Derivatives of Human Complement Component C3 for Therapeutic Complement Depletion: A Novel Class of Therapeutic Agents

  • David C. Fritzinger
  • Brian E. Hew
  • June Q. Lee
  • James Newhouse
  • Maqsudul Alam
  • John R. Ciallella
  • Mallory Bowers
  • William B. Gorsuch
  • Benjamin J.  Guikema
  • Gregory L.  Stahl
  • Carl-Wilhelm Vogel
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 632)


To obtain proteins with the complement-depleting activity of Cobra Venom Factor (CVF), but with less immunogenicity, we have prepared human C3/CVF hybrid proteins, in which the C-terminus of the α-chain of human C3 is exchanged with homologous regions of the C-terminus of the β -chain of CVF. We show that these hybrid proteins are able to deplete complement, both in vitro and in vivo. One hybrid protein, HC3-1496, is shown to be effective in reducing complement-mediated damage in two disease models in mice, collagen-induced arthritis and myocardial ischemia/reperfusion injury. Human C3/CVF hybrid proteins represent a novel class of biologicals as potential therapeutic agents in many diseases where complement is involved in the pathogenesis.


Complement Activation Complement Component Paroxysmal Nocturnal Hemoglobinuria Hybrid Protein Complement Deficiency 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



Part of the research was supported by Incode Biopharmaceutics Corporation, Lahaina, Hawaii, USA.


  1. Andrä, J., Halter, R., Kock, M. A., Niemann, H., Vogel, C.-W., and Paul, D. (2002). Generation and characterization of transgenic mice expressing cobra venom factor. Mol Immunol 39, 357–365PubMedCrossRefGoogle Scholar
  2. Brand, D. D., Myers, L. K., Terato, K., Whittington, K. B., Stuart, J. M., Kang, A. H., and Rosloniec, E. F. (1994). Characterization of the T cell determinants in the induction of autoimmune arthritis by bovine alpha 1(II)-CB11 in H-2q mice. J Immunol 152, 3088–3097Google Scholar
  3. Carroll, M. C. (2004). The complement system in B cell regulation. Mol Immunol 41, 141–146PubMedCrossRefGoogle Scholar
  4. Cochrane, C. G., Müller-Eberhard, H. J., and Aikin, B. S. (1970). Depletion of plasma complement in vivo by a protein of cobra venom: its effect on various immunologic reactions. J Immunol 105, 55–69PubMedGoogle Scholar
  5. Costa, C., Zhao, L., Burton, W. V., Rosas, C., Bondioli, K. R., Williams, B. L., Hoagland, T. A., Dalmasso, A. P., and Fodor, W. L. (2002). Transgenic pigs designed to express human CD59 and H-transferase to avoid humoral xenograft rejection. Xenotransplantation 9, 45–57PubMedCrossRefGoogle Scholar
  6. de Bruijn, M. H. and Fey, G. H. (1985). Human complement component C3: cDNA coding sequence and derived primary structure. Proc Natl Acad Sci U S A 82, 708–712PubMedCrossRefGoogle Scholar
  7. Fodor, W. L., Rollins, S. A., Guilmette, E. R., Setter, E., and Squinto, S. P. (1995). A novel bifunctional chimeric complement inhibitor that regulates C3 convertase and formation of the membrane attack complex. J Immunol 155, 4135–4138PubMedGoogle Scholar
  8. Fritzinger, D. C., Petrella, E. C., Connelly, M. B., Bredehorst, R., and Vogel, C.-W. (1992). Primary structure of cobra complement component C3. J Immunol 149, 3554–3562PubMedGoogle Scholar
  9. Fritzinger, D. C., Bredehorst, R., and Vogel, C.-W. (1994). Molecular cloning and derived primary structure of cobra venom factor. Proc Natl Acad Sci U S A 91, 12775–12779PubMedCrossRefGoogle Scholar
  10. Fritzinger, D. C., Hew, B. E., Thorne, M., and Vogel, C.-W. (2004). Functional characterization of cobra venom factor/cobra C3 hybrid proteins. Mol Immunol 41, 230Google Scholar
  11. Fritzinger, D. C., Hew, B. E., Pangburn, M. K., and Vogel, C.-W. (2008). Generation of human C3 derivatives with CVF-like function for therapeutic complement depletion. FASEB J 19, A324Google Scholar
  12. Fritzinger, D. C., Hew, B. E., Pangburn, M. K., Janssen, B. J. C., Gros, P., and Vogel, C.-W. (2008). Human C3/cobra venom factor hybrid proteins with potential therapeutic applications. Mol Immunol 43, 141–142CrossRefGoogle Scholar
  13. Fritzinger, D. C., Hew, B. E., Lee, J. Q., and Vogel, C.-W. (2007). Human C3/cobra venom factor hybrid proteins for therapeutic complement depletion: in vivo activity and fine mapping of important domains. Mol Immunol 44, 3945CrossRefGoogle Scholar
  14. Fritzinger, D. C., Hew, B. E., Thorne, M., Pangburn, M. K., Janssen, B. J., Gros, P., and Vogel, C.-W. (2007b). Human C3 derivatives with Cobra venom factor-like functions for therapeutic complement depletion. Dev Comp Immunol, in pressGoogle Scholar
  15. Fujita, T., Matsushita, M., and Endo, Y. (2004). The lectin-complement pathway – its role in innate immunity and evolution. Immunol Rev 198, 185–202PubMedCrossRefGoogle Scholar
  16. Gowda, D. C., Schultz, M., Bredehorst, R., and Vogel, C.-W. (1992). Structure of the major oligosaccharide of cobra venom factor. Mol Immunol 29, 335–342PubMedCrossRefGoogle Scholar
  17. Gowda, D. C., Petrella, E. C., Raj, T. T., Bredehorst, R., and Vogel, C.-W.. (1994)Immunoreactivity and function of oligosaccharides in cobra venom factor. J Immunol 152, 2977–2986PubMedGoogle Scholar
  18. Gowda, D. C., Glushka, J., Halbeek, H., Thotakura, R. N., Bredehorst, R., and Vogel, C.-W. (2001). N-linked oligosaccharides of cobra venom factor contain novel alpha(1-3)galactosylated Le(x) structures. Glycobiology 11, 195–208PubMedCrossRefGoogle Scholar
  19. Grier, A. H., Schultz, M., and Vogel, C.-W. (1987). Cobra venom factor and human C3 share carbohydrate antigenic determinants. J Immunol 139, 1245–1252PubMedGoogle Scholar
  20. Guo, R. F. and Ward, P. A. (2005). Role of C5a in inflammatory responses. Annu Rev Immunol 23, 821–852PubMedCrossRefGoogle Scholar
  21. Hebell, T., Ahearn, J. M., and Fearon, D. T. (1991). Suppression of the immune response by a soluble complement receptor of B lymphocytes. Science 254, 102–105PubMedCrossRefGoogle Scholar
  22. Hew, B. E., Thorne, M., Fritzinger, D. C., and Vogel, C.-W. (2004). Humanized cobra venom factor (CVF): Generation of Human C3 derivitives with CVF-like function. Mol Immunol 41, 244–244Google Scholar
  23. Hillmen, P., Hall, C., Marsh, J. C., Elebute, M., Bombara, M. P., Petro, B. E., Cullen, M. J., Richards, S. J., Rollins, S. A., Mojcik, C. F., and Rother, R. P. (2004). Effect of eculizumab on hemolysis and transfusion requirements in patients with paroxysmal nocturnal hemoglobinuria. N Engl J Med 350, 552–559PubMedCrossRefGoogle Scholar
  24. Jacobson , M. P., Pincus, D. L., Rapp, C. S., Day, T. J., Honig, B., Shaw, D. E., and Friesner, R. A. (2004). A hierarchical approach to all-atom protein loop prediction. Proteins 55, 351–367PubMedCrossRefGoogle Scholar
  25. Janssen, B. J., Huizinga, E. G., Raaijmakers, H. C., Roos, A., Daha, M. R., Nilsson-Ekdahl, K., Nilsson, B., and Gros, P. (2005). Structures of complement component C3 provide insights into the function and evolution of immunity. Nature 437, 505–511PubMedCrossRefGoogle Scholar
  26. Janssen, B. J., Christodoulidou, A., McCarthy, A., Lambris, J. D., and Gros, P. (2006). Structure of C3b reveals conformational changes that underlie complement activity. Nature 444, 213–216PubMedCrossRefGoogle Scholar
  27. Kock, M. A., Hew, B. E., Bammert, H., Fritzinger, D. C., and Vogel, C. W. (2004). Structure and function of recombinant cobra venom factor. J Biol Chem 279, 30836–30843PubMedCrossRefGoogle Scholar
  28. Kölln, J., Spillner, E., Andrä, J., Klensang, K., and Bredehorst, R. (2004a). Human C3 derivatives engineered for decomplementation by forming stable C3 convertases. Mol Immunol 41, 259CrossRefGoogle Scholar
  29. Kölln, J., Spillner, E., Andrä, J., Klensang, K., and Bredehorst, R. (2004b). Complement inactivation by recombinant human C3 derivatives. J Immunol 173, 5540–5545Google Scholar
  30. Kölln, J., Bredehorst, R., and Spillner, E. (2005). Engineering of human complement component C3 for catalytic inhibition of complement. Immunol Lett 98, 49–56PubMedCrossRefGoogle Scholar
  31. Lachmann, P. J. and Halbwachs, L. (1975). The influence of C3b inactivator (KAF) concentration on the ability of serum to support complement activation. Clin Exp Immunol 21, 109–114PubMedGoogle Scholar
  32. Maillard, J. L., and Zarco, R. M. (1968). [Decomplementization by a factor extracted from cobra venom. Effect on several immune reactions of the guinea pig and rat]. Ann Inst Pasteur (Paris) 114, 756–774Google Scholar
  33. Medicus, R. G., Götze, O., and Müller-Eberhard, H. J. (1976). The serine protease nature of the C3 and C5 convertases of the classical and alternative complement pathways. Scand J Immunol 5, 1049–1055PubMedCrossRefGoogle Scholar
  34. Mollnes, T. E. and Kirschfink, M. (2006). Strategies of therapeutic complement inhibition. Mol Immunol 43, 107–121PubMedCrossRefGoogle Scholar
  35. Morgan, B. P. and Harris, C. L. (2003). Complement therapeutics; history and current progress. Mol Immunol 40, 159–170PubMedCrossRefGoogle Scholar
  36. Nagaki, K., Iida, K., Okubo, M., and Inai, S. (1978). Reaction mechanisms of beta1H globulin. Int Arch Allergy Appl Immunol 57, 221–232PubMedCrossRefGoogle Scholar
  37. Nelson, R. A., Jr. (1966). A new concept of immunosuppression in hypersensitivity reactions and in transplantation immunity. Surv Ophthalmol 11, 498–505PubMedGoogle Scholar
  38. Pangburn, M. K., Schreiber, R. D., and Müller-Eberhard, H. J. (1977). Human complement C3b inactivator: isolation, characterization, and demonstration of an absolute requirement for the serum protein beta1H for cleavage of C3b and C4b in solution. J Exp Med 146, 257–270PubMedCrossRefGoogle Scholar
  39. Pangburn, M. K. and Müller-Eberhard, H. J. (1986). The C3 convertase of the alternative pathway of human complement. Enzymic properties of the bimolecular proteinase. Biochem J 235, 723–730PubMedGoogle Scholar
  40. Sahu, A. and Lambris, J. D. (2000). Complement inhibitors: a resurgent concept in anti-inflammatory therapeutics. Immunopharmacology 49, 133–148PubMedCrossRefGoogle Scholar
  41. Shernan, S. K., Fitch, J. C., Nussmeier, N. A., Chen, J. C., Rollins, S. A., Mojcik, C. F., Malloy, K. J., Todaro, T. G., Filloon, T., Boyce, S. W., Gangahar, D. M., Goldberg, M., Saidman, L. J., and Mangano, D. T. (2004). Impact of pexelizumab, an anti-C5 complement antibody, on total mortality and adverse cardiovascular outcomes in cardiac surgical patients undergoing cardiopulmonary bypass. Ann Thorac Surg 77, 942–949; discussion 949–950PubMedCrossRefGoogle Scholar
  42. Till, G. O., Johnson, K. J., Kunkel, R., and Ward, P. A. (1982). Intravascular activation of complement and acute lung injury. Dependency on neutrophils and toxic oxygen metabolites. J Clin Invest 69, 1126–1135PubMedCrossRefGoogle Scholar
  43. Till, G. O., Morganroth, M. L., Kunkel, R., and Ward, P. A. (1987). Activation of C5 by cobra venom factor is required in neutrophil-mediated lung injury in the rat. Am J Pathol 129, 44–53PubMedGoogle Scholar
  44. Vogel, C.-W. (1991). Cobra venom factor, the complement-activating protein of cobra venom. In: Anthony Tu (ed.). Handbook of Natural Toxins: Reptile and Amphibian Venoms, Marcel Dekker, New York, pp. 147–188Vol. 5.Google Scholar
  45. Vogel, C.-W., and Müller-Eberhard, H. J. (1982). The cobra venom factor-dependent C3 convertase of human complement. A kinetic and thermodynamic analysis of a protease acting on its natural high molecular weight substrate. J Biol Chem 257, 8292–8299PubMedGoogle Scholar
  46. Vogel, C.-W., and Müller-Eberhard, H. J. (1984). Cobra venom factor: improved method for purification and biochemical characterization. Journal of Immunology Methods 73, 203–220CrossRefGoogle Scholar
  47. Vogel, C.-W. and Fritzinger, D. C. (2007). Humanized cobra venom factor: experimental therapeutics for targeted complement activation and complement depletion. Curr Pharm Des 13, 2916–2926PubMedCrossRefGoogle Scholar
  48. Vogel, C.-W., Smith, C. A., and Müller-Eberhard, H. J. (1984). Cobra venom factor: structural homology with the third component of human complement. J Immunol 133, 3235–3241PubMedGoogle Scholar
  49. Vogel, C.-W., Bredehorst, R., Fritzinger, D. C., Grunwald, T., Ziegelmüller, P., and Kock, M. A. (1996). Structure and function of cobra venom factor, the complement-activating protein in cobra venom. Adv Exp Med Biol 391, 97–114PubMedCrossRefGoogle Scholar
  50. Vogel, C. W., Fritzinger, D. C., Hew, B. E., Thorne, M., and Bammert, H. (2004). Recombinant cobra venom factor. Mol Immunol 41, 191–199PubMedCrossRefGoogle Scholar
  51. Walsh, M. C., Bourcier, T., Takahashi, K., Shi, L., Busche, M. N., Rother, R. P., Solomon, S. D., Ezekowitz, R. A., and Stahl, G. L. (2005). Mannose-binding lectin is a regulator of inflammation that accompanies myocardial ischemia and reperfusion injury. J Immunol 175, 541–546PubMedGoogle Scholar
  52. Wehrhahn, D., Meiling, K., Fritzinger, D. C., Bredehorst, R., Andrä, J., and Vogel, C.-W. (2000). Analysis of the structure/function relationship of Cobra Venom Factor (CVF) and C3: generation of CVF/cobraC3 hybrid proteins. Immunopharmacology 49, 94CrossRefGoogle Scholar
  53. Weisman, H. F., Bartow, T., Leppo, M. K., Marsh, H. C., Jr., Carson, G. R., Concino, M. F., Boyle, M. P., Roux, K. H., Weisfeldt, M. L., and Fearon, D. T. (1990). Soluble human complement receptor type 1: in vivo inhibitor of complement suppressing post-ischemic myocardial inflammation and necrosis. Science 249, 146–151PubMedCrossRefGoogle Scholar
  54. Whaley, K., and Ruddy, S. (1976). Modulation of the alternative complement pathways by beta1H globulin. J Exp Med 144, 1147–1163PubMedCrossRefGoogle Scholar
  55. Zhou, C. Y., McInnes, E., Copeman, L., Langford, G., Parsons, N., Lancaster, R., Richards, A., Carrington, C., and Thompson, S. (2005). Transgenic pigs expressing human CD59, in combination with human membrane cofactor protein and human decay-accelerating factor. Xenotransplantation 12, 142–148PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • David C. Fritzinger
    • 1
  • Brian E. Hew
  • June Q. Lee
  • James Newhouse
    • 2
  • Maqsudul Alam
    • 3
  • John R. Ciallella
    • 4
  • Mallory Bowers
  • William B. Gorsuch
    • 5
  • Benjamin J.  Guikema
  • Gregory L.  Stahl
  • Carl-Wilhelm Vogel
  1. 1.Cancer Research Center of HawaiiUniversity of Hawaii at ManoaHonoluluUSA
  2. 2.Maui High Performance Computing CenterKiheiUSA
  3. 3.Advanced Studies in Genomics, Proteomics, and BioinformaticsUniversity of Hawaii at ManoaHonoluluUSA
  4. 4.Melior Discovery CorporationExtonUSA
  5. 5.Brigham and Women’s Hospital, Center for Experimental Therapeutics and Reperfusion InjuryHarvard UniversityBostonUSA

Personalised recommendations