High Affinity Binders to EphA2 Isolated from Abdurin Scaffold Libraries; Characterization, Binding and Tumor Targeting

PLoS One. 2015 Aug 27;10(8):e0135278. doi: 10.1371/journal.pone.0135278. eCollection 2015.

Abstract

Abdurins are a novel antibody-like scaffold derived from the engineering of a single isolated CH2 domain of human IgG. Previous studies established the prolonged serum half-life of Abdurins, the result of a retained FcRn binding motif. Here we present data on the construction of large, diverse, phage-display and cell-free DNA display libraries and the isolation of high affinity binders to the cancer target, membrane-bound ephrin receptor tyrosine kinase class A2 (EphA2). Antigen binding regions were created by designing combinatorial libraries into the structural loops and Abdurins were selected using phage display methods. Initial binders were reformatted into new maturation libraries and low nanomolar binders were isolated using cell-free DNA display, CIS display. Further characterization confirmed binding of the Abdurins to both human and murine EphA2 proteins and exclusively to cell lines that expressed EphA2, followed by rapid internalization. Two different EphA2 binders were labeled with 64Cu, using a bifunctional MeCOSar chelator, and administered to mice bearing tumors from transplanted human prostate cancer cells, followed by PET/CT imaging. The anti-EphA2 Abdurins localized in the tumors as early as 4 hours after injection and continued to accumulate up to 48 hours when the imaging was completed. These data demonstrate the ability to isolate high affinity binders from the engineered Abdurin scaffold, which retain a long serum half-life, and specifically target tumors in a xenograft model.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Agents / chemistry*
  • Cell Line, Tumor
  • Humans
  • Mice
  • Peptide Library
  • Positron-Emission Tomography
  • Protein Engineering / methods*
  • Receptor, EphA2 / metabolism*
  • Tomography, X-Ray Computed
  • Xenograft Model Antitumor Assays

Substances

  • Antineoplastic Agents
  • Peptide Library
  • Receptor, EphA2

Grants and funding

This study was funded and sponsored by Research Corporation Technologies, Inc. and the research activities directed by KRG. KRG and the authors made the decision to publish this work. Research Corporation Technologies, Inc., Isogenica, Ltd., IRBM Science Park, Baker IDI, Clarity Pharmaceuticals and VTU Technology Ltd. provided support in the form of salaries for the authors but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of all authors are articulated in the ‘author contributions’ section.