Generation and characterization of a diabody targeting the αvβ6 integrin

PLoS One. 2013 Sep 4;8(9):e73260. doi: 10.1371/journal.pone.0073260. eCollection 2013.

Abstract

The αvβ6 integrin is up-regulated in cancer and wound healing but it is not generally expressed in healthy adult tissue. There is increasing evidence that it has a role in cancer progression and will be a useful target for antibody-directed cancer therapies. We report a novel recombinant diabody antibody fragment that targets specifically αvβ6 and blocks its function. The diabody was engineered with a C-terminal hexahistidine tag (His tag), expressed in Pichia pastoris and purified by IMAC. Surface plasmon resonance (SPR) analysis of the purified diabody showed affinity in the nanomolar range. Pre-treatment of αvβ6-expressing cells with the diabody resulted in a reduction of cell migration and adhesion to LAP, demonstrating biological function-blocking activity. After radio-labeling, using the His-tag for site-specific attachment of (99m)Tc, the diabody retained affinity and targeted specifically to αvβ6-expressing tumors in mice bearing isogenic αvβ6 +/- xenografts. Furthermore, the diabody was specifically internalized into αvβ6-expressing cells, indicating warhead targeting potential. Our results indicate that the new αvβ6 diabody has a range of potential applications in imaging, function blocking or targeted delivery/internalization of therapeutic agents.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibody Specificity
  • Antigens, Neoplasm / immunology*
  • Antigens, Neoplasm / metabolism
  • Cell Line
  • Female
  • Gene Expression Regulation, Neoplastic
  • Humans
  • Immunoglobulin Fragments / chemistry
  • Immunoglobulin Fragments / genetics*
  • Immunoglobulin Fragments / immunology*
  • Integrins / immunology*
  • Integrins / metabolism
  • Isotope Labeling
  • Mice
  • Pichia / genetics
  • Protein Engineering*
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics*
  • Recombinant Proteins / immunology*
  • Skin Neoplasms / metabolism
  • Technetium / chemistry

Substances

  • Antigens, Neoplasm
  • Immunoglobulin Fragments
  • Integrins
  • Recombinant Proteins
  • integrin alphavbeta6
  • Technetium