Abstract 4311: Defining the GRB7 interactome in triple-negative breast cancer.
Authors: Calvo Veronica A. , Giricz Orsolya , Kenny Paraic A. 
Published: August 2014

Triple-negative breast cancers (TNBC) lack expression of Estrogen Receptor, Progesterone Receptor, and HER2 and, as such, are non-responsive to currently available targeted therapies. Thus, more insight is needed into the molecular mechanisms underlying this disease in order to identify new therapeutic targets. GRB7 is an SH2-domain containing adaptor protein, which couples upstream activated receptor tyrosine kinases to downstream signaling intermediates. Most of the interest in GRB7 has been as a result of its co-amplification with the HER2 oncogene in HER2-overexpressing breast tumors, in which GRB7/HER2 co-overexpression is associated with a particularly poor prognosis. However, our recent large study of 764 patients, which included 246 TNBC cases, revealed that high levels of GRB7 in TNBC patients was the strongest positive predictor of tumor recurrence in this population. Additional recent studies in our lab have shown that GRB7 inhibition reduces cell motility and invasion and promotes apoptosis in TNBC cell lines and so suggest that GRB7 may be an important therapeutic target in this disease.

Due to the lack of available non-peptide drugs specifically targeting GRB7, our aim is to identify the signaling pathways that GRB7 couples in TNBC, which may be more easily targetable in breast tumors. For this purpose, we are analyzing the GRB7-binding partners and the signaling pathways mediated by GRB7 in TNBC cell lines. We have established triple-negative MDA-MB-231, MDA-MB-468, MDA-MB-436, HCC1806, HCC1937, Hs578T and BT549 cells stably overexpressing FLAG-GRB7 protein. So far, we have observed that GRB7 overexpression leads to increased AKT/mTOR, MAPK/ERK and SAPK/JNK pathways activation, which might explain the enhanced survival and invasion related to GRB7 activity in TNBC cell lines.

We determined the GRB7-interacting partners in MDA-MB-231 cells by co-IP and LC-MS/MS. These GRB7 immunoprecipitates include proteins regulating microtubule dynamics like MAP1B, MAP4, Spectrin, RRBP1, LRPPRC and CKAP5. We also detected the previously described interaction with STAU1, and found other proteins that might be also related to stress granules and mRNA translation control like SERBP1 and IGF2BP1. Some of the interacting proteins we have found have previously been related to breast cancer like the splicing factor kinase SRPK1, the transcription factor chaperone THOC4, CKAP5 and the adaptor protein 14-3-3. Other interactors are involved in DNA replication like DNA topoisomerase 1 and XRCC6. We are currently evaluating the roles of these proteins in triple-negative breast cancer cell lines.

This study is funded, in part, by a postdoctoral fellowship from Fundacion Alfonso Martin Escudero to VC.

Citation Format: Veronica A. Calvo, Orsolya Giricz, Paraic A. Kenny. Defining the GRB7 interactome in triple-negative breast cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4311. doi:10.1158/1538-7445.AM2013-4311