Abstract
Life-threatening thrombocytopenia and bleeding, common side effects of clinically available αIIbβ3 antagonists, are associated with the induction of ligand-induced integrin conformational changes and exposure of ligand-induced binding sites (LIBSs). To address this issue, we examined intrinsic mechanisms and structure–activity relationships of purified disintegrins, from Protobothrops flavoviridis venom (i.e., Trimeresurus flavoviridis), TFV-1 and TFV-3 with distinctly different pro-hemorrhagic tendencies. TFV-1 with a different αIIbβ3 binding epitope from that of TFV-3 and chimeric 7E3 Fab, i.e., Abciximab, decelerates αIIbβ3 ligation without causing a conformational change in αIIbβ3, as determined with the LIBS antibody, AP5, and the mimetic, drug-dependent antibody (DDAb), AP2, an inhibitory monoclonal antibody raised against αIIbβ3. Consistent with their different binding epitopes, a combination of TFV-1 and AP2 did not induce FcγRIIa-mediated activation of the ITAM–Syk–PLCγ2 pathway and platelet aggregation, in contrast to the clinical antithrombotics, abciximab, eptifibatide, and disintegrin TFV-3. Furthermore, TFV-1 selectively inhibits Gα13-mediated platelet aggregation without affecting talin-driven clot firmness, which is responsible for physiological hemostatic processes. At equally efficacious antithrombotic dosages, TFV-1 caused neither severe thrombocytopenia nor bleeding in FcγRIIa-transgenic mice. Likewise, it did not induce hypocoagulation in human whole blood in the rotational thromboelastometry (ROTEM) assay used in perioperative situations. In contrast, TFV-3 and eptifibatide exhibited all of these hemostatic effects. Thus, the αIIbβ3 antagonist, TFV-1, efficaciously prevents arterial thrombosis without adversely affecting hemostasis.
Highlights
Integrin αIIb β3 is a member of the integrin family of adhesion receptors and remains in an inactive state in normal circulation, preventing undesirable thrombus formation [1]
We previously reported that monoclonal antibodies (mAbs) 7E3 shares the same binding site with RGD-containing αIIb β3 antagonists rhodostomin and trigramin [5,23], which cause thrombocytopenia and bleeding owing to their effects on a conformational change of integrin αIIb β3
We previously reported that upon drug-induced ligand-induced binding sites (LIBSs) exposure, the mimetic drug-dependent antibody (DDAb) mAb, AP2, which was raised against the epitopes of IIb 3, recruited Fc RIIa [19]
Summary
Integrin αIIb β3 is a member of the integrin family of adhesion receptors and remains in an inactive state in normal circulation, preventing undesirable thrombus formation [1]. Disintegrins are Arg-Gly-Asp (RGD)/Lys-Gly-Asp (KGD)-containing, cysteine-rich proteins in many snake venoms [3]. Based on the RGD/KGD mimetic sequence, disintegrins acts as αIIb β3 antagonists and potential antithrombotic agents [4,5,6]. Three clinically available αIIb β3 antagonists, represented by abciximab, eptifibatide, and tirofiban, are used as potent antithrombotics for their rapid action and high efficacy. Their use is primarily limited to patients undergoing percutaneous coronary intervention because of significant bleeding risk [7,8]
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