Diana A. Gorog

Antiplatelet drug 'resistance'. Part 1: mechanisms and clinical measurements

Antiplatelet drug therapy has become one of the cornerstones of treatment for patients with cardiovascular disease. Large clinical trials have shown that antiplatelet medications have important clinical benefits and prevent adverse outcomes in patients with coronary artery disease. Recurrent adverse cardiovascular events still occur in a substantial proportion of patients on standard dual antiplatelet therapy, however, which has been attributed to nonresponsiveness to this treatment. Both pharmacological and pharmacokinetc mechanisms are involved in variability in responsiveness to antiplatelet agents, and include drug bioavailability, medication noncompliance, drug–drug interactions, cytochrome P450 activity, and genetic polymorphisms. Numerous observational studies have consistently shown an association between antiplatelet drug nonresponsiveness and adverse clinical outcomes. However, these studies are limited by varying antiplatelet drug dosing regimens, heterogeneous laboratory assessments for ex vivo platelet function, and wide interindividual variation in platelet responses. Only within the last 2 years have randomized clinical trials indicated that increased dosing with antiplatelet drugs could reduce adverse clinical outcomes. Nonetheless, large clinical trials with standardized laboratory methods and well-defined protocols are needed that will definitively determine the association between antiplatelet drug nonresponsiveness and clinical events, and establish therapeutic strategies to overcome blunted antiplatelet effects.

Prognostic Value of Plasma Fibrinolysis Activation Markers in Cardiovascular Disease

The pivotal role of hypoactive endogenous fibrinolysis in the occurrence of thrombotic cardiovascular events is now well-recognized. To evaluate the diagnostic and prognostic role of impaired fibrinolysis, plasma fibrinolysis markers have been investigated in large prospective studies in both healthy individuals and patients with established coronary disease. Antigen and activity levels of components of the fibrinolytic system were measured by immunoassays, which replaced earlier global fibrinolysis tests. This review covers 45 studies in nearly 50,000 subjects, examining the association between plasma markers of fibrinolysis and coronary artery disease, to establish the usefulness of these markers in predicting future cardiovascular events. The predictive value of plasma levels of tissue-type plasminogen activator, platelet activator inhibitor-1, plasmin-antiplasmin complex, D-dimer, thrombin activatable fibrinolysis inhibitor, and lipoprotein(a) for major adverse cardiac events is highly variable and conflicting, especially after adjusting for conventional risk factors, judging from the published data in the last decade. The value of fibrinolysis activity markers is very limited in aiding diagnosis and risk stratification in the individual patient, on the basis of the weak prognostic values obtained in some studies and the lack of power in others. The physiological limitations of such markers in reflecting endogenous fibrinolysis is discussed. The emerging novel global assays of fibrinolysis will require large-scale clinical trials before their prognostic power or superiority to multiple biomarker measurements can be evaluated.

Platelet Function Tests in Clinical Cardiology: Unfulfilled Expectations

This review is a critical evaluation of publications in the past decade on the usefulness of platelet function tests (PFTs) in clinical cardiology, in aiding diagnosis, predicting risk, and monitoring therapy. The ideal PFT should: 1) detect baseline platelet hyperreactivity; 2) allow individualization of antiplatelet medication; 3) predict thrombotic risk; and 4) predict bleeding risk. The practicalities of clinical cardiology demand rapid, accurate, and reliable tests that are simple to operate at the bedside and available 24 h a day, 7 days a week. Point-of-care PFTs most widely evaluated clinically include PFA-100 and VerifyNow. None of these tests can reliably detect platelet hyperreactivity and thus identify a prothrombotic state. Identification of antiplatelet nonresponsiveness or hyporesponsiveness is highly test specific, and does not allow individualization of therapy. The power of PFTs in predicting thrombotic events for a given individual is variable and often modest, and alteration of antithrombotic treatment on the basis of the results of PFTs has not been shown to alter clinical outcome. PFTs in current mainstream use cannot reliably assess bleeding risk. These tests have been in use for over a decade, but the hopes raised by PFTs in clinical practice remain unfulfilled. Although physiologically relevant measurement of platelet function now is more important than ever, a critical reappraisal of available techniques in light of clinical requirements is needed. The use of native blood, global stimulus instead of individual agonists, contribution of thrombin generation by activated platelets to the test results, and establishment of a PFT therapeutic range for each antiplatelet drug should be considered and is discussed.

Platelet membrane glycoprotein Ibα gene −5T/C Kozak sequence polymorphism as an independent risk factor for the occurrence of coronary thrombosis

Objective: To explore the potential of the GPIbα gene variable number tandem repeat (VNTR) and −5T/C Kozak polymorphisms to act as independent risk factors for myocardial infarction. Methods: 256 patients aged 33–80 years (180 caucasian, 76 Indian Asian) were recruited at cardiac catheterisation for any diagnostic indication, and divided into two groups: group A, with confirmed previous myocardial infarction evident on ECG or ventriculogram (88 patients, 79 men, 9 women) and group B, with no evidence of myocardial infarction (168 patients, 101 men, 67 women). Results: There was no significant difference in race, age, hypertension, smoking status, or family history between the infarct and non-infarct groups, though there was a significant difference in sex (89.8% male in group A, 60.1% male in group B, p < 0.001). Genotype analysis showed a strong association between the GPIbα Kozak homozygous TT genotype and the occurrence of myocardial infarction (group A: TT 85.2%, TC 12.5%, CC 2.3%; group B: TT 67.3%, TC 32.7%, p = 0.001). No significant association was found between myocardial infarction and the GPIbα VNTR, although analysis of the CC VNTR genotype against all other GPIbα VNTR genotypes showed a marginal association with myocardial infarction (p = 0.059). There was no association between the Kozak sequence polymorphism (p = 0.797) or GPIbα VNTR (p = 0.714) and the degree of vessel disease. Conclusions: The homozygous TT Kozak genotype may be a significant factor in the outcome of coronary artery disease completed by myocardial infarction. Conversely, the Kozak C allele in the heterozygous state TC may confer some protection against myocardial infarction.

State-of-the-Art PaperPlatelet Function Tests in Clinical Cardiology: Unfulfilled Expectations

This review is a critical evaluation of publications in the past decade on the usefulness of platelet function tests (PFTs) in clinical cardiology, in aiding diagnosis, predicting risk, and monitoring therapy. The ideal PFT should: 1) detect baseline platelet hyperreactivity; 2) allow individualization of antiplatelet medication; 3) predict thrombotic risk; and 4) predict bleeding risk. The practicalities of clinical cardiology demand rapid, accurate, and reliable tests that are simple to operate at the bedside and available 24 h a day, 7 days a week. Point-of-care PFTs most widely evaluated clinically include PFA-100 and VerifyNow. None of these tests can reliably detect platelet hyperreactivity and thus identify a prothrombotic state. Identification of antiplatelet nonresponsiveness or hyporesponsiveness is highly test specific, and does not allow individualization of therapy. The power of PFTs in predicting thrombotic events for a given individual is variable and often modest, and alteration of antithrombotic treatment on the basis of the results of PFTs has not been shown to alter clinical outcome. PFTs in current mainstream use cannot reliably assess bleeding risk. These tests have been in use for over a decade, but the hopes raised by PFTs in clinical practice remain unfulfilled. Although physiologically relevant measurement of platelet function now is more important than ever, a critical reappraisal of available techniques in light of clinical requirements is needed. The use of native blood, global stimulus instead of individual agonists, contribution of thrombin generation by activated platelets to the test results, and establishment of a PFT therapeutic range for each antiplatelet drug should be considered and is discussed.

Endogenous Fibrinolysis: An Important Mediator of Thrombus Formation and Cardiovascular Risk.

Most acute cardiovascular events are attributable to arterial thrombosis. Plaque rupture or erosion stimulates platelet activation, aggregation, and thrombosis, whilst simultaneously activating enzymatic processes that mediate endogenous fibrinolysis to physiologically maintain vessel patency. Interplay between these pathways determines clinical outcome. If proaggregatory factors predominate, the thrombus may propagate, leading to vessel occlusion. However, if balanced by a healthy fibrinolytic system, thrombosis may not occur or cause lasting occlusion. Despite abundant evidence for the fibrinolytic system regulating thrombosis, it has been overlooked compared with platelet reactivity, partly due to a lack of techniques to measure it. We evaluate evidence for endogenous fibrinolysis in arterial thrombosis and review techniques to assess it, including biomarkers and global assays, such as thromboelastography and the Global Thrombosis Test. Global assays, simultaneously assessing proaggregatory and fibrinolytic pathways, could play a role in risk stratification and in identifying impaired fibrinolysis as a potential target for pharmacological modulation.

First direct comparison of platelet reactivity and thrombolytic status between Japanese and Western volunteers: Possible relationship to the “Japanese paradox”

OBJECTIVE To determine and compare thrombotic and endogenous thrombolytic status in Japanese and Western populations. BACKGROUND Incidence of coronary heart disease (CHD) and AMI in Japan remains lower than in Western countries. Primary genetic effects are unlikely, given the increased CHD in Japanese migrants. For men, cholesterol and blood pressure have been similar in Japan and the U.S. Dietary factors are implicated, but how these effect CHD is unclear. We postulated that differences in thrombotic and/or thrombolytic status may contribute. METHODS We measured thrombotic and thrombolytic status in 100 healthy Japanese (J) from Japan and 100 healthy Westerners (W) from the U.K. using the Global Thrombosis Test (GTT). The GTT employs non-anticoagulated blood to create platelet-rich thrombi under high shear (occlusion time OT; seconds), and then measures the restart of blood flow, due to spontaneous thrombolysis (lysis time LT; seconds). RESULTS OT was longer in (J) compared to (W) (545 vs. 364, p<0.0001). LT was longer in (J) than in (W) (1753 vs. 1052, p<0.0001). Distribution of LT in (J) did not conform to a normal population, with markedly impaired thrombolytic status (LT>3,000 s) in 18%, compared to none of the Westerners (p<0.0001). CONCLUSIONS There are marked differences in thrombotic and thrombolytic status, with (J) having less prothrombotic (longer OT) but less favourable endogenous thrombolytic profile (longer LT). This may be important in the aetiology of thrombotic events. Since platelets and thrombolysis were both inhibited in (J) and yet incidence of AMI is lower, OT would seem more important than LT as a determinant of overall thrombotic risk in this population.

Impaired thrombolysis: a novel cardiovascular risk factor in end-stage renal disease

AIMS End-stage renal disease (ESRD) patients have an excess cardiovascular risk, above that predicted by traditional risk factor models. Prothrombotic status may contribute to this increased risk. Global thrombotic status assessment, including measurement of occlusion time (OT) and thrombolytic status, may identify vulnerable patients. Our aim was to assess overall thrombotic status in ESRD and relate this to cardiovascular risk. METHODS AND RESULTS Thrombotic and thrombolytic status of ESRD patients (n = 216) on haemodialysis was assessed using the Global Thrombosis Test. This novel, near-patient test measures the time required to form (OT) and time required to lyse (lysis time, LT) an occlusive platelet thrombus. Patients were followed-up for 276 ± 166 days for major adverse cardiovascular events (MACE, composite of cardiovascular death, non-fatal MI, or stroke). Peripheral arterial or arterio-venous fistula thrombosis was a secondary endpoint. Occlusion time was reduced (491 ± 177 vs. 378 ± 96 s, P < 0.001) and endogenous thrombolysis was impaired (LT median 1820 vs.1053 s, P < 0.001) in ESRD compared with normal subjects. LT ≥ 3000 s occurred in 42% of ESRD patients, and none of the controls. Impaired endogenous thrombolysis (LT ≥ 3000 s) was strongly associated MACE (HR = 4.25, 95% CI = 1.58-11.46, P = 0.004), non-fatal MI and stroke (HR = 14.28, 95% CI = 1.86-109.90, P = 0.01), and peripheral thrombosis (HR = 9.08, 95% CI = 2.08-39.75, P = 0.003). No association was found between OT and MACE. CONCLUSION Impaired endogenous thrombolysis is a novel risk factor in ESRD, strongly associated with cardiovascular events.

Novel oral anticoagulants in acute coronary syndrome

Coronary artery disease (CAD) is a leading cause of morbidity and mortality worldwide with a prevalence that has now reached pandemic levels as a consequence of the rapid modernization of the developing world. Its presentation as an acute coronary syndrome (ACS) is a frequent reason for hospital admission and of profound implications for personal, societal and global health. Despite improvements in the management of ACS with anti-platelet and anticoagulant therapy and revascularization techniques, many patients continue to suffer recurrent ischemic events. The need to reduce future cardiovascular events has led to the development of novel therapies to prevent coronary thrombosis, targeting thrombin-mediated pathways. These include direct Xa inhibitors (apixaban, rivaroxaban and darexaban), direct thrombin inhibitors (dabigatran) and PAR 1 antagonists (vorapaxar and atopaxar). This article critically reviews the comparative mechanisms of action, the risks and benefits, together with the clinical evidence base for the use of these novel oral agents in the management of ACS patients.

Global Thrombosis Test (GTT) can detect major determinants of haemostasis including platelet reactivity, endogenous fibrinolytic and thrombin generating potential

BACKGROUND Detection of both thrombosis and bleeding risk are essential in clinical cardiology. Thrombin generated by activated platelets and from the extrinsic coagulation pathway is the major determinant of thrombogenesis and hemostasis. Although novel oral anticoagulants further increase the bleeding risk of antiplatelet drugs, platelet function tests do not reliably predict hemorrhagic complications. It seems that in addition to platelet aggregation, true assessment of bleeding risks requires the measurement of both platelet and plasma derived thrombin activity. OBJECTIVE To adapt a novel, near-patient test for the assessment of both antithrombotic and anticoagulant effects of oral thrombin inhibitors. METHODS The point-of-care Global Thrombosis Test (GTT), which measures platelet reactivity to shear-activation in native blood, was used. Thrombin, generated from activated platelets (procoagulant activity) plays a pivotal role in GTT measurement. In order to assess endogenous thrombin potential, in a separate blood sample thrombin generation was induced by microparticles formed during hypotonic hemolysis. Thus two blood samples were tested to measure simultaneously platelet reactivity (occlusion time, OT) and hemolysis (microparticles)-induced endogenous thrombin potential (OT-H). RESULTS In healthy subjects (n=32), OT measured in native blood was reduced in hemolysed blood (100% vs. 43 ± 4%; OT vs. OT-H respectively). Shortening of OT in hemolysed blood (OT-H) was dose-dependently inhibited by the in vitro added thrombin inhibitor argatroban. In patients receiving dabigatran (n=27), OT and, to a lesser extent, OT-H was prolonged, compared to healthy volunteers. Intra-assay variation of OT-H was low (4.5%), but interindividual variation was great, both in healthy subjects (61%) and in patients on dabigatran (65%). Thrombin inhibitors argatroban, heparin (in vitro) and dabigatran (in vivo) all prolonged both OT and OT-H. There was no correlation between the measured OT and OT-H data. CONCLUSIONS Microparticles shed from erythrocytes during hypotonic lysis of native blood considerably shortened OT. In a direct proportion to the applied concentrations, various thrombin inhibitors prolonged both OT (antithrombotic effect) and to a lesser extent, OT-H (anticoagulant effect). Further large studies are required to evaluate the usefulness of this technique in a clinical setting, in assessing the anticoagulant and antithrombotic effects of medication and relating GTT results with observed thrombotic and bleeding events.