Radiation and cetuximab are therapeutics used in the management of head and neck squamous cell carcinoma (HNSCC). Despite clinical success with these modalities, development of both intrinsic and acquired resistance is an emerging problem in the management of this disease. Thus, identifying and targeting the molecules that drive resistance to radiation or cetuximab is essential for improving efficacy of treatment approaches. Previously, we identified AXL as a logical molecular target in HNSCC and indicated that it may play a pivotal role in resistance to radiation and cetuximab. In our current study, we advanced these early findings into pre-clinical models using patient-derived xenografts (PDXs). We found bemcentinib which is a small molecule inhibitor AXL, can enhance therapy in radiation- or cetuximab- resistant PDXs. Next, we investigated the molecular mechanisms of how AXL signaling which leads to resistance. Three tyrosine residues of AXL (Y779, Y821, Y866) were mutated and examined for their sensitivity to cetuximab and/or radiation. Overexpression of AXL wild type rendered radiation- or cetuximab- sensitive HNSCC line resistant, but cell lines overexpressing AXL-Y821F retained their sensitivity. Advancing this line of investigation in vivo indicated that tumors overexpressing AXL were resistant to cetuximab whereas tumors harboring the Y821 mutation were sensitive. These results demonstrated that signals emanating from Y821 may be critical for cetuximab- and radiation- resistant pathways. Further, reverse phase protein array (RPPA) was employed to analyze the proteomic architecture of signaling pathways in these cell lines. The c-Abl expression level was higher in HN30-AXL compared to HN30-vector or HN30-AXL-Y821F mutant. Further, immunoprecipitation indicated that c-Abl and AXL were highly associated in HN30-AXL cells, while this association was absent in vector or AXL-Y821F mutant cells. Finally, targeting c-Abl, using imatinib, in cetuximab resistant PDXs led to complete tumor regression and further, up to 3 months after cessation of treatment, these tumors exhibited no recurrence. Collectively, the studies presented herein suggest that tyrosine 821 of AXL mediates resistance to cetuximab by activation of c-Abl in HNSCC and targeting both EGFR and c-Abl leads to a robust anti-tumor response. These results provide rationale for the clinical targeting of c-Abl to enhance the therapeutic modalities used in treating HNSCC.

Citation Format: Nellie K McDaniel, Mari Iida, Kwangok P Nickel, Tamara S Rodems, Adam D Swick, Prashanth J Prabakaran, Jaimee C Eckers-Kubatzke, Noah B Welke, Carlene Kranjac, Alison E Schulz, Randy J Kimple, Ravi Salgia, Deric L Wheeler. AXL mediates cetuximab resistance through tyrosine 821 and activation of c-Abl kinase in head and neck cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A128. doi:10.1158/1535-7163.TARG-19-A128