Conversion of murine antibodies to human antibodies and their optimization for ovarian cancer therapy targeted to the folate receptor

Mariangela Figini, Franck Martin, Renata Ferri, Elena Luison, Elena Ripamonti, Alberto Zacchetti, Mimosa Mortarino, Vito Di Cioccio, Giovanni Maurizi, Marcello Allegretti, Silvana Canevari

Research output: Contribution to journalArticle

Abstract

We previously developed murine and chimeric antibodies against a specific epithelial ovarian carcinoma (EOC) marker, named folate receptor (FR), and promising results were obtained in phase II trials. More recently, we successfully generated a completely human Fab fragment, C4, by conversion of one of the murine anti-FR antibodies to human antibody using phage display and guided selection. However, subsequent efforts to obtain C4 in a dimer format, which seems especially desirable for EOC locoregional treatment, resulted in a highly heterogeneous product upon natural dimerization and in a very poor production yield upon chemical dimerization by a non-hydrolyzable linker to a di-Fab-maleimide (DFM). We therefore designed, constructed and characterized a large Fab dual combinatorial human antibody phage display library obtained from EOC patients and potentially biased toward an anti-tumor response in an effort to obtain new anti-FR human antibodies suitable for therapy. Using this library and guiding the selection on FR-expressing cells with murine/human antibody chains, we generated four new human anti-FR antibody (AFRA) Fab fragments, one of which was genetically and chemically manipulated to obtain a chemical dimer, designated AFRA-DFM5.3, with high yield production and the capability for purification scaled-up to clinical grade. Overall affinity of AFRA-DFM5.3 was in the 2-digit nanomolar range, and immunohistochemistry indicated that the reagent recognized the FR expressed on EOC samples. 131I-AFRA-DFM5.3 showed high immunoreactivity, in vitro stability and integrity, and specifically accumulated only in FR-expressing tumors in subcutaneous preclinical in vivo models. Overall, our studies demonstrate the successful conversion of murine to completely human anti-FR antibodies through the combined use of antibody phage display libraries biased toward an anti-tumor response, guided selection and chain shuffling, and point to the suitability of AFRA5.3 for future clinical application in ovarian cancer.

Original languageEnglish
Pages (from-to)531-546
Number of pages16
JournalCancer Immunology, Immunotherapy
Volume58
Issue number4
DOIs
Publication statusPublished - Apr 2009

Keywords

  • Antibody fragment
  • Chemical dimerization
  • Folate receptor
  • Ovarian cancer
  • Phage display

ASJC Scopus subject areas

  • Cancer Research
  • Oncology
  • Immunology
  • Immunology and Allergy

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  • Cite this

    Figini, M., Martin, F., Ferri, R., Luison, E., Ripamonti, E., Zacchetti, A., Mortarino, M., Di Cioccio, V., Maurizi, G., Allegretti, M., & Canevari, S. (2009). Conversion of murine antibodies to human antibodies and their optimization for ovarian cancer therapy targeted to the folate receptor. Cancer Immunology, Immunotherapy, 58(4), 531-546. https://doi.org/10.1007/s00262-008-0575-5