Advertisement

Molecular Biotechnology

, Volume 35, Issue 1, pp 51–63 | Cite as

Novel short peptides isolated from phage display library inhibit vascular endothelial growth factor activity

  • Berrin Erdag
  • Koray B. Balcioglu
  • Asli Kumbasar
  • Omur Celikbicak
  • Gabrielle Zeder-Lutz
  • Danièle Altschuh
  • Bekir Salih
  • Kemal Baysal
Research

Abstract

Signal transduction through the vascular endothelial growth factor (VEGF)-VEGF receptor (VEGFR) pathway has a pivotal importance in angiogenesis, and has therefore become a prime target in antitumor therapy. In search for peptides antagonizing VEGF binding to its receptors, we screened a random heptamer library displayed on phage for peptides that bind the whole VEGF165 molecule and inhibit VEGF dependent human umbilical vein endothelial cell (HUVEC) proliferation. Two selected peptides with sequences WHLPFKC and WHKPFRF were synthesized. Biacore and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis indicated that these peptides bind the VEGF homodimer in a concentration-dependent manner, with micromolar affinity, and with a 2:1 peptide: VEGF stoichiometry. They inhibited HUVEC proliferation in vitro by 77 and 55%, respectively. Taken together, our results indicate that these peptides could be potent inhibitors of angiogenesis. Furthermore, we show that the peptide-VEGF binding properties can be quantified, a prerequisite for the further optimization of binders.

Index Entries

Vascular endothelial growth factor angiogenesis peptide phage display HUVEC 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Folkman, J. (1995) Angiogenesis in cancer, vascular, rheumatoid and other disease. Nat. Med. 1, 27–31.PubMedCrossRefGoogle Scholar
  2. 2.
    Carmeliet P. (2003) Angiogenesis in health and discease. Nature. Med. 9, 653–660.PubMedCrossRefGoogle Scholar
  3. 3.
    Carmeliet, P., Ferriera, V., Breier, G., et al. (1996) Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele. Nature 380, 435–439.PubMedCrossRefGoogle Scholar
  4. 4.
    Tammela, T., Enholm, B., Alitalo, K., and Paavonen, K. (2005) The biology of vascular endothelial growth factors. Cardiovasc. Res. 65, 550–563.PubMedCrossRefGoogle Scholar
  5. 5.
    De-Vries, C., Escobedo, J. A., Ueno H., Houck, K., Ferrara, N., and Williams, L. T. (1992) The fms-like tyrosine kinase, a receptor for vascular endothelial growthy factor. Science 255, 989–991.PubMedCrossRefGoogle Scholar
  6. 6.
    Ferrara, N., Gerber, H. P., and LeCouter, J. (2003) The biology of VEGF and its receptors. Nat. Med. 9, 669–676.PubMedCrossRefGoogle Scholar
  7. 7.
    Hicklin, D. J., Witte, L., Zhu, Z., et al. (2001) Monoclonal antibody strategies to block angiogenesis. DDT 6, 517–528.PubMedGoogle Scholar
  8. 8.
    Kim, K. J., Li, B., Winer, J., et al. (1993) Inhibition of vascular endothelial growth factor induced angiogenesis suppresses tumor growth in vivo Nature 362, 841–844.PubMedCrossRefGoogle Scholar
  9. 9.
    Lander, R. C., Sato, A. K., Gorzelany J., and Souza, M. (2004) Phage display derived peptides as therapeutic alternatives to antibodies. DDT 9, 525–529.Google Scholar
  10. 10.
    Bikfalvi, A. and Bicknell, R. (2002) Recent advances in angiogenesis anti-angiogenesis and vascular targeting. Trends Pharmacol. Sci. 23, 576–582.PubMedCrossRefGoogle Scholar
  11. 11.
    Smith, G. P. and Scott, J. K. (1993) Libraries of peptides and proteins displayed on Filamentous phage. Methods Enzymol. 217, 228–257.PubMedCrossRefGoogle Scholar
  12. 12.
    Fairbrother W. F., Christinger, H. S., Cochran, A. G., et al. (1998) Novel peptides selected to bind Vascular Endothelial growth factor target the Receptor binding site. Biochemistry 37, 17,754–17,764.CrossRefGoogle Scholar
  13. 13.
    Asai, T., Magatsuka, M., Kuromi, K., et al. (2002) Suppression of tumor growth by novel peptides homing to tumor-derived new blood vessels. FEBS Lett. 510, 206–210.PubMedCrossRefGoogle Scholar
  14. 14.
    Binetruy-Tournaire, R., Demongel, C., Mulavaud, B., et al. (2000) Identification of a peptide blocking vascular endothelial growth factor (VEGF) mediated angiogenesis. EMBO J. 19, 1527–1533.CrossRefGoogle Scholar
  15. 15.
    Hetian, L., Ping, A., Shumei S. et al. (2002) A novel peptide isolated from a phase display library inhibits tumor growth and metastasis by blocking the binding of vascular endothelial growth factor to its kinase domain receptor. J. Biol. Chem., 277, 43,137–43,14.CrossRefGoogle Scholar
  16. 16.
    Piossek C., Scheider-Mergener, J., Schirer, M., Vakalopoulou, E., Germeroth, L., and Thierauch, K. H. (1999) Vascular endothelial growth factor (VEGF) receptor II-derived peptides inhibit VEGF. J. Biol. Chem. 274, 5612–5619.PubMedCrossRefGoogle Scholar
  17. 17.
    Nakamura, T., Takasugi, H., Aizawa, T. et al. (2005) Peptide mimics of epidermal growth factor with antagonistic activity. J. Biotechnol. 116, 211–219.PubMedCrossRefGoogle Scholar
  18. 18.
    Scott, J. K. and Smith, G. P. (1990) Searching for peptide ligands with an epitope library. Science 249, 389–395.Google Scholar
  19. 19.
    Jaffe, E. A., Nachmann, R. L., Becker, C. G., and Minick, C. R. (1973) Culture of human endothelial cells from umbilical veins. Identification by morphologic and immunologic criteria. J. Clin. Invest. 52, 2745–2756.PubMedGoogle Scholar
  20. 20.
    Maciag, T., Cerundolo, J., Ilsley S., Kelley, P. R., and Forand, R. (1979) An endothelial cell growth factor from bovine hypothelamus identification and partial characterization. Proc. Natl. Acad. Sci. USA 76, 5674–5678.PubMedCrossRefGoogle Scholar
  21. 21.
    Rousch, M., Lutgerink, J. T., Coote, J., de Bruine, A., Arends, J. W., and Hoogenboom, H. R. (1998) Somatostatin displayed on filamentous phage as a receptor-specific. Br. J. Pharmacol. 125, 5–16.PubMedCrossRefGoogle Scholar
  22. 22.
    Hoogenboom, H. R., Lutgerink, J. T., Pelsers, M. M., et al (1999) Selection-dominant and nonaccessible epitopes on cell-surface receptors revealed by cell-panning with a large phage antibody library. Eur J. Biochem. 260, 774–784.PubMedCrossRefGoogle Scholar
  23. 23.
    Ueda, Y., Yamagishi, T., Samata, K., et al. (2003) A novel low molecular weight antagonist of vascular endothelial growth factor receptor binding: VGA 1155. Mol. Cancer Ther. 2, 1105–1111.PubMedGoogle Scholar
  24. 24.
    Karlsson, R., Michaelsson, A., and Mattsson, L. (1991) Kinetic analysis of monoclonal antibody-antigen interactions with a new biosensor based analytical system. J. Immunol. Methods 145, 229–240.PubMedCrossRefGoogle Scholar
  25. 25.
    Monk, B. J., Choi, D. C., Pugmire, G., and Burger, R. A. (2005) Activity of bevacizumab (rhuMAB VEGF) in advanced refractory epithelial ovarian cancer. Gynecologic Oncology 96, 902–905.PubMedCrossRefGoogle Scholar
  26. 26.
    Lien, S. and Lowman, H. B. (2003) Therapeutic peptides. Trends Biotechnol. 21, 556–562.PubMedCrossRefGoogle Scholar
  27. 27.
    An, P., Lei, H., Zhang, J., et al. (2004) Suppression of tumor growth and metastasis by a VEGFR-1 antagonizing peptide identified from a phage display library. Int. J. Cancer 111, 165–173.PubMedCrossRefGoogle Scholar
  28. 28.
    Koivunen, E., Arap, W., Valtanen, et al. (1999). Tumor targeting with a selective gelatinase inhibitor. Nature Biotechnology, 17, 768–774.PubMedCrossRefGoogle Scholar
  29. 29.
    Kim, M. Y., Byeon, C. W., Hong, K. H., Han, K. H., and Jeong, S. (2005) Inhibition of the angiogenesis by the MCP-1 (monocyte chemoattactant protein-1) binding peptide FEBS Lett. 579, 1597–1601.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc 2006

Authors and Affiliations

  • Berrin Erdag
    • 1
  • Koray B. Balcioglu
    • 1
  • Asli Kumbasar
    • 1
  • Omur Celikbicak
    • 2
  • Gabrielle Zeder-Lutz
    • 3
  • Danièle Altschuh
    • 3
  • Bekir Salih
    • 2
  • Kemal Baysal
    • 1
  1. 1.TUBITAK Research Institute for Genetic Engineering & BiotechnologyKocaeliTurkey
  2. 2.Department of ChemistryHacettepe UniversityAnkaraTurkey
  3. 3.UMR 7175-LC1, CNRS/ULP, ESBS, Pare d'InnovationIllkirch CedexFrance

Personalised recommendations