Abstract 637: Ligand-Binding Induced Epitopes on GPIIb/IIIa Allow Highly Efficient Clot-Targeted Anticoagulation without Bleeding Time Prolongation: A Biotechnological Approach Based on the Single-Chain Antibody Technology
Therapeutic anticoagulation is widely used but limitations in efficacy and bleeding complications cause an ongoing search for new agents. However, with new agents developed it seems to be an inherent problem that increased efficiency is accompanied by an increase in bleeding complications. We investigate whether targeting of anticoagulants to activated platelets provides a means to break this association. Ligand-induced binding sites (LIBS) on fibrinogen/ fibrin-binding GPIIb/IIIa represent an abundant clot-specific target. We cloned an anti-LIBS single-chain antibody (scFv-anti-LIBS) and genetically fused it with a potent, direct factor Xa (fXa) inhibitor, tick anticoagulant peptide (TAP). Specific antibody binding of the fusion molecule scFv-anti-LIBS-TAP was proven in flow cytometry; anti-fXa activity was demonstrated in chromogenic assays. In vivo anticoagulative efficiency was determined by Doppler-flow in a ferric chloride-induced carotid artery thrombosis model in mice. ScFv-anti-LIBS-TAP prolonged occlusion time comparable to enoxaparine, recombinant TAP, and non-targeted mutant-scFv-TAP. ScFv-anti-LIBS-TAP revealed antithrombotic effects at low doses at which the non-targeted mutant-scFv-TAP failed. In contrast to the other anticoagulants tested, bleeding time as measured by tail transection was not prolonged by scFv-anti-LIBS-TAP. The novel clot-targeting approach of anticoagulants via single-chain antibody directed against a LIBS epitope on GPIIb/IIIa promises effective anticoagulation with reduced bleeding risk.