Variation in platelet response to thrombin may affect the safety and efficacy of PAR antagonism. The Thr120 variant of the common single nucleotide polymorphism (SNP) rs773902 in the protease-activated receptor (PAR) 4 gene is associated with higher platelet aggregation compared to the Ala120 variant. We investigated the relationship between the rs773902 SNP with major bleeding and ischemic events, safety, and efficacy of PAR1 inhibition in 6177 NSTE ACS patients in the TRACER trial. There was a lower rate of GUSTO moderate/severe bleeding in patients with the Thr120 variant. The difference was driven by a lower rate in the smaller homozygous group (recessive model, HR 0.13 [0.02–0.92] P = 0.042). No significant differences were observed in the ischemic outcomes. The excess in bleeding observed with PAR1 inhibition was attenuated in patients with the Thr120 variant, but the interactions were not statistically significant. In summary, lower major bleeding rates were observed in the overall TRACER cohort with the hyperreactive PAR4 Thr120 variant. The increase in bleeding with vorapaxar was attenuated with the Thr120 variant, but we could not demonstrate an interaction with PAR1 inhibition. These findings warrant further exploration, including those of African ancestry where the A allele (Thr120) frequency is ~65%.

Very low–dose (VLD) factor Xa (FXa) inhibition, in combination with acetylsalicylic acid (ASA) and clopidogrel, is associated with improved outcomes in patients with acute coronary syndrome (ACS) with a tolerable bleeding risk profile. To date, there are no data documenting platelet inhibition and the anticoagulatory effects of VLD FXa inhibition on top of guidelineadherent dual-antiplatelet therapy (DAPT) in patients with ACS. Patients with non–ST-elevation myocardial infarction (NSTEMI) receiving oral DAPT (ASA 1 clopidogrel, n 5 20; or ASA 1 ticagrelor, n 5 20) were prospectively enrolled in a nonrandomized study. Coagulation- and platelet-dependent thrombin generation (TG), measured by means of the calibrated automated thrombogram, were significantly decreased after in vitro and in vivo addition of rivaroxaban. As shown by a total thrombus-formation analysis approach, rivaroxaban treatment led to a significantly decreased coagulation-dependent (AR-chip) thrombus formation in patients treated with ASA plus P2Y12 inhibitor (clopidogrel/ticagrelor), whereas the pure platelet-dependent (PL-chip) thrombus formation was not affected at all. Adjunctive rivaroxaban therapy was not associated with significant differences in platelet aggregation assessed by light-transmission aggregometry (LTA). Nevertheless, according to fluorescence-activated cell sorter analysis, VLD rivaroxaban treatment resulted in a significantly reduced expression of platelet HMGB-1, whereas P-selectin exposure was not affected. Furthermore, an enhanced effect of rivaroxaban on total thrombus formation and TG was observed in particular in clopidogrel nonresponder patients defined as adenosine 59-diphosphate-induced LTA $40%. VLD rivaroxaban reduces thrombus formation and platelet-dependent TG in patients with ACS receiving DAPT, which can be of potential ischemic benefit. This trial was registered at www.clinicaltrials.gov as #NCT01417884.

We investigated the effect of sequential treatment escalation with dapagliflozin and saxagliptin on beta cell function in patients with T2DM insufficiently controlled on metformin monotherapy during a hyperglycaemic clamp investigation. Twenty-six patients (19 males, age 63.5±7.0 years; duration of diabetes 8.8±4.7 years; HbA1c 63.9±15.8 mmol/mol; mean±SD) were enrolled in the study. During a first treatment period (TP1) all patients received 10 mg dapagliflozin for one month, followed by the addition of 5 mg saxagliptin or placebo for another month (TP2). At baseline and at the end of each treatment period, fasting glucose and insulin levels were analysed, and a hyperglycaemic clamp with the measurement of plasma C-peptide, insulin, proinsulin, and glucagon was performed. 

The platelet P2Y12 receptor is involved in ADP mediated platelet activation. A variety of methods and assays are available to measure P2Y12 function, such as the VerifyNow® (Instrumentation Laboratory), light transmission aggregometry (LTA), and the Cy-Quant VASP/P2Y12 ELISA (Stago). The purpose of this study was to evaluate suitability of batch processing of samples and to determine effects of sample stability for the Cy-Quant VASP/P2Y12 ELISA method.

• factors that contribute toward inadequate antiplatelet drug responsiveness;
• personalizing antiplatelet therapy to improve clinical outcomes;
• the limitations of platelet function testing;
• what is the optimal timing for platelet function testing: at the time of ACS or post-treatment;
• the role of ROTEM and TEG in addressing clot integrity and platelet function; and
• the current role for platelet function testing in evaluating patients and outcomes.

Reliable shipment of clinical blood, serum and plasma samples from remote sites to central labs for standardized analyses is increasingly important in medicnal product trials. This study investigates the benefits of dedicated packaging for protecting such samples from extreme seasonal temperature fluctuations during shipment.

In this paper we describe the thermal properties of MLM's Safeguard Box which was tested with respect to the stability of real samples under stress conditions.

Thrombin is the final product of the intrinsic and extrinsic pathways, converting soluble fibrinogen into fibrin. It also serves as a potent activator of platelet function. As a key enzyme in hemostasis and thrombosis, it is imperative that reliable monitoring of thrombin be performed in study subjects during pharmacologic safety trials, and in patients dosed with hemostasis-modulating medication.

Circulating stimuli, including inflammatory cytokines and some therapeutics, activate platelets and endothelial cells. Measurement of circulating mciroparticles serves as an indicator of in vivo vascular function, and in clinical studies is used to identify risk of thrombosis and bleeding.

Aggregometry remains the gold standard for evaluating platelet function due to the capability to tailor testing and to adequately assess drug dose response. It is the reference method used clinically to test for platelet disorders.

Flow cytometry is a sensitive technique for assessing specific components of platelet function, including degranulation, receptor signaling and surface protein activation.

Increasing evidence implicates platelets as key cellular mediators of inflammation and vascular disease. Platelets express and secrete a variety of molecules and inflammatory mediators, such as platelet factor 4, P-selectin, CD40L, vWF, CD63, thrombospondins, MMPs, polyphosphates, ADP, thromboxane, nitric oxide, IL-1β and GPIb. Many of these mediators promote recruitment of leukocytes to sites of endothelial damage or tissue inflammation and this pathway likely contributes to pathogenesis of inflammatory and cardiovascular diseases, such as atherosclerosis, diabetes, and rheumatoid arthritis. 

Platelets are the intended, and sometimes unintended, targets of countless therapeutics. There is also an increasing appreciation that pathologies such as coronary artery disease, diabetes, and cancer alter platelet function. 

Inflammation, endothelial damage, and platelet activation trigger secondary hemostasis, comprising initiation and propagation of the coagulation cascade and the formation of insoluble fibrin to consolidate and stabilize the thrombus.

This paper on the new FDA draft validation guidelines is the second from MLM.For many years, the FDA guidance on bioanalytical method validation (BMV) issued by the CDER in 20011, has been the Holy Grail for laboratories which deal with the pharmacokinetic analysis of drugs and their metabolites in clinical trials. A revised version has been expected at least since the EMA issued its guideline on BMV in 20122, which is much more explicit and detailed in its requirements compared to the FDA guidance from 2001. The new FDA draft guidance is summarised in this editorial, and its implications for clinical studies are discussed.