1197P - Loss of Amplified Egfr Gene with Mutation as a Novel Mechanism of Acquired Resistance to Egfr-Tkis in Egfr Mutated Nsclc Cells

Abstract

ABSTRACT Background The epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) show notable effects for non-small cell lung cancers (NSCLC) harboring EGFR activating mutations. However, almost all patients eventually acquire resistance to EGFR-TKIs within several years. The major mechanisms of acquired resistance including 2nd mutation of T790M and amplification of MET have been identified. However, the mechanisms of other resistance remain unclear. In this study, we established novel erlotinib-resistant NSCLC cells and examined their resistant mechanisms. Methods NSCLC HCC827 cells have an EGFR Exon 19 deletion and gene amplification and are highly sensitive to erlotinib. The resistant cells were established by continuously exposing HCC827 cells to 0.1, 1 or 10 µM of erlotinib in 96 well plates for three months. The resistant mechanisms were determined by direct sequence analysis and EGFR FISH analysis. Results The fourteen and three resistant cells were established by 0.1 µM and 1 µM of erlotinib exposure, respectively. No resistant cells appeared in the wells of 10 µM of erlotinib. The IC50 values of these resistant cells were more than 25-fold higher than that of parental cells. No 2nd mutation of T790M was detected in any of the resistant cells and MET gene amplification was detected in only one resistant cells. Instead, we found that 13/17 resistant cells were dominated by EGFR not amplified cells and one of resistant cells B10 consisted of more than 99.9% of EGFR not amplified cells, and that the parental cells consisted of only 2.5% of EGFR not amplified cells and 97.5% of EGFR amplified cells. EGFR not amplified clone 4D8 isolated from parental cells and showed resistance to erlotinib comparable to the resistant cells B10. Furthermore, we found that EGFR not amplified cells were constantly emerged from EGFR amplified clone isolated from parental cells under normal cell culture condition. Conclusion Loss of amplified EGFR gene with mutation causes acquired resistance in HCC827 cells when exposed to relatively low concentration of erlotinib while high concentration of erlotinib deprives HCC827 cells of the chance of emergence of resistant cells. Disclosure All authors have declared no conflicts of interest.