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A bivalent disulfide-stabilized Fv with improved antigen binding to erbB2

T K Bera1, M Onda, U Brinkmann

  • 1National Cancer Institute, National Institutes of Health, Building 37, Bethesda, MD, 20892-4255, USA.

Journal of Molecular Biology
|August 12, 1998
PubMed
Summary

Researchers engineered a stable, bivalent antibody fragment (dsFv)2 targeting erbB2. This engineered antibody fragment, when fused to a toxin, shows enhanced cancer cell killing and stability, making it promising for immunotherapy.

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Area of Science:

  • Biotechnology
  • Immunology
  • Oncology

Background:

  • Monoclonal antibody e23 targets the erbB2 antigen.
  • Developing effective cancer immunotherapies requires stable and potent therapeutic agents.

Purpose of the Study:

  • To engineer a stable, bivalent disulfide-stabilized Fv (dsFv)2 molecule from the anti-erbB2 antibody e23.
  • To create and evaluate a bivalent dsFv(2)-based immunotoxin for enhanced cytotoxicity and stability.

Main Methods:

  • Protein engineering was used to create a disulfide-stabilized bivalent Fv (dsFv)2 molecule with a flexible linker.
  • The dsFv(2) molecule was fused to a truncated Pseudomonas exotoxin to form an immunotoxin.
  • The immunotoxin was expressed in E. coli, refolded, purified, and tested for binding affinity and cytotoxicity.

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Main Results:

  • The bivalent dsFv(2) molecule exhibited higher affinity for erbB2 compared to monovalent dsFv and similar affinity to the parental antibody.
  • The dsFv(2) immunotoxin demonstrated 5- to 20-fold greater cytotoxicity against erbB2-positive cancer cell lines than the monovalent version.
  • The bivalent dsFv(2) molecule showed high stability, retaining 94% activity after 24 hours in human serum at 37°C.

Conclusions:

  • The engineered bivalent dsFv(2) molecule is a stable and effective format for antibody-based therapeutics.
  • The dsFv(2) immunotoxin shows significant potential as a reagent for cancer immunotherapy and diagnosis due to its enhanced efficacy and stability.