Prolonged in-vivo half-life of factor VIIa by fusion to albumin

Thomas Weimer; Wilfried Wormsbächer; Ulrich Kronthaler; Wiegand Lang; Uwe Liebing; Stefan Schulte

doi:10.1160/TH07-08-0525

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2008; 99(04): 659-667
DOI: 10.1160/TH07-08-0525

Blood Coagulation, Fibrinolysis and Cellular Haemostasis

Schattauer GmbH

Prolonged in-vivo half-life of factor VIIa by fusion to albumin

Thomas Weimer

¹CSL Behring GmbH, Marburg, Germany

,

Wilfried Wormsbächer

¹CSL Behring GmbH, Marburg, Germany

,

Ulrich Kronthaler

¹CSL Behring GmbH, Marburg, Germany

,

Wiegand Lang

¹CSL Behring GmbH, Marburg, Germany

,

Uwe Liebing

¹CSL Behring GmbH, Marburg, Germany

,

Stefan Schulte

¹CSL Behring GmbH, Marburg, Germany

› Author Affiliations

Abstract

Summary

For the treatment of haemophilia patients with inhibitors, recombinant factor VIIa (rFVIIa) is available as a therapeutic option to control bleeding episodes with a good balance of safety and efficacy. However, the short in-vivo half-life of approximately 2.5 hours makes multiple injections necessary, which is inconvenient for both physicians and patients. Here we describe the generation of a recombinant FVIIa molecule with an extended half-life based on genetic fusion to human albumin. The recombinant FVII albumin fusion protein (rVII-FP) was expressed in mammalian cells and upon activation displayed a FVII activity close to that of wild type FVIIa. Pharmacokinetic studies in rats demonstrated that the half-life of the activated recombinant FVII albumin fusion protein (rVIIa-FP) was extended six- to sevenfold compared with wild type rFVIIa. The in-vitro and in-vivo efficacy was evaluated and was found to be comparable to a commercially available rFVIIa (NovoSeven^®). The results of this study demonstrate that it is feasible to develop a half-life extended FVIIa molecule with haemostatic properties very similar to the wild-type factor.

Keywords

FVIIa - albumin fusion - rVIIa-FP - coagulation

Full Text

References

References
1 Greenberg DL, Davie EW. Blood coagulation factors: Their complementary DNAs, genes and expression. Hemostasis and Thrombosis.. Philadelphia: Lippincott Williams & Wilkins; 2001: 135-156.
2 Wildgoose P, Nemerson Y, Hansen LL. et al. Measurement of basal levels of factor VIIa in hemophilia A and B patients. Blood 1992; 80: 25-28.
3 Hedner U. Recombinant activated factor VII as universal haemostatic agent. Blood Coag Fibrinolysis 1998; 9 (Suppl. 01) S147-152.
4 Lusher JM, Roberts HR, Davignon G. et al. A randomized, double-blind comparison of two dosage levels of recombinant factor VIIa in the treatment of joint, muscle and mucocutaneous haemorrhages in persons with haemophilia A and B, with and without inhibitors. Haemophilia 1998; 4: 790-798.
5 Butenas S, Brummel KE, Bouchard BA. et al. How factor VIIa works in hemophilia. J Thromb Haemost 2003; 1: 1158-1160.
6 Roberts HR, Monroe DR, White GC. The use of recombinant factor VIIa in the treatment of bleeding disorders. Blood 2004; 104: 3858-3864.
7 Erhardtsen E. Pharmacokinetics of recombinant activated factor VII (VIIa). Semin Thromb Hemost 2000; 26: 385-391.
8 Key NS, Aledort LM, Beardsley D. et al. Home treatment of mild to moderate bleeding episodes using recombinant Factor VIIa (Novoseven) in hemophiliacs with inhibitors. Thromb Haemost 1998; 80: 912-918.
9 Astermark J, Donfield SM, Di Michele DM. et al. A randomized comparison of bypassing agents in hemophilia complicated by an inhibitor: the FEIBA Novo-Seven Comparative (FENOC) Study. Blood 2006; 109: 546-551.
10 Lusher JM. Recombinant factor VIIa (NovoSeven) in the treatment of internal bleeding in patients with factor VIII and IX inhibitors. Hemostasis 1996; 26 (Suppl. 01) 124-130.
11 Shapiro AD, Gilchrist GS, Hoots WK. et al. Prospective, randomized trial for two doses of rFVIIa (Novoseven) in haemophilia patients with inhibitors undergoing surgery. Thromb Haemost 1998; 80: 773-778.
12 Dumont JA, Low SC, Peters RT. et al. Monomeric Fc fusions: Impact on pharmacokinetics and biological activity of protein therapeutics. BioDrugs 2006; 20: 151-160.
13 Chuang VTG, Kragh-Hansen U, Otagiri M. Pharmaceutical strategies utilizing recombinant human serum albumin. Pharm Res 2002; 5: 569-577.
14 Sterling K. The turnover rate of serum albumin in man as measured by I-131-tagged albumin. J Clin Invest 1957; 30: 1228-1237.
15 Sheffield WP, Smith IJ, Syed S. et al. Prolonged in vivo anticoagulant activity of a hirudin-albumin fusion protein secreted from Pichia pastoris. Blood Coag Fibrinol 2001; 12: 433-443.
16 Duttaroy A, Kanakaraj P, Osborn BL. et al. Development of a long-acting insulin analog using albumin fusion technology. Diabetes 2005; 54: 251-258.
17 Wang W, Ou Y, Shi Y. AlbuBNP, a recombinant B-type natriuretic peptide and human serum albumin fusion hormone, as a long-term therapy of congestive heart failure. Pharm Res 2004; 21: 2105-2111.
18 Osborn BL, Olsen HS, Nardelli B. et al. Pharmacokinetic and pharmacodynamic studies of a human serum albumin-interferon-α fusion protein in Cynomolgus monkeys. J Pharmacol Exp Ther 2002; 303: 540-548.
19 Bain VG, Kaita KD, Yoshida EM. et al. A phase 2 study to evaluate the antiviral activity, safety, and pharmacokinetics of recombinant human albumin-interferon alfa fusion protein in genotype 1 chronic hepatitis C patients. J Hepatol 2006; 44: 671-678.
20 Balan V, Nelson DR, Sulkowski MS. et al. A Phase I/II study evaluating escalating doses of recombinant human albumin-interferon-alpha fusion protein in chronic hepatitis C patients who have failed previous interferon-alpha-based therapy. Antivir Ther 2006; 11: 35-45.
21 Sheffield WP, Mamdani A, Hortelano G. et al. Effects of genetic fusion of factor IX to albumin on in vivo clearance in mice and rabbits. Br J Haematol 2004; 126: 565-573.
22 Chiou WL. Critical evaluation of the potential error in pharmacokinetic studies of using the linear trapezoidal rule method for the calculation of the area under the plasma level-time curve. J Pharmacokinet Biopharm 1978; 6: 539-546.
23 Bjoern S, Thim L. Activation of coagulation factor VII to VIIa. Res Disclosure. 1986: 564-565.
24 Pedersen AH, Lund-Hansen T, Bisgaard-Frantzen H. et al. Autoactivation of human recombinant coagulation FVII. Biochemistry 1989; 28: 9331-9336.
25 Mollerup I, Bayne S, Halkjaer E. et al. The use of RP-HPLC for measuring activation and cleavage of rFVIIa during purification. Biotechnol Bioeng 1995; 48: 501-505.
26 Mann KG, Butenas S, Brummel K. The dynamics of Thrombin formation. Arterioscler Thromb Vasc Biol 2003; 23: 17-25.
27 Hoffmann M, Monroe DM. Coagulation 2006: A modern view of hemostasis. Hematol Oncol Clin N Am 2007; 21: 1-11.
28 Dickneite G. Prothrombin complex concentrate versus recombinant factor VIIa for reversal of coumarin anticoagulation. Thromb Res 2007; 119: 643-651.
29 Persson E, Kjalke M, Olsen OH. Rational design of coagulation factor VIIa variants with substantially increased intrinsic activity. Proc Natl Acad Sci USA 2001; 98: 13583-13588.
30 Persson E, Bak H, Olsen OH. Substitution of Valine for Leucine 305 in Factor VIIa increases the intrinsic enzymatic activity. J Biol Chem 2001; 276: 29195-29199.
31 Waldmann TA, Strober W. Metabolism of immunoglobulins. Prog Allergy 1969; 13: 1-110.
32 Chaudhury C, Mehnaz S, Robinson JM. et al. The major histocompatibility complex-related Fc receptor for IgG (FcRn) binds albumin and prolongs its lifespan. J Exp Med 2003; 197: 315-322.
33 Anderson CL, Chaudhury C, Kim J. et al. Perspective – FcRn transports albumin: relevance to immunology and medicine. Trends in Immunology 2006; 7: 343-348.
34 Ewenstein BM, Joist JH, Shapiro AD. et al. Pharmacokinetic analysis of plasma-derived and recombinant FIX concentrates in previously treated patients with moderate or severe hemophilia B. Transfusion 2002; 42: 190-197.
35 Ghosh S, Pendurthi UR, Steinoe A. et al. Endothelial cell protein C receptor acts as a cellular receptor for factor VIIa on endothelium. J Biol Chem 2007; 282: 11849-11857.
36 Wolberg AS, Stafford DW, Erie DA. Human FIX binds to specific sites on the collagenous domain of collagen IV. J Biol Chem 1997; 272: 16717-16720.
37 Cheung WF, van den Born J, Kühn K. et al. Identification of the endothelial binding site for FIX. Proc Natl Acad Sci USA 1996; 93: 11068-11073.
38 Gui T, Lin HF, Jin DY, Hoffmann M. et al. Circulating and binding characteristics of wild-type factor IX and certain Gla domain mutants in vivo. Blood 2002; 100: 153-158.