Ciprofol is a novel short-acting intravenous anaesthetic developed in China that is mainly metabolized by cytochrome P450 2B6 (CYP2B6) and uridine diphosphate glucuronosyltransferase 1A9 (UGT1A9). Currently, insufficient evidence is available to support drug‒drug interactions between ciprofol and CYP2B6 inactivators. Here, we established a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method to assess the concentration of ciprofol and investigated the effects of psoralen and clopidogrel on the metabolism of ciprofol in liver microsomes and rats. In rat and human liver microsomes, the median inhibitory concentration (IC50) values of psoralen were 63.31 μmol·L-1 and 34.05 μmol·L-1, respectively, showing mild inhibitory effects on ciprofol metabolism, whereas the IC50 values of clopidogrel were 6.380 μmol·L-1 and 2.565 μmol·L-1, respectively, with moderate inhibitory effects. SD rats were randomly divided into three groups: psoralen (27 mg·kg-1), clopidogrel (7.5 mg·kg-1), and the same volume of 0.5% carboxy methyl cellulose. After 7 days, all rats were injected with 2.4 mg·kg-1 ciprofol. Compared with the control group, the AUC and MRT values of ciprofol in the psoralen and clopidogrel groups were significantly greater, whereas the CL values were significantly lower. In addition, the durations of loss of righting reflex (LORR) in the psoralen and clopidogrel groups were 16.1% and 23.0% longer than that in the control group, respectively. In conclusion, psoralen and clopidogrel inhibit ciprofol metabolism to different degrees and prolong the duration of LORR in rats.

Clopidogrel is widely used worldwide as an antiplatelet therapy in patients with acute coronary disease. Genetic factors influence interindividual variability in response. Some studies have explored the polygenic contributions in the drug response, generating pharmacogenomic risk scores (PgxPRS). Importantly, these factors are less explored in underrepresented populations, such as Latin-American countries. Identifying patients at risk of high-on-treatment platelet reactivity (HTPR) is highly valuable in translational medicine. In this study we used a custom next-generation sequencing (NGS) panel composed of 91 single nucleotide polymorphisms (SNPs) and 28 genes related to clopidogrel metabolism, to analyze 70 patients with platelet reactivity values, assessed through closure time (CT). Our results demonstrated the association of SNPs with HTPR and non-HTPR, revealing the strongest associations with rs2286823 (OR: 5,0; 95% CI: 1,02-24,48; p: 0,03), rs2032582 (OR: 4,41; 95% CI: 1,20-16,12; p: 0,019), and rs1045642 (OR: 3,38; 95% CI: 0,96-11,9; p: 0,05). Bivariate regression analysis demonstrated the significant association of several SNPs with the CT value, a \"surrogate\" biomarker of clopidogrel response. Exploratory results from the LASSO regression model showed a high discriminatory capacity between HTPR and non-HTPR patients (AUC: 0,955), and the generated PgxPRS demonstrated a significant negative association between the risk score, CT value, and the condition of HTPR and non-HTPR. To our knowledge, our study addresses for the first time the analysis of the polygenic contribution in platelet reactivity using NGS and establishes PgxPRS derived from the LASSO model. Our results demonstrate the polygenic implication of clopidogrel response and offer insights applicable to the translational medicine of antiplatelet therapy in an understudied population.

Antiplatelet therapy is an important factor influencing the postterm patency rate of carotid artery stenting (CAS). Clopidogrel is a platelet aggregation inhibitor mediated by the adenosine diphosphate receptor and is affected by CYP2C19 gene polymorphisms in vivo. When the CYP2C19 gene has a nonfunctional mutation, the activity of the encoded enzyme will be weakened or lost, which directly affects the metabolism of clopidogrel and ultimately weakens its antiplatelet aggregation ability. Therefore, based on network pharmacology, analyzing the influence of CYP2C19 gene polymorphisms on the antiplatelet therapeutic effect of clopidogrel after CAS is highly important for the formulation of individualized clinical drug regimens. The effect of the CYP2C19 gene polymorphism on the antiplatelet aggregation of clopidogrel after CAS was analyzed based on network pharmacology. A total of 100 patients with ischemic cerebrovascular disease who were confirmed by the neurology department and required CAS treatment were studied. CYP2C19 genotyping was performed on all patients via a gene chip. All patients were classified into the wild-type (WT) group (*1/*1), heterozygous mutation (HTM) group (CYP2C19*1/*2, CYP2C19*1/*3), and homozygous mutation (HMM) group (CYP2C19*2/*2, CYP2C19*2/*3, and CYP2C19*3/*3). High-performance liquid chromatography (HPLC) with tandem mass spectrometry (MS/MS) was used to detect the blood concentration of clopidogrel and the plasma clopidogrel clearance (CL) rate in different groups of patients before and after clopidogrel treatment. The platelet aggregation rate of patients with different genotypes was measured by turbidimetry. The incidences of clopidogrel resistance (CR) and stent thrombosis in different groups after three months of treatment were analyzed. The results showed that among the different CYP2C19 genotypes, patients from the HTM group accounted for the most patients, while patients from the HTM group accounted for the least patients. Similarly, the clopidogrel CL of patients in the HMM group was lower than that of patients in the WT group and HTM group (P < 0.01). The platelet inhibition rate of patients in the HMM group was evidently inferior to that of patients in the WT group and HTM group (P < 0.01). The incidence of CR and stent thrombosis in the WT group was notably lower than that in the HTM and HMM groups (P < 0.01). These results indicate that the CYP2C19 gene can affect CR occurrence and stent thrombosis after CAS by influencing clopidogrel metabolism and platelet count.

A recently discovered haplotype-CYP2C:TG-determines the ultrarapid metabolism of several CYP2C19 substrates. The platelet inhibitor clopidogrel requires CYP2C19-mediated activation: the risk of ischemic events is increased in patients with a poor (PM) or intermediate (IM) CYP2C19 metabolizer phenotype (vs. normal, NM; rapid, RM; or ultrarapid, UM). We investigated whether the CYP2C:TG haplotype affected efficacy/bleeding risk in clopidogrel-treated patients. Adults (n = 283) treated with clopidogrel over 3-6 months were classified by CYP2C19 phenotype based on the CYP2C19*2*17 genotype, and based on the CYP2C19/CYP2C cluster genotype, and regarding carriage of the CYP2:TG haplotype, and were balanced on a number of covariates across the levels of phenotypes/haplotype carriage. Overall, 45 (15.9%) patients experienced ischemic events, and 49 (17.3%) experienced bleedings. By either classification, the incidence of ischemic events was similarly numerically higher in PM/IM patients (21.6%, 21.8%, respectively) than in mutually similar NM, RM, and UM patients (13.2-14.8%), whereas the incidence of bleeding events was numerically lower (13.1% vs. 16.6-20.5%). The incidence of ischemic events was similar in CYP2C:TG carries and non-carries (14.1% vs. 16.1%), whereas the incidence of bleedings appeared mildly lower in the former (14.9% vs. 20.1%). We observed no signal to suggest a major effect of the CYP2C19/CYP2C cluster genotype or CYP2C:TG haplotype on the clinical efficacy/safety of clopidogrel.

The main goal of the study was to improve the compliance and convenience of patients by designing and development of an immediate release (IR) fixed-dose combination (Clopidogrel bisulphate and Aspirin) tablets. The proposed combination product utilizes Clopidogrel to protect the moisture-sensitive aspirin component, enhancing its stability against atmospheric conditions. Response-surface approach (Design Expert vs. 13) was used to generate this IR tablet by calculating the right composition of independent variables such as Microcrystalline cellulose 102, pregelatinized starch and Hydroxypropyl cellulose. 32 factorial design was used to estimate the effects of these independent variables on the responses of dependent variables (disintegration & friability) and constructed a total of nine (9) formulations. Pre and Post formulation, quality control parameters were investigated as per pharmacopeia. A systematic approach was used for the optimization process and a prototype checkpoint batch (CPB) based on the better contrast of independent variables was prepared. In vitro analysis of formulations was carried out to estimate the responses. Friability was found in the range of 0.088-1.076%w/w, except F1 = 1.076 all are within limits (NMT 1.0%). Disintegration time was recorded 7.3 ± 1.20 as lower and 24.5 ± 1.63 min was the highest. The release of drugs from their dosage form was fast and rapid, for clopidogrel after 15min was 70.42-96.82% with SD ± 8.71 and aspirin was 69.88-91.49% in 15 min with SD ± 6.41, all the tablets were released more than 80% in 20 min. The stability outcomes of CPB tablets after 15 days of stress study (60 ± 2°C and 75 ± 5%) indicated good compatibility and stability of APIs with excipients. It was concluded that the direct compression method can be preferred to prepare a combination product with cost-effectiveness. It was also concluded that the proposed methodology could increase Aspirin\'s stability and allow for an aqueous coating system to finish the product with a film coating. By using Design Expert software, the best composition of the formulation can be selected and optimized in a short period of time with minimum trial and errors. The results also demonstrated that the use of a fixed-dose combination tablet instead of the individual is expected to be more convenient to patients and thus improves patient compliance and decreases the occurrence of adverse effects and side effects.

Dual antiplatelet therapy (DAPT) with the combination of clopidogrel and aspirin is recommended for preventing secondary ischemic events in patients with acute coronary syndrome (ACS) or acute ischemic stroke (AIS). Proton pump inhibitors (PPIs) are suggested as preventive treatment for these patients. Due to clopidogrel-PPI interactions, separating their administration might be considered. However, a paucity of studies has been conducted to investigate the outcome differences between concurrent and interval-based use in ACS and AIS patients. Our study aimed to evaluate clinical outcomes based on administration timing. This study included patients with ACS or AIS onset or recurrence of within the last month. Patients who were expected to receive DAPT for at least 6 months and who were currently taking or planning to take esomeprazole were included. Patients were divided into Group 1 (interval administration group, IA group) and Group 2 (concurrent administration group, CA group) according to the interval between esomeprazole and DAPT administration. The time interval was based on 12 hours. The primary outcome was the occurrence of major adverse cardiocerebrovascular events (MACCEs), and safety outcomes were defined as major bleeding, minor bleeding and gastrointestinal bleeding and intracranial hemorrhage. A total of 3600 patients completed this study. The proportions of patients in the 2 groups were as follows: CA group, 99% (n = 3489) and IA group, 1% (n = 111). The primary outcome occurred in 0.9% of patients in the IA group and 1.8% of patients in the CA group (P = .51). There was no significant distinction in the overall bleeding risk of the CA group compared to that of the IA group (2.75% in the CA group and 2.70% in the IA group). Additionally, there was no significant difference observed between the 2 groups for safety outcomes. This multicenter, prospective, observational study that enrolled patients with ACS or AIS demonstrated that there was no significant difference in the occurrence of MACCEs and bleeding issues within 6 months according to the medication administration interval. The majority of patients with DAPT were taking PPIs simultaneously in real-world practice.

It is controversial whether hemodialysis affects the efficacy of the antiplatelet agents. We aimed to investigate the impact of hemodialysis on efficacies of the antiplatelet agents in coronary artery disease (CAD) patients complicated with end-stage renal disease (ESRD). 86 CAD patients complicated with ESRD requiring hemodialysis were consecutively enrolled. After 5-day treatment with aspirin and clopidogrel or ticagrelor, the platelet aggregations induced by arachidonic acid (PLAA) or adenosine diphosphate (PLADP), and the P2Y12 reaction unit (PRU) were measured before and after hemodialysis. The propensity matching score method was adopted to generate a control group with normal renal function from 2439 CAD patients. In patients taking aspirin, the PLAA remained unchanged after hemodialysis. In patients taking clopidogrel, the PLADP (37.26 ± 17.04 vs. 31.77 ± 16.09, p = 0.029) and corresponding clopidogrel resistance (CR) rate (23 [48.9%] vs. 14 [29.8%], p = 0.022) significantly decreased after hemodialysis, though PRU remained unchanged. Subgroup analysis indicated that PLADP significantly decreased while using polysulfone membrane (36.8 ± 17.9 vs. 31.1 ± 14.5, p = 0.024). In patients taking ticagrelor, PLADP, and PRU remained unchanged after hemodialysis. ESRD patients had higher incidences of aspirin resistance (AR) and CR compared to those with normal renal function (AR: 16.1% vs. 0%, p = 0.001; CR: 48.4% vs. 24.8%, p = 0.024). Hemodialysis does not have negative effect on the efficacies of aspirin, clopidogrel and ticagrelor in ESRD patients with CAD. ESRD patients have higher incidences of AR and CR compared with those with normal renal function.Trial registration ClinicalTrials.gov Identifier: NCT03330223, first registered January 4, 2018.

CYP2C19 gene has multiple single nucleotide polymorphism (SNP), which is the major determinant for clopidogrel treatment responses. Therefore, CYP2C19 SNP detection is essential for predicting clopidogrel efficacy. Currently, there is still no quick and effective method for routine detection of common CYP2C19 SNPs in clinical laboratories, which is critically needed prior to clopidogrel treatment. AllGlo™ based quantitative PCR was used to develop a novel genotyping method for CYP2C19 SNP detection, termed CyPAllGlo. The performance of CyPAllGlo was compared with that of the commonly used fluorescence in situ hybridization (FISH) method, and the data was verified by DNA sequencing. CyPallGlo was used to identify CYP2C19 polymorphisms in 363 patients with coronary heart disease. The univariate analysis was used to access the antiplatelet efficacy of clopidogrel in patients. The associations between CYP2C19 polymorphisms and clopidogrel efficacy were analyzed. Using CyPAllGlo to detect CYP2C19*2 and CYP2C19*3 alleles was highly specific and fast. The detection limit was approximately 0.07 µg/µl and 0.7 µg/µl for CYP2C19*2 and CYP2C19*3, respectively. The consistency between FISH and CyPAllGlo were 98.07% for CYP2C19*2 and 99.17% for CYP2C19*3. DNA sequencing showed that the accuracy of CyPAllGlo was 100%. The analysis time for the whole CyPAllGlo procedure was approximately 60 min. Univariate analysis showed that the anticoagulation efficacy of clopidogrel was related to patient age, CYP2C19 genotype, metabolic phenotype, and LDL level. The logistic regression analysis showed that the genotype of CYP2C19 and metabolic phenotype was the two risk factors for clopidogrel antiplatelet ineffectiveness. This novel CyPAllGlo is a rapid and accurate method for detection of CYP2C19 SNP. The specificity and consistency of CyPAllGlo are comparable with that of widely used DNA sequencing. These findings provide valuable rapid method for predicting clopidogrel efficacy, which can be quickly translated to improve personalized precision medicine for coronary heart disease treatment.

Most proton pump inhibitors (PPIs) inhibit the bioactivation of clopidogrel to its active metabolite. There is controversy concerning whether PPIs alter the effectiveness of clopidogrel in reducing the risk of ischemic stroke (IS). We therefore aimed to examine the risk of IS associated with concomitant use of clopidogrel and omeprazole, a PPI commonly used in clinical settings. We conducted a retrospective cohort study using the National Health Insurance Research Database of Taiwan dated from 2000 to 2013. The study cohorts comprised 407 patients diagnosed with acute coronary syndrome (ACS) and with concomitant use of clopidogrel and omeprazole (the exposed cohort), 814 ACS patients with single use of clopidogrel (the comparison cohort), and 230 ACS patients with concurrent use of clopidogrel and pantoprazole (the reference cohort). The primary outcome was incident IS. The hazard ratios (HRs) and 95% confidence intervals (CIs) derived from the time-dependent Cox regression model were used to assess the association between concomitant use of clopidogrel and omeprazole and the risk of IS. The incidence rate of IS was significantly higher in the exposed cohort (81.67 per 1000 person-years) than in the comparison cohort (57.45 per 1000 person-years), resulting in an adjusted HR of 1.39 (95% CI 1.03-1.74). By contrast, there was no significant difference in the risk of IS between the exposed and reference cohorts (adjusted HR 1.11; 95% CI 0.81-1.52). The present study revealed that patients taking both clopidogrel and omeprazole was associated with an increased risk of IS.

In this study, we aimed to investigate the association between gut microbiota and high on-treatment platelet reactivity (HTPR) in patients with acute ischemic stroke (AIS).
We enrolled a total of 48 AIS patients, including 19 HTPR patients and 29 non-high on-treatment platelet reactivity (NHTPR) patients, along with 10 healthy controls. Clinical and laboratory data, as well as stool samples, were collected from all participants. The composition and function of gut microbiota were assessed using 16S rRNA sequencing. Differences in the gut microbiota between the two groups were analyzed, and a diagnostic model based on the gut microbiota was established using random forest model.
HTPR patients exhibited a decreased microbial richness compared to NHTPR patients. Additionally, the relative abundance of unidentified_Clostridia and Ralstonia was lower in HTPR patients. Significant differences in biological functions, such as toxoplasmosis, were observed between the two groups. The combination of Ralstonia, unidentified-Clostridia, Mailhella, Anaerofustis, and Aggregatibacter showed excellent predictive ability for HTPR occurrence (AUC=0.896). When comparing AIS patients with healthy controls, alterations in the microbiota structure were observed in AIS patients, with imbalances in short-chain fatty acid-producing bacteria and pathogenic bacteria. Significant differences in biological functions, such as oxidative phosphorylation, were noted between the two groups. The combination of Alloprevotella, Terrisporobacter, Streptococcus, Proteus, and unidentified_Bacteria exhibited strong predictive power for AIS occurrence (AUC=0.994).
This study is the first to uncover the microbial characteristics of HTPR in AIS patients and demonstrate the predictive potential of specific bacterial combinations for HTPR occurrence.