In Vivo Studies of Activated Porcine Factor VIII

Summary

The haemostatic effectiveness of activated FVIII was compared to that of non-activated FVIII in a cross-over study in a canine model of haemophilia. Activation of FVIII in porcine concentrate was achieved by the addition of 3 × 10⁻⁵ IU thrombin per ml of concentrate, which gave consistent increases in 1-stage FVIII activity of 13- to 14-fold and slow decay. The haemostatic effect was monitored by measurements of the cuticle bleeding time 10 and 45 min after infusion and there were no consistent differences between the activated and non-activated concentrates. One-stage factor VIII assays on plasmas 5 min after infusion showed identical mean values for activated and non-activated concentrates, indicating that most of the higher activity observed in vitro had disappeared rapidly from the circulation.

These results suggest that controlled activation of FVIII by thrombin, which increases its activity in 1-stage assays, is unlikely to be of therapeutic benefit. For therapeutic concentrates which may contain small amounts of activated FVIII, the 1-stage assay may be an unreliable guide to their therapeutic effect.

• References

• 1 Biggs R, Macfarlane RG, Denson KWE, Ash BJ. Thrombin aids the interaction of factors VIII and IX. Br J Haematol 1965; 11: 276-295
• 2 Niemetz J, Nossel HL. Activated coagulation factors: in vivo and in vitro studies. Br J Haematol 1969; 16: 337-351
• 3 Mertens K, van Wijngaarden A. Bertina RM The role of factor VIII in the activation of human blood coagulation factor X by activated factor IX. Thromb Haemost 1985; 54: 654-660
• 4 Eaton D, Rodriguez H, Vehar GA. Proteolytic processing of human factor VIII. Correlation of specific cleavages by thrombin, factor Xa and activated protein C with activation and inactivation of factor VIII coagulant activity. Biochemistry 1986; 25: 505-512
• 5 Barrowcliffe TW, Edwards SJ, Kemball-Cook G, Thomas DP. Factor VIII degradation products in heated concentrates. Lancet 1986; 1: 1448-1449
• 6 Barrowcliffe TW, Watton J, Tubbs JE, Harman A, Kemball-Cook G. Potency of high purity factor VIII concentrates. Lancet 1990; 2: 124
• 7 Barrowcliffe TW, Dawson NJ, Kemball-Cook G. Activated factor VIII in concentrates prepared by monoclonal antibody and recombinant technology. Thromb Haemost 1989; 62: 198
• 8 Kemball-Cook G, Tubbs JE, Dawson NJ, Barrowcliffe TW. The behaviour of different factor VIII concentrates in a chromogenic factor X-activating system. Br J Haematol 1993; 84: 273-278
• 9 Giles AR, Tinlin S, Hoogendoorn H, Greenwood R. A canine model of hemophilic (factor VIII:C deficiency) bleeding. Blood 1982; 60: 727-730
• 10 Lollar P, Parker ET, Fay PJ. Coagulant properties of hybrid human/porcine factor VIII molecules. J Biol Chem 1992; 267: 23652-23657
• 11 Littlewood JanetD, Barrowcliffe TW. The development and characterisation of antibodies to human factor VIII in haemophilic dogs. Thromb Haemost 1987; 57: 314-321
• 12 Giles AR, Tinlin S, Hoogendoorn H, Greenwood P, Greenwood R. Development of factor VIII:C antibodies in dogs with hemophilia A (factor VIII:C deficiency). Blood 1984; 63: 451-456
• 13 Littlewood JD. Factor VUI-phospholipid mixtures and factor VIII inhibitors: studies in haemophilic dogs. PhD Thesis University of Cambridge; 1988
• 14 Pijnappels MIM, Briet E, van der Zweet GTh, Huisden R, van Tilburg NH, Eulderink F. Evaluation of the cuticle bleeding time in canine haemophilia A. Thromb Haemost 1986; 55: 70-73
• 15 Kirkwood TBL, Snape TJ. Biometric principles in clotting and clot lysis assays. Clin Lab Haematol 1980; 2: 155-167
• 16 Brinkhous KM, Sandberg H, Garris JB, Mattsson C, Palm M, Griggs T, Read SM. Purified human factor VIII procoagulant protein: comparative hemostatic response after infusion into hemophilic and von Willebrand disease dogs. Proc Natl Acad Sci USA 1985; 82: 8752-8756
• 17 Altieri DC, Capitanio AM, Mannucci PM. Von Willebrand factor contaminating porcine factor VIII concentrate (Hyate C) causes platelet aggregation. Br J Haematol 1986; 63: 703-711