BackgroundRecombinant factor VIIa (rFVIIa) is approved for use in controlling bleeding episodes in people with hemophilia who have developed inhibitors to replacement therapy. Due to its short half-life (t(1/2)), frequent injections are required, limiting its use as a prophylactic treatment. A novel, recombinant fusion protein linking coagulation factor VIIa with albumin (rVIIa-FP) has been developed to extend the t(1/2) of rFVIIa.ObjectivesThe aim of our studies was to investigate the pharmacokinetic/pharmacodynamic characteristics of rVIIa-FP in preclinical animal species.MethodsPharmacokinetic (PK) parameters were derived after single intravenous dosing in hemophilia A mice, rats, rabbits and monkeys. PK analysis was based on human FVII plasma levels determined by measuring FVII antigen levels by ELISA in mice and rats, and FVIIa activity using STACLOT((R)) VIIa-rTF in rabbits and monkeys. Induction of thrombin generation was investigated in mice, while hemostatic activity was assessed by thrombus formation in rabbits.ResultsCompared with rFVIIa, rVIIa-FP displayed a prolonged t(1/2), enhanced in vivo recovery and reduced clearance in all species investigated. In mice, 16h after treatment with rVIIa-FP, thrombin levels were quantifiable, indicating prolonged efficacy, whereas values had approached baseline at this time after treatment with rFVIIa. After 12h, hemostatic efficacy was negligible in rFVIIa-treated rabbits, but sustained in animals receiving rVIIa-FP.ConclusionsThese studies indicate that the longer t(1/2) of rVIIa-FP compared with rFVIIa translates into extended activity. These findings suggest that rVIIa-FP has the potential to be administered less frequently than rFVIIa-containing concentrates in clinical use.

The prophylactic treatment of haemophilia B and the management of haemophilia A or B with inhibitors demand frequent administrations I of coagulation factors due to the suboptimal half-lives of the products commercially available and currently in use, e.g. recombinant factor IX (rFIX) and recombinant factor Vila (rFVIIa), respectively. The extension of the half-lives of rFIX and rFVIIa could allow for longer intervals between infusions and could thereby improve adherence and clinical outcomes and may improve quality of life. Albumin fusion is one of a number of different techniques currently being examined to prolong the half-life of rFIX and rFVIIa. Results from a phase I clinical trial demonstrated that the recombinant fusion protein linking FIX to albumin (rIX-FP) has a five-times longer half-life than rFIX, and preclinical studies with the recombinant fusion protein linking FVIIa to albumin (rVIIa-FP) suggest that rVIIa-FP possesses a significantly extended half-life versus rFVIIa. In this review, we describe albumin fusion technology and examine the recent progress in the development of rIX-FP and rVIIa-FP.

Abstract Abstract 1115 Recombinant factor VIIa (rFVIIa) is approved to control bleeding in hemophilia A and B patients who have developed inhibitory antibodies to replacement therapy and in acquired hemophilia. rFVIIa is rapidly eliminated with a terminal half-life of approximately 2.5 hours in humans. This short half life necessitates frequent injections and considerably limits prophylactic use. A recombinant fusion protein linking activated factor VII with albumin (rVIIa-FP) was engineered to extend the half life of rFVIIa. The present studies were conducted to gather knowledge about the pharmacokinetic and pharmacodynamic (PK/PD) properties of rVIIa-FP in preclinical animals models to enable a more precise estimate of the procoagulant activity of rVIIa-FP for the future clinical use in haemophilia patients with inhibitors to FVIII and FIX. Single intravenous doses of either rVIIa-FP or the licensed rFVIIa comparator, NovoSeven® were administered to mice, rabbits and monkeys. Subsequently, plasma samples were harvested at different time-points after study drug administration and systemic levels of rVIIa-FP or NovoSeven® were assessed either by FVIIa activity measurements using the Staclot® assay system or by FVII antigen measurements using an enzyme linked immunoabsorbance assay (ELISA). When investigating dose responses or duration of hemostatic activity, rVIIa-FP and NovoSeven® were administered to mice or rabbits before tail clip or induction of venous stasis, respectively. Following a single intravenous bolus application the procoagulant activity of both study drugs was assessed in both models at different time points after administration up to 24 hours to proof prolonged procoagulant effects of rVIIa-FP compared to NovoSeven®. Intriguingly, in all animal species tested the systemic bioavailability, clearance and in addition the terminal half-life (t1/2β), of rVIIa-FP were significantly better in comparison to NovoSeven®. Based on FVIIa activity measurements in plasma samples derived from monkeys the 11 fold higher area under the curve (AUC) was accompanied with a 4-fold longer t1/2β, while in vivo recovery (IVR) of rVIIa-FP exceeded that of NovoSeven® by a 70 %. In rabbits the 20 fold higher AUC was associated with a 5-fold longer t1/2β and a 4-fold higher IVR when measuring specific FVIIa activity. In mice measurements of FVII antigen revealed a 11 fold higher AUC, while t1/2β was 4 times longer and IVR was found to be doubled in comparison to NovoSeven®. The assessment of pharmacodynamic activity after dosing based on specific FVIIa activity adjusted by the Staclot® assay revealed, that the procoagulant effect of rVIIa-FP was not different from NovoSeven®, when measuring total blood loss under acute bleeding conditions after tail clip. However, when assessing late time-points after treatment the prolonged systemic availability of rVIIa-FP translated into a longer and sustained hemostatic activity of rVIIa-FP compared to NovoSeven® in both animal models. Intriguingly, ex vivo studies in hemophilia A mice showed that the 4 fold prolonged systemic availability of rVIIa-FP in plasma translates into sustained FVIIa activity when recorded as thrombin generation activity compared to NovoSeven® at 16 hours after treatment. Furthermore in rabbits, the 5 fold longer half-life translated into a significantly greater procoagulant activity as measured by thrombus formation in both jugular veins after venous stasis. In conclusion, the recombinant albumin fusion technology was successfully applied to human recombinant FVIIa for a significantly improved of PK/PD profile as observed in different pre-clinical animal species. Future clinical studies can proof whether the observed improved PK/PD characteristics will also translate into a half-life extension and a longer hemostatic effect in hemophilia patients with inhibitors to FVIII and FIX. Allowed: The entire body of the abstract, including text and tables, must not exceed 3,800 characters. Spaces are not included in this number; title, authors' names, and affiliations are counted separately. Figures are not included in the character count. Disclosures: Zollner: CSL Behring GmbH: Employment. Schuermann:CSL Behring GmbH: Employment. Raquet:CSL Behring GmbH: Employment. Mueller-Cohrs:CSL Behring GmbH: Employment. Weimer:CSL Behring GmbH: Employment. Pragst:CSL Behring GmbH: Employment. Dickneite:CSL Behring GmbH: Employment. Schulte:CSL Behring GmbH: Employment.