Abstract 1960: T790M-independent EGFR TKI resistance is associated with a broad multi-drug resistant phenotype but selective vulnerabilities to spindle assembly complex (SAC) and CDK inhibitors
Authors: Nilsson Monique B. , Robichaux Jacqueline , Sun Huiying , Tong Pan , Diao Lixia , Hofstad Mia , Fan YouHong , Wang Jing , Heymach John 
Published: August 2018

While EGFR mutant NSCLC patients are initially responsive to EGFR targeted therapies, resistant disease inevitably emerges and in nearly half of resistance cases, tumors lack secondary EGFR mutations such as T790M and are refractory to 2nd and 3rd generation EGFR tyrosine kinase inhibitors (TKI). The identification of treatment regimens with efficacy against T790M-negative resistance remains a major clinical challenge. To address this unmet need, we derived a panel of NSCLC cell lines with acquired resistance to the EGFR TKI, erlotinib. A subset of EGFR TKI resistant variants were negative for secondary EGFR mutations, were resistant to 2nd and 3rd generation EGFR TKIs including osimertinb, afatinib, and dacomitinib, and had undergone epithelial to mesenchymal transition (EMT) as demonstrated by loss of E-cadherin, enhanced expression of N-cadherin and Axl and an increased invasive phenotype as determined by Boyden chamber assay. Proteomic profiling revealed that although EGFR TKI resistant cells displayed similar mesenchymal and invasive phenotypes, there was significant heterogeneity in protein expression and pathway activation among resistant variants derived from the same parental cell line. To identify therapeutic agents with activity against EMT-associated EGFR TKI resistance, we performed high-throughput drug screening to test the efficacy of 1,321 compounds. EMT-associated EGFR TKI resistance was accompanied by the acquisition of broad spectrum drug resistance. Compared to parental cells, mesenchymal EGFR TKI resistant cells were significantly more resistant to chemotherapeutic agents used to treat NSCLC including pemetrexed, irinotecan, vinblastine, and gemcitabine. EGFR TKI resistant cells displayed acquired resistance to 147 other tyrosine kinase and serine/threonine kinase inhibitors. In contrast, both parental cells and mesenchymal EGFR TKI resistant variants were highly sensitive to CDK inhibitors and agents targeting spindle assembly checkpoint (SAC) components including PLK1, Aurora, KSP, and survivin. These finding were validated by MTS and clonogenic assays. Treatment with SAC inhibitors induced the accumulation of cells in G2/M phase, enlarged nuclear size, and polyploidy. To clinically validate these findings, we established a cell line (MDA-011) from the pleural effusion of an EGFR mutant NSCLC patient with T790M-negative resistance to erlotinib. In vitro, MDA-011 cells were resistant to erlotinib and osimertinib. MDA-011 cells were highly sensitive to CDK inhibitors and SAC inhibitors as determined by MTS and clonogenic assays. These data indicate that EMT-associated resistance to EGFR TKIs is associated with broad spectrum drug resistance but vulnerabilities to CDK and SAC inhibition which can potentially be exploited to overcome resistant disease in NSCLC patients.

Citation Format: Monique B. Nilsson, Jacqueline Robichaux, Huiying Sun, Pan Tong, Lixia Diao, Mia Hofstad, YouHong Fan, Jing Wang, John Heymach. T790M-independent EGFR TKI resistance is associated with a broad multi-drug resistant phenotype but selective vulnerabilities to spindle assembly complex (SAC) and CDK inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1960.