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Best Practice & Research Clinical Endocrinology & Metabolism 23 (2009) 375–388

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Aspirin and clopidogrel: efficacy and resistance in diabetesmellitus

Dominick J. Angiolillo, MD, PhD, FACC, FESC, FSCAI *, Siva Suryadevara, MDDivision of Cardiology, University of Florida College of Medicine – Jacksonville, 655 West 8th Street, Jacksonville, FL 32209, USA

Keywords:plateletaspirinclopidogreldiabetes mellitusresistance

* Corresponding author. Tel.: þ1 904 244 3933,E-mail address: [email protected]

1521-690X/$ – see front matter � 2008 Elsevier Ldoi:10.1016/j.beem.2008.12.001

Diabetes mellitus patients are characterized by enhanced plateletreactivity which exposes them to an increased risk of athero-thrombotic events in the setting of acute coronary syndromes orpercutaneous coronary interventions. Although aspirin and clopi-dogrel, used either solely or in combination, are associated withimproved clinical outcomes in high-risk patients, diabeticspatients treated with antiplatelet agents remain at higher risk ofrecurrent ischemic events. Recent laboratory findings suggest thatthis observation may be related to a reduced responsiveness or‘resistance’ to these agents. In this chapter the efficacy of currentlyavailable oral antiplatelet agents in preventing ischemic events isreviewed. In addition, the antiplatelet ‘resistance’ phenomenon inthe diabetic population and its impact on clinical outcomes issummarized. Finally, future developments in the field directedtowards individualized treatment strategies and novel antiplateletagents are examined.

� 2008 Elsevier Ltd. All rights reserved.

Diabetes mellitus is a global pandemic which currently affects more than 150 million peopleworldwide.1 This figure is expected to double over the next 20 years, almost exclusively due to anincrease in the prevalence of type-2 diabetes mellitus (T2DM).1 Accelerated atherosclerosis and athe-rothrombosis remain the leading causes of morbidity and mortality in patients with diabetes mellitus.2

In fact, individuals with diabetes mellitus have a 2–4-fold increased risk of developing atheroscleroticcardiovascular disease.3 Patients with diabetes mellitus also have a higher rate of recurrent athero-thrombotic events compared to patients without diabetes mellitus.4,5 Mortality in diabetic patients

Fax: þ1 904 244 3102.(D.J. Angiolillo).

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D. J. Angiolillo, S. Suryadevara / Best Practice & Research Clinical Endocrinology & Metabolism 23 (2009) 375–388376

without a history of myocardial infarction (MI) is comparable to that seen in non-diabetic patients witha history of MI.6 For these reasons, diabetes mellitus has been classified as a coronary risk equivalent.7

Several mechanisms account for the increased risk of atherothrombosis in patients with diabetesmellitus.1–6 Although these patients typically often have other cardiovascular risk factors – includinghypertension, hyperlipidemia, and physical inactivity – this accounts for no more than 25% of theircardiovascular risk.8 Therefore, other variables intrinsic to the diabetic population may contribute totheir increased cardiovascular risk. These include hyperglycemia, insulin resistance, and pro-inflam-matory and prothrombotic states.9–11 The prothrombotic state seen in T2DM is related to endothelialdysfunction, impaired fibrinolysis, increased levels of coagulation factors, and high platelet reactivity(HPR). These mechanisms are summarized in Table 1. Given that platelet aggregation plays a centralrole in the development of atherothrombotic events, it is not surprising that HPR has been associatedwith short- to mid-term atherothrombotic complications in patients with T2DM, and that antiplatelettherapy is one of the cornerstones of both primary and secondary prevention of atherothromboticevents in patients with T2DM.12

Despite antiplatelet therapy, diabetic patients remain at increased risk of recurrent ischemic eventscompared to non-diabetic patients. Of concern is the observation that persistent HPR following evendual antiplatelet therapy is more frequent in diabetic compared to non-diabetic individuals.9,13,14 Thisrelates to the emerging issue of inadequate responsiveness or ‘resistance’ to antiplatelet agents, whichappears to have a greater prevalence among diabetic patients. This review focuses on diabetes-asso-ciated platelet dysfunction, the role of oral antiplatelet therapy in patients with diabetes mellitus, andthe resistance to standard antiplatelet regimens observed in this population.

Platelet dysfunction in diabetes mellitus

Platelet dysfunction in diabetes mellitus is related to several mechanisms – including metabolicderangements, oxidative stress, and endothelial dysfunction – which lead to enhanced platelet reac-tivity.9–11 In fact, platelets in diabetics respond more frequently even to subthreshold stimuli, aredissipated and consumed more rapidly, and thus contribute to accelerated thrombopoiesis comparedto platelets in non-diabetic individuals.15

Metabolic derangements

Chronic hyperglycemia has been identified as a causal factor for platelet hyperreactivity and in-vivoplatelet activation in patients with diabetes mellitus.16,17 Enhanced thromboxane biosynthesis wasamong the earliest characterized abnormalities in platelets of diabetic individuals and has been shownto be related to the level of glycemic control. Indeed, tight glycemic control leads to a reduction inthromboxane levels.18 Acute hyperglycemia also induces increased platelet activation when plateletsare exposed to high shear stress conditions.19 Additionally, short-term hyperglycemia leads to

Table 1Mechanisms leading to the prothrombotic state in diabetes mellitus patients.

Impaired fibrinolysisIncreased PAI-1 and a-2 antiplasminDecreased t-PA

Impaired platelet functionIncreased adhesion, aggregation, and activation (GP IIb/IIIa, Psel, CD40L)

Impaired coagulationIncreased fibrinogenIncreased vWFIncreased thrombinIncreased FVII, FVIIIDecreased AT-IIIDecreased sulfated heparins

Endothelial dysfunctionIncreased adhesion molecules (VCAM, etc)Increased Leukocyte–endothelial interactionIncreased oxidative cell stress (induction NFkB)Impaired vasodilation (increased ET-1, decreased NO)Impaired endothelial regeneration

AT-III, anti-thrombin III; CD40L, CD40 ligand; ET-1, endothelin; FVII, FVIII, coagulation factors VII and VIII; GP IIb/IIIa, glycoproteinIIb/IIIa; NFkB: nuclear factor kB; NO, nitric oxide; PAI-1, plasminogen activator inhibitor 1; Psel, P-selectin; t-PA, tissue plas-minogen activator; VCAM, vascular cell adhesion molecule-1; vWF, von Willebrand factor.

D. J. Angiolillo, S. Suryadevara / Best Practice & Research Clinical Endocrinology & Metabolism 23 (2009) 375–388 377

hypersensitivity to agonists due to impaired calcium homeostasis, activation of protein kinase C (PKC),decreased production of platelet-derived nitric oxide, and increased formation of superoxide.9–11,20,21

Furthermore, hyperglycemia causes non-enzymatic glycation of platelet membrane proteins whichresults in changes in protein structure and conformation, as well as alterations of membrane lipiddynamics.9–11 Altered platelet membrane dynamics may itself convey an increased surface expressionof P-selectin and glycoprotein receptors and may modulate cell–cell interactions.9–11 In particular,platelets of diabetic patients demonstrate enhanced interaction with red blood cells. The resultantincrease in adenosine diphosphate (ADP) contributes to platelet hyperreactivity.22 Hyperglycemia alsoinduces an increase in non-enzymatic glycation of low-density lipoprotein (glycLDL) which rendersplatelets more susceptible to oxidative stress.23–25 Additionally, glycLDL may induce plateletdysfunction by increasing the intracellular calcium concentration and inhibiting platelet membranesodium–potassium-ATP activity.23–25

Despite the lack of an insulin-responsive glucose transporter in platelets, insulin exerts non-clas-sical effects on the platelet as platelets retain a functional insulin receptor. In-vitro studies have shownthat insulin reduces platelet aggregation by inhibiting the P2Y12 pathway.26–28 This is attributed to lossof Gi-mediated inhibitory effects on intracellular signaling pathways. This in turn reduces suppressionof cyclic adenosine monophosphate (cAMP), thus inhibiting P2Y12 signaling and reducing plateletreactivity.28,29 However, platelets of type-2 diabetics are victims of the insulin resistance phenomenonwhich characterizes T2DM and results in reduced sensitivity to insulin.27 The net result is up-regu-lation of the P2Y12 pathway and increased platelet reactivity in diabetic patients.

Oxidative stress

Chronic hyperglycemia can result in reactive oxygen species (ROS) production directly viaglucose metabolism and auto-oxidation, and indirectly through the formation of advanced glycationend products and their receptor binding. ROS may then activate signaling molecules within endo-thelial cells, including PKC and nuclear factor-kB (NF-kB), resulting in transcription of redox-sensitive genes which switch to pro-inflammatory and prothrombotic phenotypes.29 ROS may alsoinduce structural and functional changes in coagulative proteins. Finally, ROS may lead to theformation of 8-iso-PGF2a, a non-enzymatic oxidation product of circulating LDL and arachidonicacid, which mediates vasoconstriction and platelet hyperreactivity.30–33 Not surprisingly, improvedglycemic control is associated with a significant reduction in both lipid peroxidation and plateletactivation in vivo.17

Endothelial dysfunction

The endothelium produces mediators of both vasoconstriction (i.e. endothelin, superoxide anion,angiotensin II, and thromboxane) and vasodilation (i.e. nitric oxide and prostacyclin).34 Patients withT2DM exhibit a shift in the balance between these mediators in favor of vasoconstriction.35 Hyper-glycemia plays a pivotal role in this dysfunction. It inhibits the production of nitric oxide and increasesthe production of ROS, which as alluded to earlier leads to the expression of numerous genes thatregulate pro-inflammatory cytokines and platelet adhesion molecules. This may result in increasedproduction of tissue factor, the major procoagulant found in atherosclerotic plaque, as well as alter-ations in coagulation and fibrinolytic factors.36,37 Overall, endothelial dysfunction and disturbances inthe coagulation pathway may lead to initial platelet adhesion, activation, and subsequent aggregation,while intrinsic platelet abnormalities, which result from metabolic derangements and/or changes inintraplatelet signaling pathways, contribute to the platelet hyperreactivity seen in T2DM.

Role of oral antiplatelet agents: aspirin and clopidogrel

A variety of antiplatelet agents, administered either orally or intravenously, are used for theprevention and treatment of arterial thrombotic disorders. Aspirin and clopidogrel are the two mostcommonly used oral antiplatelet agents in patients with diabetes mellitus, and these are discussedbelow.


Aspirin

Aspirin selectively acetylates cyclooxygenase-1 (COX-1), thereby blocking the formation of throm-boxane A2 in platelets.38 This effect is irreversible, as platelets are enucleate and thus are unable to re-synthesize COX-1. For several decades, aspirin was the sole antiplatelet option for the prevention andtreatment of manifestations of cardiovascular disease. Today, this compound remains the antiplateletagent of choice for secondary prevention of ischemic events in patients with atherosclerotic disease,including those with diabetes mellitus. Aspirin mayalso be used for primary prevention of ischemic events.Although this indication in the general population is controversial, there is an overall expert consensus onthe appropriateness of aspirin for primary prevention in patients with diabetes mellitus. Aspirin asa primary and secondary prevention strategy in patients with diabetes mellitus is described below.

Aspirin as a primary prevention strategy in diabetes mellitusThe American Diabetes Association (ADA) recommends the use of low-dose aspirin (75–162 mg/

day) as a primary prevention strategy in patients with type-1 or type-2 diabetes mellitus at increasedcardiovascular risk, including those >40 years of age or with additional risk factors (family history ofcardiovascular disease, hypertension, smoking, dyslipidemia, or albuminuria).39 However, aspirintherapy should not be recommended for patients under the age of 21 years because of the increasedrisk of Reye’s syndrome associated with aspirin use in this population. Of note, people under the age of30 have generally not been studied. The American Heart Association (AHA) also recommends the use ofaspirin (75–160 mg/day) as a primary prevention strategy in high-risk individuals, defined as thosewith a 10-year risk of coronary heart disease �10%, which is a threshold exceeded by most diabetics,particularly those with T2DM.40 In contrast to the recommendations from the United States, in theEuropean guidelines there is no mention of aspirin for the primary prevention of myocardial infarctionor cardiovascular death, whereas it is recommended for the prevention of stroke.41

There are four controlled clinical trials which have evaluated the efficacy of aspirin in diabeticpatients. The US Physicians’ Health Study was a primary prevention trial which enrolled 22,701 healthymen including 533 men with diabetes mellitus.42 Among diabetic patients, the risk of MI declined from10.1% in those treated with placebo to 4.0% in those treated with aspirin 325 mg every other day (RR0.39; P¼ 0.01). In the Early Treatment of Diabetic Retinopathy Study (ETDRS), aspirin produceda significant 28% reduction in risk of MI over a 5-year period.43 This was the largest trial assessing theefficacy of aspirin designed to explore various photocoagulation approaches for pre-proliferativeretinopathy and/or maculopathy. It included 3711 patients with type-1 and -2 diabetes mellitus. Theeffect of aspirin (650 mg/day) versus placebo therapy on retinopathy as well as cardiovascular eventswas studied. The results showed that in the first 5 years of follow-up, the relative risk for MI was 0.72(99% CI 0.55–0.95, P� 0.01) in the aspirin group. An important safety finding was that there was noincrease in the risk of retinal, vitreous, or serious gastrointestinal bleeding in patients assigned toaspirin treatment. The Hypertension Optimal Treatment (HOT) study examined antihypertensivetherapy in 18,790 hypertensive individuals, 1501 of whom had diabetes mellitus.44 Low-dose aspirintherapy (75 mg/day) resulted in a further 15% reduction in the risk of cardiovascular events beyond thatseen with antihypertensive therapy (P¼ 0.03). Of note, the incidence of fatal bleeding, includingintracerebral hemorrhage, was similar in both aspirin-treated and placebo-treated patients, whereasnon-fatal bleeding was more common with aspirin therapy. Finally, the Primary Prevention Projectevaluated the efficacy of low-dose aspirin (100 mg/day) and/or vitamin E for primary prevention ofcardiovascular events in patients (n¼ 4495) with one or more risk factors (hypertension, hypercho-lesterolemia, diabetes mellitus, obesity, family history of premature MI, or old age).45 Treatment withaspirin was associated with a significantly lower incidence of cardiovascular death (from 1.4 to 0.8%; RR0.56; CI 0.31–0.99) and total cardiovascular events (from 8.2 to 6.3%; RR 0.77; CI 0.62–0.95). However,a lower, and statistically insignificant, effect was seen among diabetic patients, whereas non-diabeticpatients demonstrated consistent benefit. This may be attributed to the small number of diabeticsenrolled. In fact, in order to have adequate power, 4000 people with diabetes were planned to bestudied. However, recruitment was stopped prematurely (n¼ 1031 patients) because of the benefitseen in the entire cohort. Thus, the diabetes analysis was not adequately powered to detect an effect ofaspirin on vascular events. The ongoing ASCEND (A Study of Cardiovascular Events iN Diabetes) trial is


currently randomizing at least 10,000 diabetic individuals to aspirin 100 mg/day or placebo in theprimary prevention setting and will allow definition of the role of aspirin for this indication(clinicaltrials.gov NCT00135226).

Aspirin as a secondary prevention strategy in diabetes mellitusThe ADA recommends the use of low-dose aspirin (75–162 mg/day) for secondary prevention of

cardiovascular events in all diabetic patients with atherosclerotic disease.39 This position is supportedby the results of two large meta-analyses of major secondary prevention trials by the AntithromboticTrialists’ Collaboration (ATC), which concluded that oral antiplatelet agents (mostly aspirin) wereprotective in patients at high risk for cardiovascular disease, including those with diabetes mellitus.5,46

The ATC meta-analysis of 287 secondary prevention trials involved 212,000 high-risk patients withacute or prior vascular disease or another condition that increased their risk of vascular occlusion.5

Aspirin, in doses ranging from 75 to 325 mg/day, was the most frequently used agent. In the majorhigh-risk groups (acute MI, past history of MI, past history of stroke or transient ischemic attack, acutestroke, and other relevant history of vascular disease), antiplatelet therapy reduced the incidence ofvascular events by 23%. Of note, a low dose of aspirin (75–150 mg/day) was found to be at least aseffective as higher daily doses. Furthermore, bleeding complications were reduced with the lowerdoses. In the >4500 diabetic patients studied in the ATC, the incidence of vascular events was alsoreduced, from 23.5% in the control group to 19.3% in the group treated with antiplatelet therapy(P< 0.01), compared to 17.2 to 13.7% (P< 0.00001) in the w42,000 non-diabetic patients.46 Althoughthe overall incidence of vascular events was much higher in diabetic patients, the benefit of antiplatelettherapy in both diabetic and non-diabetic patients was consistent (42 vascular events were preventedfor every 1000 diabetic patients and 35 events for every 1000 non-diabetic patients).

Clopidogrel

Clopidogrel, a second-generation oral thienopyridine, is a selective, irreversible antagonist of theplatelet P2Y12 ADP receptor. Clopidogrel is currently the thienopyridine of choice as it has a morefavorable safety profile than ticlopidine, which is a first-generation thienopyridine.47 There is muchevidence to support the superiority of clopidogrel compared to aspirin in patients with diabetesmellitus. The Clopidogrel versus Aspirin in Patients at Risk of Ischemic Events (CAPRIE) trial examinedthe effects of clopidogrel (75 mg/day) versus aspirin (325 mg/day) in a large secondary preventionpopulation consisting of 19,185 patients with a history of recent ischemic stroke, recent MI, orestablished peripheral arterial disease (PAD).48 Approximately 20% of the study population was dia-betic. The annual incidence of the primary endpoint (combined incidence of vascular death, MI, orischemic stroke) was 5.32% with clopidogrel and 5.83% with aspirin, representing an 8.7% RR reductionwith clopidogrel compared to aspirin (P¼ 0.043). Bhatt et al retrospectively analyzed the results of thediabetic subgroup in the CAPRIE study. Of the 1914 diabetic patients randomized to clopidogrel, 15.6%had the composite vascular primary endpoint versus 17.7% of those randomized to aspirin therapy(P¼ 0.042).49 This led to 21 vascular events prevented for every 1000 diabetic patients treated. Thisfigure increased to 38 among patients treated with insulin at baseline. Interestingly, in non-diabeticpatients the reduction in the composite vascular primary endpoint with clopidogrel (11.8%) versusaspirin (12.7%) was statistically insignificant.

The Clopidogrel in Unstable Angina to Prevent Recurrent Events (CURE) study examined outcomeswith clopidogrel plus aspirin versus aspirin alone in 12,562patients with unstable angina or non-ST-elevation MI.50 Patients were randomized to treatment with either clopidogrel (300 mg loading doseand 75 mg/day thereafter) or placebo in addition to aspirin (75–325 mg/day) for up to 1 year. Patientstreated with clopidogrel and aspirin demonstrated a significant 20% reduction in the first primaryoutcome (composite vascular death, MI, or stroke) compared to patients treated with aspirin andplacebo (P< 0.001). The diabetic subpopulation represented 2840 patients. Within this group, patientstreated with clopidogrel and aspirin experienced a w17% reduction in the first primary outcomecompared to those treated with aspirin and placebo (CI 0.70–1.02). Therefore, the effect in the diabeticsubgroup was in the same direction as in the overall study population, but was of borderline statisticalsignificance. Of note, the event rate was much higher in the diabetic cohort of patients. In fact, the


primary composite cardiovascular endpoint occurred in 14.2% of patients treated with clopidogrelversus 16.7% of those treated with placebo.

These high event rates may be in part attributed to the persistence of high platelet reactivity indiabetic patients even when on dual antiplatelet therapy. In support of this, several functional studieshave shown that both in the acute phase, following administration of a 300-mg loading dose of clo-pidogrel, and in the maintenance phase, while on 75 mg/day, patients with T2DM have a lower degreeof platelet inhibition compared to non-diabetic patients.13,14 This phenomenon is even more marked ininsulin-requiring diabetics.13,14 Additionally, in a recent functional study performed in a selected cohortof patients with T2DM and high platelet reactivity, the use of a higher maintenance dose of clopidogrel(150 mg/day) resulted in improvement in platelet function profiles, although the majority of patientsremained above the predefined therapeutic threshold.51 In the CURE study, although there wasa higher incidence of major bleeding with dual antiplatelet therapy, no difference in life-threateningbleeding was observed. Of note, the increased risk of major bleeding with combination antiplatelettherapy has been attributed to the dose of aspirin used, whereas the use of aspirin doses <100 mg/daywas associated with the lowest risk of bleeding without any compromise in clinical benefit.52

On the basis of these and other clinical studies, the ADA currently recommends the use of clopi-dogrel as an adjunctive therapy in very-high-risk diabetic patients or as an alternative therapy inaspirin-intolerant patients.39 In addition, the current American College of Cardiology (ACC)/AmericanHeart Association (AHA) guidelines for the management of unstable angina and non-ST-elevation MIrecommend the addition of clopidogrel (300 mg loading dose and 75 mg/day maintenance dose) toaspirin in patients presenting with unstable angina and non-ST-elevation MI.53 Furthermore, clopi-dogrel should be used regardless of the treatment strategy adopted (invasive or non-invasive) andshould be continued ideally for up to 1 year.

Recently, the use of clopidogrel in patients with ST-elevation MI has been approved by the US Foodand Drug Administration. This position is mirrored in the current ACC/AHA guidelines on themanagement of patients with ST-elevation MI.54 The CLARITY (Clopidogrel as Adjunctive ReperfusionTherapy) study and the PCI-CLARITY sub-study were designed to address whether a beneficial effectwith clopidogrel, including a loading dose, would be attained among STEMI patients treated withthrombolytic therapy and undergoing coronary angiography during the index hospitalization.55,56 Atotal of 3491 patients who presented within 12 hours following the onset of STEMI were randomized totreatment with clopidogrel (300 mg loading dose followed by 75 mg/day maintenance dose) orplacebo. Patients were scheduled to undergo coronary angiography after 48 hours, and those whounderwent PCI during the index hospitalization formed the basis of PCI-CLARITY (n¼ 1863).56 Clopi-dogrel pretreatment significantly reduced the incidence of cardiovascular death or ischemic compli-cations both before and after PCI (at 30 days), without a significant increase in major or minor bleeding.In the COMMIT (ClOpidogrel and Metoprolol in Myocardial Infarction Trial) trial 45,852 patients withacute MI were studied.57 The addition of clopidogrel 75 mg/day to aspirin and other standard treat-ments, including fibrinolytic therapy, safely reduced in-hospital mortality and major vascular events.Of note, in both studies the benefit observed with treatment with clopidogrel was consistent across allsubgroups, including patients with diabetes mellitus.

In contrast to the clear benefit seen with dual antiplatelet therapy across the spectrum of patientswith ACS, including those undergoing PCI, the results of the CHARISMA (Clopidogrel for High Athe-rothrombotic Risk and Ischemic Stabilization, Management, and Avoidance) trial showed that in high-risk but non-acute, patients (n¼ 15,603) with either clinically evident cardiovascular disease ormultiple cardiovascular risk factors, clopidogrel plus aspirin was not significantly more effective thanaspirin alone in reducing the rate of MI, stroke, or death from cardiovascular causes, and was actuallyharmful in patients without documented atherosclerotic disease.58 Of note, a large number of patientsenrolled in the latter subgroup were diabetic. Therefore, dual antiplatelet therapy cannot be advocatedin the primary prevention setting for diabetic individuals.

Resistance to aspirin and clopidogrel: an emerging clinical issue

Antiplatelet therapy with aspirin and/or clopidogrel is the cornerstone of treatment forpatients with stable and unstable atherosclerotic cardiovascular disease, including those with


diabetes mellitus. However, a proportion of patients experience recurrent atherothrombotic eventsdespite single and even dual antiplatelet therapy. This finding led to the concept of ‘resistance’ toantiplatelet agents. It is important to underscore that ‘resistance’ is a laboratory finding thatconsists of failure of an antiplatelet agent to adequately block its specific target on the platelet.This relates to – but is not equal to – a clinical failure, which is defined by the recurrence of anischemic event. For aspirin, resistance involves inadequate inhibition of the COX-1-mediatedthromboxane A2 pathway, while for clopidogrel, resistance involves P2Y12 receptor signaling.59,60

Importantly, thrombotic events cannot be attributed to drug ‘resistance’ if the efficacy of theantiplatelet agent was not tested in the affected patient. Without such confirmation, this occur-rence should be considered ‘treatment failure’ rather than ‘resistance’ to the antiplatelet agent inquestion.

Aspirin resistance

A number of clinical studies have correlated aspirin ‘resistance’ with long-term adverse clinicaloutcomes not only in patients with coronary artery disease but also in individuals with ischemic strokeor peripheral arterial disease.61–66 The prevalence of aspirin resistance reported by investigators variesconsiderably. Such disparate estimates are largely due to differences in the definition of resistance, typeof assay used, dose of aspirin, and patient population under consideration. However, when tests thatspecifically assess COX-1 activity (e.g. serum thromboxane B2 or light transmittance aggregometryusing stimuli with arachidonic acid) are examined, aspirin produces resistance only infrequently.67 Infact, using COX-1-specific assays, the ASPECT (Aspirin-Induced Platelet Effect) study and an observa-tional study conducted by Frelinger et al reported aspirin resistance in less than 5% of patients.68,69 Inclinical practice, the foremost reason for higher prevalence of aspirin resistance when using assays thatspecifically assess COX-1 activity is poor patient compliance.59,67 The disparate estimates reported inlaboratory studies are largely assay-dependent. In many of these studies, assays that are not specific toCOX-1 signaling (e.g. PFA-100) may have been used. In fact, although many available platelet functiontests are able to detect aspirin-induced effects, the results obtained are not all specific to the degree ofCOX-1 inhibition and may be reflective of other platelet-signaling pathways. Further, the overallprevalence of inadequate aspirin effects may be influenced by the patient population under investi-gation. Patients with diabetes and ACS, who are typically characterized by hyperreactivity, also presentwith high platelet reactivity while receiving aspirin therapy. Interactions with drugs, such as ibuprofen,which interfere with aspirin-induced COX-1 acetylation, may also be a cause of inadequate or absenteffects of aspirin. The latter may also be responsible for an increased risk of ischemic effects despiteaspirin use.70

The Aspirin-Induced Platelet Effect (ASPECT) study, which tested stepwise concentrations ofarachidonic acid and different aspirin concentrations in conjunction with sophisticated analyticalmethods, confirmed that the occurrence of aspirin resistance has been greatly overestimated.68

However, when arachidonic acid is not used as the stimulant, a higher prevalence of ‘resistance’ may beobserved. Patients with diabetes in this study were more likely to be ‘resistant’ using these alternativeagonists.71 These findings underscore the importance of the methodology used when defining andtesting for antiplatelet effects induced by aspirin. Even though the significance of different assaystesting aspirin sensitivity (specific and non-specific for COX-1 inhibition) still needs to be betterdefined, meta-analyses using various laboratory assays support the poor prognostic implications ofinadequate aspirin-induced effects.72,73

Although the concept of aspirin resistance in diabetes mellitus has existed for more than 20 years,only a limited number of studies has investigated the potential mechanisms of aspirin resistance thatare intrinsic to diabetic patients.74 Diabetic individuals are characterized by increased platelet reac-tivity and an increased level and activity of prothrombotic clotting factors as alluded to earlier in thisreview, which may explain their predisposition for aspirin resistance. Hyperglycemia has beenconsidered to have a role in aspirin resistance and may explain why no differences in aspirin resistancehas been shown when comparing patients with type-1 and type-2 diabetes mellitus.75 Furthermore, aninteraction between glycation and acetylation has been repeatedly demonstrated. Increased glycationof platelet and coagulation factor proteins may interfere with the acetylation process, thereby


contributing to aspirin resistance in the presence of diabetes mellitus. It remains unclear whetherimproved glycemic control enhances the efficacy of aspirin, or if increased doses of aspirin are bene-ficial in the presence of poor glycemic control.76,77

Unfortunately, there are no published studies specifically designed to evaluate the clinical efficacyof aspirin and the implications of biochemical aspirin resistance in patients with diabetes mellitus.However, some studies have correlated aspirin resistance with adverse clinical outcomes in patientswith known atherosclerotic disease.61–66,72,73

Currently, there is a lack of evidence supporting specific strategies to overcome aspirin resistancein diabetes mellitus, including the use of a higher dose or the concomitant use of another antiplateletagent such as clopidogrel. Picotamide has been suggested as a treatment alternative to aspirin.78

Since picotamide inhibits both thromboxane A2 (TxA2) synthase and TxA2 receptors, it is able toblock the effect of TxA2 that is generated through COX-1 escape mechanisms, which may representa pathway of aspirin resistance in diabetic patients. In the Drug Evaluation in Atherosclerotic VascularDisease in Diabetics (DAVID) study, a total of 1209 adults aged 40–75 years with T2DM and peripheralarterial disease were randomized to receive picotamide (600 mg twice a day) or aspirin (320 mg oncea day) for 24 months.78 The cumulative incidence of the 2-year overall mortality was significantlylower among patients who received picotamide (3.0%) than in those who received aspirin (5.5%), witha relative risk ratio for picotamide versus aspirin of 0.55 (95% CI: 0.31–0.98%). However, although thecombined endpoint of mortality and morbidity had a slightly lower incidence in the picotamidegroup, this difference did not reach statistical significance. Other thromboxane receptor antagonistsare also currently under clinical investigation. Ultimately, further clinical research is needed to helpformulate guidelines for the diagnosis and treatment of aspirin resistance in patients with diabetesmellitus.

Clopidogrel resistance

Clopidogrel, a second generation oral thienopyridine, is a specific, irreversible antagonist of theplatelet P2Y12 ADP receptor and thus inhibits platelet aggregation in a manner entirely distinctfrom that of aspirin. Similarly to aspirin, the prevalence of clopidogrel resistance reported in theliterature varies considerably and is related to differences in the definitions used, type of assayused, dose of clopidogrel, and patient population.60 Nonetheless, inter-individual variability inplatelet response to clopidogrel is now a well-established concept. Clopidogrel non-responsivenessis more prevalent in patients with diabetes mellitus than non-diabetic patients.13,14,79,80 In partic-ular, T2DM patients undergoing elective PCI have approximately a four-fold increase in the numberof non-responders at 24 hours following a standard 300-mg loading dose.13 Also, platelet reactivityis persistently elevated in T2DM patients even in the maintenance phase of clopidogrel and ishighest among those patients requiring insulin therapy.14 These findings may explain why diabeticpatients, particularly those in the most advanced stage of their disease (e.g. insulin-requiringdiabetics), continue to have a high risk of atherothrombotic events.52 This may also explain whystudies with drug-eluting stents have identified diabetes as a predictor of stent thrombosis.81–84

Accumulating data have emerged suggesting that a correlation may exist between clopidogrel non-responsiveness and adverse clinical outcomes, including stent thrombosis.60,85 Of note, plateletfunction profiling selectively performed among T2DM patients on aspirin and clopidogrel therapyshowed that there is a broad range of responsiveness, although with higher platelet reactivitycompared to non-diabetic patients.12 T2DM with HPR, defined as the upper quartile of maximalADP-induced platelet aggregation following 20 mmol/L stimuli, had the highest risk of 2-yearadverse outcomes (see Fig. 1).12 Of note, platelet dysfunction was independent of glycemic controland inflammatory status.

Numerous mechanisms may account for this observation, but up-regulation of the P2Y12 pathwayappears to be of particular importance in patients with T2DM. In-vitro studies have shown that insulinreduces platelet aggregation by inhibition of the P2Y12 pathway.11,27,28 As alluded to earlier in thisreview, human platelets are targets of insulin, which interacts with its own receptor on the surface ofthe platelet leading to loss of Gi activity. This results in suppression of cAMP, inhibition of P2Y12

signaling, and reduced platelet reactivity.27,28 However, platelets of diabetic patients are victims of the

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Fig. 1. Impact of platelet function profiles on clinical outcomes in type-2 diabetes mellitus patients with coronary artery disease.Type-2 diabetes mellitus patients (n¼ 173) on aspirin and clopidogrel therapy were divided into four groups according to quartiledistribution of maximum platelet aggregation induced by ADP (20 mmol/L). Patients in the upper quartile of post-treatment plateletreactivity had the highest 2-year incidence of major adverse cardiovascular events (MACE).


insulin resistance phenomenon that characterizes T2DM and results in decreased sensitivity toinsulin.11 The net result is up-regulation of the P2Y12 pathway and increased platelet reactivity indiabetic patients. Other mechanisms linked to clopidogrel non-responsiveness in patients with T2DMinclude increased exposure to ADP, increased cytosolic levels of calcium, and increased plateletturnover.11

Several studies have focused on how to overcome clopidogrel non-responsiveness. Currentguidelines provide only a weak recommendation (class IIB, level of evidence C) for increasing themaintenance dose of clopidogrel to 150 mg/day in those patients in whom stent thrombosis may becatastrophic or lethal (unprotected left main, bifurcating left main, and last patent coronary vessel) if<50% inhibition of platelet aggregation is demonstrated.86 In line with this, the Optimizing anti-Platelet Therapy In diabetes MellitUS (OPTIMUS) study selectively examined T2DM patients with HPRwhile in their chronic phase of treatment with clopidogrel.51 The use of a 150-mg clopidogrelmaintenance dose resulted in marked platelet inhibition as determined by numerous platelet func-tion measures compared to a 75-mg dose. However, a significant number of patients remained abovethe therapeutic threshold of post-treatment platelet reactivity adopted in this study. Unfortunately,this study was not sufficiently powered to assess safety and clinical efficacy. Another approach toimprove P2Y12 inhibition in T2DM patients treated with clopidogrel is with the adjunctive use ofcilostazol, as assessed in the OPTIMUS-2 study.87 This study showed that the increase in intraplateletcAMP levels induced by cilostazol led to enhanced P2Y12 inhibitory effects.87 This may explain whyclinical studies have shown that triple oral antiplatelet therapy (aspirin, clopidogrel, and cilostazol) isassociated with better clinical outcomes than with dual therapy, particularly in patients with diabetesmellitus.88,89

Glycoprotein IIb/IIIa inhibitors have been shown to be beneficial in diabetic patients, particularlywhen presenting with an ACS.90 However, the fact that these agents are administered intravenouslyand are suitable only for acute use makes them a non-viable treatment alternative to overcomeinadequate antiplatelet drug responsiveness in the maintenance phase of treatment. This underscoresthe need for alternative and more potent antiplatelet agents. There are several P2Y12 receptor antag-onists under advanced clinical investigation.60,91 These include prasugrel, ticagrelor, and cangrelor.Prasugrel and ticagrelor are administered orally, while cangrelor is for intravenous use. Prasugrel is anirreversible agent, while ticagrelor and cangrelor are reversible. All three agents are characterized byincreased potency, and are associated with less response variability compared to clopidogrel.Encouraging clinical outcome data from large-scale phase-III testing is available for prasugrel, which isa third-generation thienopyridine.92

The Trial to Assess Improvement in Therapeutic Outcomes by Optimizing Platelet Inhibitionwith Prasugrel (TRITON) TIMI 38 was a randomized, double-blind, parallel-group, multinational

0

5

10

15

20

25

30HR=0.70

P<0.001

HR=0.63

P=0.009

HR=0.74

P=0.009

Diabeticson insulin

(N=776)

Diabetics noton insulin

(N=2370)

Overalldiabetics

(N=3146)

Fig. 2. Impact of diabetic status on the primary endpoint in the Trial to Assess Improvement in Therapeutic Outcomes by OptimizingPlatelet Inhibition with Prasugrel (TRITON), comparing prasugrel with clopidogrel. The major adverse cardiovascular events (MACE)rate representing the primary endpoint (cardiovascular death, non-fatal myocardial infarction, or non-fatal stroke) was reducedsignificantly with prasugrel compared to clopidogrel among diabetic subjects (n¼ 3146); a benefit for prasugrel was observedamong diabetic subjects on insulin (n¼ 776) and those not on insulin (n¼ 2370).


trial comparing prasugrel with clopidogrel in patients (n¼ 13,608) with moderate- to high-riskACS who underwent PCI.93 Patients were randomized to treatment with either a prasugrel 60-mgloading dose followed by a 10-mg/day maintenance dose, or a clopidogrel 300-mg loading dosefollowed by a 75-mg/day maintenance dose for a median duration of 14.5 months. The primaryendpoint (composite of cardiovascular death, non-fatal MI, or non-fatal stroke) occurred in 12.1%of patients treated with clopidogrel versus 9.9% of patients treated with prasugrel (HR, 0.81; 95%CI, 0.73–0.90; P< 0.001). However, there was an increased risk of major bleeding which wasobserved in 2.4% of the patients receiving prasugrel compared to 1.8% of the patients receivingclopidogrel (HR, 1.32; 95% CI, 1.03–1.68; P¼ 0.03). Despite the increased bleeding risk, the pre-specified net clinical benefit analysis, defined as the composite of efficacy and bleeding endpoints(death from any cause, non-fatal MI, non-fatal stroke, and TIMI major hemorrhage), still favoredprasugrel (13.9% of patients in the clopidogrel group versus 12.2% in the prasugrel group; HR 0.87;95% CI, 0.79–0.95; P¼ 0.004). Of note, the patients who benefited most from prasugrel therapywere diabetics.94 There were 3146 subjects with a preexisting history of diabetes, of which 776receiving insulin therapy. The primary endpoint was reduced significantly with prasugrel amongdiabetic subjects (12.2% versus 17.0%; HR, 0.70; P< 0.001). A benefit for prasugrel was observedamong diabetic subjects on insulin (14.3% versus 22.2%; HR, 0.63; P¼ 0.009) and those not oninsulin (11.5% versus 15.3%; HR, 0.74; P¼ 0.009) (Fig. 2). Myocardial infarction was reduced withprasugrel by 40% among diabetic subjects (8.2% versus 13.2%; HR, 0.60; P< 0.001). Similar TIMImajor hemorrhage rates were observed in diabetic subjects for clopidogrel and prasugrel (2.6%versus 2.5%; HR, 1.06; P¼ 0.81).The net clinical benefit with prasugrel was greater for subjectswith diabetes (14.6% versus 19.2%; HR, 0.74; P< 0.001) than for subjects without diabetes (11.5%versus 12.3%; HR, 0.92; P¼ 0.16)., in whom there was a 30% relative risk reduction in the primaryendpoint in favor of prasugrel (12.2 versus 17.0%, HR 0.70, P< 0.001) without differences in majorbleeding rates (2.6 versus 2.5%, HR 1.06, P¼ 0.81).

Ultimately, the development of more potent antiplatelet agents, including those that inhibittargets other than COX-1 and P2Y12 such as thrombin, are warranted in order to overcome themultitude of stimuli leading to enhanced platelet reactivity which characterizes diabetic patients. Ofnote, thrombin is the most potent platelet stimulus, and thrombin generation is pronounced indiabetic patients. Several thrombin receptor antagonists that block the PAR-1 receptor subtype(E5555, SCH 530348) are currently under clinical investigation.95 The advent of novel antiplateletagents will set the basis for the development of individualized and tailored antithrombotic treatmentstrategies in diabetic patients.

Research agenda

� Novel and more potent antiplatelet agents are currently under advanced clinical develop-ment and may represent future therapeutic alternatives to reduce the risk of developingischemic events in diabetes mellitus patients.

Practice points

� Diabetes mellitus patients are characterized by enhanced platelet reactivity which contrib-utes to their elevated atherothrombotic risk.� Oral antiplatelet therapy, including aspirin and clopidogrel, are key to reducing the long-term

risk of ischemic events in high risk patients, including diabetics.� Despite the use of oral antiplatelet agents, diabetic patients continue to have a high risk of

adverse cardiovascular outcomes.� Recent laboratory findings have shown that diabetic patients have a greater prevalence of

inadequate platelet inhibition or ‘‘resistance’’ to aspirin and clopidogrel compared to non-diabetics, which may contribute to their enhanced atherothrombotic risk.� Individualized antithrombotic treatment strategies may be warranted in diabetes mellitus

patients.


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