Today, the development of inhibitory antibodies against FVIII during substitution therapy constitutes the most serious complication amongst patients with severe haemophilia A and occurs in previously untreated patients with incidences ranging from 20 to 33% (Ehrenforth et al, 1992; Ljung et al, 1992; Kreuz et al, 1999; Scharrer et al, 1999; Wight & Paisley, 2003). The presence of high titre inhibitors (>5 Bethesda Units (BU)/ml) often require the use of bypassing agents, such as plasma-derived activated prothrombin complex concentrate (aPCC) or recombinant FVIIa (rFVIIa) to obtain haemostasis (Ingerslev et al, 1991; Sørensen & Ingerslev, 2004a). Currently, rFVIIa is successfully used for the treatment of bleeds in patients with congenital severe haemophilia with inhibitors (Abshire & Kenet, 2004). However, in a few haemophilia patients with inhibitors, the haemostatic response to rFVIIa, as well as to aPCC, appears rather low (Martinowitz et al, 2005). Moreover, considerable inter-individual response variations indicate a possible need for individualised dose titration (Sørensen & Ingerslev, 2004a). Hence, in a subgroup of difficult-to-treat haemophilia patients with inhibitors, a novel form or formulation of rFVIIa with optimised efficacy and a more predictable haemostatic response profile would appear attractive.
During recent years innovations have resulted in the development of novel, potent rFVIIa analogues that exhibit an increased intrinsic activity, as defined by an enhanced ability of tissue factor (TF)-independent activation of factor X (FX) on the phospholipid surface of activated platelets (Persson et al, 2001a,b; Persson & Olsen, 2002; Persson et al, 2004). Some of these analogues were reported to possess an enhanced potential to facilitate haemostasis, as evaluated by studies in FVIII-deficient plasma (Lisman et al, 2003) and haemophilia A mice models (Tranholm et al, 2003). Although the mechanisms of action of rFVIIa are subject to debate, accumulating data stress the importance of TF (Butenas et al, 2003a) and phospholipids as well as the existence of a TF-independent, platelet-dependent activation of FX (Hoffman & Monroe, 2001). The importance of the latter is illustrated by data on the rFVIIa analogues, which have a significantly higher intrinsic activity when not bound to TF (Persson et al, 2001b). Appreciating the important interactions of platelets and white blood cells in the overall regulation of haemostasis (Polgar et al, 2005), whole blood (WB) seems the optimal medium for in vitro evaluation of haemostatic capacity. Moreover, previous studies performed with rFVIIa in animal models have documented a need for very high doses to achieve a haemostatic effect in certain species, such as rodents. Thus, WB coagulation analyses on blood from patients represent the most appropriate means of exploring the haemostatic potential of rFVIIa and analogues of rFVIIa under various clinical conditions. Our laboratory has developed a thrombelastographic model of continuous WB coagulation, employing activation with minute amounts of TF, and novel data processing of the raw coagulation data signal to provide dynamic parameters of clot formation (Sørensen et al, 2003a). So far, a comprehensive evaluation of the model has been performed in various conditions of abnormal haemostasis, demonstrating the modified thrombelastographic method as an attractive and applicable approach for the characterisation of haemostatic deterioration and prediction of the effect of rFVIIa, aPCC, and other types of haemostatic intervention (Ingerslev et al, 2003; Sørensen & Ingerslev, 2003; Sørensen et al, 2003a; Sørensen et al, 2003b; Sørensen & Ingerslev, 2004a,b; Fenger-Eriksen et al, 2005; Sørensen et al, 2005a). Hence, in addition to detailed and important measurements of thrombin generation (Rand et al, 1996; Cawthern et al, 1998; Butenas et al, 2003a,b), analyses of thrombelastographic WB clot formation and WB clot stability in blood from patients with severe haemophilia A may provide desirable information about the haemostatic potential of rFVIIa and rFVIIa analogues. The present study hypothesised that a rFVIIa analogue (Mutations: V158D/E296V/M298Q-FVIIa, designated: NN1731) would provide a significantly accelerated WB clot formation when compared with rFVIIa, and that NN1731 would significantly improve WB clot stability compared to rFVIIa. Utilising the above-mentioned thrombelastographic model (Sørensen et al, 2003a; Sørensen & Ingerslev, 2004a), the present study aimed to evaluate the haemostatic potential of NN1731 (Novo Nordisk, Bagsvaerd, Denmark) in comparison with wild-type rFVIIa (NovoSeven®, Novo Nordisk) on clot formation characteristics and clot resistance to fibrinolysis in WB from patients with severe haemophilia A.