Identification of microRNAs involved in gefitinib resistance of non‑small‑cell lung cancer through the insulin‑like growth factor receptor 1 signaling pathway

Multiple clinical and experimental studies have suggested that epidermal growth factor receptor tyrosine kinase inhibitors (EGFR‑TKIs) may be effective at treating advanced non‑small cell lung cancer (NSCLC), however, the molecular basis of primary resistance to EGFR‑TKIs in NSCLC remains unclear. In the current study, the insulin‑like growth factor 1 receptor (IGF‑1R) gene in the gefitinib‑resistant human lung adenocarcinoma epithelial cell line A549 (A549/GR) was silenced using small interfering RNA (siRNA) in order to determine the role of microRNA (miRNA) in the development of resistance against epidermal growth factor receptor tyrosine kinase inhibitors (EGFR‑TKIs) in lung adenocarcinoma. The relative gefitinib‑resistant capacity in A549 and A549/GR cells was determined using a cell counting kit 8. A549/GR cells were transfected with chemically synthesized siRNA to silence the IGF‑1R gene. A total of 48 h after siRNA transfection, IGF‑1R expression in A549/GR cells was evaluated using reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) and western blotting. miRNA expression in A549/GR cells and A549/GR cells with silenced IGF‑1R was analyzed using a miRNA microarray. The microarray results of 10 miRNAs were then compared with the results of RT‑qPCR. The results demonstrated that the gefitinib‑resistance capacity of A549/GR cells was six times higher than that of A549 cells. Additionally, RT‑qPCR and western blotting demonstrated that the IGF‑1R gene in A549/GR cells was successfully silenced by siRNA. The highest silencing rate (72%) of the IGF‑1R gene was obtained using siRNA‑2. The microarray identified 72 miRNAs with significantly different expression in A549/GR cells with silenced IGF‑1R compared with A549/GR cells. Of the 72 differentially expressed miRNAs, 13 miRNAs (including miR‑497‑3p and miR‑1273c) were up‑regulated and 59 miRNAs (including miR‑361‑3p and miR‑345‑3p) were down‑regulated in A549/GR cells with silenced IGF‑1R compared with A549/GR cells. The changes in the expression of 10 different miRNAs were confirmed by RT‑qPCR. Thus, the present study successfully established an A549/GR cell line with silenced IGF‑1R. The results suggest that a number of miRNAs associated with the IGF‑1R signaling pathway, including miR‑497‑3p and miR‑144‑5p, were involved in the development of resistance against EGFR‑TKIs in A549 cells. These miRNAs may provide novel targets to treat lung adenocarcinoma exhibiting resistance against EGFR-TKIs.