Download PDF
Semin Thromb Hemost 2001; 27(4): 395-404
DOI: 10.1055/s-2001-16892
Copyright © 2001 by Thieme Medical Publishers, Inc., 333 Seventh Avenue, New York, NY 10001, USA. Tel.: +1(212) 584-4662

Recombinant von Willebrand Factor: Preclinical Development

Barbara Plaimauer1 , Uwe Schlokat1 , Peter L. Turecek2 , Artur Mitterer1 , Wolfgang Mundt1 , Wilfried Auer2 , Ludwig Pichler2 , Herbert Gritsch,2 Friedrich Dorner1 , Hans-Peter Schwarz2
  • 1Biomedical Research Center, Baxter AG, Hyland-Immuno Division, Orth/Donau, Austria
  • 2Baxter AG, Hyland-Immuno Division, Vienna, Austria
Further Information

Publication History

Publication Date:
31 August 2001 (online)

    Permissions and Reprints

ABSTRACT

Von Willebrand factor (vWF) is a multimeric glycoprotein (GP) that attracts platelets to the site of vascular injury, mediates platelet-platelet interaction, and stabilizes factor VIII (FVIII) in the circulation. Quantitative and qualitative defects of vWF result in von Willebrand disease (vWD), manifested by modest to severe bleeding episodes. Substitution therapy, with plasma-derived FVIII/vWF complex concentrates, is used for patients suffering the more severe forms of vWD. Efficacy of these preparations is often unsatisfactory because inadvertent proteolytic degradation during the manufacturing process causes them to lack the hemostatically most active high-molecular-weight multimers. In contrast, recombinant vWF (r-vWF), which is constitutively expressed at high yields in Chinese hamster ovary (CHO) cells and secreted into the conditioned medium under perfusion fermentation in ``protein-free'' medium, has high-molecular-weight multimers of extraordinary structural integrity. Functional analysis has shown that r-vWF promotes ristocetin cofactor-mediated platelet aggregation, collagen interaction and FVIII binding, and platelet-collagen adhesion under shear stress. Infusing vWF-deficient animals with r-vWF corrected vWF concentration and reduced blood loss, subsequently stabilizing endogenous FVIII associated with the reduction of bleeding time. Compared with plasma-derived vWF preparations, r-vWF was found to have a prolonged half-life, further enhancing the potential value of r-vWF as a therapeutic agent for treating patients suffering from vWD.

KEYWORD

von Willebrand factor - recombinant expression - multimers - biological activity - animal model

REFERENCES

  • 1 Sadler J E. Biochemistry and genetics of von Willebrand factor.  Annu Rev Biochem . 1998;  67 395-424
    CrossrefPubMedGoogle Scholar
  • 2 Ruggeri Z M. Structure and function of von Willebrand factor.  Thromb Haemost . 1999;  82 576-584
    PubMedGoogle Scholar
  • 3 Federici A B. Diagnosis of von Willebrand disease.  Haemophilia . 1998;  4 654-660
    CrossrefPubMedGoogle Scholar
  • 4 Ginsburg D. The molecular biology of von Willebrand disease.  Haemophilia . 1999;  5 19-27
    PubMedGoogle Scholar
  • 5 Sadler J E. A revised classification of von Willebrand disease.  Thromb Haemost . 1994;  71 520-525
    Article in Thieme ConnectPubMedGoogle Scholar
  • 6 Sadler J E, Mannucci P M, Berntorp E. Impact, diagnosis and treatment of von Willebrand disease.  Thromb Haemost . 2000;  84 160-174
    Article in Thieme ConnectPubMedGoogle Scholar
  • 7 Mannucci P M. Treatment of von Willebrand disease.  Haemophilia . 1998;  4 661-664
    CrossrefPubMedGoogle Scholar
  • 8 Mannucci P M. Desmopressin (DDAVP) in the treatment of bleeding disorders: the first twenty years.  Haemophilia . 2000;  6 60-67
    PubMedGoogle Scholar
  • 9 Berntorp E. Plasma product treatment in various types of von Willebrand's disease.  Haemostasis . 1994;  24 289-297
    PubMedGoogle Scholar
  • 10 Dobrkovska A, Krzensk U, Chediak J R. Pharmacokinetics, efficacy and safety of Humate-P in von Willebrand disease.  Haemophilia . 1998;  4 33-39
    PubMedGoogle Scholar
  • 11 Berntorp E. Von Willebrand factor containing factor VIII concentrates.  Haemophilia . 1999;  5 60-63
    CrossrefPubMedGoogle Scholar
  • 12 Mannucci P M, Tenconi P M, Castaman G. Comparison of four virus-inactivated plasma concentrates for treatment of severe von Willebrand disease: a cross-over randomized trial.  Blood . 1992;  79 3130-3137
    PubMedGoogle Scholar
  • 13 Mannucci P M. Biochemical characteristics of therapeutic plasma concentrates used in the treatment of von Willebrand disease.  Haemostasis . 1994;  24 285-288
    PubMedGoogle Scholar
  • 14 Meyer D, Girma J-P. Von Willebrand factor: structure and function.  Thromb Haemost . 1993;  70 99-104
    PubMedGoogle Scholar
  • 15 Furlan M. Von Willebrand factor: molecular size and functional activity.  Ann Hematol . 1996;  72 341-348
    CrossrefPubMedGoogle Scholar
  • 16 Wise R J, Barr P J, Wong P A. Expression of a human proprotein processing enzyme: Correct cleavage of the von Willebrand factor precursor at a paired basic amino acid site.  Proc Natl Acad Sci USA . 1990;  87 9378-9382
    PubMedGoogle Scholar
  • 17 Himmelspach M, Plaimauer B, Dorner F. Furin as a tool for the endoproteolytic maturation of susceptible recombinant biopharmaceuticals. In: von Broekhoven A, Shapiro F, Anné J, eds. Novel Frontiers in the Production of Compounds for Biomedical Use Dordrecht, The Netherlands: Kluwer; 2001: 219-248
    Google Scholar
  • 18 Birch K A, Pober J S, Zavoico G B. Calcium/calmodulin transduces thrombin-stimulated secretion: studies in intact and minimally permeabilized human umbilical vein endothelial cells.  J Cell Biol . 1992;  118 1501-1510
    CrossrefPubMedGoogle Scholar
  • 19 Ribes J A, Francis C W, Wagner D D. Fibrin induces release of von Willebrand factor from endothelial cells.  J Clin Invest . 1987;  79 117-123
    PubMedGoogle Scholar
  • 20 Sporn L A, Marder V J, Wagner D D. Inducible secretion of large, biologically potent von Willebrand factor multimers.  Cell . 1986;  46 185-190
    CrossrefPubMedGoogle Scholar
  • 21 Ewenstein B M, Warhol M J, Handin R I. Composition of the von Willebrand factor storage organelle (Weibel-Palade body) isolated from cultured human umbilical vein endothelial cells.  J Cell Biol . 1987;  104 1423-1433
    CrossrefPubMedGoogle Scholar
  • 22 Sporn L A, Marder V J, Wagner D D. Von Willebrand factor released from Weibel-Palade bodies binds more avidly to extracellular matrix than that secreted constitutively.  Blood . 1987;  69 1531-1534
    PubMedGoogle Scholar
  • 23 Tsai H M, Nagel R L, Hatcher V B. Multimeric composition of endothelial cell-derived von Willebrand factor.  Blood . 1989;  73 2074-2076
    PubMedGoogle Scholar
  • 24 Dent J A, Berkowitz S D, Ware J. Identification of a cleavage site directing the immunochemical detection of molecular abnormalities in type IIA von Willebrand factor.  Proc Natl Acad Sci USA . 1990;  87 6306-6310
    PubMedGoogle Scholar
  • 25 Dent J A, Galbusera M, Ruggeri Z M. Heterogeneity of plasma von Willebrand factor multimers resulting from proteolysis of the constituent subunit.  J Clin Invest . 1991;  88 774-782
    CrossrefPubMedGoogle Scholar
  • 26 Furlan M, Robles R, Affolter D. Triplet structure of von Willebrand factor reflects proteolytic degradation of high molecular weight multimers.  Proc Natl Acad Sci USA . 1993;  90 7503-7507
    PubMedGoogle Scholar
  • 27 Fischer B E, Thomas K B, Schlokat U. Triplet structure of human von Willebrand factor.  Biochem J . 1998;  331 483-488
    PubMedGoogle Scholar
  • 28 Furlan M, Robles R, Lamie B. Partial purification and characterization of a protease from human plasma cleaving von Willebrand factor to fragments produced by in vivo proteolysis.  Blood . 1996;  87 4223-4234
    CrossrefPubMedGoogle Scholar
  • 29 Sadler J E, Shelton-Inloes B B, Sorace J M. Cloning of cDNA and genomic DNA for human von Willebrand factor.  Cold Spring Harb Symp Quant Biol . 1986;  51 515-523
    PubMedGoogle Scholar
  • 30 Bonthron D T, Handin R I, Kaufman R J. Structure of pre-pro-von Willebrand factor and its expression in heterologous cells.  Nature . 1986;  324 270-273
    CrossrefPubMedGoogle Scholar
  • 31 Meulien P, Nishino M, Mazurier C. Processing and characterization of recombinant von Willebrand factor expressed in different cell types using a vaccinia virus vector.  Thromb Haemost . 1992;  67 154-160
    PubMedGoogle Scholar
  • 32 Fischer B, Mitterer A, Schlokat U. Structural analysis of recombinant von Willebrand factor: identification of hetero- and homo-dimers.  FEBS Lett . 1994;  351 345-348
    CrossrefPubMedGoogle Scholar
  • 33 Schlokat U, Fischer B, Mitterer A. Herstellung und Charakterisierung von rekombinantem von Willebrand- Faktor zur therapeutischen Anwendung. In: Scharrer I, Schramm W, eds. 26 Hämophilie-Symposium Hamburg 1995 Berlin, Germany: Springer Verlag Publishers 1996: 147-158
    Google Scholar
  • 34 Fischer B E, Schlokat U, Mitterer A. Structural analysis of recombinant von Willebrand factor produced at industrial scale fermentation of transformed CHO cells co-expressing recombinant furin.  FEBS Lett . 1995;  375 259-262
    CrossrefPubMedGoogle Scholar
  • 35 Fischer B E, Schlokat U, Reiter M. Biochemical and functional characterization of recombinant von Willebrand factor produced on a large scale.  Cell Mol Life Sci . 1997;  53 943-950
    CrossrefPubMedGoogle Scholar
  • 36 Rehemtulla A, Dorner A J, Kaufman R J. Regulation of PACE4 propeptide-processing activity: Requirement for a post-endoplasmic reticulum compartment and autoproteolytic activation.  Proc Natl Acad Sci USA . 1992;  89 8235-8239
    PubMedGoogle Scholar
  • 37 Rehemtulla A, Kaufman R J. Preferred sequence requirements for cleavage of pro-von Willebrand factor by propeptide-processing enzymes.  Blood . 1992;  79 2349-2355
    PubMedGoogle Scholar
  • 38 Schlokat U, Fischer B E, Herlitschka S. Production of highly homogeneous and structurally intact recombinant von Willebrand factor multimers by furin-mediated propeptide removal in vitro.  Biotechnol Appl Biochem . 1996;  24 257-267
    PubMedGoogle Scholar
  • 39 Hatsuzawa K, Nagahama M, Takahashi S. Purification and characterization of Furin, a Kex2-like processing endoprotease, produced in Chinese hamster ovary cells.  J Biol Chem . 1992;  267 16094-16099
    PubMedGoogle Scholar
  • 40 Vidricaire G, Denault J-B, Leduc R. Characterization of a secreted form of human furin endoprotease.  Biochem Biophys Res Commun . 1993;  195 1011-1018
    CrossrefPubMedGoogle Scholar
  • 41 Vey M, Schäfer W, Berghöfer S. Maturation of trans-Golgi network protease Furin: Compartmentalization of propeptide removal, substrate cleavage, and COOH-terminal truncation.  J Cell Biol . 1994;  127 1829-1842
    CrossrefPubMedGoogle Scholar
  • 42 Preininger A, Schlokat U, Mohr G. Strategies for recombinant furin employment in a biotechnological process: complete target protein precursor cleavage.  Cytotechnol . 1999;  30 1-15
    CrossrefPubMedGoogle Scholar
  • 43 Fischer B, Mitterer A, Dorner F. Comparison of N-glycan pattern of recombinant human coagulation factors II and IX expressed in Chinese hamster ovary (CHO) and African green monkey (Vero) cells.  J Thromb Thrombolysis . 1996;  3 57-62
    PubMedGoogle Scholar
  • 44 Plaimauer B, Schlokat U, Himmelspach M. Posttranslational modifications of recombinant von Willebrand factor: limitations and experimental improvement at high yield expression. In: Ikura K, Nagao M, Masuda S, Sasaki R, eds. Animal Cell Technology: Challenges for the 21st Century Dordrecht, The Netherlands: Kluwer 1999: 105-109
    Google Scholar
  • 45 Schwarz H P, Turecek P L, Pichler L. Recombinant von Willebrand factor.  Thromb Haemost . 1997;  78 571-576
    PubMedGoogle Scholar
  • 46 Siekmann J, Turecek P L, Schwarz H P. The determination of von Willebrand factor activity by collagen binding assay.  Haemophilia . 1998;  4 15-24
    PubMedGoogle Scholar
  • 47 Fischer B E, Kramer G, Mitterer A. Effect of multimerization of human and recombinant von Willebrand factor on platelet aggregation, binding to collagen and binding of coagulation factor VIII.  Thromb Res . 1996;  84 55-66
    CrossrefPubMedGoogle Scholar
  • 48 Malmqvist M. BIACORE: an affinity biosensor system for characterization of biomolecular interactions.  Biochem Soc Trans . 1999;  27 335-340
    PubMedGoogle Scholar
  • 49 Vlot A J, Koppelman S J, van den Berg H M. The affinity and stoichiometry of binding of human factor VIII to von Willebrand factor.  Blood . 1995;  85 3150-3157
    CrossrefPubMedGoogle Scholar
  • 50 Leyte A, Verbeet M P, Brodniewicz-Proba T. The interaction between human blood-coagulation factor VIII and von Willebrand factor. Characterization of a high-affinity binding site on factor VIII.  Biochem J . 1989;  257 679-683
    PubMedGoogle Scholar
  • 51 Wise R J, Dorner A J, Krane M. The role of von Willebrand factor multimers and propeptide cleavage in binding and stabilization of Factor VIII.  J Biol Chem . 1991;  266 21948-21955
    PubMedGoogle Scholar
  • 52 Bendetowicz A V, Morris J A, Wise R J. Binding of factor VIII to von Willebrand factor is enabled by cleavage of the von Willebrand factor propeptide and enhanced by formation of disulfide-linked multimers.  Blood . 1998;  92 529-538
    CrossrefPubMedGoogle Scholar
  • 53 Turecek P, Siekman J, Pichler L. Rekombinanter von Willebrand-Faktor: Präklinik. In: Scharrer I, Schramm W, eds. 26 Hämophilie-Symposium Hamburg 1995 Berlin, Germany: Springer Verlag Publishers 1996: 159-168
    Google Scholar
  • 54 Turecek P L, Gritsch H, Richter G. Assessment of bleeding for the evaluation of therapeutic preparations in small animal models of antibody-induced hemophilia and von Willebrand disease.  Thromb Haemost . 1997;  77 591-599
    PubMedGoogle Scholar
  • 55 Turecek P L, Gritsch H, Pichler L. In vivo characterization of recombinant von Willebrand factor in dogs with von Willebrand disease.  Blood . 1997;  90 3555-3567
    PubMedGoogle Scholar
  • 56 Turecek P L, Pichler L, Auer W. Evidence for extracellular processing of pro-von Willebrand factor after infusion in animals with and without severe von Willebrand disease.  Blood . 1999;  94 1637-1647
    PubMedGoogle Scholar
  • 57 Roussi J, Turecek P L, Andre P. Effects of human recombinant, plasma-derived and porcine von Willebrand factor in pigs with severe von Willebrand disease.  Blood Coagul Fibrinolysis . 1998;  9 361-372
    CrossrefPubMedGoogle Scholar
  • 58 Borchiellini A, Fijnvandraat K, ten Cate W J. Quantitative analysis of von Willebrand factor propeptide release in vivo: effect of experimental endotoxemia and administration of 1-deamino-8-D-arginine vasopressin in humans.  Blood . 1996;  88 2951-2958
    CrossrefPubMedGoogle Scholar