A combinatorial library strategy for the rapid humanization of anticarcinoma BR96 Fab

Mae Joanne Rosok, Dale E. Yelton, Linda J. Harris, Jürgen Bajorath, Karl Erik Hellström, Ingegerd Hellström, Gina A. Cruz, Karin Kristensson, Huey C Lin, William D. Huse, Scott M. Glaser

Research output: Contribution to journalArticle

58 Scopus citations

Abstract

We have used a combinatorial mutagenesis strategy to humanize BR96, a monoclonal antibody that binds to the Lewis Y class of tumor antigens. This approach allows simultaneous assessment of hundreds of humanized variable regions to identify the molecules that best preserve affinity, thus overcoming the major drawback of current humanization procedures, the requirement to construct and analyze each humanized antibody separately. Murine residues of BR96 were mutated to human if they were solvent-exposed residues that did not participate in the formation of the antigen binding site and were not at the interface of the light and heavy chain. At positions that might be involved in binding to antigen, the choice between the murine and human residue was more difficult. Murine and human alternatives were incorporated into a combinatorial library at positions representing buried residues that might affect the structural integrity of the antigen binding site. By encoding this library of humanized BR96 Fabs in an M13 phage vector, we rapidly identified several candidates with nearly identical antigen binding, within 2.fold, of the chimeric Fab. Additional mutagenesis directed at sites suggested in the literature as potentially important for antigen binding in a similar anti-Lewis Y antibody yielded no further improvements.

Original languageEnglish (US)
Pages (from-to)22611-22618
Number of pages8
JournalJournal of Biological Chemistry
Volume271
Issue number37
DOIs
StatePublished - 1996
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry

Fingerprint Dive into the research topics of 'A combinatorial library strategy for the rapid humanization of anticarcinoma BR96 Fab'. Together they form a unique fingerprint.

  • Cite this

    Rosok, M. J., Yelton, D. E., Harris, L. J., Bajorath, J., Hellström, K. E., Hellström, I., Cruz, G. A., Kristensson, K., Lin, H. C., Huse, W. D., & Glaser, S. M. (1996). A combinatorial library strategy for the rapid humanization of anticarcinoma BR96 Fab. Journal of Biological Chemistry, 271(37), 22611-22618. https://doi.org/10.1074/jbc.271.37.22611