Prolonged enoxaparin therapy compared with standard-of-care antithrombotic therapy in opiate-treated patients undergoing primary percutaneous coronary intervention.

A novel enoxaparin regimen consisting of intra-arterial bolus (0.75 mg/kg) followed by intravenous infusion (0.75 mg/kg/6 hours) has been developed as a possible solution to the delayed absorption of oral P2Y12 inhibitors in opiate-treated ST-elevation myocardial infarction (STEMI) patients undergoing primary angioplasty. We aimed to study the feasibility of this regimen as an alternative to standard-of-care treatment (SOC) with unfractionated heparin ± glycoprotein IIb/IIIa antagonist (GPI). One hundred opiate-treated patients presenting with STEMI and accepted for primary angioplasty were randomized (1:1) to either enoxaparin or SOC. Fifty patients were allocated enoxaparin (median age 61, 40% females) and 49 allocated SOC (median age 62, 22% females). One developed stroke before angiography and was withdrawn. One SOC patient had a gastrointestinal bleed resulting in 1 g drop in hemoglobin and early cessation of GPI infusion. Two enoxaparin patients had transient minor bleeding: one transient gingival bleed and one episode of coffee ground vomit with no hemoglobin drop or hemodynamic instability. Two SOC and no enoxaparin group patients had acute stent thrombosis. These preliminary data support further study of this novel 6-hour enoxaparin regimen in opiate-treated PPCI patients.

Offset of ticagrelor prior to coronary artery bypass graft surgery for acute coronary syndromes: effects on platelet function and cellular adenosine uptake.

Ticagrelor is an antagonist of both platelet adenosine diphosphate (ADP) receptor P2Y12 and equilibrative nucleoside transporter-1. Optimal timing of ticagrelor cessation prior to coronary artery bypass grafting (CABG) remains unclear. We characterized the offset of ticagrelor's effects on platelets and cellular adenosine uptake in ticagrelor-treated patients (n = 13) awaiting CABG. Blood was drawn prior to CABG at multiple timepoints 2 to 120 (h) after the last dose of ticagrelor. Platelet function (n = 13) was assessed with multiple electrode aggregometry (MEA), expressed as arbitrary units (U) derived from area-under-the-curve (AUC) in response to ADP, and inhibition of adenosine uptake by high-performance liquid chromatography (n = 7). Mean±SD AUC was 20.3 ± 8.2 U (2 h post-ticagrelor), 33.0 ± 18.3U (24 h), 56.6 ± 30.6U (48 h), 61.4 ± 20.2U (72 h), 82.8 ± 24.2U (96 h) and 96.0 ± 15.3U (120 h). There was a significant difference between 72 h and 120 h (p = .007), but not between 96 h and 120 h (p > .99). By 96 h, all patients had AUC >31U, an accepted cutoff below which surgical bleeding risk is increased. Adenosine uptake showed no significant differences between the timepoints. These data suggest it takes 4 days for platelet reactivity to recover sufficiently after cessation of ticagrelor to avoid the excess risk of CABG-related bleeding. Discontinuing ticagrelor had no measurable effect on cellular adenosine uptake.

Circulating MicroRNA Levels Indicate Platelet and Leukocyte Activation in Endotoxemia Despite Platelet P2Y12 Inhibition.

There is evidence for the effects of platelet inhibition on innate immune activation. Circulating microRNAs (miRNAs) have been implicated as markers of platelet and leukocyte activation. In the present study, we assessed the effects of P2Y12 inhibitors on platelet and leukocyte miRNAs during endotoxemia. Healthy volunteers were randomly assigned to receive oral ticagrelor (n = 10), clopidogrel (n = 8) or no drug (n = 8) for one week, followed by an intravenous bolus of 2 ng/kg endotoxin. Serum was collected at baseline, after one week of antiplatelet treatment and 6 and 24 h after endotoxin administration. MiRNAs were screened using LNA-based qPCR, followed by TaqMan-qPCR validation of candidates. Clinical validation was performed in 41 sepsis patients. Platelet-enriched miR-197, miR-223 and miR-223* were decreased in volunteers following antiplatelet therapy. Endotoxin increased platelet miRNAs, whilst the opposite effect was seen for leukocyte-enriched miR-150. Neither of these endotoxin-mediated effects were altered by P2Y12 inhibitors. Sepsis patients with fatal outcomes (n = 12) had reduced miR-150 levels compared with survivors (n = 29). In conclusion, we show that miR-150 is downregulated in experimental endotoxemia and can predict survival in sepsis but is unaffected by P2Y12 inhibition. While P2Y12 inhibition reduces platelet-associated miRNAs in healthy volunteers, it fails to attenuate the response of platelet miRNAs to endotoxemia.

Study of Two Dose Regimens of Ticagrelor Compared With Clopidogrel in Patients Undergoing Percutaneous Coronary Intervention for Stable Coronary Artery Disease.

BACKGROUND: Ticagrelor has superior efficacy to clopidogrel in the management of acute coronary syndromes but has not been assessed in patients undergoing percutaneous coronary intervention for stable coronary artery disease. We compared the pharmacodynamic effects of ticagrelor and clopidogrel in this stable population. METHODS: One hundred eighty aspirin-treated stable coronary artery disease patients, who were planned to undergo elective percutaneous coronary intervention in a single center, were randomized 1:1:1 to either a standard clopidogrel regimen or 1 of 2 regimens of ticagrelor, either 90 mg (T90) or 60 mg twice daily (T60), both with a 180 mg loading dose. Cellular adenosine uptake was assessed, at the time of the procedure and pre- and postdose at 1 month, by adding adenosine 1 µmol/L to aliquots of anticoagulated whole blood and mixing with a stop solution at 0, 15, 30, and 60 seconds, then measuring residual plasma adenosine concentration by high-performance liquid chromatography. Systemic plasma adenosine concentration and platelet reactivity were assessed at the same timepoints. High-sensitivity troponin T was measured pre- and 18 to 24 hours postpercutaneous coronary intervention. RESULTS: One hundred seventy-four patients underwent an invasive procedure, of whom 162 received percutaneous coronary intervention (mean age 65 years, 18% female, 21% with diabetes mellitus). No effect on in vitro adenosine uptake was seen postdose at 1 month for either ticagrelor dose compared with clopidogrel (residual adenosine at 15 seconds, mean±SD: clopidogrel 0.274±0.101 µmol/L; T90 0.278±0.134 µmol/L; T60 0.288±0.149 µmol/L; P=0.37). Similarly, no effect of ticagrelor on in vitro adenosine uptake was seen at other timepoints, nor was plasma adenosine concentration affected (all P>0.1). Both maintenance doses of ticagrelor achieved more potent and consistent platelet inhibition than clopidogrel (VerifyNow P2Y12 reaction units, 1 month, mean±SD: predose, T60: 62±47, T90: 40±38, clopidogrel 181±44; postdose, T60: 34±30, T90: 24±21, clopidogrel 159±57; all P<0.0001 for ticagrelor versus clopidogrel). High platelet reactivity was markedly less with both T60 and T90 compared with clopidogrel (VerifyNow P2Y12 reaction units>208, 1 month postdose: 0%, 0%, and 21%, respectively). Median (interquartile range) high-sensitivity troponin T increased 16.9 (6.5-46.9) ng/L for clopidogrel, 22.4 (5.5-53.8) ng/L for T60, and 17.7 (8.1-43.5) ng/L for T90 (P=0.95). There was a trend toward less dyspnea with T60 versus T90 (7.1% versus 19.0%; P=0.09). CONCLUSIONS: Maintenance therapy with T60 or T90 had no detectable effect on cellular adenosine uptake at 1 month, nor was there any effect on systemic plasma adenosine levels. Both regimens of ticagrelor achieved greater and more consistent platelet inhibition than clopidogrel but did not appear to affect troponin release after percutaneous coronary intervention. CLINICAL TRIAL REGISTRATION: URL: https://www. CLINICALTRIALS: gov. Unique identifier: NCT02327624.

Very-low-dose twice-daily aspirin maintains platelet inhibition and improves haemostasis during dual-antiplatelet therapy for acute coronary syndrome.

Higher aspirin doses may be inferior in ticagrelor-treated acute coronary syndrome (ACS) patients and reducing bleeding risk whilst maintaining antithrombotic benefits could improve outcomes. We characterized the pharmacodynamics of a novel dual-antiplatelet-therapy regimen consisting of very-low-dose twice-daily (BD) aspirin with standard-dose ticagrelor. A total of 20 ticagrelor-treated ACS patients entered a randomized crossover to take aspirin 20 mg BD (12-hourly) during one 14-day period and 75 mg once-daily (OD) in the other. After 14 days of treatment, serum thromboxane (TX)B2 and light-transmittance aggregometry were assessed pre- and 2 h post-morning-dose, bleeding time was measured post-dose, and TXA2 and prostacyclin stable metabolites were measured in urine collected 2 h post-morning-dose. Data are expressed as mean ± SD. After 14 days treatment, serum TXB2 levels were significantly greater 2 h post-dosing with aspirin 20 mg BD vs. 75 mg OD (3.0 ± 3.6 ng/mL vs. 0.8 ± 1.9 ng/mL; p = 0.018) whereas pre-dosing levels were not significantly different (3.5 ± 4.1 ng/mL vs. 2.5 ± 3.1 ng/mL, p = 0.23). 1-mmol/L arachidonic acid-induced platelet aggregation was similarly inhibited by both regimens pre-dose (8.5 ± 14.3% vs. 5.1 ± 3.6%, p = 0.24) and post-dose (8.7 ± 14.2% vs. 6.6 ± 5.3%; p = 0.41). Post-dose bleeding time was shorter with 20 mg BD (680 ± 306 s vs. 834 ± 386 s, p = 0.02). Urinary prostacyclin and TX metabolite excretion were not significantly different. In conclusion, compared to aspirin 75 mg OD, aspirin 20 mg BD provided consistent inhibition of platelet TXA2 release and aggregation, and improved post-dose hemostasis, in ticagrelor-treated ACS patients. Further studies are warranted to assess whether this regimen improves the balance of clinical efficacy and safety.

Monitoring Antiplatelet Therapy.

The increasing use of antiplatelet therapy, particularly aspirin and oral P2Y12 inhibitors, in the prevention and management of arterial thrombosis, has stimulated extensive pharmacodynamic studies and research into tailored antiplatelet regimens. Many different methodologies have been studied for monitoring antiplatelet drugs and some are now well validated and used in clinical practice. However, clinical studies of tailored antiplatelet therapy have not convincingly demonstrated a benefit of this approach in patients treated with aspirin and clopidogrel, coupled with the fact that more potent antiplatelet therapies have more consistent effects compared with clopidogrel and so may reduce the rationale for monitoring. On the other hand, the optimum timing of urgent surgery after cession of oral antiplatelet therapy may be informed by platelet function testing. This review discusses the different methodologies that have been used to monitor the effects of antiplatelet therapy and highlights the current position of platelet function testing in clinical practice.

Platelet P2Y12 Inhibitors Reduce Systemic Inflammation and Its Prothrombotic Effects in an Experimental Human Model.

OBJECTIVE: Clinical studies suggest that platelet P2Y12 inhibitors reduce mortality from sepsis, although the underlying mechanisms have not been clearly defined in vivo. We hypothesized that P2Y12 inhibitors may improve survival from sepsis by suppressing systemic inflammation and its prothrombotic effects. We therefore determined whether clopidogrel and the novel, more potent P2Y12 inhibitor, ticagrelor, modify these responses in an experimental human model. APPROACH AND RESULTS: We randomized 30 healthy volunteers to ticagrelor (n=10), clopidogrel (n=10), or no antiplatelet medication (controls; n=10). We examined the effect of P2Y12 inhibition on systemic inflammation, which was induced by intravenous injection of Escherichia coli endotoxin. Both P2Y12 inhibitors significantly reduced platelet-monocyte aggregate formation and peak levels of major proinflammatory cytokines, including tumor necrosis factor α, interleukin-6, and chemokine (C-C motif) ligand 2. In contrast to clopidogrel, ticagrelor also significantly reduced peak levels of IL-8 and growth colony-stimulating factor and increased peak levels of the anti-inflammatory cytokine IL-10. In addition, ticagrelor altered leukocyte trafficking. Both P2Y12 inhibitors suppressed D-dimer generation and scanning electron microscopy revealed that ticagrelor also suppressed prothrombotic changes in fibrin clot ultrastructure. CONCLUSIONS: Potent inhibition of multiple inflammatory and prothrombotic mechanisms by P2Y12 inhibitors demonstrates critical importance of platelets as central orchestrators of systemic inflammation induced by bacterial endotoxin. This provides novel mechanistic insight into the lower mortality associated with P2Y12 inhibitors in patients with sepsis in clinical studies.

Cangrelor inhibits the binding of the active metabolites of clopidogrel and prasugrel to P2Y12 receptors in vitro.

Cangrelor is a rapid-acting, direct-binding, and reversible P2Y12 antagonist which has been studied for use during percutaneous coronary intervention (PCI) in patients with or without pretreatment with an oral P2Y12 antagonist. As cangrelor is administered intravenously, it is necessary to switch to an oral P2Y12 antagonist following PCI, such as the thienopyridines clopidogrel, and prasugrel or the non-pyridine ticagrelor. Previous studies have suggested a negative pharmacodynamic interaction between cangrelor and thienopyridines. This in vitro study evaluated the receptor-level interaction between cangrelor and the active metabolite (AM) of clopidogrel or prasugrel by assessing functional P2Y12 receptor number using a (33)P-2MeSADP binding assay. All P2Y12 antagonists studied resulted in strong P2Y12 receptor blockade (cangrelor: 93.6%; clopidogrel AM: 93.0%; prasugrel AM: 97.9%). Adding a thienopyridine AM in the presence of cangrelor strongly reduces P2Y12 receptor blockade by the AM (clopidogrel AM: 7%, prasugrel AM: 3.2%). The thienopyridine AMs had limited ability to compete with cangrelor for binding to P2Y12 (% P2Y12 receptor blockade after co-incubation with cangrelor 1000 nmol/L: 11.7% for clopidogrel AM 3 µmol/L; 34.1% for prasugrel AM 3 µmol/L). In conclusion, in vitro cangrelor strongly inhibits the binding of clopidogrel and prasugrel AMs to the P2Y12 receptor, consistent with the previous observation of a negative pharmacodynamic interaction. Care may need to be taken to not overlap exposure to thienopyridine AMs and cangrelor in order to reduce the risk of thrombotic complications following PCI.

PAR1 antagonists inhibit thrombin-induced platelet activation whilst leaving the PAR4-mediated response intact.

Thrombin-induced platelet activation is initiated by PAR1 and PAR4 receptors. Vorapaxar, a PAR1 antagonist, has been assessed in patients with acute coronary syndromes (ACS) and stable atherosclerotic disease in addition to standard-of-care treatment. In clinical trials, vorapaxar has been observed to reduce the frequency of ischaemic events in some subgroups though in others has increased the frequency of bleeding events. Among patients undergoing CABG surgery, which is associated with excess thrombin generation, bleeding was not increased. The aim of these studies was to investigate the effects of selective PAR1 antagonism on thrombin-induced platelet activation in patients receiving vorapaxar or placebo in the TRACER trial and to explore the roles of PAR1 and PAR4 in thrombin-induced platelet activation in healthy volunteers. ACS patients receiving vorapaxar or placebo in the TRACER trial were studied at baseline and 4 hours, 1 and 4 months during drug administration. Thrombin-induced calcium mobilisation in platelet-rich plasma was assessed by flow cytometry. In vitro studies were performed in healthy volunteers using the PAR1 antagonist SCH79797 or PAR4 receptor desensitisation. Vorapaxar treatment significantly inhibited thrombin-induced calcium mobilisation, leaving a residual, delayed response. These findings were consistent with calcium mobilisation mediated via the PAR4 receptor and were reproduced in vitro using SCH79797. PAR4 receptor desensitization, in combination with SCH79797, completely inhibited thrombin-induced calcium mobilisation confirming that the residual calcium mobilisation was mediated via PAR4. In conclusion vorapaxar selectively antagonises the PAR1-mediated component of thrombin-induced platelet activation, leaving the PAR4-mediated response intact, which may explain why vorapaxar is well tolerated in patients undergoing CABG surgery since higher thrombin levels in this setting may override the effects of PAR1 antagonism through PAR4 activation, thus preserving haemostasis. Further assessment may be warranted.

Evolving pattern of platelet P2Y12 inhibition in patients with acute coronary syndromes.

Dual antiplatelet therapy consisting of clopidogrel in addition to aspirin has previously been the standard of care for patients with acute coronary syndromes (ACS) but international guidelines have been evolving over the last 4 years with the introduction of prasugrel and ticagrelor. In October 2009, prasugrel was approved in the UK by the National Institute of Health and Clinical Excellence (NICE) for use in patients with ST-elevation myocardial infarction (STEMI) undergoing primary percutaneous coronary intervention (PCI), diabetic patients with non-ST-elevation (NSTE) ACS undergoing PCI and patients with stent thrombosis while other ACS patients were to continue receiving clopidogrel. Ticagrelor was approved in October 2011 by NICE for use in patients with moderate-to-high risk NSTE ACS and STEMI undergoing primary PCI and was recommended in preference to clopidogrel in European guidelines. These recommendations were adopted in our region, constituting a population of 1.8 million. We studied the effect of changing patterns of P2Y12 inhibitor usage on levels of platelet inhibition during maintenance therapy. Patients admitted to Northern General Hospital, Sheffield, with NSTE ACS or STEMI managed with primary PCI were enrolled over two periods of time: May 2010 to November 2011 (T1); and October 2012 to February 2013 (T2). Venous blood samples were obtained at 1 month after the onset of ACS. Light transmittance aggregometry (LTA) was performed and maximum aggregation response to ADP 20 µM was determined. A total of 116 patients were enrolled in T1 of whom 82 were receiving clopidogrel and 34 were receiving prasugrel. Twenty-nine patients were enrolled in T2, all of whom were receiving ticagrelor. Mean LTA results according to treatment with clopidogrel, prasugrel and ticagrelor were 57 ± 18%, 41 ± 20%, and 31 ± 12%, respectively. Prasugrel was associated with significantly lower platelet aggregation responses than clopidogrel (p < 0.001) and ticagrelor was associated with significantly lower platelet aggregation responses than both prasugrel (p = 0.015) and clopidogrel (p < 0.001). We conclude that international guidelines and NICE approval have led to increasing levels of P2Y12 inhibition in ACS patients in this UK centre between May 2010 and February 2013. Ticagrelor was associated with significantly greater P2Y12 inhibition than both clopidogrel and prasugrel during maintenance therapy.

Influence of body weight and body mass index on the chronic pharmacokinetic and pharmacodynamic responses to clinically available doses of ticagrelor in patients with chronic coronary syndromes.

Ticagrelor has multiple indications, including for some patients with chronic coronary syndromes (CCS) at high risk of ischaemic events. Body mass can potentially affect pharmacodynamics (PD) and pharmacokinetics (PK). We investigated the influence of body mass (range 53-172 kg, 20.8-46.9 kg/m2) on PD/PK in 221 CCS patients receiving ticagrelor 60 mg or 90 mg twice-daily (BD) during two randomised-controlled trials. Correlations between body weight (BW) or body mass index (BMI) and PD/PK measurements obtained during maintenance treatment at trough ('pre-dose') and peak effect ('post-dose') were assessed. BW and BMI correlated with P2Y12 reactivity units at pre-dose (e.g. BW:R = 0.26, p = 0.008) but not post-dose timepoints. BW affected ticagrelor active metabolite (TAM) levels (e.g. 60 mg BD, post-dose:R = -0.39, p < 0.0001) and there was evidence of an inverse power law relationship between BW and TAM-to-ticagrelor ratio. PK with ticagrelor 60 mg correlated significantly with BMI. BW and BMI did not affect the chance of high platelet reactivity, which remained very low across the whole cohort. There was no difference in PRU between the two doses of ticagrelor within each weight or BMI group. Body mass has modest effects on the PK/PD response to ticagrelor in patients with CCS but currently-used regimens appear adequate across the range of BW/BMI studied.

Ticagrelor effectively and reversibly blocks murine platelet P2Y12-mediated thrombosis and demonstrates a requirement for sustained P2Y12 inhibition to prevent subsequent neointima.

OBJECTIVE: Our goal was to study the effects of ticagrelor on murine platelet function and thrombosis and characterize the time course of P2Y(12) inhibition required to inhibit neointima formation following vascular injury. METHODS AND RESULTS: Mice were treated with ticagrelor or vehicle. Platelet aggregation and P-selectin expression were assessed over time, and thrombus formation was assessed in laser-injured cremasteric arterioles of P2Y(12)+/+ and P2Y(12)-/- mice. Neointima formation in FeCl(3)-injured carotid artery was assessed in C57BL/6 mice treated with different regimens of ticagrelor. Ticagrelor inhibited platelet aggregation and P-selectin expression in a dose-dependent, reversible manner. Ticagrelor inhibited thrombus formation to the same extent as seen in P2Y(12)-/- mice. Neointima formation was markedly reduced in mice treated with ticagrelor before and 4 hours after injury (neointima area: control, 39 921±22 749 µm(2), versus ticagrelor, 3705±2600 µm(2); P<0.01), whereas administration of ticagrelor either before injury only or from 4 hours postinjury was ineffective. CONCLUSIONS: Ticagrelor effectively and reversibly inhibits P2Y(12)-mediated platelet function and thrombosis in mice. P2Y(12) inhibition is required both at the time of and after injury to effectively inhibit neointima formation. Additional studies are warranted to evaluate the role of P2Y(12) inhibition in preventing restenosis.

The active metabolite of prasugrel effectively blocks the platelet P2Y12 receptor and inhibits procoagulant and pro-inflammatory platelet responses.

The aim of these studies was to investigate the extent of platelet P2Y(12) receptor inhibition by the thienopyridine active metabolite of prasugrel, R-138727. Blood was taken from healthy volunteers and pre-incubated with R-138727 or cangrelor (AR-C66931MX). Platelet aggregation was assessed in platelet rich plasma (PRP) and whole blood (WB). Vasodilator stimulated phosphoprotein (VASP) phosphorylation, platelet procoagulant activity (annexin V binding and microparticle formation) and calcium mobilisation were measured by flow cytometry. Platelet-leukocyte co-aggregate formation and sCD40L release, both pro-inflammatory responses of platelets, were measured by flow cytometry and ELISA, respectively. P2Y(12) receptor antagonism was determined using a radioligand binding assay ((33)P 2-MeSADP) in resting and stimulated platelets and the effects of clopidogrel administration were also assessed. R-138727 yielded concentration-dependent inhibition of platelet aggregation, VASP phosphorylation inhibition, procoagulant activity and pro-inflammatory responses. In the presence of R-138727 or cangrelor there was increased calcium reuptake following agonist stimulation. R-138727 30 micromol/L and cangrelor 1 micromol/L completely inhibited (33)P 2-MeSADP binding, compared to partial inhibition following clopidogrel administration. Platelet activation and granule secretion did not expose an additional pool of P2Y(12) receptors. Prasugrel's active metabolite effectively blocks the P2Y(12) receptor with the highest concentrations tested yielding complete inhibition of P2Y(12)-mediated amplification of several important platelet responses.

Pharmacodynamic Effects of a 6-Hour Regimen of Enoxaparin in Patients Undergoing Primary Percutaneous Coronary Intervention (PENNY PCI Study).

Delayed onset of action of oral P2Y12 inhibitors in ST-elevation myocardial infarction (STEMI) patients may increase the risk of acute stent thrombosis. Available parenteral anti-thrombotic strategies, to deal with this issue, are limited by added cost and increased risk of bleeding. We investigated the pharmacodynamic effects of a novel regimen of enoxaparin in STEMI patients undergoing primary percutaneous coronary intervention (PPCI). Twenty patients were recruited to receive 0.75 mg/kg bolus of enoxaparin (pre-PPCI) followed by infusion of enoxaparin 0.75 mg/kg/6 h. At four time points (pre-anti-coagulation, end of PPCI, 2-3 hours into infusion and at the end of infusion), anti-Xa levels were determined using chromogenic assays, fibrin clots were assessed by turbidimetric analysis and platelet P2Y12 inhibition was determined by VerifyNow P2Y12 assay. Clinical outcomes were determined 14 hours after enoxaparin initiation. Nineteen of 20 patients completed the enoxaparin regimen; one patient, who developed no-reflow phenomenon, was switched to tirofiban after the enoxaparin bolus. All received ticagrelor 180 mg before angiography. Mean (± standard error of the mean) anti-Xa levels were sustained during enoxaparin infusion (1.17 ± 0.06 IU/mL at the end of PPCI and 1.003 ± 0.06 IU/mL at 6 hours), resulting in prolonged fibrin clot lag time and increased lysis potential. Onset of platelet P2Y12 inhibition was delayed in opiate-treated patients. No patients had thrombotic or bleeding complications. In conclusion, enoxaparin 0.75 mg/kg bolus followed by 0.75 mg/kg/6 h provides sustained anti-Xa levels in PPCI patients. This may protect from acute stent thrombosis in opiate-treated PPCI patients who frequently have delayed onset of oral P2Y12 inhibition.

Consistent platelet inhibition with ticagrelor 60 mg twice-daily following myocardial infarction regardless of diabetes status.

Diabetes increases cardiovascular risk and reduces pharmacodynamic response to some oral antiplatelet drugs. This study aimed to determine whether ticagrelor 60 mg twice daily (bid) provided potent and consistent platelet inhibition in patients with vs without diabetes in the PEGASUS-TIMI 54 platelet function substudy. Out of 180 patients studied, 58 patients were randomised to and had received at least four weeks of ticagrelor 60 mg bid, with 20 (34 %) having diabetes, 58 patients received ticagrelor 90 mg bid, with 12 (21 %) having diabetes, and 64 patients received placebo, with 18 (28 %) having diabetes. Blood was sampled pre- and 2 hours post-maintenance dose. In patients treated with ticagrelor 60 mg bid, on-treatment platelet reactivity to ADP, as determined by light transmission aggregometry (LTA), VerifyNow and VASP, was similar in patients with vs without diabetes (LTA post-dose, ADP 20 µM: 29 ± 14 vs 34 ± 10 %, respectively; p = 0.19). A consistent inhibitory effect of ticagrelor 60 mg bid was observed pre- and post-dose regardless of diabetes status, even in insulin-treated patients. Patients with diabetes did not have an increased incidence of high platelet reactivity in either ticagrelor group. Platelet reactivity was similar in patients with diabetes treated with ticagrelor 60 mg vs 90 mg bid. Pharmacokinetics of ticagrelor were not affected by diabetes status. In conclusion, ticagrelor 60 mg bid is equally effective at reducing platelet reactivity in patients with and without diabetes, yielding a consistently high level of platelet inhibition regardless of diabetes status.

Utility of a whole blood single platelet counting assay to monitor the effects of tirofiban in patients with acute coronary syndromes scheduled for coronary intervention.

This study aimed to establish the utility of a whole-blood single-platelet counting (WBSPC) assay, a measure of microaggregation, in monitoring the effects of tirofiban, comparing this with optical aggregometry (OA) and the Ultegra TRAP cartridge system (UTC), measures of macroaggregation. Fifty-nine patients with acute coronary syndrome scheduled for coronary angiography +/- angioplasty were studied. WBSPC assay (ADP 0.3-100 microM, Sysmex KX21 analyzer), OA (ADP 20 microM) and UTC were performed: before starting tirofiban; 30 min, 4 and 24 h after starting tirofiban; and 1 and 2 h after stopping tirofiban. Thirty minutes after starting tirofiban, there was substantial inhibition of platelet aggregation (40 +/- 30%; WBSPC, 2 minutes after addition of ADP 30 microM) and this remained stable at 4 and 24 h. OA (86 +/- 17%; inhibition of maximal aggregation, ADP 20 microM) and UTC (93 +/- 7%) showed marked inhibition with less inter-individual variation. There was no significant correlation between OA and UTC results (R(2) = 0.006), but fair correlation between OA and WBSPC results (R(2) = 0.37). Greater inhibition of macroaggregation (OA and UTC) was seen compared to microaggregation (WBSPC) such that WBSPC was more discriminating in the therapeutic range when macroaggregation was often completely inhibited. A WBSPC assay of platelet microaggregation shows promise for monitoring GPIIb/IIIa antagonists.

Morphine delays the onset of action of prasugrel in patients with prior history of ST-elevation myocardial infarction.

Delays in the onset of action of prasugrel during primary percutaneous coronary intervention (PPCI) have been reported and could be related to the effects of morphine on gastric emptying and subsequent intestinal absorption. The study objective was to determine whether morphine delays the onset of action of prasugrel in patients with a prior history of ST-elevation myocardial infarction (STEMI) treated with PPCI. This was a crossover study of 11 aspirin-treated patients with prior history of STEMI treated with PPCI, for which prasugrel and morphine had been previously administered. Patients were randomised to receive either morphine (5 mg) or saline intravenously followed by 60 mg prasugrel. Blood samples were collected before randomised treatment and over 24 hours after prasugrel administration. The inhibitory effects of prasugrel on platelets were determined using the VerifyNow P2Y12 assay and light transmission aggregometry. Plasma levels of prasugrel and prasugrel active metabolite were measured. Platelet reactivity determined by VerifyNow PRU, VerifyNow % Inhibition and LTA was significantly higher at 30-120 minutes (min) when morphine had been co-administered compared to when saline had been co-administered. Morphine, compared to saline, significantly delayed adequate platelet inhibition after prasugrel administration (158 vs 68 min; p = 0.006). Patients with delayed onset of platelet inhibition also had evidence of delayed absorption of prasugrel. In conclusion, prior administration of intravenous morphine significantly delays the onset of action of prasugrel. Intravenous drugs may be necessary to reduce the risk of acute stent thrombosis in morphine-treated STEMI patients undergoing PPCI.

Platelet Inhibition With Ticagrelor 60 mg Versus 90 mg Twice Daily in the PEGASUS-TIMI 54 Trial.

BACKGROUND: The PEGASUS-TIMI 54 (Prevention of Cardiovascular Events in Patients with Prior Heart Attack Using Ticagrelor Compared to Placebo on a Background of Aspirin-Thrombolysis In Myocardial Infarction 54) trial studied 2 doses of ticagrelor, 90 mg twice a day (bid) and 60 mg bid, for long-term prevention of ischemic events in patients with prior myocardial infarction. Both doses similarly reduced the rate of ischemic events versus placebo. The pharmacokinetics and pharmacodynamics of ticagrelor 60 mg bid have not been studied. OBJECTIVES: In this study, the authors sought to study the pharmacokinetics and pharmacodynamics for ticagrelor 60 mg compared with 90 mg bid. METHODS: A total of 180 patients who received >4 weeks of study medication had blood sampling in the morning pre-maintenance dose and again 2 h post-dose. All patients received aspirin. Plasma levels of ticagrelor and its active metabolite AR-C124910XX were determined. P2Y12 inhibition was assessed by the VerifyNow P2Y12 assay (Accumetrics, Inc., San Diego, California) (P2Y12 reaction units [PRU]), light transmittance aggregometry (adenosine diphosphate 5 and 20 µmol/l and arachidonic acid), and vasodilator-stimulated phosphoprotein phosphorylation assays. VerifyNow Aspirin assays and serum thromboxane B2 measurements were performed. RESULTS: Mean pre- and post-dose plasma levels of ticagrelor were 35% and 38% lower, respectively, with 60 mg versus 90 mg. Both doses achieved high levels of platelet inhibition pre- and post-dose, with numerically slightly more variability with 60 mg: mean (SD) pre-dose PRU values were 59 ± 63 and 47 ± 43 for ticagrelor 60 and 90 mg, respectively (p = 0.34). High platelet reactivity, determined as PRU >208, was rare with the 60-mg pre-dose and was absent post-dose. Platelet reactivity pre- and post-dose, as measured by light transmittance aggregometry or vasodilator-stimulated phosphoprotein assays, was numerically but not significantly lower with 90 mg than with 60 mg. Aspirin response was not affected by either dose. CONCLUSIONS: Ticagrelor 60 mg bid achieved high levels of peak and trough platelet inhibition in nearly all patients, similar to that with 90 mg bid, helping to explain the efficacy of the lower ticagrelor dose in PEGASUS-TIMI 54.

Agonists of toll-like receptor (TLR)2 and TLR4 are unable to modulate platelet activation by adenosine diphosphate and platelet activating factor.

Inappropriate platelet activation is a feature of acute and chronic diseases such as disseminated intravascular coagulation (DIC) and atherosclerosis. Since proinflammatory microbial-derived agonists can be involved in the pathogenesis of these diseases, we examined the potential role of TLR4 (mediating responses to LPS) and TLR2 (which responds to bacterial lipopeptides) in platelet activation. Our data suggested low-level expression of TLR2 and TLR4 on platelets, determined by flow cytometry, and we also observed expression of TLR4 on a megakaryocytic cell line by both flow cytometry and immunohistochemistry. Stimulation of the platelets with the TLR4 agonist LPS, and the synthetic TLR2 agonist Pam3CSK4, resulted in no platelet aggregation, no increase in CD62P surface expression and no increase in the cytosolic concentration of Ca2+. The TLR agonists were also unable to directly activate platelets primed with epinephrine, or pretreated with a low concentration of ADP or PAF. Pretreatment of platelets with LPS or Pam3CSK4 also failed to modulate the platelet response to submaximal concentrations of the classical platelet agonists ADP and PAF. We conclude that the TLR agonists LPS and Pam3CSK4 have no direct effect on platelet activation and that platelet TLRs may be a remnant from megakaryocytes. TLR2 and TLR4 agonists are thought to have a significant role in diseases such as atherosclerosis and DIC, but our research suggests that this is through a mechanism other than direct platelet activation or by modification of platelet responses to other agonists.