Paving the way to factor X deficiency therapies

In this issue of Blood, Verhenne and colleagues¹ describe a murine model of coagulation factor X (FX) deficiency and utilize this model to evaluate fitusiran as a therapeutic option. FX deficiency is a rare bleeding disorder, affecting ∼1 in 1 million individuals, which is divided into 2 subtypes.² Type I is characterized by low FX protein and activity, and individuals with type II deficiency have normal protein concentration but reduced activity. Lower FX concentration or activity is associated with worse symptoms, including a risk for intracranial hemorrhage in the most severe patients. Thus, effective treatment options are critical, but the current options are limited and consist primarily of blood products, including virally inactivated plasma and purified plasma proteins, such as cryoprecipitate, prothrombin complex concentrate, or FX itself.

By contrast, hemophilia A, a more common bleeding disorder caused by deficiency or dysfunction of coagulation factor VIII (FVIII), has seen a recent boon of therapeutic options, some of which have been approved for use and others that are in different stages of development.^3,4 These include emicizumab, a bispecific antibody that mimics the function of FVIII; recombinant factor VIIa, which directly promotes the activation of FX (FXa); and an array of inhibitors that target components of the anticoagulant system. Targeted anticoagulants include tissue factor pathway inhibitor (TFPI), protein C, protein S, and antithrombin, all of which either downregulate FX activation or directly inhibit FXa procoagulant function. Thus, a variety of approaches are used with the common goal of promoting FXa and subsequent thrombin generation.

The development of these novel hemophilia A treatments has been possible because of the availability of a mouse model. The FVIII knockout mouse is viable,⁵ and thus it is possible to test whether use of a specific therapeutic or inhibition of a specific target is safe and effective for restoring hemostasis in vivo before starting a clinical trial. Until now, a similar tool has not been available for preclinical development of treatments for type I FX deficiency. The FX-knockout mouse is not viable; ∼50% of F10^−/− mice die during embryonic development, and the remainder die shortly after birth.⁶ In 2008, Tai et al developed a transgenic mouse model of type II FX deficiency, which expresses a mutant form of FX with reduced specific activity.⁷ These mice have normal FX antigen concentration but ∼5% activity and have a mild bleeding phenotype.

Verhenne et al provide the first viable model of type I FX deficiency, utilizing the Mx1-Cre system, in which excision of the targeted gene is induced by polyinosinic:polycytidylic acid (pI:pC) treatment. This resulted in >99% loss of FX antigen and activity. Interestingly, the mice did not present with any spontaneous bleeding phenotype, though they did exhibit severely impaired coagulation in injury models that mimic either hemostasis (tail clip or saphenous vein puncture) or thrombosis (cremaster muscle laser injury). The authors next used this model to test the effectiveness of fitusiran to treat FX deficiency. Fitusiran is a small interfering RNA, which inhibits production of antithrombin and is currently under development for hemophilia A and hemophilia B treatment.⁸ By decreasing antithrombin concentration, fitusiran prolongs the activity of factors IXa and Xa and thrombin, all of which are inhibited by antithrombin. Fitusiran effectively restored hemostatic competence in FX-deficient mice, in both in vivo and ex vivo assessments, demonstrating the utility of this new model system. Other therapeutics currently being developed for hemophilia can also be tested in this system. For example, TFPI also inhibits FXa activity⁹; thus, agents that target TFPI may restore hemostasis in these mice. Similarly, agents that target the protein C and S system may promote hemostasis in these mice by stabilizing factor V, the essential cofactor for FXa.¹⁰ 

Outside of FX deficiency itself, these mice also provide an opportunity to test the role of FXa in other disease processes, particularly when used in conjunction with the type II FX deficiency mouse, which has normal protein but low coagulant activity. Alterations in coagulation factors, antithrombin, and other components of the hemostatic system are common in many inflammatory conditions, though it is not always clear what role coagulation plays in driving the pathology. The use of these 2 complementary mouse models would allow investigators to assess the importance of FX/Xa in these processes, as well as the specific role of its procoagulant activity. In short, Verhenne and colleagues have created a much-needed model for FX deficiency that will be invaluable in the assessment of potential therapeutics and will be useful in understanding the broader function of FX in physiology and pathology.

Conflict-of-interest disclosure: J.P.W. received an investigator-initiated grant through Pfizer, Inc.