Selective depletion of plasma prekallikrein or coagulation factor XII inhibits thrombosis in mice without increased risk of bleeding

Abstract

Recent studies indicate that the plasma contact system plays an important role in thrombosis, despite being dispensable for hemostasis. For example, mice deficient in coagulation factor XII (fXII) are protected from arterial thrombosis and cerebral ischemia-reperfusion injury. We demonstrate that selective reduction of prekallikrein (PKK), another member of the contact system, using antisense oligonucleotide (ASO) technology results in an antithrombotic phenotype in mice. The effects of PKK deficiency were compared with those of fXII deficiency produced by specific ASO-mediated reduction of fXII. Mice with reduced PKK had ϳ 3-fold higher plasma levels of fXII, and reduced levels of fXIIa-serpin complexes, consistent with fXII being a substrate for activated PKK in vivo. PKK or fXII deficiency reduced thrombus formation in both arterial and venous thrombosis models, without an apparent effect on hemostasis. The amount of reduction of PKK and fXII required to produce an antithrombotic effect differed between venous and arterial models, suggesting that these factors may regulate thrombus formation by distinct mechanisms. Our results support the concept that fXII and PKK play important and perhaps nonredundant roles in pathogenic thrombus propagation, and highlight a novel, specific and safe pharmaceutical approach to target these contact system proteases. (Blood. 2011; 118(19):5302-5311) IntroductionThe blood coagulation system responds to vascular injury with local production of a clot formed of fibrin mesh and activated platelets. While this process is essential for hemostasis, dysregulated coagulation can lead to blood vessel occlusion (thrombosis), precipitating life-threatening events such as myocardial infarction, stroke and venous thromboembolism. In the classic view of blood coagulation, thrombin generation and fibrin formation can be initiated by 2 distinct mechanisms referred to as the extrinsic and intrinsic pathways. 1,2 The extrinsic pathway involves binding of plasma factor VIIa (fVIIa) to extravascular tissue factor (TF) at a site of vessel injury. 3 The first step in the intrinsic pathway requires the surface-dependent activation of plasma factor XII (fXII) to fXIIa in a process called contact activation. 4,5 Contact activation involves 2 other proteins, prekallikrein (PKK) and high molecular weight kininogen (HK). HK serves as a docking molecule for PKK on the contact surface. PKK is cleaved by fXIIa to form the protease ␣-kallikrein, which in turn cleaves fXII to generate additional fXIIa. Collectively, fXII, PKK and HK comprise the plasma contact system. FXIIa generated by contact activation can activate factor XI (fXI) to fXIa, triggering a series of proteolytic cleavage events that culminates in thrombin generation and fibrin clot formation.While the contact system can clearly trigger coagulation in vitro, it is not required for hemostasis. Humans and other animals deficient in a contact activation protein are largely asymptomatic. 4,[6][7][8] However, the contact system may play an important r...

Recent insights into the role of the contact pathway in thrombo-inflammatory disorders

“…40 Comparable results have been obtained using PKknockout mice, which indicates that PK inhibition could be a potential target for anticoagulation. 41 However, there are some contradictory results: inhibition of plasma kallikrein with different inhibitors produced a prothrombotic state, 42 which is a relevant observation for the further development of kallikrein inhibitors.…”

“…Similar to what was seen for factor XI, antisense oligonucleotides to factor XII reduced thrombus formation in several mouse thrombosis models in both venous and arterial beds. 40 To our knowledge, no human trials have been performed with factor XII-inhibiting agents.…”

Inhibition of factor XI activation attenuates inflammation and coagulopathy while improving the survival of mouse polymicrobial sepsis

“…15,16 In various animal models, decreasing or eliminating FXI procoagulant activity through gene knockout, pharmacologic inhibition, or antisense oligonucleotidemediated knockdown is also antithrombotic without significantly impairing hemostasis, [17][18][19][20][21] suggesting that FXI is an important driver of pathologic coagulation with only a supportive function in normal hemostasis. Interestingly, FXII and prekallikrein have also been shown to contribute to the development of experimental thrombosis in mice, 22,23 despite the normal to possibly prothrombotic phenotype associated with deficiency of either of these proteins (Hagemen trait and Fletcher trait, respectively) in humans. 14,[24][25][26] We previously demonstrated that FXI deficiency was associated with improved survival and reduced coagulopathy relative to wild-type mice during polymicrobial peritoneal sepsis.…”

Putting polyphosphates to the test: evidence against platelet-induced activation of factor XII

“…[4][5][6] These studies provide strong evidence for a role for FXII in pathological thrombosis in mice, although clinical evidence for an analogous role in humans is still lacking.…”