Selection and characterization of anti-MUC-1 scFvs intended for targeted therapy

Michelle D. Winthrop, Sally J. DeNardo, Huguette Albrecht, Gary R. Mirick, Linda A. Kroger, Kathleen R. Lamborn, Çeslovas Venclovas, Michael E. Colvin, Patricia A. Burke, Gerald L Denardo

Research output: Contribution to journalArticle

20 Scopus citations

Abstract

Purpose: The selection and characterization of anti-MUC-1 single-chain antibody fragments (scFv) is a first step toward the construction of new anticancer molecules designed for optimal blood clearance and tumor penetration. The mucin MUC-1 was chosen as an antigen because it is abundantly expressed on epithelial cancers in an aberrantly glycosylated form, making it structurally and antigenically distinct from MUC-1 expressed on normal cells. Experimental Design: A previously constructed anti-MUC-1 phage display library from hyperimmunized mice, with 5 × 105 calculated variants, was screened for the selection of anti-MUC-1 scFvs. Selection criteria were high binding to a MUC-1 peptide containing 4 tandem repeats of 20 amino acids and to MUC-1-positive MCF-7 (human breast cancer) cell lysates in ELISA. Results: Six anti-MUC-1 scFv clones were selected and characterized. Nucleotide sequencing showed that four of them were full length scFv genes (variable heavy chain + variable light chain), whereas the remaining two contained either a variable heavy chain or a variable light chain alone. Their binding affinities (K a) range between 8 × 107 and 109 M -1. Immunohistopathology demonstrated reactivity with breast cancer cells (MCF-7 and BT20) and human breast biopsy tissue. Molecular modeling revealed high structural similarity of the anti-MUC-1 scFvs with the X-ray-determined structure of the anti-CEA scFv (MFE-23). Conclusions: In vitro antigen binding was demonstrated for the selected anti-MUC-1 scFvs. The binding affinities of these scFvs are in a promising range for efficient in vivo antigen binding. These anti-MUC-1 scFvs will be evaluated as antigen-binding modules in new multifunctional agents for the detection and therapy of cancer.

Original languageEnglish (US)
JournalClinical Cancer Research
Volume9
Issue number10 II
StatePublished - Oct 1 2003

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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    Winthrop, M. D., DeNardo, S. J., Albrecht, H., Mirick, G. R., Kroger, L. A., Lamborn, K. R., Venclovas, Ç., Colvin, M. E., Burke, P. A., & Denardo, G. L. (2003). Selection and characterization of anti-MUC-1 scFvs intended for targeted therapy. Clinical Cancer Research, 9(10 II).