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Dissecting early AXL signaling regulators and associated phenotypes in erlotinib-treated EGFR mutant lung cancer by phophosite perturbations

ABSTRACT: Targeted therapies for receptor tyrosine kinases are effective but invariably limited by drug resistance. In EGFR-mutant lung cancer, AXL activation drives resistance to erlotinib by restoring cell survival and migration. To map these signaling mechanisms, we generated a panel of lung adenocarcinoma PC9 cell lines with phenylalanine substitutions at each intracellular AXL tyrosine residue. By integrating phosphorylation data with phenotypic changes via multivariate modeling, we found that AXL signaling organizes into two clusters enriched for Abl1 and SFK substrate motifs. A peptide specificity screen identified FAK1 as a top proximal substrate of AXL. Downstream, AXL-mediated YAP1 activation was found to sustain drug tolerance, while combined inhibition eliminated persister cells in vitro. These AXL and YAP pathways correlate with disease progression and poor clinical outcomes in erlotinib-treated patients. Collectively, this study dissects the specific signaling regulators by which AXL drives erlotinib resistance.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Lung Non-squamous Non-small Cell Carcinoma

SUBMITTER: Jacqueline Gerritsen

LAB HEAD: Forest White

PROVIDER: PXD079076 | Pride | 2026-06-14

REPOSITORIES: Pride

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			Action	DRS
	20190404_JG_AM_PC9_AXLL_TMT10_AC35.mzML	Mzml
	20190404_JG_AM_PC9_AXLL_TMT10_AC35.mzid	Mzid
	20190404_JG_AM_PC9_AXLL_TMT10_AC35.raw	Raw
	20190419_JG_AM_PC9_BR3_TR2_AXL_TMT10_AC35.mzML	Mzml
	20190419_JG_AM_PC9_BR3_TR2_AXL_TMT10_AC35.mzid	Mzid

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Axl mediates acquired resistance of head and neck cancer cells to the epidermal growth factor receptor inhibitor erlotinib

Project description:Elevated expression and activity of the epidermal growth factor receptor (EGFR) is associated with development and progression of head and neck cancer (HNC) and a poor prognosis. Clinical trials with EGFR tyrosine kinase inhibitors (TKIs; eg. erlotinib) have been disappointing in HNC. To investigate the mechanisms mediating resistance to these agents, we developed a HNC cell line (HN5-ER) with acquired erlotinib resistance. In contrast to parental HN5 HNC cells, HN5-ER cells exhibited an epithelial-mesenchymal (EMT) phenotype with increased migratory potential, reduced E-cadherin and epithelial-associated miRNAs, and elevated vimentin expression. Phosphorylated RTK profiling identified Axl activation in HN5-ER cells. Growth and migration of HN5-ER cells was blocked with a specific Axl inhibitor, R428, and R428 re-sensitized HN5-ER cells to erlotinib. Microarray analysis of HN5-ER cells confirmed the EMT phenotype associated with acquired erlotinib resistance, and identified activation of gene expression associated with cell migration and inflammation pathways. Moreover, increased expression and secretion of interleukin (IL)-6 and IL-8 in HN5-ER cells suggested a role for inflammatory cytokine signaling in EMT and erlotinib resistance. Expression of the tumor suppressor miR-34a was reduced in HN5-ER cells and increasing its expression abrogated Axl expression and reversed erlotinib resistance. Finally, analysis of 302 HNC patients revealed that high tumor Axl mRNA expression was associated with poorer survival (HR 1.66, p=0.007). In summary, our results identify Axl as a key mediator of acquired erlotinib resistance in HNC and suggest that therapeutic inhibition of Axl by small molecule drugs or specific miRNAs might overcome anti-EGFR therapy resistance. Differential gene expression between parental and acquired erlotinib resistant head and neck cancer cell lines of HN5.

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