hTERT expression is elevated in EGFRm NSCLC cell lines and patient tumor tissues with acquired resistance to osimertinib. (A) RNA-seq data for hTERT upregulation in osimertinib-resistant PC-9 cells. Each column is the mean ± SD of triplicate treatments. (B) RT-qPCR detection of hTERT mRNA levels in the indicated cell lines. Each column is the mean ± SD of triplicate treatments. (C–E) Western blotting detection of hTERT protein from the indicated cell lines with or without exposure to 200 nM osimertinib (Osim) or HS-10296 for 16 h (C and D) or basal levels of TERT in the indicated cell lines (E). P, parental; AR, AZD9291-resistant; HSR, HS-10296-resistant. (F) IF detection of hTERT in the indicated cell lines exposed to 100 nM osimertinib for 24 h. (G and H) Telomerase activity (G) and telomere length (H) in the tested cell lines were detected using the Telomerase Activity Quantification qPCR Assay and Absolute Human Telomere Length Quantification qPCR Assay kits, respectively. The data are the means ± SDs of four replicate determinations. (I and J) hTERT in human EGFRm NSCLC issues were detected with IHC and representative pictures of hTERT staining are shown (J). WI, weight index, which represents % positive stain × intensity score (0–3). Statistical differences were assessed with a two-sided unpaired Student’s t test. Source data are available for this figure: SourceData F3.