PROJECT SUMMARY One of the most deployed molecular targeting agents in the treatment of head and neck cancer (HNC) is the Epidermal Growth Factor Receptor (EGFR)-targeted monoclonal antibody cetuximab (CTX). Cetuximab is a central component of the first-line EXTREME regimen and in combination with radiation therapy and is the only drug that exhibits efficacy in both locally advanced and recurrent/metastatic HNC. Although many HNC patients see CTX over the course of their cancer therapy, research over the past decade has revealed that many patients that do respond initially will eventually become refractory to CTX. To identify drivers of CTX resistance (CTXR) in HNC, our lab utilized HNC human tumor specimens, patient-derived xenografts (PDXs) and a series of acquired and intrinsically resistant in vitro and in vivo model systems. This work led to the identification of the receptor tyrosine kinase (RTK) AXL as a central driver of CTXR. To define the molecular mechanisms of how AXL signaling leads to CTXR, three tyrosine residues of AXL (Y779, Y821, Y866) were mutated and mapped for their sensitivity to CTX. Both in vitro and in vivo analysis revealed that 1) CTXR emanates from signaling from Y821 of AXL via the c-Abl kinase, 2) pre-clinical modeling (PDX) indicated that the combination of CTX plus imatinib (IMT), a c-Abl kinase inhibitor, lead to complete tumor regression without recurrence in HNC models and 3) preliminary data for this application suggests that AXL, via Y821, sequesters c-Abl in the cytosol and prevents it from moving to the nucleus, which is induced by IMT, to promote apoptosis. Collectively, we hypothesize that AXL plays a critical role in CTXR by sequestering c-Abl in the cytosol and that therapeutic targeting c-Abl in combination with CTX will lead to profound responses in HNC patients. We plan to conduct this interdisciplinary project, spanning from basic biology to a pilot clinical trial through the three following aims: 1) To investigate if a) cytoplasmic sequestration of c-Abl is responsible for AXL mediated CTXR and if b) if AXL and c-Abl association is a predictive biomarker of CTX response, 2) determine if targeting EGFR and c-Abl simultaneously in CTXR PDXs can enhance and re- sensitize to CTX, and 3) To perform a pilot window-of-opportunity study of CTX plus imatinib in patients with recurrent or metastatic HNC. Ultimately, our goal is to improve treatment outcomes for HNC patients. Successful pursuit of these investigations has the potential to significantly improve and refine current HER family-centric therapeutic approaches in HNC by understanding the molecular mechanisms of how Axl and c-Abl impart CTXR. In addition, this project has high significance by repurposing two FDA approved drugs in a new cancer setting to improve outcomes. Collectively, this work will impact future clinical treatment paradigms and identify patients most likely to benefit from targeting EGFR and c-Abl in HNC.