In vitro measurement of platelet glycoprotein IIb/IIIa receptor blockade by abciximab: interindividual variation and increased platelet secretion

Background and Objectives. Inhibition of soluble fibrinogen binding to acti vated platelets represents the target of pharmacologic approach with antago nists of the glyco-protein IIb/IIIa (GPIIb/IIIa) complex. In this study we assessed the effects of abciximab, a recombinant chimeric Fab fraction of t he antibody against GPIIb/IIIa, on several markers of platelet activation. Design and Methods. The platelet surface expression of GPIIb/IIIa was measu red by a flow cytometry technique using a two-colour assay. GPIIb/IIIa was detected by FITC-conjugated antibodies in whole blood, either unstimulated or exposed to platelet stimuli. The following antibodies were used: CD41, w hich recognizes the IIb/IIIa complex both in activated and non-activated co nformers, and PAC-1, which is directed toward the activated conformer of GP IIb/IIIa. In addition, the same blood sample was incubated with CD62 antibo dy to measure P-selectin, as a marker of a-granule degranulation. The effec t of abciximab was also assessed by experiments carried out on shear stress -induced platelet aggregation, a test that appears to be a predictor of pla telet hemostatic function. Results. Abciximab inhibited CD41 binding to glycoprotein IIb (GPIIb) in a concentration-dependent manner and also inhibited the binding of PAC-1 to a ctive GPIIb/IIIa. In contrast, membrane-associated P-selectin was significa ntly increased by the drug, which suggests that blockade of GPIIb/IIIa rece ptors results in an increased platelet degranulation in response to agonist s. Shear stress-induced platelet aggregation was inhibited by abciximab, wi th a more pronounced effect on blood filtration, which represents an index of platelet aggregate formation. Interpretation and Conclusions. Our results indicate that GPIIb/IIIa blocka de by abciximab is accompanied by an increase of a-granule secretion, sugge sting that different mechanisms regulate these aspects of platelet activati on. The described flow cytometry technique, that allows the simultaneous in vitro detection of several platelet markers, is a suitable method for asse ssing the effects of agents which interfere with platelet function. (C) 200 1, Ferrata Storti Foundation.