Poster Presentation The 42nd Lorne Conference on Protein Structure and Function 2017

Developing potent and specific inhibitors of the Grb7 breast cancer target (#255)

Gabrielle M Watson 1 , Ketav Kulkarni 1 , Jianrong Sang 1 , Matthew Wilce 1 , Jackie Wilce 1
  1. Biomedicine Discovery Institute, Monash University, Clayton, VIC

Growth factor receptor bound protein 7 (Grb7) is an intracellular signalling protein that is overexpressed in a number of cancers, including HER2+ and triple negative breast cancer. Grb7 interacts with phosphorylated tyrosine kinases via its C-terminal SH2 domain leading to migratory and proliferative signalling, and it is this domain that is a therapeutic target for the development of anti-cancer agents.1

A non-phosphorylated cyclic peptide, G7-18NATE, was found to bind specifically to the Grb7-SH2, blocking Grb7 interactions with upstream signalling molecules.2 When attached to the cell penetrating peptide Penetratin, it has been shown to inhibit pancreatic cell migration, reduce cellular growth and migration in breast cancer cell lines, and inhibit tumor metastasis in a mouse model of pancreatic cancer.1,3 However, G7-18NATE binds to the Grb7-SH2 with moderate affinity (KD = 18.1 μM); therefore, to improve on this, we have developed G7-18NATE derivatives using a structure-based drug design approach.

 

We have developed bicyclic derivatives that bind with nanomolar affinity whilst maintaining specificity for Grb7-SH2 over the closely related SH2 domains of Grb2 and Grb10. However, these derivatives require high levels of phosphate for high affinity binding. To overcome this phosphate dependency, we incorporated phosphotyrosine (pY) mimetics with the affinity improving 140-fold in physiological concentrations of phosphate. The X-ray crystal structures of Grb7-SH2/peptide complexes reveal precisely the molecular interactions that define this improved potency and specificity. Lastly, when attached to Penetratin, we have shown that the optimised inhibitors effectively block Grb7 mediated interactions in a HER2+ breast cancer cell line.

 

It is envisaged that the development of these specific and potent Grb7 targeting peptides will both provide tools to further explore the role of Grb7 in healthy and cancerous cells, and also provide proof-of-concept evidence that Grb7 can be successfully targeted for the development of novel anti-cancer agents.

  1. Pradip, D., Bouzyk, M., Dey, N., and Leyland-Jones, B. (2013) Dissecting GRB7-mediated signals for proliferation and migration in HER2 overexpressing breast tumor cells: GTP-ase rules. Am. J. Cancer Res. 3, 173-95.
  2. Pero, S. C., Oligno, L., Daly, R. J., Soden, A. L., Liu, C., Roller, P. P., Li, P., and Krag, D. N. (2002) Identification of novel non-phosphorylated ligands, which bind selectively to the SH2 domain of Grb7. J. Biol. Chem. 277, 11918-11926.
  3. 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, 491-8.