Osimertinib suppresses hTERT expression accompanied by telomerase/telomere inhibition and TIF induction. (A and B) RNA-seq data were generated from PC-9 cells treated with DMSO or 100 nM osimertinib (Osim) for 14 h. Each column is the mean ± SD of triplicate treatments. FPKM, fragments per kilobase per million. (C) RT-qPCR data for hTERT suppression by osimertinib in the indicated cell lines exposed to DMSO or 100 nM osimertinib for 14 h. Each column is the mean ± SD of triplicate treatments. (D–G) The given cell lines were exposed to different concentrations of osimertinib as indicated for 24 h (D), 200 nM osimertinib for varied times as indicated (E), 200 nM indicated EGFR-TKIs for 24 h (F), or 500 nM osimertinib for 24 h (G). The proteins of interest were detected with western blotting. (H) IF was used to detect hTERT in PC-9 tumors treated with 15 mg/kg osimertinib for 9 days. (I and J) The indicated cell lines were exposed to DMSO or 100 nM osimertinib for 24 h and then subject to telomerase (I) and telomere length (J) assays using the Telomerase Activity Quantification qPCR Assay and Absolute Human Telomere Length Quantification qPCR Assay kits, respectively. The data are means ± SDs of four replicate treatments. (K and L) Both PC-9 and HCC827 cell lines were treated with 200 nM osimertinib for 24 h followed with the TIF assay. TIFs were counted from 20 cells for each treatment and represented as means ± SEs. Arrows indicate TIFs (colocalization of TRF2 and γ-H2AX). Statistical differences were assessed with two-sided unpaired Student’s t test. Source data are available for this figure: SourceData F1.