Eptifibatide-induced thrombocytopenia and thrombosis in humans require FcγRIIa and the integrin β3 cytoplasmic domain

Abstract

Thrombocytopenia and thrombosis following treatment with the integrin αIIbβ3 antagonist eptifibatide are rare complications caused by patient antibodies specific for ligand-occupied αIIbβ3. Whether such antibodies induce platelet clearance by simple opsonization, by inducing mild platelet activation, or both is poorly understood. To gain insight into the mechanism by which eptifibatide-dependent antibodies initiate platelet clearance, we incubated normal human platelets with patient serum containing an αIIbβ3-specific, eptifibatide-dependent antibody. We observed that in the presence of eptifibatide, patient IgG induced platelet secretion and aggregation as well as tyrosine phosphorylation of the integrin β3 cytoplasmic domain, the platelet FcγRIIa Fc receptor, the protein-tyrosine kinase Syk, and phospholipase Cγ2. Each activation event was inhibited by preincubation of the platelets with Fab fragments of the FcγRIIa-specific mAb IV.3 or with the Src family kinase inhibitor PP2. Patient serum plus eptifibatide did not, however, activate platelets from a patient with a variant form of Glanzmann thrombasthenia that expressed normal levels of FcγRIIa and the αIIbβ3 complex but lacked most of the β3 cytoplasmic domain. Taken together, these data suggest a novel mechanism whereby eptifibatide-dependent antibodies engage the integrin β3 subunit such that FcγRIIa and its downstream signaling components become activated, resulting in thrombocytopenia and a predisposition to thrombosis.

The pyrrolidinoindoline alkaloid Psm2 inhibits platelet aggregation and thrombus formation by affecting PI3K/Akt signaling

“…The activation of the GPIIb/IIIa receptor is a common final step of platelet aggregation in response to various agonists, after which αIIbβ3 binds to soluble fibrinogen and bridges platelets, leading to platelet aggregation [34] . Antagonists of the GPIIb/IIIa receptor inhibit platelet aggregation induced by various agonists by preventing fibrinogen from binding to activated GPIIb/IIIa receptors [35] . Psm2 inhibited ADP-, thrombin-, U46619-, and collagen-induced platelet aggregation and blocked platelet adhesion to immobilized fibrinogen and collagen without affecting the binding of the GPIIb/IIIa receptor to fibrinogen, which suggested that Psm2 affects inside-out platelet signaling.…”

Approach to the Diagnosis and Management of Drug-Induced Immune Thrombocytopenia

“…The mechanism of thrombocytopenia is thought to be the formation of a neoepitope on GPIIbIIIa, which becomes the target of naturally-occurring antibodies or antibodies induced by prior exposure to the drug [23]. In addition to causing platelet clearance, eptifibatidedependent anti-platelet antibodies have been shown to induce platelet activation through platelet FcγRIIa receptor binding, which may explain the occurrence of thrombosis in some patients with eptifibatide-thrombocytopenia [24].…”

Heparin promotes platelet responsiveness by potentiating αIIbβ3-mediated outside-in signaling

“…19,20 All other blood samples from healthy volunteers who had not taken any anticoagulant in the past 2 weeks were donated as approved by the Institutional Review Board of the BloodCenter of Wisconsin, and informed consent was obtained in accordance with the Declaration of Helsinki. Blood was drawn into sodium citrate, pH 7.4, supplemented with 50 ng/mL prostaglandin E1 (PGE 1 ) and spun at 200g for 10 minutes.…”

“…Pretreatment with eptifibatide and/or abciximab also suppressed Akt phosphorylation in platelets exposed to either soluble heparin ( Figure 4B) or immobilized heparin (supplemental Figure 4), as well as their ability to spread on immobilized fondaparinux (supplemental Figure 2C). Finally, platelets from a patient with Glanzmann thrombasthenia that expressed 100% levels of a mutant ␣IIb␤3 complex lacking much of the ␤3 cytoplasmic domain 19,20 failed to increase the phosphorylation of Akt on exposure to soluble UFH ( Figure 4C). Taken together, these data demonstrate that the mechanism by which heparin potentiates platelet activation is via outside-in signaling through the major platelet integrin, ␣IIb␤3 ( Figure 5).…”