Quantifying GPIIb/IIIa Receptor Binding Using 2 Monoclonal Antibodies

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Abstract
Background —Dosing of glycoprotein (GP) IIb/IIIa receptor antagonists is frequently based on the inhibition of platelet aggregation, which may be influenced by the agonist used or concurrent medications. Here we describe a monoclonal antibody-based technique to quantify total and ligand-occupied GPIIb/IIIa receptors. Methods and Results —In vitro binding of monoclonal antibodies, LYP18 (Mab1) and 4F8 (Mab2), to the GPIIb/IIIa complex, was characterized using purified receptor and to platelets by flow cytometry. Patients undergoing coronary angioplasty received a single 20 mg dose of the oral GPIIb/IIIa antagonist, xemilofiban, or matching placebo, and antibody binding was compared with inhibition of platelet aggregation. Mab1 and Mab2 were bound to purified GPIIb/IIIa and to unoccupied, inactivated receptor on platelets. Mab2 identified the β3 subunit, whereas Mab1 was complex-specific. Neither antibody interfered with the other’s binding, suggesting that they identified distinct sites. Mab1 identified 53 300±5423 GPIIb/IIIa sites per platelet, whereas Mab2 identified 50 120±5066 sites per platelet. Mab1 binding was inhibited by abciximab in a dose dependent manner (IC 50 , 0.85±0.1 μg/mL), whereas Mab2 binding was unaffected. In contrast, the 2 small molecular weight antagonists, SC-57101A (IC 50 , 0.22±0.06 μmol/L) and eptifibatide (IC 50 , 0.35±0.14 μmol/L) inhibited Mab2 but not Mab1 binding. In patients treated with xemilofiban, Mab1 binding was unaltered but Mab2 binding decreased from 37 930±2061 sites per platelet at baseline to 8318±870 sites per platelet 6 hours after dosing ( P P Conclusions —Mab1 and Mab2 bind to GPIIb/IIIa and are differentially displaced by abciximab and small molecular weight antagonists. These antibodies may be used to monitor receptor number and occupancy during administration of a GPIIb/IIIa antagonist.

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