Antagonists to human and mouse vascular endothelial growth factor receptor 2 generated by directed protein evolution in vitro

Chem Biol. 2006 May;13(5):549-56. doi: 10.1016/j.chembiol.2005.12.009.


Using directed in vitro protein evolution, we generated proteins that bound and antagonized the function of vascular endothelial growth factor receptor 2 (VEGFR2). Binders to human VEGFR2 (KDR) with 10-200 nM affinities were selected by using mRNA display from a library (10(13) variants) based on the tenth human fibronectin type III domain (10Fn3) scaffold. Subsequently, a single KDR binding clone (K(d) = 11 nM) was subjected to affinity maturation. This yielded improved KDR binding molecules with affinities ranging from 0.06 to 2 nM. Molecules with dual binding specificities (human/mouse) were also isolated by using both KDR and Flk-1 (mouse VEGFR2) as targets in selection. Proteins encoded by the selected clones bound VEGFR2-expressing cells and inhibited their VEGF-dependent proliferation. Our results demonstrate the potential of these inhibitors in the development of anti-angiogenesis therapeutics.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Line
  • Cell Proliferation / drug effects
  • Directed Molecular Evolution*
  • Humans
  • Mice
  • Mutagenesis
  • Oligopeptides / chemistry
  • Oligopeptides / metabolism
  • Oligopeptides / pharmacology
  • Protein Binding
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Vascular Endothelial Growth Factor Receptor-2 / antagonists & inhibitors*
  • Vascular Endothelial Growth Factor Receptor-2 / metabolism


  • Oligopeptides
  • Recombinant Proteins
  • Vascular Endothelial Growth Factor Receptor-2