PUB021 SLK Causes Resistance to EGFR-TKI in Lung Cancer

Abstract

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors, such as gefitinib and dacomitinib are effective for non-small cell lung cancer with activating EGFR mutations. However, relapse typically occurs after an average of 6-12 months of continuous treatment. Although some mechanism(s) of resistance have been identified, approximately 30% of acquired resistance cases remain unknown. Targeted therapies that inhibit EGFR tyrosine kinase activity (EGFR-TKI, Gefitinib) are of particular interest in lung adenocarcinomas with activating mutation of EGFR. However, all patients will escape after several months of treatment and ultimately develop progressive disease. A better understanding of the molecular mechanisms underlying resistance to EGFR-TKIs is a critical challenge as it can help design new drugs and therapeutic strategies to overcome resistance. The aim of this study was to explore additional mechanisms of EGFR-TKI resistance in lung tumors. We generated resistant clones of the sensitive EGFR exon 19 mutant NSCLC cell lines PC9 and HCC827 by exposing them to increasing concentrations of gefitinib or dacomitinib for several months. We identified a specific deregulation of SLK in resistant clones and unraveled a SRSF2/SLK pathway as a new mechanism of lung tumor progression following gefitinib treatment. Generation of Gefitinib and Dacomitinib resistant PC9 and HCC827 clones Cell growth assay, Apoptosis Western blotting Small interfering RNA (siRNA) transfection, RNA extraction and RT/qPCR analysis Indirect Immunofluorescence Statistical Analysis. Some resistant clones had absence of known mechanisms of resistance including the EGFRT790M mutation, MET gene amplification and Small-cell lung cancer histology but exhibited a lower growth kinetic compared sensitive cells. Focusing on these clones we found that some of them had undergone Epithelial to Mesenchymal Transition (EMT). In one clone with EMT, overexpression of a truncated SLK protein was detected. Overexpression of normal sized SLK protein was observed in the others independently of EMT status. Neutralization of the splicing factor SRSF2 sensitized resistant cells to gefitinib-induced apoptosis and strongly inhibited SLK expression. SLK: a potential target of SRSF2 These results demonstrated for the first time that a SRSF2-dependent pathway is an important determinant of acquired resistance of lung tumors to EGFR-TKI Collectively these data highlighted the SRSF2/SLK module as a new component of EGFR signaling network and strongly suggested that activation of a survival SRSF2/SLK pathway is a new mechanism of lung tumor progression following gefitinib treatment.