Figure 3.

Evaluating erlotinib sensitivity of EGFR mutants in the context of YFP-EGFR-ICD. A. Representative examples of MCF-7 cells expressing YFP-EGFR-ICD wild type, Del746 or Del746/T790M (green), stained for endogenous phosphorylated Akt (red). Cells were treated for 20 hours with the indicated concentration of erlotinib. The different response of each EGFR variant to erlotinib treatment is readily visualized by immunoflourescence. YFP-EGFR-ICD wild type does not induce Akt phosphorylation, and relocates into thick cytoplasmic fibrils at 10 μM erlotinib. One thousand-fold lower concentration of the drug (10 nM) inhibited Del746-induced Akt phosphorylation, and caused fibrilar relocation of the ectopic protein. The double mutant Del746/T790M did not form fibrils and induced Akt phosphorylation even in the presence of 10 μM erlotinib. B. Images show that YFP-EGFR-ICD Del746 (green) remains phosphorylated at Y1092 (red) after relocating into fibrils in the presence of erlotinib.