Combination Antithrombotic Therapies

Abstract

HomeCirculationVol. 121, No. 4Combination Antithrombotic Therapies Free AccessResearch ArticlePDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessResearch ArticlePDF/EPUBCombination Antithrombotic Therapies Paul A. Gurbel and Udaya S. Tantry Paul A. GurbelPaul A. Gurbel From the Sinai Center for Thrombosis Research, Baltimore, Md. Search for more papers by this author and Udaya S. TantryUdaya S. Tantry From the Sinai Center for Thrombosis Research, Baltimore, Md. Search for more papers by this author Originally published2 Feb 2010https://doi.org/10.1161/CIRCULATIONAHA.109.853085Circulation. 2010;121:569–583Atherothrombosis is the major pathophysiological process responsible for the occurrence of severe ischemic events in patients with cardiovascular diseases. In the United States, atherothrombosis strongly influenced mortality in 2004: One in 2.8 deaths was due to CVD, 1 in 5 deaths to coronary heart disease, and 1 in 17 deaths to stroke.1 Because cardiovascular disease is a progressive and systemic disease, long-term antithrombotic therapies that effectively target the entire arterial vasculature and modulate the key components responsible for thrombus generation are essential to improve patient outcomes. Because platelet activation is determined by multiple receptor-mediated signaling pathways, clinical studies have evaluated the efficacy of multidrug administration in the prevention of atherothrombotic complications.2,3 The major concern with these therapies is the critical balance between anti-ischemic effect and bleeding risk. This review summarizes our understanding of the role of combination antiplatelet therapies in the treatment and prevention of atherothrombosis.Pathophysiology of AtherothrombosisPlatelet activation and aggregation play a pivotal role in the generation of occlusive thrombus at the site of coronary arterial plaque rupture. In addition, platelets influence various endothelial and inflammatory responses during the initiation and progression of atherosclerosis. Under normal conditions, anucleate circulating platelets are in a quiescent state. Healthy vascular endothelium prevents adhesion and activation of platelets by producing antithrombotic factors such as CD39 (ectoADPase), prostaglandin I2, nitric oxide, heparin, matrix metalloproteinase-9, protein S, and thrombomodulin.3,4 Endothelial activation and denudation and frank atherosclerotic plaque rupture expose the subendothelial matrix and release prothrombotic factors during acute coronary syndromes (ACS) and percutaneous interventions. These processes result in localized platelet adhesion and platelet activation. After adhesion to the exposed subendothelial matrix, platelets are activated by shear and the soluble agonists thromboxane A2 (TxA2), ADP, and thrombin. TxA2 is produced from arachidonic acid, which originates from membrane phospholipids and binds to Tx receptors; ADP is secreted from dense granules and binds to P2Y12 and P2Y1 receptors. These 2 secondary agonists, through an autocrine and paracrine fashion, produce sustained activation of glycoprotein IIb/IIIa receptors, leading to stable platelet-rich thrombus generation. The ADP-P2Y12 interaction contributes most to ADP-induced aggregation measured by conventional aggregometry.5 Platelet activation also results in the membrane exposure of phosphatidyl serine, providing binding sites for coagulation factors. The coagulation process results in the generation of thrombin and subsequent platelet-fibrin clot formation.3 Endogenous phosphodiesterase (PDE) activity that affects intraplatelet cAMP levels also modulates platelet function (the Figure). Finally, isoprostanes derived from membrane arachidonic acid through peroxidation have been shown to induce platelet aggregation by activating the receptor for TxA2.6 Inhibition of PDE and cyclooxygenase (COX) may play a more important role in the treatment of peripheral and cerebrovascular disease, whereas antiplatelet therapy with clopidogrel and aspirin is the cornerstone of ACS and poststent treatment.7 Finally, the relative contribution of each pathway (ADP-platelet, TxA2-platelet, thrombin-platelet, coagulation, and PDE activity) to the development of thrombus formation in the individual patient is unknown. Therefore, despite immense research, determination of the optimal antiplatelet and antithrombotic therapy remains an elusive goal. The side effects observed during contemporary antithrombotic therapy may be related in part to the uniform dosing regimens used that ignore the inherent variability in the antiplatelet and antithrombotic response. Download figureDownload PowerPointFigure. Mechanism of action of oral antiplatelet drugs. Ticlopidine, clopidogrel, and prasugrel irreversibly block the P2Y12 receptor, whereas ticagrelor and elinogrel irreversibly block the P2Y12 receptor. Aspirin irreversibly inhibits the COX-1 enzyme and thereby attenuates TxA2 production and TxA2-medicated platelet response. SCH530348 and E5555 block the PAR-1 receptor, thereby inhibiting thrombin-mediated platelet response. PDE inhibitors such as cilostazol and dipyridamole stimulate synthesis of prostaglandin I2 (PGI2) and nitric oxide (NO) in endothelial cells and inhibit the active transport of adenosine into cells, particularly red blood cells. By stimulating adenylyl/guanylyl cyclase (AC/GC) activity in platelets, PGI2, NO, and adenosine all increase intracellular cAMP levels and thus attenuate platelet aggregation. AA indicates arachidonic acid; PI3K, phosphatidylinositol 3 kinase; ATP/GTP, adenosine triphosphate/guanosine triphosphate; VASP-P, vasodilator-stimulated phosphoprotein phosphorylation; GP, glycoprotein; PKA, phosphokinase A; and A2a receptor, adenosine 2a receptor.AspirinAspirin is the primary component of combination antiplatelet therapy. The antiplatelet effect of aspirin is attributed primarily to the irreversible inhibition of platelet COX-1 by acetylation of serine residue 529, resulting in a downstream reduction in the synthesis of TxA2 and consequent TxA2-induced platelet activation/aggregation. However, recent studies indicate that the antiplatelet effect of aspirin may also involve inhibition of pathways distinct from COX-1 (non–COX-1 pathways). In addition, aspirin is known to reduce thrombin generation, to enhance fibrin clot permeability and clot lysis, and to promote nitric oxide production in platelets. Aspirin also has antiinflammatory properties that may enhance its antithrombotic effect.8Recent pharmacodynamic studies have demonstrated that low-dose aspirin has variable effects in inhibiting platelet non–COX-1 pathways. The term “aspirin nonresponsiveness or resistant” has been used to describe selected patients, especially high-risk patients and diabetics, exhibiting high platelet function as measured by assays that are nonspecific in their assessment of COX-1 activity.9,10 Ischemic event recurrence is also higher in patients found to be aspirin resistant by COX-1–nonspecific assays.11,12 The dose dependence of aspirin in the inhibition of non–COX-1 pathways suggests that selected patients require a dose >81 mg to achieve an optimal antiplatelet effect. However, this observation conflicts with the results of meta-analyses demonstrating no conclusive clinical benefits of high-dose aspirin therapy.12,13Aspirin Dose and BleedingIn a recent meta-analysis of randomized controlled trials of low-dose aspirin (75 to 325 mg/d), aspirin treatment was associated with a 0.13% absolute increase in any major bleeding and a 0.12% increase in major gastrointestinal bleeding compared with placebo in cardiovascular disease patients. There was an increased relative risk (RR) of major bleeding (RR, 1.71; 95% confidence interval [CI], 1.41 to 2.08), major gastrointestinal bleeding (RR, 2.07; 95% CI, 1.61 to 2.66), and intracranial bleeding (RR, 1.65; 95% CI, 1.06 to 5.99). No difference in bleeding was observed in patients treated with 75 to 162.5 mg/d versus >162.5 to 325 mg/d.14ThienopyridinesBecause ADP plays a critical role in the amplification of platelet aggregation and the genesis of a stable occlusive thrombus, the inhibition of ADP-induced platelet activation was an early focus of alternative antiplatelet treatment. Moreover, drug intolerance, gastrointestinal bleeding, and treatment failure associated with aspirin therapy also stimulated the development of new antiplatelet agents. Thienopyridines are prodrugs that require metabolic activation by the cytochrome P450 pathway. The active metabolite forms a disulfide bond with the P2Y12 receptor, rendering the receptor unable to bind ADP and thus attenuating the subsequent response of the platelet to ADP.Aspirin Monotherapy Versus Thienopyridine MonotherapyTiclopidine was the first thienopyridine to be widely used clinically. In the Ticlopidine Aspirin Stroke Study (TASS), ticlopidine treatment (500 mg daily) was superior to aspirin (1300 mg QD) in reducing nonfatal stroke or death at 3 years (17% versus 19%; RR reduction [RRR], 12%; P=0.048) in patients with previous stroke.15 In the Ticlopidine Versus Aspirin After Myocardial Infarction (STAMI) trial, 1470 patients with acute myocardial infarction (MI) treated with thrombolysis received either aspirin (160 mg QD) or ticlopidine (500 mg QD) therapy. In that trial, there was no difference between the 2 treatment arms in primary combined end point of death, recurrent MI, stroke, or angina.16 In the Clopidogrel Versus Aspirin in Patients at Risk of Ischemic Events (CAPRIE) trial, aspirin (325 mg QD) was compared with clopidogrel therapy (75 mg/d) in patients at risk for ischemic events, and an RRR of 8.7% was observed in clopidogrel-treated patients in the composite end point of MI, ischemic stroke, or vascular death17 (the Table). Most of the benefit was observed in patients with peripheral arterial disease. Although the overall safety profile was similar between the 2 groups, the aspirin-treated group had a greater rate of gastrointestinal bleeding (2.66% versus 1.99; RR, 1.34; 95% CI, 1.11 to 1.61; P<0.05).17 Subgroup analysis revealed that clopidogrel reduced the risk of acute MI (≈19%) more significantly than aspirin in both low- and high-risk patients.18 On the basis of the favorable results of the CAPRIE trial, the US Food and Drug Administration (FDA) approved the use of a 75-mg daily dose of clopidogrel for patients with a history of recent heart attack, recent stroke, or established peripheral arterial disease. Table. Important Clinical Trials Evaluating Clinical Efficacy of Clopidogrel and Aspirin TherapyTrialPatients (n)ProtocolEnd PointsOutcomeSafety OutcomeCLP indicates clopidogrel; ASA, aspirin; IS, ischemic stroke; GI, gastrointestinal; LD, loading dose; CV, cardiovascular; CAD, coronary artery disease; CVD, CV disease; PAD, peripheral vascular disease; US, unstable angina; and DM, diabetes mellitus.CAPRIE17Symptomatic atherosclerotic disease (19 185)75 mg CLP vs 325 mg ASA for median of 1.9 yFirst occurrence of IS, MI, or vascular deathRRR=8.7%; 95% CI=0.3–16.5; P=0.043No significant difference in rate of a bleeding disorder (9.28% vs 9.27%); ASA recipients had a greater rate of GI bleeding (2.66% vs 1.9%; P<0.05)CURE27ACS (12 562)300 mg LD+75 mg/d CLP-vs placebo+75–325 mg ASA for 3 to 12 moCV death, nonfatal MI, or strokeRR=0.8; 95% CI=0.72–0.90; P<0.001Significant increase in major bleeding (RR=1.38; 95% CI=1.13–1.67; P=0.0001) and minor bleeding (RR=2.12; 95% CI=1.75–2.56; P<0.001)PCI-CURE28PCI (2658)CLP vs placebo+ASACV death, nonfatal MI, or stroke4.5% vs 6.5%; RRR=30%; P=0.003CREDO29Symptomatic CAD patients (n=2116) undergoing PCI with and without stenting300 mg CLP vs placebo; 75 mg/d CLP+325/d ASA for 28 d; 75 mg CLP or placebo+81–325 mg ASA for 12 mo28-d CV death, MI, and urgent vessel revascularization; 1-y CV death, MI, and stroke; safety end point: modified TIMI major, minor, or insignificant28-d outcome (RRR=18.5%; 95% CI=−4.2–41.8; P=0.23); patients with CLP LD >6 h before PCI (RRR=38.6%; 95% CI=−1.6–62.9; P=0.051); 1-year outcome (RRR=26.9%; 95% CI=3.9–44; P=0.02)A trend toward increased major bleeding with CLP (8.8% vs 6.7%; P=0.07)CLARITY-TIMI 2830Patients within 12 h of STEMI receiving lytic therapy (3491)300 mg LD+75 mg MD CLP or placebo+ASA+ fibrinolyitc+heparinOccluded infarct artery, death, or recurrent MI (before angiography)RRR=36%; 95% CI=0.53–0.76; P<0.001TIMI-defined major bleeding and intracranial hemorrhage and TIMI minor bleedings were not statistically differentPCI-CLARITY31Patients treated with PCI (1863)Same as above30-d CV death, recurrent MI, or recurrent ischemia leading to urgent revascularizationRR=0.80; 96% CI=0.65-0.97; P=0.03.COMMIT32STEMI patients within 24 h of symptom onset (45 852)72 mg/d CLP+162 mg/d ASA+metoprolol vs 162 mg/d ASA+metoprolol for 4 wkDeath, infarction, or strokeOR=0.91; 95% CI=0.87–0.99; P=0.03No significant difference in composite of all transfused, fatal, or cerebral bleedings (0.58% vs 0.555%; P=0.59), but CLP increased minor bleeding (3.6% vs 3.1%; P=0.005)CHARISMA34Documented CAD, CVD, or PAD or ≥3 atherothrombotic risk factors (15 603)75–162 mg ASA+75 mg CLP vs 75–162 mg/d ASA+placebo–media for 28 moPrimary end point of first occurrence of MI, IS, or vascular death; secondary end point of hospitalization for UA, TIA, or revascularization procedurePrimary end point: RR=0.93; 95% CI=0.83–1.05; P=0.22; secondary end point: RR=0.92; 95% CI=0.85–0.995; P=0.004Increase in severe bleeding according GUSTO definition (RR=1.25; 95% CI=0.97–1.61; P=0.09) and rate moderate bleeding (RR=1.62; 95% CI=1.27–2.08; P<0.001)Posthoc analysis of CHARISMA35Patients with documented prior MI, IS, or symptomatic PAD75–162 mg ASA+75 mg CLP vs 75–162 mg/d ASA+placeboSame as aboveRRR=17%; 95% CI=4–28; P=0.01Nonsignificant increase in severe bleeding (RR=1.11; 95% CI=0.81–1.57; P=0.509)MATCH37Recent history of IS and TIA and previous MI, angina, DM, or symptomatic PAD (7599)75 mg/d CLP vs 75 mg/d CLP+75 mg/d ASA for 18 moIS, MI, vascular death, or rehospitalization for acute ischemic eventRRR=6.5%; 95% CI=−4.6–16.3; P=0.244More common major (2% vs 1%) and life-threatening (3% vs 1%) bleedings with CLP+ASAThienopyridine Plus Aspirin: StentingGiven the important contributions of P2Y12 and COX-1 to the amplification of platelet aggregation mediated by ADP and TxA2, respectively, it was hypothesized that simultaneous inhibition of both pathways would provide a superior antithrombotic effect compared with single pathway inhibition. Nonhuman primate investigations using exteriorized arteriovenous shunts treated with metallic endovascular stents demonstrated the enhanced antithrombotic effect of clopidogrel with the addition of aspirin.19 Subsequent investigations in human volunteers treated with aspirin and clopidogrel similarly demonstrated the synergistic effect of dual antiplatelet therapy compared with aspirin monotherapy in a perfusion chamber assay measuring ex vivo thrombus formation.20 However, the results of large-scale clinical trials of dual antiplatelet therapy confirmed what was expected from the laboratory observations and changed the treatment paradigm for many cardiovascular disease states. In these trials, thienopyridines added to aspirin therapy demonstrated the potent effect in the inhibition of ischemic events across the spectrum of cardiovascular disease settings.2In the Stent Anticoagulation Restenosis Study (STARS), a significant decrease in the combined end point of death, MI, acute MI, angiographically evident thrombosis, and revascularization of the target vessel within 30 days was observed in patients undergoing stenting randomized to aspirin plus ticlopidine therapy (0.55%) compared with aspirin therapy alone (3.6%) and aspirin plus warfarin therapy (2.7%; P<0.001 for the comparison of all groups).21 This important trial led the way for dual antiplatelet therapy to become the standard of care in patients undergoing coronary arterial stenting. Other studies of ticlopidine plus aspirin versus aspirin plus anticoagulation demonstrated concordant effects.22–24 All of these studies influenced a change in the antithrombotic treatment paradigm, and dual antiplatelet therapy became the standard of care for the poststent patient. However, unfavorable side effects (neutropenia, bone marrow aplasia, and thrombotic thrombocytopenic purpura) were associated with ticlopidine treatment, which led to the development of the second-generation thienopyridine, clopidogrel.25In the Clopidogrel Aspirin Stent Interventional Cooperative Study (CLASSICS), the safety of clopidogrel was compared with that of ticlopidine after successful coronary stenting. Treatment with clopidogrel 75 mg QD with or without a loading dose was associated with fewer major bleeding complications, less neutropenia, less thrombocytopenia, and early discontinuation of study drug because of noncardiac adverse events (4.6% versus 9.1%; P=0.005).26 Similar recurrent ischemic event rates were observed between groups. The results of CLASSICS influenced the nearly uniform use of clopidogrel as the thienopyridine of choice that continues today.Clopidogrel Plus Aspirin and Versus Aspirin MonotherapyAcute Coronary SyndromesThe major clinical trials evaluating the efficacy and safety of dual antiplatelet with clopidogrel and aspirin versus aspirin monotherapy in ACS are summarized in the Table. The first large-scale trial was the Clopidogrel in Unstable Angina to Prevent Recurrent Events (CURE) study. Dual antiplatelet therapy of clopidogrel (300-mg loading dose followed by 75 mg/d) and aspirin (75 to 325 mg/d) in patients with unstable angina and non–ST-elevation acute MI (NSTEMI) was associated with a superior reduction in 9-month cardiovascular mortality, nonfatal MI, and stroke across a broad range of risk groups.27 In a subset analysis of the CURE study (PCI-CURE), there was a reduction in the risk of MI before percutaneous coronary intervention (PCI) and cardiovascular death or MI 4 weeks after PCI in patients receiving clopidogrel and aspirin pretreatment for up to 10 days and continuing on long-term treatment.28 The results of the CURE trial strongly influenced cardiologists to adopt the strategy of a 300-mg clopidogrel loading dose during PCI in all patients as the standard of care. This strategy was approved by the FDA for the treatment of patients with ACS. In the Clopidogrel for the Reduction of Events During Observation (CREDO) trial, there was a significantly reduced risk of adverse ischemic events with long-term clopidogrel therapy. However, substantial benefits of a prestent clopidogrel loading dose (300 mg) were seen only when administered >6 hours before PCI.29The Clopidogrel as Adjunctive Reperfusion Therapy–Thrombolysis in Myocardial Infarction (CLARITY-TIMI) 28 study investigated the efficacy of a 300-mg clopidogrel load/75 mg QD administered to patients within 12 hours of onset of an STEMI treated with fibrinolytic therapy. Clopidogrel therapy was associated with a 20% reduction (P=0.03) in the composite end point of cardiovascular death, reinfarction, or recurrent ischemia requiring urgent revascularization at 30 days.30 The PCI-CLARITY study examined 57% of the patients from CLARITY-TIMI 28 who underwent PCI and showed that clopidogrel pretreatment was also associated with a 46% reduction in the odds of cardiovascular death, recurrent MI, or stroke within 30 days with no significant increase in the incidence of bleeding complications. This benefit was observed regardless of glycoprotein IIb/IIIa inhibitor treatment or a loading dose of open-label clopidogrel at the time of PCI.30 It is also interesting that patients who were pretreated with a daily dose of 75 mg clopidogrel and received an additional loading dose of 300 mg at the time of PCI had the maximum protection against death, reinfarction, or stroke.31,32In the Clopidogrel and Metoprolol in Myocardial Infarction Trial (COMMIT) trial of acute STEMI patients, adjunctive clopidogrel therapy was associated with a modest but significant RRR in the primary end point of death, reinfarction, or stroke at hospital discharge and was not associated with an increased risk of major bleeding. The benefit of clopidogrel therapy was observed within the first 24 hours.33Primary and Secondary PreventionIn the Clopidogrel for High Atherothrombotic Risk and Ischemic Stabilization, Management, and Avoidance (CHARISMA) trial, the efficacy and safety of long-term treatment with clopidogrel (75 mg QD) in addition to aspirin (75 to 162 mg QD) were compared with aspirin monotherapy (75 to 162 mg QD) in patients at high risk for cardiovascular events. Overall, there was a nonsignificant decrease in the primary end point of first occurrence of MI, stroke, or cardiovascular death, and a significant decrease in the secondary end point of hospitalization for unstable angina, transient ischemic attack (TIA), or a revascularization procedure (RR, 0.92; 95% CI, 0.85 to 0.995; P=0.04) in patients treated with dual antiplatelet therapy compared with aspirin monotherapy. At the same time, dual antiplatelet therapy was associated with a marginal increase in the primary safety end point of severe bleeding and a significant increase in the rate of moderate bleeding compared with aspirin monotherapy according to the Global Utilization of Streptokinase and tPa for Occluded Arteries (GUSTO) definition. In the predefined subgroup of patients with clinically evident atherothrombosis (symptomatic group), there was a statistically significant decrease in the primary end point observed with dual antiplatelet therapy compared with aspirin monotherapy but not in a subgroup of patients with multiple risk factors (asymptomatic group). In that analysis, moderate bleeding was increased nonsignificantly in asymptomatic patients but significantly in symptomatic patients (P<0.001).34 In another subgroup analysis of patients with documented prior MI, ischemic stroke, or symptomatic peripheral arterial disease (CAPRIE-like cohort), there was a significant reduction in the primary end point of cardiovascular death, MI, or stroke (hazard ratio [HR], 0.83; 95% CI, 0.72 to 0.96; P=0.01) associated with long-term clopidogrel plus aspirin treatment. Although there was no significant difference in the rate of severe bleeding, moderate bleeding was significantly increased with clopidogrel plus aspirin treatment (HR, 1.60; 95% CI, 1.16 to 2.20; P=0.0004).35Finally, a meta-analysis of 5 randomized trials (CURE, CREDO, CLARITY, COMMIT, and CHARISMA) in 79 624 patients demonstrated that the incidence of all-cause mortality was 6.3% in the aspirin plus clopidogrel group versus 6.7% in the aspirin group (odds ratio [OR], 0.94; 95% CI, 0.89 to 0.99; P=0.026). The incidence of MI was 2.7% and 3.3% (OR, 0.82; 95% CI, 0.75 to 0.89; P<0.0001) and of stroke was 1.2% and 1.4% (OR, 0.82; 95% CI, 0.73 to 0.93; P=0.002) in the clopidogrel plus aspirin versus aspirin group, respectively. Similarly, the incidence of major bleeding was 1.6% and 1.3% (OR, 1.26; 95% CI, 1.11 to 1.41; P<0.0001) and of fatal bleeding was 0.28% and 0.27% (OR, 1.04; 95% CI, 0.76 to 1.43; P=0.79) in the aspirin plus clopidogrel versus aspirin group alone, respectively.36Clopidogrel Plus Aspirin Versus Clopidogrel Monotherapy: TIA and StrokeIn the Management of Atherothrombosis With Clopidogrel in High-Risk Patients (MATCH) trial, there was no additional value of adding aspirin to clopidogrel in patients with a recent history of TIA or ischemic stroke who had >1 additional cardiovascular risk factor (ie, history of MI, angina, peripheral arterial disease, or diabetes mellitus; RRR, 6.4%; 95% CI, 4.6 to 16.3; P=0.24). However, significantly higher life-threatening bleeding (3% versus 1%; P<0.0001), specifically intracranial hemorrhage, was observed with clopidogrel therapy.37Thus, a clear net benefit of dual antiplatelet therapy was observed in a wide range of patients with ACS and in patients undergoing coronary stenting as a secondary prevention strategy. However, current data do not support its use in the primary prevention or treatment of cerebrovascular disease.Limitations of Clopidogrel Plus AspirinEarly pharmacodynamic studies evaluating the antiplatelet response during clopidogrel and aspirin therapy, especially in patients undergoing stenting treated with a 300-mg loading dose and a 75-mg maintenance dose of clopidogrel, revealed various limitations: a delayed antiplatelet response and overall modest degree of platelet inhibition (≈30% to 50%), normally distributed response variability, nonresponsiveness in a substantial percentage of patients, and irreversible platelet inhibition and interindividual variability in the recovery of platelet function that may affect the outcomes of patients needing urgent surgery with exposure to an unpredictable risk of bleeding and ischemia. The prevalence of clopidogrel resistance varies from ≈8% to 30% and is dependent on dose and time of measurement in relation to dosing.38 In addition, various methods, including point-of-care methods reflecting P2Y12 reactivity, have been used to measure clopidogrel responsiveness. Although a good correlation has been demonstrated among these methods, measurement of vasodilator-stimulated phosphoprotein phosphorylation assay by flow cytometry has been considered by some investigators to be the most specific method to indicate P2Y12 reactivity.38,39 More recently described limitations include potentially important drug-drug interactions, interactions with cigarette smoking, and the influence of CYP450 genotype. Numerous reports associate the occurrence of post-PCI ischemic events, including stent thrombosis, with clopidogrel nonresponsiveness indicated by high on-treatment platelet reactivity.40Clopidogrel is metabolized in a 2-step process. The thienopyridine ring of clopidogrel is oxidized to 2-oxo-clopidogrel, which is subsequently hydrolyzed to a highly labile active metabolite, R-130964.41 It has been reported that CYP2C19, CYP1A2, and CYP2B6 participate the first step, whereas CYP2C19, CYP2C9, CYP2B6, and CYP3A participate in the second step.42In the Intracoronary Stenting and Antithrombotic Regimen: Choose Between 3 High Oral Doses for Immediate Clopidogrel Effect (ISAR-CHOICE) study, the effects of higher loading doses were investigated. An increase in the clopidogrel loading dose from 300 to 600 mg increased platelet inhibition. However, 900 mg did not result in further suppression of ADP-induced platelet aggregation and was associated with a reduced increase in plasma concentration of the active metabolite. These data suggest that intestinal absorption limited the amount of the 900-mg load accessible to the liver for conversion to the active metabolite.43 Similarly, only a moderate and nonsignificant increase in antiplatelet effects was observed after a 900-mg clopidogrel loading dose compared with 600 mg in the Assessment of the Best Loading Dose of Clopidogrel to Blunt Platelet Activation, Inflammation and Ongoing Necrosis (ALBION) study.44Multiple lines of evidence strongly suggest that insufficient active metabolite generation is the primary explanation for clopidogrel response variability and nonresponsiveness.45 Variable levels of active metabolite generation after clopidogrel administration have been attributed to variability in intestinal absorption influenced by polymorphism of an ABCB1 gene that encodes p-glycoprotein. Moreover, the ABCB1 gene variant has been identified with decreased clopidogrel active metabolite generation and a 70% relative increased risk of 1-year cardiovascular events.46,47 Functional variability in P450 isoenzyme activity has also been associated with variable levels of active metabolite generation. In earlier studies, coadministration of lipophilic statins (which compete with clopidogrel for CYP3A4) such as atorvastatin or simvastatin has been shown to attenuate the antiplatelet effect of clopidogrel in pharmacodynamic studies.48 However, this interaction has not been proven to affect clinical outcomes significantly in patients treated with dual antiplatelet therapy. However, a trend toward a better clinical outcome in patients coadministered clopidogrel and non–CYP3A4-metabolized statins compared with CYP3A4-metabolized statins has been demonstrated in some retrospective analyses.49Similar to statins, various proton pump inhibitors (PPIs), frequently used with clopidogrel and aspirin, are also metabolized by CYP450 isoenzymes. In a randomized controlled study involving patients undergoing stenting, coadministration of omeprazole with aspirin and clopidogrel significantly reduced the antiplatelet effect of clopidogrel as measured by the vasodilator-stimulated phosphoprotein phosphorylation assay.50 Coadministration of clopidogrel with PPIs, especially CYP2C19-metabolized PPIs such as omeprazole, lansoprazole, and rabeprazol, has been shown to be associated with reduced clinical efficacy in some but not all retrospective analyses.51,52 At this time, there is no conclusive evidence that PPIs influence clopidogrel metabolism and attenuate the clinical benefits of clopidogrel treatment. However, it has been recommended by some that PPIs be used in patients administered clopidogrel only when there are solid clinical indications.Cigarette smoking has been reported to induce CYP1A2 activity, thereby potentially affecting clopidogrel metabolism. In the CREDO trial, a larger reduction in clinical events occurred in patients receiving clopidogrel who were smokers compared with nonsmokers receiving clopidogrel treatment.53 A recent retrospective analysis of patients undergoing elective stenting treated with clopidogrel and aspirin reported that current smokers had greater platelet inhibition