Abstract
Aim
The genetic polymorphism of CYP2C19 influences clopidogrel metabolism and resistance. Aim was to assess the association between CYP2C19 loss of function variation, clopidogrel resistance based on platelet reactivity units and clinical outcomes.
Methods
A total of 668 patients of Acute Coronary Sundrome (ACS) who underwent Percutaneous Coronary Intervention (PCI) were subjected to genetic screening and 143 patients undrewent platelet function test to study the association between drug metabolization and its effects based on platelet reactivity unit values.
Results
Clopidogrel resistance with CYP2C 19 loss of function variation was noted in 54.64% of patients. Clinical outcomes, such as target vessel revascularization, target lesion revascularization, in-stent restenosis, and stent thrombosis, were also studied.
Conclusion
CYP2C19 loss of function variation is strongly associated with clopidogrel resistance and adverse clinical outcomes.
Keywords: Acute coronary syndrome, CYP2C19, Clopidogrel resistance
1. Introduction
CYP2C19 is a highly polymorphic enzyme that gives varied responses to the clopidogrel drug in patients with acute coronary syndrome (ACS).1 Loss of function (LOF) variations is associated with reduced efficacy or no activity of clopidogrel. The association between LOF variants and attenuated platelet inhibition is clinically actionable. Patient demographic factors, genetics, and drug-related factors are used to examine the inter-patient response to clopidogrel.2 The prevalence of clopidogrel resistance in India ranges from 5 to 44 %.3 By selecting antiplatelet therapy based on the patient's platelet function testing (PFT) and genetics, ischemic and bleeding events may be reduced.4 The present study is aimed to assess the association between CYP2C19 metabolization status, clopidogrel resistance by PFT based on platelet reactivity units (PRUs), and clinical outcomes.
2. Methods
A total of 668 individuals undergoing PCI were included in this study after seeking ethical approval from the Institutional Review Board (IRB) and obtaining informed consent from the subjects. The samples were collected after informed consent according to clinical study guidelines laid down by ICMR and the institution. All ACS patients were included in this study. Patients with stable ischemic heart disease and unable to come for follow up were excluded as per our earlier studies.5,6
The DNA was isolated using the QIAGEN DNA Mini Kit, followed by qualitative and quantitative evaluation. Genotyping was done as per standard protocol.6 For studying the platelet function test, 'Verify PRU' (Warfen, Barcelona, Spain) was used according to the manufacturer's protocol.
3. Results
A total of 668 subjects were included in this study, 78 % (n = 518) were males, 34 % were hypertensive, 28 % were diabetic, 19 % consumed alcohol, 19 % were smokers, and 39 % were overweight.
The CYP2C19 loss of function variants were noted in 54.64 % (n = 365) of the patients and were categorized as poor metabolizers for clopidogrel while the rest of individuals belonged to ultra (18.26 %) and extensive (27.1 %) metabolizers. Further, to examine the association between the genetic testing results based on clopidogrel metabolization status and PRU values, random testing for PFT was done for 143 individuals, where 45 %, 12 % and 43 % had good ( PRU value < 160 U), intermediate (PRU 160-200 U) and no drug effect (PRU > 200U) respectively. Significant statistical correlation was observed between genetic test results and PRU values with Odds Ratio (OR) of 10.4 (p-value 0.02) (Table 1).
Table 1.
Chi-square test to establish relation between the genetic clopidogrel metabolizes status and drug effect calculated based on PRU values.
| Good drug effect (in percentage) | Intermediate drug effect (in percentage) | Poor drug effect (in percentage) | Chi square (p value) | |
|---|---|---|---|---|
| GEMa | 53 | 50 | 31 | 10.4 (0.02) |
| GIMb | 41 | 50 | 51 | |
| GPMc | 6 | 0 | 18 |
GEM = Genetic extensive metabolizer observed using CYP2C19 (*1/*2*/17) alleles.
GIM = Genetic Intermediate metabolizer observed using CYP2C19 (*1/*2*/17) alleles.
GPM = Genetic poor metabolizer observed using CYP2C19 (*1/*2*/17) alleles.
The relationship between clopidogrel resistance and clinical outcomes in patients who underwent PCI was evaluated (Table 2). In TVR and TLR cases, 75 % and 88.85 % were clopidogrel resistant (Intermediate Metabolizers and Poor Metabolizers) respectively. Among TLR patients , 88.85 % of the individuals had PRU values above 160, indicating increased platelet reactivity were observed in TLR individuals. Among patients with ISR, 80 % had PRU values above 160 and were found to be clopidogrel resistant. All individuals of stent thrombosis were clopidogrel resistant and all of these individuals had PRU values above 160.
Table 2.
Association of TVR, TLR, ISR & stent thrombosis with genetic profile of the patients-clopidogrel metabolizer status.
| Re-vascularization Event | Number of patients | Percentage of patients (n = 668) | Normal metabolizer | Intermediate metabolizer | Poor metabolizer |
|---|---|---|---|---|---|
| TVRa | 16 | 2.40 % | 25 % | 31.20 % | 43.75 % |
| TLRb | 9 | 1.34 % | 11.10 % | 33.30 % | 55.50 % |
| ISRc | 5 | 0.75 % | 20 % | 40 % | 40 % |
| STd | 4 | 0.60 % | – | 25 % | 75 % |
Target vessel re-vascularization.
Target lesion re-vascularization.
Instent restenosis.
Stent Thrombosis.
12 patients died due to cardiovascular disease, 75 % were clopidogrel resistant as per genetic testing and 83.32 % had more than 160 PRU value. Our analysis focused on the *2, *3, and *17 and statistical power for these specific single nucleotide polymorphisms (SNPs) exceeds 0.80, a critical threshold, considering the anticipated OR and effective allele frequencies derived from our study performed in the Indian cohort.5
4. Discussion
The present study highlights the clinical significance of clopidogrel resistance in ACS among studied subjects and Impact of CYP2C19 variants on enzyme activity and clopidogrel metabolism (Fig. 1). This is consistent with various in vitro studies and randomized trials.7,8 In the studied cohort, the prevalence of LOF variants is 54.64 % in ACS patients, which aligns with previously reported prevalence of these variants in Indian populations9,10 and emphasizes on relevance of genetic testing. It has been observed that patients with inadequate response to antiplatelet medications are significantly at a greater risk of stent thrombosis and myocardial infarction as evident by a study conducted on 2208 patients where individuals with CYP2C19 LOF alleles had a greater probability of cardiovascular events.11
Fig. 1.
Impact of CYP2C19 variants on enzyme activity and clopidogrel metabolism. Figure illustrates the impact of CYP2C19 genetic variants on enzyme activity and clopidogrel metabolism. The image depicts four metabolizer phenotypes: Normal (Ultra and extensive metaboliser * 1/17,*17/*17, *1/*1), Intermediate (*1/*2 or *1/*3), and Poor (*2/*2, *2/*3, or *3/*3). Enzyme activity progressively decreases from normal to poor metabolizers, represented by a color gradient from green to red. The figure demonstrates the conversion efficiency of clopidogrel to its active metabolite for each phenotype. Normal metabolizers exhibit efficient conversion and platelet inhibition, while poor metabolizers show minimal conversion and inadequate inhibition. This visual representation effectively explains why individuals with certain CYP2C19 variants may experience reduced clopidogrel efficacy, potentially increasing their risk of adverse cardiovascular events in acute coronary syndrome patients undergoing percutaneous coronary intervention.
Moreover, the present study showed a significant association between CYP2C19 genetic variants in ACS patients and physiological PFT-PRU values, which can act as a marker for clinical outcomes. Mortality due to cardiovascular causes was in 12 patients (1.79%) of which 75% had LOF variants. All individuals who experienced stent thrombosis in our study were poor or intermediate metabolizers of clopidogrel. The limitation of the present study was that the follow-up period was only six months. It is recommended to conduct a follow-up study with a longer duration, such as 1–2 years, in order to assess their long-term outcomes. However, we acknowledge that concomitant medications could also potentially influence clopidogrel efficacy and resistance. These interactions could potentially confound the relationship between genetic polymorphisms and clopidogrel response. While we did not specifically analyze concomitant medications in this study, we recognize this as a limitation. Future research should include a comprehensive analysis of patients' medication histories to better understand the complex interplay between genetic factors, drug interactions, and clopidogrel resistance.
The implementation of pharmacogenomics (PGX) -based genotype guided clopidogrel medication is a complex challenge.12 There are non-genetic factors involved which require intervention of cardiovascular expert to decide suitable antiplatelet treatment. Hence, for pharmacogenomic based therapy, the incorporation of such factors into algorithms (to generate PGX report for suitable antiplatelet therapy) will enhance the precision and efficacy of genotype-guided clopidogrel medication.
5. Conclusion
This study highlights the correlation between CYP2C19 LOF polymorphisms, clopidogrel resistance as measured by PRUs and adverse clinical outcomes in ACS patients post PCI. This underscores the importance of personalized treatment based on genetic findings for optimizing antiplatelet therapy and to improve the outcomes.
Declaration of competing interest
All authors declare no competing interest. Dr. Anuka Sharma and Dr. Varun Sharma are the employees of NMC Genetics India Private Limited.
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