Impact of esomeprazole on platelet reactivity and clinical outcome ...

Thrombosis Research 135 (2015) 1081–1086

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Regular Article

Impact of esomeprazole on platelet reactivity and clinical outcome according to CYP2C19 genotype in coronary heart disease patients during dual antiplatelet therapy Seiji Hokimoto a,⁎, Tomonori Akasaka a, Noriaki Tabata a, Yuichiro Arima a, Kenichi Tsujita a, Kenji Sakamoto a, Koichi Kaikita a, Kazunori Morita b, Naoki Kumagae b, Eiichiro Yamamoto a, Kentaro Oniki b, Kazuko Nakagawa b, Hisao Ogawa a a b

Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan Division of Pharmacology and Therapeutics, Graduate School of Medical and Pharmaceutical Sciences, and Center for Clinical Pharmaceutical Sciences, Kumamoto University, Kumamoto, Japan

a r t i c l e

i n f o

Article history: Received 28 December 2014 Received in revised form 12 February 2015 Accepted 30 March 2015 Available online 11 April 2015 Keywords: Cardiovascular disease Clopidogrel Pharmacogenetics Polymorphism Platelets

a b s t r a c t Objectives: The aim of this study was to investigate the effect of CYP2C19 polymorphism and co-therapy with esomeprazole on the antiplatelet efficacy of clopidogrel. Background: The antiplatelet efficacy of clopidogrel depends on CYP2C19 polymorphism or the co-administration of some kind of proton pump inhibitor (PPI). Methods: CYP2C19 genotype and the residual platelet reactivity (RPR) were measured in 361 coronary heart disease patients (male, mean age 69 yrs), and the risk of cardiovascular events over a 3-month follow-up was assessed to evaluate the impact of co-administration of esomeprazole during dual antiplatelet therapy with aspirin and clopidogrel. Results: The values of RPR did not differ between esomeprazole and non-esomeprazole groups (4389 ± 1112 versus 4079 ± 1355 AU · min, P = 0.103). RPR value was higher in intermediate metabolizers (IM) than in extensive metabolizers (EM) (4089 ± 1252 versus 3697 ± 1215 AU · min P = 0.012) and, similarly, higher in poor metabolizers (PM) than in IM (4884 ± 1027 versus 4089 ± 1252 AU · min, P b 0.001). There were no differences in RPR between esomeprazole and non-esomeprazole groups according to CYP2C19 genotype (EM, 3954 ± 1192 versus 3645 ± 1220 AU · min, P = 0.361; IM, 4401 ± 1063 versus 4051 ± 1271 AU · min, P = 0.293; PM, 4917 ± 669 versus 4876 ± 1099 AU · min, P = 0.907, respectively). There was also no difference in clinical outcomes between esomeprazole and non-esomeprazole groups in the three-month follow-up (0% versus 0.92%, P = 0.487). Conclusions: These results suggest that concomitant use of esomeprazole with clopidogrel is not associated with reduced antiplatelet efficacy of clopidogrel or increased risk of cardiovascular events, irrespective of CYP2C19 genotype. © 2015 Elsevier Ltd. All rights reserved.

Introduction Dual antiplatelet therapy (DAPT) with aspirin and clopidogrel is currently recommended for the prevention of atherothrombotic events in patients with acute coronary syndrome (ACS) who undergo percutaneous coronary intervention (PCI) [1–3]. Concern about bleeding risk, especially gastrointestinal bleeding, has emerged in patients with ACS or PCI requiring DAPT[4]. So, co-administration of PPI with DAPT is recommended in consensus report of the American College of Cardiology Foundation (ACCF), American College of Gastroenterology (ACG), and ⁎ Corresponding author at: Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kumamoto University, 1-1-1, Honjo, Chuo-ku, Kumamoto 860–8556, Japan. Tel.: +81 96 373 5175; fax: +81 96 362 3256. E-mail address: [email protected] (S. Hokimoto).

http://dx.doi.org/10.1016/j.thromres.2015.03.033 0049-3848/© 2015 Elsevier Ltd. All rights reserved.

American Heart Association (AHA) [5]. However, several studies have indicated that concomitant use of clopidogrel and a PPI is associated with reduced antiplatelet efficacy of clopidogrel, and increased adverse clinical outcomes after stent placement in patients with ACS [6–10]. One possibility is that both clopidogrel and PPI are metabolized by CYP isoenzymes, but to varying degrees, and that negative effect might be due in part to competitive inhibition of the CYP2C19 enzyme [11]. There is another problem on DAPT. Clopidogrel is a prodrug that requires hepatic biotransformation and has to be converted to an active metabolite [12]. Others and we have reported that CYP2C19 is an important factor in this activation process and that the presence of CYP2C19 loss-of-function (LOF) alleles is associated with high residual platelet reactivity (RPR) and adverse clinical outcomes in patients with coronary heart disease (CHD), especially patients following ACS and PCI [13–17]. In short, we need to consider the two points under the concomitant use

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of clopidogrel and PPI; drug-drug interaction with clopidogrel and PPI, and CYP2C19 polymorphism, which both could affect the antiplatelet efficacy of clopidogrel. We previously reported that the concomitant use of rabeprazole during dual antiplatelet therapy with clopidogrel and aspirin is not associated with high RPR [16] because rabeprazole is mostly metabolized by non-enzymatic pathways, and shows the least potent inhibition of CYP2C19 in vitro [18]. For other PPI, especially omeprazole, which is a major substrate and mostly metabolized by CYP2C19, there is concern about concomitant use with clopidogrel, although this is controversial [6,11,19]. Esomeprazole is an optical isomer of omeprazole, and less metabolized by CYP2C19 [20]; however, the impact of esomeprazole on platelet reactivity or clinical outcome is unknown in Japanese patients, who have a high incidence of CYP2C19 LOF alleles compared with Caucasians, during dual antiplatelet therapy with aspirin and clopidogrel. Thus, we investigated whether CYP2C19 polymorphism or concomitant use of esomeprazole and clopidogrel is associated with on-treatment platelet reactivity and clinical outcomes in Japanese patients with CAD following stent implantation. Methods Study Population This study was conducted in accordance with the Declaration of Helsinki and was approved by the ethics committee of the institution, and written informed consent was obtained from each patient or the family of the subject. We enrolled 361 patients with CHD undergoing PCI from the PCI list in Kumamoto University Hospital. The exclusion criteria were: 1) written informed consent could not be obtained from the subject or their family; 2) already taking clopidogrel; 3) on anticoagulants, or antiplatelet agents other than clopidogrel or aspirin; and 4) already taking anticoagulants, antiplatelet agents, or PPI routinely before admission. All patients received DAPT with 100 mg/day aspirin and 75 mg/day clopidogrel as a maintenance dose, after a loading dose of clopidogrel at 300 mg, in this prospective single-center study with a 3-month follow-up. The patients were prospectively assigned to intensification of treatment with esomeprazole (10–20 mg/day, n = 50) or conventional treatment with regular gastric medicine (teprenone at 150 mg/day, or histamine H2-receptor antagonist: famotidine at 10–20 mg/day, n = 311). Gastric medicine including esomeprazole, teprenone, or famotidine was started on the same day as clopidogrel loading. The gastric medicine was primarily administered on the basis of a prior history of upper gastrointestinal ulcer or bleeding, or the presence of upper gastrointestinal symptoms, such as heartburn or epigastric pain, and prophylaxis for gastrointestinal damage. The final decision to administer esomeprazole or other gastric medicine was at the discretion of the attending physician. In this study, the occult blood test in the stool was routinely examined in all patients after hospitalization, and if the test was positive and the patient was hemodynamically stable with medical therapy, the patient was needed to take gastrointestinal endoscopy. Thus, high risks patients of gastrointestinal bleeding may be decreased due to these tests. We examined patients monthly after discharge by visiting as an outpatient or telephone interview. So, patients were checked on drug adherence monthly.

activity enzyme in the Japanese population [23]. CYP2C19 genotypes were therefore classified into three phenotypes: 1) extensive metabolizers (EM) carrying normal-function alleles (CYP2C19*1/*1); 2) intermediate metabolizers (IM) carrying one loss-of-function allele (*1/*2, *1/*3); and 3) poor metabolizers (PM) carrying two loss-offunction alleles (*2/*2, *2/*3, *3/*3). Measurement of Residual Platelet Reactivity Platelet function tests were performed 24 hours after administration of a loading dose of 300 mg of clopidogrel, followed by a 75 mg daily maintenance dose. Blood samples were collected using the doublesyringe technique, in which the first 2 to 4 ml of blood was discarded to avoid spontaneous platelet activation. The blood samples were drawn into Vacutainer tubes containing 0.5 ml of 3.2% sodium citrate (Becton-Dickinson, San Jose, CA) and processed within 60 minutes. As previously reported [15], aggregation in platelet-rich plasma induced by 20 μmol/L adenosine diphosphate (ADP; Chrono-Log) was measured using a light transmission aggregometer (MCM HEMA TRACER 313; PAM12C, LMS Inc., Japan). RPR was defined as the area under the platelet aggregation curve, which was used to express the aggregation response over the measured time (aggregation units · min, AU · min). In this study, platelet function tests were done at the start of medication, not at the interim or end of this study. Clinical Outcomes The study endpoints were residual platelet reactivity and cardiovascular death, nonfatal myocardial infarction, or ischemic stroke. Patients visited our outpatient department to receive a medical examination one month after discharge. Cardiovascular death was defined as death due to myocardial infarction, congestive heart failure, or documented sudden cardiac death. We used the universal definition of myocardial infarction in this study [24]. The diagnosis of stroke was made if a subject had clinical and radiological evidence of stroke. For subjects experiencing more than 2 acute events, only the first event was considered in the analysis. Moreover, we checked gastrointestinal bleeding at the outpatient department or by telephone interview every one month after PCI. Regarding gastrointestinal bleeding, we examined patients monthly after discharge by visiting as an outpatient or telephone interview. So, patients were checked on physical condition monthly. If findings of anemia, melena, or gastric symptoms such as heartburn or epigastric pain were observed, gastrointestinal endoscopy was performed. Statistical Analyses Continuous variables are expressed as mean ± SD. Categorical variables are expressed as frequencies and percentages. Statistical analysis was carried out exclusively by an independent statistician. The comparison between groups was analyzed using Pearson’s Chi-square test for categorical variables and unpaired t-test or Mann–Whitney U test for continuous variables. A p-value b 0.05 was regarded as significant. The SPSS 22.0 software (IBM Institute Inc., USA) was used for all statistical analyses. Results

Genotyping Patients’ Characteristics According to Esomeprazole Use Genomic DNA was extracted from whole blood using a DNA Extractor WB kit (Wako Pure Chemical Industries Ltd., Osaka, Japan) following a modified version of the protocol of Richards et al. [21]. Polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP) analyses for CYP2C19*2 (681G N A) and CYP2C19*3 (636G N A) were performed as described previously [22,23]. CYP2C19*2 and *3 are considered to account for N99% of alleles generating the null-

Clinical characteristics of subjects taking and not taking esomeprazole are shown in Table 1. There was no difference in baseline characteristics between the two groups, except for the prevalence of ACS. The prevalence of ACS was higher in esomeprazole group than in non-esomeprazole group (40.0 versus 26.4 %, P = 0.023). There were also no significant differences in the distribution of the CYP2C19


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Table 1 Baseline Characteristics between Esomeprazole and Non-Esomeprazole Groups.

Male (%) Age (years) Number of vessels treated Single (%) Multiple (%) ACS STEMI NSTEMI Current smoker (%) Hypertension (%) Dyslipidemia (%) Diabetes (%) Previous MI (%) eGFR (ml-min/1.73 m2) LVEF (%) Statin (%) Beta-blocker (%) ACEI/ARB (%) Ca-blocker (%) Gastroduodenal lesion (%) CYP2C19 EM (%) IM (%) PM (%) Esomeprazole 10 mg Esomeprazole 20 mg

Esomeprazole N = 50

Non-Esomeprazole N = 311

P-Value

36(72.0) 70.6 ± 10.4

229(73.6) 68.7 ± 9.9

0.808 0.202

17(34.0) 33(66.0) 21(40.0) 8(16.0) 4(8.0) 8(16.0) 41(82.0) 39(78.0) 28(56.0) 7(14.0) 57.4 ± 25.6 58.9 ± 11.6 48(96.0) 39(78.0) 35(70.0) 25(50.0) 5(10.0)

106(34.1) 205(65.9) 82(26.4) 32(10.3) 13(4.2) 56(18.0) 226(72.7) 231(74.3) 184(59.2) 56(18.0) 64.2 ± 25.9 59.2 ± 9.5 274(88.1) 250(80.4) 221(71.1) 182(58.5) 24(7.7)

0.991 0.991 0.023 0.232 0.237 0.730 0.163 0.574 0.673 0.488 0.088 0.855 0.095 0.695 0.878 0.258 0.581

18(36.0) 21(42.0) 11(22.0) 15(30.0) 35(70.0)

105(33.8) 154(49.5) 52(16.7) -

0.757 0.324 0.361

ACS = acute coronary syndromes; STEMI = ST-segment elevation myocardial infarction; NSTEMI = non-ST-segment elevation myocardial infarction; MI = myocardial infarction; eGFR = estimated glomerular filtration rate; EM = extensive metabolizers; IM = intermediate metabolizers; PM = poor metabolizers.

genotypes between the two groups (esomeprazole versus nonesomeprazole: EM, 36.7% versus 33.8%, P = 0.683; IM, 42.9% versus 49.5%, P = 0.386; PM, 20.4% versus 16.7%, P = 0.525, respectively). The prevalence of the CYP2C19 genotypes showed the same tendency as the ratio in our previous report (37, 44, and 19%, respectively) [15]. Platelet Reactivity According to Esomeprazole Use Fig. 1 shows RPR between esomeprazole and non-esomeprazole groups. There was no difference in RPR between both groups (4389 ± 1112 versus 4079 ± 1355 AU · min, P = 0.103). Table 2 and Fig. 2 show a comparison of RPR between esomeprazole and non-esomeprazole groups according to the CYP2C19 genotypes. In the analysis including all patients, RPR value was higher in IM than in EM (4089 ± 1252 versus 3697 ± 1215 AU · min P = 0.012) and, similarly, higher in PM than in IM (4884 ± 1027 versus 4089 ± 1252 AU · min, P b 0.001). Moreover, we performed a comparison of RPR between esomeprazole and non-esomeprazole groups according to the CYP2C19 genotypes (Table 3, Fig. 3). There were no significant differences in RPR between esomeprazole and non-esomeprazole groups in EM, IM, and PM (non-esomeprazole versus esomeprazole: EM, 3645 ± 1220 versus 3954 ± 1192 AU · min, P = 0.361; IM, 4051 ± 1271 versus 4401 ± 1063 AU · min, P = 0.293; PM, 4876 ± 1099 versus 4917 ± 669 AU · min, P = 0.907, respectively).

Fig. 1. Comparison of platelet reactivity between esomeprazole and nonesomeprazole groups. There was no significant difference in residual platelet reactivity between esomeprazole and non-esomeprazole groups (4389 ± 1112 versus 4079 ± 1355 AU · min, P = 0.103).

While, one patient had a gastrointestinal bleeding at one and half months after PCI in non-esomeprazole group, who was diagnosed to have gastric cancer by endoscopy (Table 4).

Discussion Wide variability in response is seen in patients administered clopidogrel [25,26], and reduced platelet inhibition by clopidogrel is associated with an increased risk of cardiovascular events [27,28]. The mechanism underlying interindividual variability in the response to clopidogrel has been linked, at least in part, to its metabolism dependent on cytochrome P450, including CYP2C19 and CYP3A4. Several domestic and overseas studies have demonstrated that patients carrying at least one non-functional allele of CYP2C19*2 show increased RPR despite conventional dual antiplatelet therapy, with a possible association with an increased risk of adverse cardiovascular events following coronary stent placement [13–17]. In contrast to people in Western countries, not only CYP2C19*2 but also *3 LOF alleles are commonly found in the Japanese population [29]. Therefore, it is meaningful to examine the interaction between PPI and clopidogrel on the basis of the CYP2C19 genotype, especially in Japanese patients. This is the first study to clarify the difference in RPR based on esomeprazole use and CYP2C19 polymorphism in CHD patients treated with stent implantation and taking dual antiplatelet agents with aspirin and clopidogrel. The important findings of the present study suggest that (1) esomeprazole did not affect RPR to clopidogrel, which is Table 2 Comparison of Platelet Reactivity in All Patients According to CYP2C19 Genotype.

Clinical Outcomes According to Esomeprazole Use Table 4 shows the details of cardiovascular events, including cardiovascular death, nonfatal myocardial infarction, and ischemic stroke, between esomeprazole and non-esomeprazole groups. There was no significant difference in the incidence of clinical outcomes between the two groups (non-esomeprazole versus esomeprazole: cardiovascular death, 0.31% versus 0%, P = 0.689; myocardial infarction, 0.62% versus 0%, P = 0.571; ischemic stroke, 0% versus 0%, P = NA, respectively).

CYP2C19 EM IM PM

All patients N = 361 (AU · min)

P-Value

3697 ± 1215 4089 ± 1252 4884 ± 1027

*0.012 ** b 0.001

Abbreviations as in Table 1. *0.012, IM versus EM; ** b 0.001, PM versus EM.

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Fig. 2. Comparison of platelet reactivity among the EM, IM, and PM groups. Residual platelet reactivity was higher in IM than in EM (4089 ± 1252 versus 3697 ± 1215 AU · min P = 0.012) and, similarly, higher in PM than in IM (4884 ± 1027 versus 4089 ± 1252 AU · min, P b 0.001). EM, extensive metabolizer; IM, intermediate metabolizer; PM, poor metabolizer.

independent of the CYP2C19 genotype, and (2) there was no increased risk of cardiovascular events regardless of esomeprazole use. We previously reported the safety of rabeprazole co-therapy in coronary stented patients during DAPT [16], and rabeprazole is mostly metabolized by non-enzymatic pathways and shows the least potent inhibition of CYP2C19 in vitro among the various PPI [18]. In the present study, RPR did not differ between esomeprazole and non-esomeprazole groups, which is consistent with previous reports from other countries [30–32]. An earlier report described increased RPR in patients with ACS receiving omeprazole with clopidogrel, presumably as a result of CYP2C19 competitive inhibition [11]. Moreover, co-therapy of omeprazole in coronary stented patients receiving DAPT was associated with adverse cardiovascular event risk in some studies [6,7]. Therefore, in Japan, there was hesitation about undertaking co-therapy of omeprazole and clopidogrel in clinical settings. Esomeprazole, which is an optical S-isomer of omeprazole and has been available since 2011 in Japan, also more effectively inhibits acid secretion and is less metabolized by CYP2C19 compared with omeprazole [20]. This involves markedly different features from omeprazole, and these results suggest that esomeprazole does not affect the antiplatelet efficacy of clopidogrel, probably due to less inhibition of the CYP2C19 enzyme. When all patients were stratified according to CYP2C19 genotype regardless of esomeprazole use, RPR varied in accordance with the number of CYP2C19 LOF alleles; in short, EM showed the most reduced RPR, IM was second, and PM showed the most increased RPR. The prevalence of the CYP2C19 genotypes differs among different ethnic groups: the prevalence of CYP2C19 PM is 2-3% in Caucasians, but it is 15-20% in Japanese [33]. CYP2C19 LOF allele carrier status and RPR levels are reported to be significant predictors of cardiovascular events in patients Table 3 Comparison of Platelet Reactivity between Esomeprazole and Non-esomeprazole Groups.

CYP2C19 EM IM PM

Esomeprazole N = 50 (AU · min)

Non-esomeprazole N = 311 (AU · min)

P-Value

3954 ± 1192 (18) 4401 ± 1063 (21) 4917 ± 669 (11)

3645 ± 1220 (105) 4051 ± 1271 (154) 4876 ± 1099 (52)

0.361 0.293 0.907

Abbreviations as in Table 1.

Fig. 3. Comparison of platelet reactivity between esomeprazole and non-esomeprazole groups according to CYP2C19 genotype. There were no significant differences in residual platelet reactivity between esomeprazole and non-esomeprazole groups in EM, IM, and PM (Non versus EPZ: EM, 3645 ± 1220 versus 3954 ± 1192 AU · min, P = 0.361; IM, 4051 ± 1271 versus 4401 ± 1063 AU · min, P = 0.293; PM, 4876 ± 1099 versus 4917 ± 669 AU · min, P = 0.907, respectively). Non, non-esomeprazole; EPZ, esomeprazole, EM, extensive metabolizer; IM, intermediate metabolizer; PM, poor metabolizer.

undergoing stent implantation on DAPT [34], although the cardiovascular event rate including thrombotic events is lower in Japan than in Western countries [35]. Therefore, the present study shows that esomeprazole co-therapy did not increase RPR on DAPT in different CYP2C19 genotypes, and that there is no increased risk of cardiovascular events regardless of CYP2C19 genotype, although some studies reported that high RPR is not always associated with future cardiovascular events [36–38]. When the patients were classified into esomeprazole use and nonuse groups in each of EM, IM, and PM, briefly, on the basis of CYP2C19 genotype, there were no differences in RPR between the two groups in each different CYP2C19 genotype. In the non-esomeprazole group alone, RPR was higher in PM than in EM or IM. Meanwhile, in the esomeprazole group alone, there was no significant difference in RPR between EM and IM because of the small number of enrolled patients receiving esomeprazole. Therefore, definitive conclusions cannot be drawn regarding the esomeprazole group. However, considering that a significant difference was detected between PM and EM, also in the esomeprazole group, the tendency is considered to be present as well as in the non-esomeprazole group. Anyway, these results demonstrate that co-administration of esomeprazole and clopidogrel does not affect the antiplatelet efficacy of clopidogrel irrespective of the CYP2C19 genotype. In this study, we compared the clinical outcome between esomeprazole and non-esomeprazole groups, albeit during a short one-month follow-up. The small sample size and low incidence of cardiovascular events might have affected the power to detect the influence on the clinical outcome; however, the above findings support co-therapy of esomeprazole during DAPT as a safe treatment. Gastrointestinal bleeding was seen in one patient of nonesomeprazole groups at one and half months after PCI. The small number of events may have been due to the short follow-up, and there is a possibility that stable CHD was included at a relatively high rate, and that the number of patients at high risk for gastrointestinal bleeding was small. In this study, follow-up period was 3 months. This period may not be enough to evaluate the risk of a PPI on clopidogrel. However, we firstly


difference in platelet function in patients with concomitant use of clopidogrel and esomeprazole [32].

Table 4 Clinical Outcomes in Esomeprazole and Non-esomeprazole Groups.

Cardiovascular death Myocardial infarction Stroke Bleeding Cardiovascular event Cardiovascular event + Bleeding

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Esomeprazole N = 50

Non-esomeprazole N = 311

P-Value

0(0.0) 0(0.0) 0(0.0) 0(0.0) 0(0.0) 0(0.0)

1(0.31) 2(0.62) 0(0.0) 1(0.31) 3(0.92) 4(1.29)

0.689 0.571 NA 0.689 0.487 0.420

Conclusions

aimed to evaluate the safety of concomitant use of clopidogrel and esomeprazole during DAPT, and we at least found that there was no increased risk of concomitant use of clopidogrel and esomeprazole. Furthermore, long-term interaction between clopidogrel and esomeprazole is unclear. Frelinger AL 3rd et al. reported that generation of clopidogrel active metabolite and inhibition of RPR were reduced in the co-administration of esomeprazole with clopidogrel compared with co-administration of lansoprazole with clopidogrel in healthy volunteers [39]. Moreover, US FDA labeling for several PPIs and clopidogrel suggests that PPI use among patients taking clopidogrel be limited to pantoprazole, rabeprazole, lansoprazole, or dexlansoprazole, and not recommend esomeprazole/omeprazole [40]. The dose of esomeprazole in the paper by Frelinger AL 3rd et al. was 40 mg, while we use 20 mg esomeprazole daily even maximum dose in Japan. Moreover, there is a racial difference in prevalence of CYP2C19 genotype, and pantoprazole is not approved in health insurance system in Japan, and is not available in clinical practice. So, we planed this study to examine the safety and efficacy of esomeprazole on Japanese patients during DAPT following stent implantation. Although there are several problems such as small sample size, short follow-up time, and low incidence of cardiovascular events, we firstly aimed to evaluate the safety of concomitant use of clopidogrel and esomeprazole during DAPT. The incidence of patients with high risk of gastrointestinal bleeding was low in this study, and we at least found that there was no increased risk of the concomitant use of clopidogrel and esomeprazole. We did not perform multivariate statistics analysis because of low incidence of cardiovascular event and no event in esomeprazole group.

Study Limitations In terms of limitations, first of all, the number of studied patients was small and the study may have been underpowered to detect a difference in RPR or clinical event rate. So, we cannot conclude that esomeprazole does not affect cardiovascular events. Especially, although there was not significant difference in RPR value in IM patients, we cannot exclude the possibility that if the number of patients increased, there would have been significant difference between non-esomeprazole and esomeprazole groups in IM patients. The number of patients between esomeprazole and non-esomeprazole groups was 50 and 311, respectively. This dissociation of the number might be due to low incidence of patients with high risk gastrointestinal bleeding, and due to dependence on the discretion of the attending physician. It is unknown about platelet reactivity before and after the initiation of DAPT, so it is not determined whether a higher RPR can be explained in terms of underlying platelet hyper-reactivity per se. PRP was measured only at the beginning of this study, and it is unknown whether or not RPR value by platelet function test in the early stage would continue in the esomeprazole treatment. In addition, we cannot rule out the effects of drugmetabolizing enzymes other than CYP2C19, such as CYP1A2, 2B6, 3A, and 2C9, on the clopidogrel response. There are racial and ethnical differences in the prevalence of CYP2C19 genotype and the results of this study may not necessarily be applicable to the other populations than Japanese. However, the recent study showed that there was no

It is possible that concomitant use of esomeprazole with clopidogrel might not be associated with reduced antiplatelet efficacy of clopidogrel and increased risk of cardiovascular events, irrespective of CYP2C19 genotype. Conflicts of Interest Statement None of the authors received any fees, honoraria, grants, or consultancies that would constitute a conflict of interest in this study. Funding This work was supported in part by grants-in-aid for scientific research from the Japanese Ministry of Education, Culture, Sports, Science and Technology. Acknowledgements The authors would like to thank medical technologist S. Iwashita of Kumamoto University Hospital for performing measurements of platelet reactivity, and medical secretaries, K. Watanabe, Y. Maeda, C. Yamamoto, R. Usui, and A. Kikuchi, Kumamoto University, for collecting the data. References [1] O'Gara PT, Kushner FG, Ascheim DD, Casey Jr DE, Chung MK, de Lemos JA, et al. ACCF/AHA guideline for the management of ST-elevation myocardial infarction: a report of the American College of Cardiology Foundation/American Heart Association Task Force on Practice Guidelines. J Am Coll Cardiol 2013;61:e78-140. [2] Jneid H, Anderson JL, Wright RS, Adams CD, Bridges CR, Casey Jr DE, et al. ACCF/AHA focused update of the guideline for the management of patients with unstable angina/non-ST-elevation myocardial infarction (updating the 2007 guideline and replacing the 2011 focused update): a report of the American College of Cardiology Foundation/American Heart Association Task Force on Practice Guidelines. J Am Coll Cardiol 2012;60:645–81. [3] Levine GN, Bates ER, Blankenship JC, Bailey SR, Bittl JA, Cercek B, et al. ACCF/AHA/ SCAI Guideline for Percutaneous Coronary Intervention. A report of the American College of Cardiology Foundation/American Heart Association Task Force on Practice Guidelines and the Society for Cardiovascular Angiography and Interventions. J Am Coll Cardiol 2011;58:e44-122. [4] Nikolsky E, Stone GW, Kirtane AJ, Dangas GD, Lansky AJ, McLaurin B, et al. Gastrointestinal bleeding in patients with acute coronary syndromes: incidence, predictors, and clinical implications: analysis from the ACUITY (Acute Catheterization and Urgent Intervention Triage Strategy) trial. J Am Coll Cardiol 2009;54:1293–302. [5] Abraham NS, Hlatky MA, Antman EM, Bhatt DL, Bjorkman DJ, Clark CB, et al. ACCF/ ACG/AHA 2010 expert consensus document on the concomitant use of proton pump inhibitors and thienopyridines: a focused update of the ACCF/ACG/AHA 2008 expert consensus document on reducing the gastrointestinal risks of antiplatelet therapy and NSAID use: a report of the American College of Cardiology Foundation Task Force on Expert Consensus Documents. Circulation 2010;122:2619–33. [6] Ho PM, Maddox TM, Wang L, Fihn SD, Jesse RL, Peterson ED, et al. Risk of adverse outcomes associated with concomitant use of clopidogrel and proton pump inhibitors following acute coronary syndrome. JAMA 2009;301:937–44. [7] Juurlink DN, Gomes T, Ko DT, Szmitko PE, Austin PC, Tu JV, et al. A population-based study of the drug interaction between proton pump inhibitors and clopidogrel. CMAJ 2009;180:713–8. [8] Huang CC, Chen YC, Leu HB, Chen TJ, Lin SJ, Chan WL, et al. Risk of adverse outcomes in Taiwan associated with concomitant use of clopidogrel and proton pump inhibitors in patients who received percutaneous coronary intervention. Am J Cardiol 2010;105:1705–9. [9] Stockl KM, Le L, Zakharyan A, Harada AS, Solow BK, Addiego JE, et al. Risk of rehospitalization for patients using clopidogrel with a proton pump inhibitor. Arch Intern Med 2010;170:704–10. [10] Ray WA, Murray KT, Griffin MR, Chung CP, Smalley WE, Hall K, et al. Outcomes with concurrent use of clopidogrel and proton-pump inhibitors: a cohort study. Ann Intern Med 2010;152:337–45. [11] Gilard M, Arnaud B, Cornily JC, Le Gal G, Lacut K, Le Calvez G, et al. Influence of omeprazole on the antiplatelet action of clopidogrel associated with aspirin: the randomized, double-blind OCLA (Omeprazole CLopidogrel Aspirin) study. J Am Coll Cardiol 2008;51:256–60.

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