Abstract
Background: Overcoming osimertinib resistance in epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer (NSCLC) is challenging due to unclear mechanisms. We previously reported a NSCLC case with EGFR mutations progressed on osimertinib therapy, revealing a novel NOTCH2-NTRK1 fusion gene in the plasma and tumor tissue. Although the NTRK gene fusion has been identified in NSCLC and a range of tumor types, the role of NOTCH2-NTRK1 in osimertinib resistance is unclear. Methods: We utilized both in vitro and in vivo models exhibiting NOTCH2-NTRK1 fusion positivity to explore the biological function of NOTCH2-NTRK1, as well as its role and mechanism in osimertinib resistance. Results: The NOTCH2-NTRK1 fusion protein has been demonstrated to transform the human bronchial epithelial cell line BEAS-2B and promote the proliferation of NSCLC cells both in vitro and in vivo. It induces osimertinib resistance by activating MAPK and PI3K-AKT pathways. Phosphoproteomic analyses revealed a significant increase in the phosphorylation level of EGFR compared to the control group. Further investigations demonstrated that the NOTCH2-NTRK1 protein is capable of interacting with the EGFR protein. Protein molecular docking studies identified seven interacting sites between NOTCH2-NTRK1 and EGFR protein. Mutations within the region encompassing these seven interaction sites effectively reversed osimertinib resistance, leading to a significant reduction in the expression of key proteins within the MAPK and PI3K-AKT pathways. Notably, the interaction between NOTCH2-NTRK1 and EGFR was maintained even with combined osimertinib and entrectinib treatment. Conclusion: Our study reveals a novel mechanism by which the NOTCH2-NTRK1 fusion confers resistance to osimertinib through its interaction with EGFR in NSCLC.