Poster Presentation Melbourne Protein Group Student Symposium 2013

Optimisation and characterisation of cyclic peptides targeting the Grb7 SH2 domain.  (#85)

Gabrielle Watson 1 , Menachem Gunzburg 1 , Ketav Kulkarni 2 , Katie Cergol 3 , Richard Payne 3 , Patrick Perlmutter 2 , Matthew Wilce 1 , Jackie Wilce 1
  1. Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia
  2. Department of Chemistry, Monash University, Clayton, VIC
  3. Department of Chemistry, University of Sydney, Darlington, NSW

Sustained proliferative signalling, an iconic hallmark of cancer, can result from constitutive activation of signalling pathways1 . Targeting these deregulated pathways is a common goal in therapeutic development with great success in breast cancer seen using the clinically available drug Herceptin. There is a continual need to develop anti-cancer agents however, with improved outcomes shown for patients when multiple signalling pathways are targeted2 .  Human growth factor receptor bound protein 7 (Grb7) is an adaptor protein that propagates signals from phosphorylated tyrosine kinases such as the receptor protein, ErbB2. Grb7 is overexpressed in a multitude of cancers, particularly pancreatic and breast cancer, and has an established role in both cancer cell proliferation and migration3 4 .

A non-phosphorylated cyclic peptide, G7-18NATE, has been shown to inhibit pancreatic cell migration, and reduce cellular growth and migration in breast cancer cell lines5 4 . The peptide binds to the Grb7 SH2 domain with micromolar affinity and is specific for the Grb7 SH2 domain compared with the closely related SH2 domains from Grb14 and Grb106 . Rational adjustments to G7-18NATE have been identified through structural analysis that are predicted to improve affinity for the target whilst maintaining specificity for Grb7. This study aims to develop and deliver optimised G7-18NATE derivatives to target the SH2 domain of Grb7. Initial experiments have identified a derivative, G7-TEM, which has increased affinity for the Grb7 SH2 domain compared with G7-18NATE under physiological buffer conditions whilst maintaining specificity.

Future experiments to be conducted on G7-TEM and further G7-18NATE derivatives include structural analysis by x-ray crystallography, and functional studies testing cellular uptake and the ability of the peptide to inhibit migration of breast cancer cells. Developing and characterising these peptides will establish fundamental methods that can be readily applied to other intracellular therapeutic targets as well as mark considerable process in Grb7 based anti-cancer drug development.

  1. Hanahan and Weinberg (2011) Hallmarks of Cancer: The Next Generation, Cell 144, 646-674.
  2. Arteaga, C.L., Sliwkowski, M.X., Osborne, C.K., Perez, E.A., Puglisi, F., and Gianni, L. (2012) Treatment of HER2-positive breast cancer: current status and future perspectives. Nat. Rev. Clin. Oncol. 9, 16
  3. Stein, D., Wu, J., Fuqua, S.A., Roonprapunt, C., Yajnik, V., D’Eustachio, P., Moskow, J., Buchberg, A.M., Osborne, C.K. and Margolis, B. (1994) The SH2 domain protein GRB-7 is co-amplified, overexpressed and in a tight complex with HER2 in breast cancer. EMBO J. 13, 1331
  4. Tanaka, S., Pero, S.C., Taguchi, K., Shimada, M., Mori, M., Krag, D.N., and Arii, S. (2006) Specific peptide ligand for Grb7 signal transduction protein and pancreatic cancer metastasis. J. Natl. Cancer Inst. 98, 491
  5. Pero, S.C., Shukla, G.S., Cookson, M. M., Flemer, S., and Krag, D.N. (2007) Combination treatment with Grb7 peptide and Doxorubicin or Trastuzumab (Herceptin) results in cooperative cell growth inhibition in breast cancer cells. Br. J. Cancer 96, 1520
  6. Gunzburg, M.J., Ambaye, N.D., Del Borgo, M.P., Pero, S.C., Krag, D.N., Wilce, M.C., Wilce, J.A. (2012) Interaction of the non-phosphorylated peptide G7-18NATE with Grb7-SH2 domain requires phosphate for enhanced affinity and specificity. J. Mol. Recognit. 25, 57