A variant of recombinant factor VIIa with enhanced procoagulant and antifibrinolytic activities in an in vitro model of hemophilia.

OBJECTIVE: Recombinant factor VIIa (rFVIIa, NovoSeven) has proven efficacy in treating bleeding in hemophilia patients with inhibitors. A rFVIIa analog with mutations V158D/E296V/M298Q (NN1731) exhibits increased procoagulant activity in in vitro and in vivo models. The aim of this work was to define the effects of NN1731 toward factor X activation, platelet activation, thrombin generation, and fibrin clot formation and stability. METHODS AND RESULTS: In a cell-based in vitro model of hemophilia, rFVIIa and NN1731 similarly increased factor X activation on tissue factor-bearing cells; however, NN1731 exhibited 30-fold higher factor Xa generation on platelets than similar rFVIIa concentrations. NN1731-mediated thrombin generation depended on platelet activation, but NN1731 did not directly activate platelets. NN1731 produced 4- to 10-fold higher maximal thrombin generation rates than equal rFVIIa concentrations. Both rFVIIa and NN1731 shortened clotting times in the absence of factors IX and VIII; however, NN1731 did so at 50-fold lower concentrations than were required of rFVIIa. In fibrinolytic conditions, both rFVIIa and NN1731 increased fibrin formation and stability; however, NN1731 was effective at 50-fold lower concentrations than were required of rFVIIa. CONCLUSIONS: By increasing factor Xa generation, NN1731 promotes the formation of thrombin and a stable clot to a greater degree than rFVIIa.

Thrombosis in children: current perspective and distinct challenges.

Coincident with a true increase in the incidence of thrombosis in childhood has come an appreciation of the impact of thromboembolic events upon children. In part, the greater awareness of childhood thrombosis reflects improved diagnostic imaging, which allows more sensitive identification of clots in infants and children. At the same time, advances in supportive care have meant that more children are being exposed to, surviving and living with thromboembolic risk factors and complications than previously. Because data regarding pediatric thrombosis have been lacking, treatment strategies have been extrapolated from experience in adults. This approach, however, results in diagnostic and therapeutic pitfalls. An understanding of issues unique to pediatric thrombosis is required; recent insights and the ongoing challenges are reviewed.

Approach to the bleeding child.

Because bruising and bleeding are normal events of childhood, the pediatrician must be able to determine whether a child's symptoms are normal or perhaps indicative of a defective hemostasis. A thorough medical history and physical examination should enable the clinician to choose those patients warranting further evaluation. Rather than referral to a hematologist at that point in time, pediatricians should be quite capable of performing the initial laboratory evaluation and making the correct diagnosis in a majority of cases.

Manipulation of prothrombin concentration improves response to high-dose factor VIIa in a cell-based model of haemophilia.

Clinical reports suggest that treatment regimens employing both activated prothrombin complex concentrates (aPCCs) and recombinant activated factor VII (rFVIIa) may control the bleeding in patients with haemophilia who fail to respond to either agent alone. We hypothesised that increased concentrations of prothrombin, as may be observed after the infusion of aPCCs, favourably influence parameters of thrombin generation in haemophilia treated with high-dose rFVIIa. We examined the effect of varied prothrombin and rFVIIa concentrations on thrombin generation in a model of haemophilia. At all concentrations of rFVIIa, increased prothrombin concentrations led to increases in the peak and rate of thrombin generation. In assays with the highest concentrations of prothrombin and rFVIIa, peak thrombin actually equalled that measured in the model of normal haemostasis. The significant impact of prothrombin concentration on the effect of rFVIIa in vitro may explain the improved haemostasis reported with concurrent use of aPCCs and rFVIIa. These results imply that persons with plasma prothrombin levels at either end of the 'normal' range could have significantly different responses to similar rFVIIa doses. Furthermore, these results suggest that increasing plasma prothrombin concentration prior to rFVIIa administration may offer advantages over the use of rFVIIa alone in the treatment of haemophilic bleeding.

High dose factor VIIa improves clot structure and stability in a model of haemophilia B.

Factor IX (FIX) deficiency results in haemophilia B and high dose recombinant activated factor VII (rFVIIa) can decrease bleeding. Previously, we showed that FIX deficiency results in a reduced rate and peak of thrombin generation. We have now used plasma and an in vitro coagulation model to examine the effect of these changes in thrombin generation on fibrin clot structure and stability. Low FIX delayed the clot formation onset and reduced the fibrin polymerisation rate. Clots formed without FIX were composed of thicker fibrin fibres than normal. rFVIIa shortened the clot formation onset time and improved the fibre structure of haemophilic clots. We also examined clot formation in the presence of a fibrinolytic challenge by including tissue plasminogen activator or plasmin in the reaction milieu. In these assays, normal FIX levels supported clot formation; however, clots did not form in the absence of FIX. rFVIIa partially restored haemophilic clot formation. These results were independent of the effects of the thrombin-activatable fibrinolysis inhibitor. Our data suggest that rFVIIa enhances haemostasis in haemophiliacs by increasing the thrombin generation rate to both promote formation of a structurally normal clot and improve clot formation and stability at sites with high endogenous fibrinolytic activities.

Recombinant activated factor VII: its mechanism of action and role in the control of hemorrhage.

PURPOSE: Recombinant activated factor VII (rFVIIa) has proven both safe and efficacious in the treatment of bleeding episodes in patients with hemophilia A or B who have developed inhibitors. More recently, a growing number of reports suggests that rFVIIa may also have indications for the treatment of bleeding in patients with other hemostatic disorders, including qualitative and quantitative platelet defects, factor deficiencies other than hemophilia, and in otherwise healthy patients with uncontrollable hemorrhage following surgery or trauma. We have attempted to reconcile the various proposed mechanisms of action of rFVIIa with its apparent efficacy in such diverse clinical settings. SOURCE: A review of the literature was performed to determine those clinical scenarios in which rFVIIa appears to have been effective in controlling associated hemorrhage. PRINCIPAL FINDINGS: Findings from our group and others have demonstrated that rFVIIa is able to directly activate factor X and increase thrombin production on the surface of activated platelets in the absence of factor VIII or IX, as well as to improve thrombin generation in thrombocytopenia, and to yield a fibrin dot more resistant to fibrinolysis in vitro. CONCLUSIONS: Through these primary mechanisms, we believe that rFVIIa may be able to compensate for a variety of defects in hemostasis and merits further investigation as a general therapeutic for uncontrollable hemorrhage.