title: Flow cytometric analysis of intraplatelet VASP phosphorylation for the detection of clopidogrel resistance in patients with ischemic cardiovascular diseases.
abstract: Interindividual variability of the inhibitory effect of clopidogrel on platelet functions leading to clopidogrel resistance has been described in some patients with ischemic cardiovascular disease. A reliable laboratory test is therefore needed to identify patients insufficiently protected by this antiplatelet treatment. The phosphorylation of vasodilator-stimulated phosphoprotein (VASP), an intraplatelet actin regulatory protein, is dependent on the level of activation of the platelet P2Y12 receptor, which is targeted by clopidogrel. The aim of this study was to use a flow cytometric VASP phosphorylation assay to evaluate the efficacy of clopidogrel therapy. The platelet reactivity index (PRI), expressed as a percentage, is the difference in VASP fluorescence intensity between resting (+PGE1) and activated (+ADP) platelets. In vitro, the PRI was strongly correlated with the inhibition of platelet aggregation induced by specific blockade of the P2Y12 receptor by the competitive antagonist AR-C69931MX (R = 0.72, P < 0.0001). Ex vivo, the PRI was 78.3 +/- 4.6% in 47 healthy donors, 79.0 +/- 4.1% in 34 patients not receiving clopidogrel and 61.1 +/- 17.0% in 33 patients treated with clopidogrel (P < 0.0001). In the clopidogrel group, the PRI values were widely dispersed (from 6.6 to 85.8%) and more than 30% of these patients had a PRI equivalent of values in patients not receiving clopidogrel. The flow cytometric analysis of VASP phosphorylation seems to be a suitable test to evaluate the efficacy of clopidogrel treatment. This assay demonstrated a wide interindividual variability of the inhibitory response of platelets to clopidogrel and showed that one-third of the patients treated appeared to be 'unprotected' by this therapy.
authors.affiliation: Institut National de la Santé et de la Recherche Médicale U.311, Etablissement Français du Sang, Alsace, Strasbourg, France.
authors.forename: B
authors.initials: B
authors.lastname: Aleil
authors.forename: C
authors.initials: C
authors.lastname: Ravanat
authors.forename: J P
authors.initials: JP
authors.lastname: Cazenave
authors.forename: G
authors.initials: G
authors.lastname: Rochoux
authors.forename: A
authors.initials: A
authors.lastname: Heitz
authors.forename: C
authors.initials: C
authors.lastname: Gachet
mesh_terms: D000199:Actins / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism; D000249:Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D000328:Adult; D000368:Aged; D001792:Blood Platelets / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000097:blood*; D000077144:Clopidogrel; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D008840:Microfilament Proteins; D008875:Middle Aged; D017202:Myocardial Ischemia / Q000097:blood* / Q000175:diagnosis*; D010750:Phosphoproteins / Q000097:blood*; D010766:Phosphorylation; D010974:Platelet Aggregation; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D018048:Receptors, Purinergic P2 / Q000378:metabolism; D058925:Receptors, Purinergic P2Y12; D013988:Ticlopidine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D013997:Time Factors; D000098662:Vasodilator-Stimulated Phosphoprotein
chemical_list: D000199:Actins; D015815:Cell Adhesion Molecules; D008565:Membrane Proteins; D008840:Microfilament Proteins; C548576:P2RY12 protein, human; D010750:Phosphoproteins; D010975:Platelet Aggregation Inhibitors; D018048:Receptors, Purinergic P2; D058925:Receptors, Purinergic P2Y12; D000098662:Vasodilator-Stimulated Phosphoprotein; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; C117446:cangrelor; D000077144:Clopidogrel; D013988:Ticlopidine