The cleavage site of C5 from man and animals as a common target for neutralizing human monoclonal antibodies

In vitro and in vivo studies

Roberto Marzari, Daniele Sblattero, Paolo Macor, Fabio Fischetti, Renato Gennaro, James D. Marks, Andrew Bradbury, Francesco Tedesco

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

The isolation of an anti-C5 single-chain fragment variable (scFv) antibody, TS-A12/22, from a human phage display library, is described. This antibody inhibits the activation of C5 and the assembly of the terminal complement complex implicated in cell and tissue damage. Using antibody-sensitized sheep erythrocytes and rabbit red cells as target cells in hemolytic assays, we found that TS-A12/22 inhibited the activation of C5 by the convertases of both classical and alternative pathways. Western blot analysis and competition experiments with synthetic peptides showed that TS-A12/22 reacted with the α chain of C5 and recognized the cleavage site of this complement component by the C5 convertase. As a result, the antibody prevented splitting of C5 and inhibited the generation of C5a and of the terminal complement complex. The identification of the TS-A12/22 recognition site as a conserved sequence in man, mouse, rat and rabbit enabled the demonstration of in vitro inhibition of complement activity in these species. The scFv TS-A12/22 was tested in a rat model of antigen-induced arthritis and proved to be effective in preventing influx of polymorphonuclear cells into the knee joint and C9 deposition on synovial tissue.

Original languageEnglish
Pages (from-to)2773-2782
Number of pages10
JournalEuropean Journal of Immunology
Volume32
Issue number10
DOIs
Publication statusPublished - Oct 2002

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compound A 12
Monoclonal Antibodies
Complement Membrane Attack Complex
Single-Chain Antibodies
Alternative Pathway Complement C5 Convertase
Classical Pathway Complement C5 Convertase
Antibodies
Complement C3-C5 Convertases
Rabbits
Conserved Sequence
Knee Joint
Bacteriophages
Libraries
Arthritis
Sheep
Erythrocytes
Western Blotting
In Vitro Techniques
Antigens
Peptides

Keywords

  • Complement
  • Inflammation
  • Phage display library
  • Single-chain fragment variable antibody

ASJC Scopus subject areas

  • Immunology

Cite this

The cleavage site of C5 from man and animals as a common target for neutralizing human monoclonal antibodies : In vitro and in vivo studies. / Marzari, Roberto; Sblattero, Daniele; Macor, Paolo; Fischetti, Fabio; Gennaro, Renato; Marks, James D.; Bradbury, Andrew; Tedesco, Francesco.

In: European Journal of Immunology, Vol. 32, No. 10, 10.2002, p. 2773-2782.

Research output: Contribution to journalArticle

Marzari, Roberto ; Sblattero, Daniele ; Macor, Paolo ; Fischetti, Fabio ; Gennaro, Renato ; Marks, James D. ; Bradbury, Andrew ; Tedesco, Francesco. / The cleavage site of C5 from man and animals as a common target for neutralizing human monoclonal antibodies : In vitro and in vivo studies. In: European Journal of Immunology. 2002 ; Vol. 32, No. 10. pp. 2773-2782.
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