Person responsible (Chair): Martine Jandrot-Perrus, Hans Deckmyn

Transcription

1 NAME OF PROJECT: Measurement of platelet activation markers: usefulness, current practices, future. Subcommittee: Platelet physiology Person responsible (Chair): Martine Jandrot-Perrus, Hans Deckmyn Design: Questionnaire survey Aim/Objective/Rationale (Needs assessment / Reason) The purpose of the survey is to come to a consensus on which biomarkers are considered as most useful/promising such that the survey can serve as an incentive to whoever to start building an easy robust test system to finally allow standardized larger studies by multiple research/clinical teams to proof or disproof the usefulness of particular markers. Determination of platelet activation markers is widely used in clinical studies with several objectives: first of all, of course, for the diagnosis, prognosis and therapeutic follow-up of prothrombotic diseases but also in a growing number of additional situations in view of the growing awareness of the involvement of platelets in multiple processes such as inflammation, repair, immune defense and cancer. Platelet activation markers is thus a generic term that encompasses platelet responses as a whole despite their extreme heterogeneous nature. However, at present there is no consensus about the usefulness of these individual markers. Hence it appears increasingly unlikely that the measurement of a single marker is sufficient to qualify the activation of platelets in vivo, however combinations of markers could allow for activation scores for each type of response. Before any attempt is made to standardize these signatures, it is necessary to make an overview, with a panel of experts, of the markers already classically used and those that would, according to them, deserve to be used more frequently, as well as the clinical objectives of these measures. The first aim is to identify biomarkers of platelet activation (BPA) helpful for patients (prevention, diagnosis, prognostic and treatment of thrombotic events). Ideal markers of in vivo platelet activation, capable to orientate physicians towards a diagnosis, treatment and predictable outcome, should be specific, unaltered by preclinical artifacts and reproducibly measured by easily performed methods The second aim is to eventually encourage the development of rapid, simple and point of care methods since a major limitation of the current markers is the feasibility of their measurement at any given time. In fact the BPA measurements should not be limited to a few academic health centers. Development of multiplexed BPA assays would be eventually encouraged.

2 Background BPA should preferentially be measured as soluble in the plasma or as platelet associated. Urinary tests could be considered despite the difficulties inherent in their implementation and of the delay in their measurement. Soluble markers are suitable for large epidemiological studies in view of the storage possibilities and include the content from dense, alpha and lysosomal granules, the secretion of which is differentially regulated depending on the nature and level of the stimulation; the secretion of TXA2 that reflects a synthesis activity, and the products of receptor shedding (amongst which sgpib (glycocalicin), sgpv, sgpvi, sp-selectin, scd40l). Elevated plasma concentrations of these soluble compounds could reflect an in vivo ongoing thrombotic state and or a contribution to an inflammatory, reparation or defense process. However, they could also reflect inappropriate in vitro release. This is why the combined measurement of markers having different half lifes may be useful to qualify the sample (such as PF4 and β-tg that have respectively a short and long half life). Platelet surface markers usually are measured by flow cytometry on fresh blood samples and thus are not possible 7/7, 24/24. However, improvement of fixation and preservation methodologies as well as of appropriate antibodies (capable e.g. to bind to fixed antigens) would be helpful for the use of these markers in large epidemiological studies. The most commonly used are determination of integrin αiibβ3 activation, fibrin(ogen) binding, and P-selectin, CD63 and phosphatidylserine exposure. They all are markers of strongly activated platelets suggesting an ongoing thrombotic state but their combination could have a different significance with respect to both the platelet activity (secretory, aggregating, procoagulant, scaffolding fibrin or even exhausted platelets) as well as to the agonist that triggered the response (thrombin, collagen, VWF, fibrin, either individually or in combination). New markers capable to detect platelet prone to be activated (primed platelets) in addition might be of great interest. Some preliminary studies suggest that the level of GPVI dimerization could be such a marker. Platelet-leukocyte aggregates may also be a sensitive indicator of primed, non aggregating platelets particularly in inflammatory states. Phosphorylated/dephosphorylated signaling proteins could also be of interest as BPA independently of the controversy on the usefulness of the VASP phosphorylation assay. Method A questionnaire survey will be performed to ask different laboratories whether they use platelet activation markers and to what clinical purpose or situation. Expected timeline: o Project stage/set up: planning o Launch: ISTH Berlin meeting, 2017 o Duration: 2 years o Finalization/analysis: 2019 o Reporting: ISTH Melbourne meeting, 2019

3 Expected outcomes (ie. publications): o Publication type (SSC Communication, Guidance document or original article): SSC communication (survey) References: Role of platelet function testing in clinical practice: Sibbing D et al Curr drug targets 2011; 155:30-44 Biomakers of platelet activation in acute coronary syndromes. Ferroni P et al. Thromb Haemost 2012;108: Platelet release of beta-thromboglobulin and platelet factor 4 and serotonin in plasma samples. Ohkawa R et al. Clin Biochem 2005; 38: Hypercoagulability, platelet function, inflammation and coronary artery disease: results of the Thrombotic RIsk progression (TRIP) study. Tantry US et al Platelets 2010;21: Platelet receptor shedding. Gardiner EE et al Methods Mol Biol. 2012;788: Regulation of platelet membrane protein shedding in health and disease. Au AE and Josefsson EC. Platelets Aug 5:1-12 Platelet glycoprotein VI, an active process inducing receptor competence is an indicator of platelet reactivity. Loyau S et al. Arterioscler Thromb Vasc Biol 2012;32:

4 Survey proposal (this will be presented in the form of an electronic web-questionnaire) Usefulness of determining biomarkers of platelet activation (BPA) a) Determination of BPA is useful for the study of subjects at risk of thrombosis for prognostic and/or prevention purposes b) Determination of BPA is useful for the study of subjects with thrombosis for treatment follow-up Determination of the following BPA is useful 1. Platelet surface markers: 1.1. Integrin alphaiibbeta3 activation 1.2. Fibrin(ogen) binding 1.3. P-selectin exposure 1.4. Phosphatidylserine exposure 2. Soluble markers 2.1. Dense granule: serotonin/5-hydroxytryptamine (5-HT) 2.2. Alpha granule: Platelet factor 4 (PF4)

5 β-thromboglobulin (β-tg) thrombospondin-1 (TSP-1) SCUBE TXA2 production Plasma TXB Plasma 11-dehydro-TXB Urinary 2,3 dinor TXB receptor shedding: glycocalicin (sgpib) sgpvi sgpv sp-selectin

6 scd40 ligand (scd40-l) 2.5. platelet-leukocyte aggregates 2.6. platelet-derived microparticles 3. Other 3.1. VASP phosphorylation 3.2. Please fill in other biomarkers that should be considered and indicate wherever possible whether this applies for a) and/or b)

7 4. Possible useful biomarker determination combinations Comb 1:. for a b Comb 2:. for a b Comb 3:. for a b.. for a b