704 Co-targeting epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase (ALK) overcomes EGFR inhibitor resistance in head and neck squamous cell carcinoma patient-derived models

Abstract

HNSCC is the sixth most common cancer worldwide. EGFR is overexpressed in up to 90% of HNSCC and associated with poor outcome. An EGFR monoclonal antibody, cetuximab, is the only approved molecular targeted therapy for HNSCC; however, resistance eventually occurs in all patients. Functional screens including a small-molecule kinase inhibitor were used to identify agents that synergized with EGFR inhibitors in reducing viability in HNSCC patient-derived tumor cells. Effective combination therapies were validated in scale-up experiments, and their true targets were evaluated using siRNAs to rule out off-target effects of the drugs. Proliferation and apoptosis were determined using immunofluorescent staining in 3D models. ALK expression and phosphorylation in tumor cells before and after anti-EGFR treatment was tested by western immunoblotting. Two ALK inhibitors on the drug screen panel showed synergistic effects with EGFR inhibitors in 6/8 HNSCC patients’ tumor cells, despite ineffectiveness of single drug. Scale-up dose-response experiments confirmed patient cell sensitivity to 4 different ALK inhibitors in combination with the EGFR inhibitor gefitinib. siRNA targeting ALK synergized with gefitinib in reducing cell viability, indicating specificity to ALK. Because of low expression of ALK mRNA in original tumors and patient tumor-derived cells, we hypothesized that EGFR inhibition induced ALK mRNA and protein. Indeed, inhibition of EGFR by gefitinib increased ALK protein expression and phosphorylation, suggesting induction of ALK expression and activation as a novel mechanism of EGFR inhibitor resistance in HNSCC amenable to combination therapy. In conclusion, we identified induction of ALK by EGFR inhibitor as a novel mechanism relevant to resistance to cetuximab in the clinic, and EGFR/ALK inhibition as a potential combination therapeutic strategy for treating EGFR inhibitor resistant HNSCC.