means for Cmax and AUClast , respectively). Therefore, concomitant administration of DS-1040 and clopidogrel or enoxaparin did not demonstrate PK drug-drug interactions.

Freeze-Dried Clopidogrel Loaded Lyotropic Liquid Crystal: Box-Behnken Optimization, In-Vitro and In-Vivo Evaluation.

BACKGROUND: Clopidogrel (CLP) suffers from extensive first pass metabolism results in a negative impact on its oral systemic bioavailability. Cubosomes are Lyotropic Liquid Crystalline (LLC) nano-systems comprising monoolein, a steric stabilizer and an aqueous system, it considered a promising carrier for different pharmaceutical compounds. Box-Behnken Design (BBD) is an efficient tool for process analysis and optimization skipping forceful treatment combinations. OBJECTIVE: The study was designed to develop freeze-dried clopidogrel loaded LLC (cubosomes) for enhancement of its oral bioavailability. METHODS: A 33 BBD was adopted, the studied independent factors were glyceryl monooleate (GMO lipid phase), Pluronic F127 (PL F127steric stabilizer) and polyvinyl alcohol powder (stabilizer). Particle Size (PS), Polydispersity Index (PDI) and Zeta Potential (ZP) were set as independent response variables. Seventeen formulae were prepared in accordance with the bottom up approach and in-vitro evaluated regarding PS, PDI and ZP. Statistical analysis and optimization were achieved using design expert software®, then the optimum suggested formula was prepared, in-vitro revaluated, freeze-dried with 3% mannitol (cryoprotectant), solid state characterized and finally packed in hard gelatin capsule for comparative in-vitro release and in-vivo evaluation to Plavix®. RESULTS: Results of statistical analysis of each individual response revealed a quadratic model for PS and PDI where a linear model for ZP. The optimum suggested formula with desirability factor equal 0.990 consisting of (200 mg GMO, 78.15 mg PL F127 and 2% PVA). LC/MS/MS study confirmed significant higher Cmax, AUC0-24h and AUC0-∞ than that of Plavix®. CONCLUSION: The results confirm the capability of developed carrier to overcome the low oral bioavailability.

Impact of low-dose prasugrel on platelet reactivity and cardiac dysfunction in acute coronary syndrome patients requiring primary drug-eluting stent implantation: A randomized comparative study.

OBJECTIVE: The aim of this study was to compare how prasugrel and clopidogrel affect platelet aggregation reactivity, cardiac enzyme release, cardiac remodeling, and the formation of in-stent thrombi after primary percutaneous coronary intervention (PCI). BACKGROUND: The advantages of using prasugrel over clopidogrel in cardiac injury following acute coronary syndrome (ACS) remain unclear. METHODS: A total of 78 ACS patients were randomly allocated into clopidogrel (300 mg loading/75 mg maintenance) or prasugrel (20 mg loading/3.75 mg maintenance) treatment groups, followed by undergoing primary PCI. Platelet reactivity and cardiac enzymes were measured before and after primary PCI. Moreover, cardiac function was measured by ultrasound echocardiography and coronary angioscopic observation was after primary PCI up to 8 months later. RESULTS: Antiplatelet reactivity in the prasugrel treatment group reached optimal levels (P2Y12 reaction units [PRU] less than 262) immediately after the administration and was maintained even at 8 months, independently of the CYP2C19 genotype. Prasugrel treatment significantly suppressed creatine kinase elevation compared to clopidogrel treatment (median value 404 IU/L to 726 IU/L vs. 189 IU/L to 1,736 IU/L, p = 0.018 for maximum values) and reduced left ventricular mass (217.2-168.8 g in prasugrel, p = 0.045; 196.9-176.4 g in clopidogrel, p = 0.061). There were no significant differences in the incidence of in-stent attached thrombi between the two groups. CONCLUSIONS: Compared to clopidogrel, prasugrel produced a stable platelet aggregation inhibitory effect in patients with ACS regardless of CYP2C19 genotype, reduced cardiac enzyme release, and prevented cardiac remodeling after ACS.

Carboxylesterase catalyzed 18O-labeling of carboxylic acid and its potential application in LC-MS/MS based quantification of drug metabolites.

LC-MS quantification of drug metabolites is sometimes impeded by the availability of internal standards that often requires customized synthesis and/or extensive purification. Although isotopically labeled internal standards are considered ideal for LC-MS/MS based quantification, de novo synthesis using costly isotope-enriched starting materials makes it impractical for early stage of drug discovery. Therefore, quick access to these isotope-enriched compounds without chemical derivatization and purification will greatly facilitate LC-MS/MS based quantification. Herein, we report a novel 18O-labeling technique using metabolizing enzyme carboxylesterase (CES) and its potential application in metabolites quantification study. Substrates of CES typically undergo a two-step oxygen exchange with H218O in the presence of the enzyme, generating singly- and doubly-18O-labeled carboxylic acids; however, unexpected hydrolytic behavior was observed for three of the test compounds - indomethacin, piperacillin and clopidogrel. These unusual observations led to the discovery of several novel hydrolytic mechanisms. Finally, when used as internal standard for LC-MS/MS based quantification, these in situ labeled compounds generated accurate quantitation comparable to the conventional standard curve method. The preliminary results suggest that this method has potential to eliminate laborious chemical synthesis of isotope-labeled internal standards for carboxylic acid-containing compounds, and can be developed to facilitate quantitative analysis in early-stage drug discovery.

Pharmacokinetics and bioequivalence of low-dose clopidogrel in healthy Chinese volunteers under fasted and fed conditions.

Clopidogrel is an antiplatelet drug whose performance at a low dose (25 mg) have not been evaluated in Chinese patients, who may be subject to different effects from Caucasians. We carried out this evaluation and compared a generic (Taijia) and a reference drug (Plavix®). We evaluated Taijia and Plavix® in 128 subjects, with 64 in a fasted state and 64 receiving a high-fat diet, and computed Cmax, AUC0-∞, and AUC0-t. Reference-scaled average bioequivalence (RSABE) methods and average bioequivalence (ABE) methods were used, and adverse events were assessed. Average maximum plasma concentrations (Cmax) of clopidogrel were significantly greater after 25 mg dose under fed conditions compared to fasted. Reference-scaled Cmax, AUC0-t, and AUC0-∞ were higher than the 0.294 cutoff during fasted, meeting RSABE criteria. Under fed conditions, SWR for AUC0-t and AUC0-∞ were lower than 0.294, permitting use of ABE. The 90% confidence intervals for AUC0-t and AUC0-∞ indicated bioequivalence. Pharmacokinetic parameters differed between fasted and fed states. The generic product was bioequivalent to the reference drug, and was safe and well tolerated. This suggests that they can be used interchangeably in a clinical setting.

Influence of statin treatment on pharmacokinetics and pharmacodynamics of clopidogrel and its metabolites in patients after coronary angiography/angioplasty.

Possible interaction between clopidogrel and CYP3A4-metabolised atorvastatin or non-CYP3A4-metabolised rosuvastatin was investigated based on pharmacokinetic parameters of clopidogrel and its metabolites as well as the platelet reactivity test in patients undergoing coronary angiography/angioplasty. The study involved 50 patients (62.7 ±â€¯7.8 years old) who underwent coronary angiography/angioplasty and were treated with clopidogrel and atorvastatin or rosuvastatin during the six months after the procedure. The P2Y12 reaction units (PRU) and pharmacokinetic parameters of clopidogrel, diastereoisomers of thiol metabolite (inactive H3 and active H4), and inactive carboxylic metabolite were measured 12-18 h and six months after the coronary angiography/angioplasty. There were no significant differences in concentrations of clopidogrel and its metabolites including the H4 active metabolite in plasma of patients co-treated with clopidogrel and atorvastatin or rosuvastatin. The use of statins did not affect the pharmacokinetic parameters of the studied compounds. A significant correlation was found between the Cmax and AUC0-t of the active H4 isomer and platelet aggregation in a group of patients treated with rosuvastatin but not in the atorvastatin group. No significant differences in PRU values were observed between the atorvastatin and rosuvastatin groups at the beginning of the study (171.4 ±â€¯54.3 vs 146.3 ±â€¯48.1 PRU, p = 0.192) as well as at six months (173.7 ±â€¯45.8 vs 157.3 ±â€¯54.9 PRU, p = 0.562). However, in a small group of patients, who were discharged from atorvastatin to rosuvastatin, an increase in the PRU values accompanied by a decreased AUC of the H4 active isomer was observed. The study confirmed that the systemic exposure to clopidogrel and its active H4 isomer of thiol metabolite, as well as the antiplatelet effect of the drug, were not negatively affected by co-administration of atorvastatin as compared with rosuvastatin.

Clopidogrel Increases Dasabuvir Exposure With or Without Ritonavir, and Ritonavir Inhibits the Bioactivation of Clopidogrel.

Dasabuvir is mainly metabolized by cytochrome P450 (CYP) 2C8 and is predominantly used in a regimen containing ritonavir. Ritonavir and clopidogrel are inhibitors of CYP3A4 and CYP2C8, respectively. In a randomized, crossover study in 12 healthy subjects, we examined the impact of clinical doses of ritonavir (for 5 days), clopidogrel (for 3 days), and their combination on dasabuvir pharmacokinetics, and the effect of ritonavir on clopidogrel. Clopidogrel, but not ritonavir, increased the geometric mean AUC0-∞ of dasabuvir 4.7-fold; range 2.0-10.1-fold (P = 8·10-7 ), compared with placebo. Clopidogrel and ritonavir combination increased dasabuvir AUC0-∞ 3.9-fold; range 2.1-7.9-fold (P = 2·10-6 ), compared with ritonavir alone. Ritonavir decreased the AUC0-4h of clopidogrel active metabolite by 51% (P = 0.0001), and average platelet inhibition from 51% without ritonavir to 31% with ritonavir (P = 0.0007). In conclusion, clopidogrel markedly elevates dasabuvir concentrations, and patients receiving ritonavir are at risk for diminished clopidogrel response.

The association between clopidogrel and 2-oxo-clopidogrel plasma levels and the long-term clinical outcome after acute myocardial infarction
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BACKGROUND AND OBJECTIVES: A significant number of ischemic events occur after acute myocardial infarction (MI), even when adhering to dual antiplatelet therapy including aspirin and clopidogrel. The aim of our study was to investigate the association between the concentration of the prodrug clopidogrel and its intermediary metabolite 2-oxo-clopidogrel plasma as well as demographic and clinical factors, and the long-term clinical outcome in patients with their first acute MI, ST-elevation myocardial infarction (STEMI) or non-ST-elevation myocardial infarction NSTEMI, treated with percutaneous coronary intervention (PCI). MATERIALS AND METHODS: This study included 172 consecutive patients with their first acute MI, 88 STEMI, and 84 NSTEMI, treated with PCI. On the third day of hospitalization, blood samples were collected from each patient to measure the concentration of clopidogrel and its metabolite 2-oxo-clopidogrel using the UHPLC-DAD-MS method. The following clinical outcomes were registered during the 28-month follow-up: mortality from cardiovascular causes, nonfatal MI, nonfatal stroke, and hospitalization for urgent myocardial revascularization or heart failure. RESULTS: Lower dose-adjusted clopidogrel concentrations (p < 0.05) were measured in NSTEMI patients with a composite of the hard clinical endpoint events of cardiovascular mortality, non-fatal MI, or a nonfatal stroke. During the follow-up, there was a 3.4 times higher risk of hard clinical endpoint events (p < 0.05) for each unit decrement of the dose-adjusted clopidogrel plasma concentration. Lower dose-adjusted concentrations of clopidogrel in these patients were associated with lower left ventricular ejection fraction (p < 0.001), and fentanyl (p < 0.001) and pantoprazole administration (p < 0.01) during the acute phase of MI. CONCLUSION: In patients with acute MI treated with PCI, lower dose-adjusted clopidogrel and dose-adjusted 2-oxo-clopidogrel plasma concentrations were associated with an increased risk of ischemic events.
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Pharmacokinetics and pharmacokinetic/pharmacodynamic relationship of vicagrel, a novel thienopyridine P2Y12 inhibitor, compared with clopidogrel in healthy Chinese subjects following single oral dosing.

BACKGROUND AND OBJECTIVES: Vicagrel, a novel thienopyridine antiplatelet agent, is an analogue of clopidogrel in development for the treatment of acute coronary syndromes. This study investigated the pharmacokinetic properties of vicagrel after single oral dosing with a direct comparison with clopidogrel in healthy Chinese subjects in the first two phase I clinical studies. The relationship between the exposure to the active metabolite and the platelet reactivity was also assessed for vicagrel. METHODS: Study A was a single-ascending-dose study of vicagrel (5-75 mg) compared with clopidogrel (75 mg) in 67 healthy volunteers. Study B was a randomized, two-period, crossover, loading-dose study of vicagrel 20 mg compared with clopidogrel 300 mg in 12 healthy subjects. Plasma concentrations of three common metabolites of vicagrel and clopidogrel, the active thiol metabolite H4, the inactive thiol metabolite H3, and the S-methylated form of H3 (SM3, the major metabolite of vicagrel), were determined using a validated UHPLC-MS/MS method. The relationship between the AUC0-t of active H4 and the P2Y12 reaction units at 4 h after administration of vicagrel was investigated. Blood concentrations of vicagrel were determined after a single oral administration of vicagrel 25 mg to two healthy Chinese subjects. RESULTS: In the single-ascending-dose study, vicagrel was metabolized rapidly with the median tmax for the three metabolites, namely, H4, H3, and SM3, ranging from 0.25-1.75 h. The pharmacokinetics of the three metabolites for vicagrel were linear across the dose range of 5-75 mg, with the mean Cmax and AUCs for H4 and H3 increasing in an approximately 1:1 dose-proportional manner and for SM3 increasing in a <1:1 dose-proportional manner. The median tmax for active H4 in the vicagrel 5 mg group was slightly shorter than that in the clopidogrel 75 mg group (0.50 versus 0.75 h). The mean AUC0-t for H4 in the vicagrel 5 mg group was similar to that in the clopidogrel 75 mg group (11.7 versus 11.8 ng∙h/mL). The AUC0-t of active H4 was apparently associated with the P2Y12 reaction units at 4 h for vicagrel. In the loading-dose study, for active H4, the median tmax was slightly shorter (0.50 versus 0.75 h) and the mean AUC0-t was 29% higher in the vicagrel 20 mg group than those in the clopidogrel 300 mg group. After a single oral administration of vicagrel 25 mg to 2 subjects, vicagrel was detected in blood but in very low concentrations. CONCLUSIONS: Vicagrel was rapidly and extensively metabolized, and the levels of the parent drug in circulation were very low. The pharmacokinetics of the three metabolites of vicagrel were linear and predictable across the dose range of 5-75 mg. The AUC of active H4 was apparently associated with the P2Y12 reaction units for vicagrel. For active H4, vicagrel 5 mg produced similar exposure (AUC) with more rapid appearance compared with clopidogrel 75 mg, and vicagrel 20 mg produced even slightly higher exposure (AUC) with more rapid appearance compared with clopidogrel 300 mg in humans. TRIAL REGISTRATION: CTR20150346, CTR20160379.

The association of clopidogrel and 2-oxo-clopidogrel plasma levels and the 40 months clinical outcome after primary PCI.

Background A significant number of ischemic events occur even when adhering to dual antiplatelet therapy including aspirin and clopidogrel. Objectives The aim of our study was to determine predictors of long-term patient clinical outcome, among variables such as prodrug clopidogrel and intermediary metabolite 2-oxoclopidogrel concentrations, as well as patients' clinical characteristics. Setting Department for the Treatment of Acute Coronary Syndrome in tertiary teaching hospital, Serbia. Methods This study enrolled 88 consecutive patients with first STEMI, treated with primary PCI, within 6 h of the chest pain onset and followed them 40 months. On the third day of hospitalization, blood samples were collected from each patient to measure clopidogrel and its metabolite 2-oxo-clopidogrel concentration by UHPLC-DAD-MS method. Main outcome measure Mortality from cardiovascular causes, nonfatal myocardial infarction, nonfatal stroke or hospitalization for urgent myocardial revascularization or heart failure. Results The composite clinical outcome of cardiovascular mortality, nonfatal myocardial infarction, nonfatal stroke, or hospitalization for urgent myocardial revascularization or heart failure, was registered in 31 patients (35.2%) during the 40-month follow-up. Lower clopidogrel (p < 0.05) and dose-adjusted clopidogrel concentrations (p < 0.05) were associated with the higher incidence of composite outcome events. Their low plasma concentrations may be predicted by fentanyl administration (p < 0.001) and creatinine clearance (p < 0.01). The decrease in dose-adjusted clopidogrel unit for each ng/ml/mg increases the risk 21.7 times (p < 0.05). Conclusion Clopidogrel dose-adjusted plasma concentration in STEMI patients, as well as multivessel coronary artery disease, showed significance in predicting an unfavorable composite clinical outcome after 40-month follow-up.

Effects of CYP2C19 Genetic Polymorphisms on the Pharmacokinetic and Pharmacodynamic Properties of Clopidogrel and Its Active Metabolite in Healthy Chinese Subjects.

PURPOSE: Some studies in the white population have shown that carriers of at least 1 loss-of-function allele in the gene that encodes the cytochrome P-450 2C19 isozyme (CYP2C19) have lower levels of the clopidogrel active metabolite (CAM) and a reduced antiplatelet effect of clopidogrel. However, data are limited regarding the association between CYP2C19 genetic variants and exposure to CAM and on the pharmacodynamic properties of CAM in the Chinese population. Data from the white population cannot be extrapolated to the Chinese population because of the marked interethnic differences in CYP2C19 variants. This study was aimed to investigate the influence of CYP2C19 genetic polymorphisms on the pharmacokinetic properties of CAM and the antiplatelet effect of clopidogrel in healthy Chinese volunteers, and to provide evidence for the role of a CYP2C19 genotyping test in predicting the antiplatelet effect of clopidogrel in the Chinese population. METHODS: Twenty healthy subjects received a single 300-mg dose of clopidogrel and were assigned to 1 of 3 groups according to CYP2C19 genotype: CYP2C19 *1/*1 (normal metabolizers [NM]; n = 8), CYP2C19 *1/*2 or *3 (intermediate metabolizers [IM]; n = 10) and CYP2C19 *2/*2 or *3 and *3/*3 (poor metabolizers [PM]; n = 2). Blood samples were collected at baseline and at 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, 6, 8, 12, and 24 hours after administration. The plasma concentrations of clopidogrel and CAM were analyzed by LC-MS/MS, and adenosine diphosphate-induced platelet aggregation was measured by light-transmittance aggregometry. FINDINGS: There were no significant differences in Cmax and AUC0-t of clopidogrel prodrug in the NM group compared with the IM and PM groups. The mean CAM Cmax value was significantly higher in the NM group than in IM and PM groups (45.39 [12.57] vs 29.15 [7.92] ng/mL [P = 0.003] and 19.55 [2.19] ng/mL [P = 0.004], respectively). The mean CAM AUC0-t value was significantly higher in the NM group than in the IM and PM groups (61.05 [21.63] vs 37.67 [11.01] ng · h/mL [P = 0.007] and 27.08 [2.72] ng · h/mL [P = 0.016]). The NM group exhibited a significantly higher percentage of inhibition of platelet aggregation than did the IM or PM group (P = 0.001). The correlations between the pharmacokinetic properties (Cmax, AUC0-t) of CAM and the pharmacodynamic data (maximal and inhibition of platelet aggregation) were significant (both, Pearson r > 0.5 and P < 0.01). IMPLICATION: In these healthy Chinese subjects, carriers of CYP2C19 loss-of-function allele(s) had significantly reduced exposure of CAM and decreased levels of inhibition of platelet aggregation with clopidogrel; these genotypes therefore might be a determinant for the formation of CAM and its antiplatelet effects. Study identifier: ChiCTR-OCH-14004382.

Clopidogrel in Critically Ill Patients.

Only limited data are available regarding the treatment of critically ill patients with clopidogrel. This trial investigated the effects and the drug concentrations of the cytochrome P450 (CYP450) activated prodrug clopidogrel (n = 43) and the half-life of the similarly metabolized pantoprazole (n = 16) in critically ill patients. ADP-induced aggregometry in whole blood classified 74% (95% confidence intervals 59-87%) of critically ill patients as poor responders (n = 43), and 65% (49-79%) responded poorly according to the vasodilator-stimulated phosphoprotein phosphorylation (VASP-P) assay. Although the plasma levels of clopidogrel active metabolite normally exceed the inactive prodrug ∼30-fold, the parent drug levels even exceeded those of the metabolite 2-fold in critically ill patients. The half-life of pantoprazole was several-fold longer in these patients compared with reference populations. The inverse ratio of prodrug/active metabolite indicates insufficient metabolization of clopidogrel, which is independently confirmed by the ∼5-fold increase in half-life of pantoprazole. Thus, high-risk patients may benefit from treatment with alternative platelet inhibitors.