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MINI 09 - Drug Resistance (ID 107)
- Event: WCLC 2015
- Type: Mini Oral
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 1
MINI09.08 - Secondary C805S Mutation in HER2 Gene Confers Acquired Resistance to HER2 Kinase Inhibitors in HER2 Mutant Lung Cancer (ID 2181)
16:45 - 18:15 | Author(s): D. Ercan
Activating mutations in the HER2 kinase domain are detected in 2-4% of non-small cell lung cancers (NSCLC), and are oncogenic in both in vitro and in vivo models. Current clinical strategies to target mutant HER2 include the use of covalent HER2 inhibitors afatinib, dacomitinib and neratinib; all of which have limited single agent activity. We evaluated how drug sensitive models of HER2 mutant lung cancer develop acquired resistance in vitro to gain biological insights and to predict how acquired resistance may develop in the clinic.
Murin Ba/F3 cells expressing duplication/insertion of four amino acids (YVMA) between codon 775 and 776 in exon 20 of HER2 gene (A775_G776insYVMA (insYVMA)) were exposed to N-ethyl-N-nitrosourea mutagenesis and expanded in the presence of neratinib and dacomitinib. Total RNAs were extracted from resistant clones and sequencing of the HER2 tyrosine kinase domain was performed. Drug resistance was confirmed with cell growth assays and western blotting.
Total 5 clones for Neratinib and 7 clones for Dacomitinib were expanded from each 300 wells. Sequencing analysis revealed that all resistant clones retained original insertion mutation and acquired same substitution of Cysteine to Serine change in codon 805 (C805S) in exon 20 of HER2 gene. This mutation is analogous to the EGFR C797S mutation that mediates resistance to 3[rd] generation EGFR inhibitors. Next, we generated Ba/F3 cells co-expressing activating mutations; insYVMA and a dacomitinib hypersensitive insertion mutation (insertion of three amino acids (WLV) after codon 774 with deletion of M774 (M774del insWLV (insWLV))), in cis with the C805S mutation. Cell growth assay revealed these double mutants were resistant to all three second generation inhibitors for EGFR family; neratinib, dacomitinib, and afatinib, compared to parental cells which only have activating mutation. They were also resistant to 3[rd] generation EGFR inhibitors; WZ40002 and AZD9291. Phosphorylation of HER2 was not completely inhibited by these drugs. Resistant cells showed moderate sensitivity to mTOR inhibitor; rapamycin alone. Combination treatment with afatinib and rapamycin effectively inhibited growth of these cells.
The C805S secondary HER2 mutation results in acquired resistance to covalent HER2 inhibitors in HER2 mutant NSCLC. Our results provide insights into drug resistance mechanisms and help predict likely clinical mechanisms of resistance to HER2 targeted therapies in HER2 mutant NSCLC.
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