Fig. 1: Claudin1 is positively correlated with EGFR-TKI resistance.

A, B Significantly downregulated and upregulated mRNAs in erlotinib-sensitive and erlotinib-resistant HCC827 cells were analyzed. Statistical comparisons were performed using a one-way analysis of variance with Dunnett’s test. n = 3, ****P < 0.0001. Volcano map: Upregulated: P < 0.05, log₂FC > 0; Downregulated: P < 0.05, log₂FC < 0. C Relative mRNA expression levels of CLDN1 in PC9 and PC9/GR cells were detected by real-time PCR. ****P < 0.0001. D Western blotting was performed to determine claudin1 expression in PC9, PC9/GR, HCC827, and H1975 cells. E Induction of gefitinib resistance in PC9 cells: PC9 cells were first treated with gefitinib at a concentration of 20 nM for 1 week. A small number of remaining cells were treated for another 2 days with a concentration of 50 nM, which was sufficient to kill nearly all parental cells. The remaining few cells were continuously cultured in the absence of gefitinib for 2 weeks. Then, cells were sequentially treated with gefitinib at concentrations of 500 nM for 1 week, 1 μM for another 1 week and 10 μM for the last 1 week. F, G Cell viability was determined by CCK-8 and claudin1 expression was detected by western blotting. H, I Significantly downregulated and upregulated CLDNs in gefitinib treated or untreated HCC827 cells. Statistical comparisons were performed using a one-way analysis of variance with Dunnett’s test. n = 3, **P < 0.01, ****P < 0.0001. Volcano map: Upregulated: P < 0.05, log₂FC > 0; Downregulated: P < 0.05, log₂FC < 0. J Significantly altered CLDNs mRNA expression levels in gefitinib-treated or untreated PC9 cells were evaluated. n = 3, **P < 0.01. K PC9 cells were treated with gefitinib (10 nM) after which claudin1 expression levels at the indicated time points were determined. L, M PC9/GR and H1975 cells were treated with the indicated concentrations of gefitinib for 24 or 48 h. Claudin1 expression was assessed by western blotting.