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Rabbit immune repertoires as sources for therapeutic monoclonal antibodies: the impact of kappa allotype-correlated variation in cysteine content on antibody libraries selected by phage display

Academic Article
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Overview

authors

  • Popkov, M.
  • Mage, R. G.
  • Alexander, C. B.
  • Thundivalappil, S.
  • Barbas III, Carlos
  • Rader, Christoph

publication date

  • January 2003

journal

  • Journal of Molecular Biology  Journal

abstract

  • The rabbit immune repertoire has long been a rich source of diagnostic polyclonal antibodies. Now it also holds great promise as a source of therapeutic monoclonal antibodies. On the basis of phage display technology, we recently reported the first humanization of a rabbit monoclonal antibody. The allotypic diversity of rabbit immunoglobulins prompted us to compare different rabbit immune repertoires for the generation and humanization of monoclonal antibodies that bind with strong affinity to antigens involved in tumor angiogenesis. In particular, we evaluated the diversity of unselected and selected chimeric rabbit/human Fab libraries that were derived from different kappa light chain allotypes. Most rabbit light chains have an extra disulfide bridge that links the variable and constant domains in addition to the two intrachain disulfide bridges shared with mouse and human kappa light chains. Here we evaluate the impact of this increased disulfide bridge complexity on the generation and selection of chimeric rabbit/human Fab libraries. We demonstrate that rabbits with mutant bas and wild-type parental b9 allotypes are excellent sources for therapeutic monoclonal antibodies. Featured among the selected clones with b9 allotype is a rabbit/human Fab that binds with a dissociation constant of 1nM to both human and mouse Tie-2, which will facilitate its evaluation in mouse models of human cancer. Examination of 228 new rabbit antibody sequences allowed for a comprehensive comparison of the LCDR3 and HCDR3 length diversity in rabbits. This study revealed that rabbits exhibit an HCDR3 length distribution more closely related to human antibodies than mouse antibodies.

subject areas

  • Animals
  • Antibodies, Monoclonal
  • Binding Sites, Antibody
  • Cysteine
  • DNA Fingerprinting
  • Disulfides
  • Flow Cytometry
  • Humans
  • Immunization
  • Immunoglobulin Allotypes
  • Immunoglobulin Fab Fragments
  • Mice
  • Peptide Library
  • Protein Binding
  • Protein Structure, Tertiary
  • Rabbits
  • Recombinant Fusion Proteins
  • Surface Plasmon Resonance
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Research

keywords

  • antibody engineering
  • antibody library
  • immunoglobulin kappa light chain
  • phage display
  • rabbit monoclonal antibodies
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Identity

International Standard Serial Number (ISSN)

  • 0022-2836

Digital Object Identifier (DOI)

  • 10.1016/s0022-2836(02)01232-9

PubMed ID

  • 12488098
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Additional Document Info

start page

  • 325

end page

  • 335

volume

  • 325

issue

  • 2

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