Abstract LB-307: Overexpression of AXL as a novel acquired resistance mechanism in EGFR-mutated non-small cell lung cancer cells

Abstract

Abstract Activating mutations of EGFR have been characterized as important mechanisms for carcinogenesis in a subset of EGFR-dependent non-small cell lung cancers (NSCLC). EGFR tyrosine kinase inhibitors (TKI), such as erlotinib and gefitinib, have dramatic clinical effects on EGFR-addicted lung cancers and are used as first-line therapy for EGFR-mutant tumors. However, this therapeutic sensitivity is invariably temporary and eventually all tumors acquire secondary resistance to the drugs and progress. Erlotinib-resistant EGFR mutations such as EGFR T790M and MET activation have been reported in drug-resistant tumors, but the mechanism of resistance remains unclear in a significant fraction of cases. Here we report a novel molecular mechanism leading to secondary resistance to erlotinib. HCC827, an EGFR-mutant and highly erlotinib-sensitive NSCLC cell line was exposed to increasing doses of erlotinib for 5 months and 2 subclonal cell lines were established, both of which grow well in the presence of high-dose erlotinib (15µM). In contrast to parental cells, the two erlotinib-resistant cell lines demonstrated sustained activation of Ras/Mek/Erk and Akt signaling pathways in the presence of erlotinib despite complete inhibition of EGFR phosphorylation. Sequencing of the entire EGFR gene sequence revealed no secondary changes, and expression of EGFR and MET, as well as MET activation did not differ from parental cells suggesting the presence of an alternative, novel mechanism of resistance. Gene expression profiling revealed a limited number of genes with altered expression levels which were verified by quantitative rt-PCR. We focused on the cell surface receptor tyrosine kinase, AXL (10-fold overexpressed in one resistant clone) given its known role in cell proliferation, angiogenesis and recent findings indicating its involvement in lapatinib-resistance in ErbB2-positive breast cancer. Prominent AXL overexpression was further confirmed in this clonal isolate on the protein level. siRNA-mediated downregulation of AXL reconstituted erlotinib-sensitivity, leading to cell death and was accompanied by inhibition of AKT activation. However, AXL knockdown in the absence of erlotinib did not have growth inhibitory effects on the cells. These results suggest that AXL signaling can serve as a backup control of proliferative pathways in our resistant cell models in the presence of EGFR inhibition serving as a novel paradigm of oncogenic switch. Mechanisms underlying overexpression of AXL and its restoration of TKI sensitivity are under investigation. Our results suggest that an oncogenic switch from EGFR-dependent to EGFR/AXL-codependent signaling can lead to secondary EGFR-TKI resistance in NSCLC, indicating that dual blockade of EGFR and AXL may serve as a novel therapeutic strategy for the treatment of EGFR-mutant lung cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-307.