Abstract 247: Understanding the role of tyrosine 1101 in the nuclear translocation of the epidermal growth factor receptor

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Abstract Triple-negative breast cancer (TNBC) is a subclass of breast cancers that are negative for the estrogen receptor, the progesterone receptor, and HER2. Currently, TNBCs have poor prognosis and very few proven molecular targets. One promising target in TNBC has been the epidermal growth factor receptor (EGFR), as it is overexpressed in as many as 80% of TNBCs. Strikingly, however, antibody-based therapy directed against the EGFR (cetuximab) in TNBCs has had modest clinical impact. Over the last decade, advances in EGFR biology have established that the EGFR functions in two distinct signaling pathways: classical membrane-bound signaling and the nuclear EGFR signaling pathway. We previously demonstrated that EGFR nuclear translocation is dependent on Src Family Kinases (SFKs) and phosphorylation of tyrosine 1101 (Y1101) on the cytoplasmic tail of the EGFR. In these studies, translocation of the EGFR to the nucleus led to activation of the nuclear EGFR signaling pathway, which resulted in cetuximab resistance. Resistance could be reversed by simultaneously targeting both the nuclear EGFR signaling pathway and the classical EGFR signaling pathway through the inhibition of its nuclear transport (blockade of SFKs) and with cetuximab, respectively. Despite the identification of nuclear EGFR as a functional molecular target in TNBC, the role of Y1101 in EGFR translocation has yet to be identified. In this study, we seek to better understand the role of Y1101 in the nuclear translocation of EGFR in TNBC. Using a battery of TNBC cell lines, we have created isogenic pairs that overexpress either wild type EGFR (EGFR-WT) or EGFR with a mutant Y1101 (EGFR-Y110F). Using electron and super-resolution confocal microscopy as well as cellular fractionation, we have shown that EGFR-Y1101 undergoes normal endocytosis upon EGF stimulation in multiple cell lines, but unlike EGFR-WT, EGFR-Y1101F deposits around the perinuclear region. These data suggest that EGFR-Y1101F is able to undergo normal translocation to the perinuclear space, but that amino acid Y1101 is necessary for direct nuclear entry, potentially due to its interactions with nucleoporins. Current work is focused on the identification of nucleoporins with which EGFR-Y1101 may interact. In addition, continued work focuses on allelic mutation using CrisprCas9. Ultimately, the long-term goal of this project is to understand how Y1101 mediates EGFR nuclear translocation. With this information, we will then aim to develop allosteric inhibitors of Y1101 that may abrogate EGFR nuclear translocation and lead to a novel therapeutic which targets the nuclear EGFR signaling pathway in TNBC. Citation Format: Bailey G. Flanigan, Mari Iida, Toni M. Brand, Deric L. Wheeler. Understanding the role of tyrosine 1101 in the nuclear translocation of the epidermal growth factor receptor. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 247.