Background Exposure of the plasma protein factor XII to an anionic surface generates activated factor XII that not only triggers the intrinsic pathway of blood coagulation through the activatio of factor XI, but also mediates various vascular responses through activation of the plasma contact system. While deficiencies of factor XII are not associated with excessive bleeding, thrombosis models in factor deficient animals have suggested that this protein contributes to stable thrombus formation. Therefore, factor XII has emerged as an attractive therapeutic target to treat or prevent pathological thrombosis formation without increasing the risk for hemorrhage. Objectives Utilizing an in vitro directed evolution and chemical biology approach, we sought to isolate a nuclease resistant RNA aptamer that binds specifically to factor XII and directly inhibits factor XII coagulant function. Methods and Results Herein, we describe the isolation and characterization of a high affinity RNA aptamer targeting factor XII/XIIa that dose dependently prolongs fibrin clot formation and thrombin generation in clinical coagulation assays. This aptamer functions as a potent anticoagulant by inhibiting the autoactivation of factor XII, as well as inhibiting intrinsic pathway activation (factor XI activation). However, the aptamer does not affect the factor XIIa-mediated activation of the proinflammatory kallikrein-kinin system (plasma kallikrein activation). Conclusions We have generated a specific and potent factor XII/XIIa aptamer anticoagulant that offers targeted inhibition of discrete macromolecular interactions involved in the activation of the intrinsic pathway of blood coagulation.

Background The plasma protease factor XIa (FXIa) has become a target of interest for therapeutics designed to prevent or treat thrombotic disorders. Methods We used a solution-based, directed evolution approach called systematic evolution of ligands by exponential enrichment (SELEX) to isolate RNA aptamers that target the FXIa catalytic domain. Results Two aptamers, designated 11.16 and 12.7, were identified that bound to previously identified anion binding and serpin bindings sites on the FXIa catalytic domain. The aptamers were non-competitive inhibitors of FXIa cleavage of a tripeptide chromogenic substrate and of FXIa activation of factor IX. In normal human plasma, aptamer 12.7 significantly prolonged the aPTT clotting time. Conclusions The results show that novel inhibitors of FXIa can be prepared using SELEX techniques. RNA aptamers can bind to distinct sites on the FXIa catalytic domain and noncompetitively inhibit FXIa activity towards its primary macromolecular substrate factor IX with different levels of potency. Such compounds can be developed for use as therapeutic inhibitors.

Understanding inherent differences between thrombosis and hemostasis in the vascular system are critical to developing safe and effective anticoagulants. To this end, constituents of the contact activated and intrinsic pathway of coagulation appear to be involved in pathological thrombus formation, but are not required for normal hemostasis. In addition to coagulation, activation of the contact system is involved in fibrinolytic, inflammatory, and angiogenic processes that can also contribute to the thrombotic environment. This review discusses the role of the contact system in these processes, and highlights the potential of FXII and FXI as safer targets for antithrombotic therapy.