Fig. 5: Dysregulation of the c-Jun/CNOT3 axis contributes to gefitinib resistance in NSCLC.

A–C PC-9 GR2 cells were treated with gefitinib for the indicated time, then cells were subjected to western blotting to assess protein expression levels. D Comparison of CNOT3 mRNA level between lung adenocarcinoma tissues with or without HER2 overexpression downloaded from TCGA database, n = 32 and 479, respectively. E PC-9, PC-9 GR1 and PC-9 GR2 cells were subjected to western blotting to assess protein expression levels. F PC-9 GR2 cells were treated with gefitinib (1 μM) and tucatinib (1 μM) for 24 h, then cells were subjected to western blotting to assess protein expression levels. G, H Comparison of CNOT3 mRNA level between gefitinib-sensitive cells and gefitinib-resistant cells downloaded from two different datasets. In Zhou cell line, gefitinib sensitive indicates IC₅₀ less than 0.4 μM (n = 5) and gefitinib resistance indicates IC₅₀ more than 4 μM (n = 35) (G). In Coldren cell line, gefitinib sensitive indicates IC₅₀ less than 0.5 μM (n = 9) and gefitinib resistance indicates IC₅₀ more than 4.5 μM (n = 11) (H). I PC-9, PC-9 GR1 and PC-9 GR2 cells were subjected to western blotting to assess protein expression levels. J–L PC-9 GR2 or H1975 cells were transfected with a control or CNOT3 siRNA. Twenty-four hours after transfection, cells were treated with gefitinib for 72 h. Cell viability was measured via the CCK-8 assay in PC-9 GR2 cells (J). PC-9 GR2 cells were subjected to western blotting to assess protein expression levels (K). H1975 cells were subjected to western blotting to assess protein expression levels (L). Data are shown as the mean±S.E.M. n = 3. D, G and H Mann–whitney U test. J One-way ANOVA with Tukey post hoc test. ^***P < 0.001, ^**P < 0.01 or ^*P < 0.05 for comparisons between the indicated groups.