The long non-coding RNA, GAS5, enhances gefitinib-induced cell death in innate EGFR tyrosine kinase inhibitor-resistant lung adenocarcinoma cells with wide-type EGFR via downregulation of the IGF-1R expression

Abstract

BackgroundEpidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) are approved for patients with recurrent non-small cell lung cancer (NSCLC). However, the efficacy of EGFR-TKIs in NSCLC therapy is limited by primary and acquired resistance. Recent studies have revealed that long non-coding RNAs (LncRNA) may be involved in EGFR-TKI resistance. Therefore, a better understanding of the interactive mechanisms underlying LncRNA-mediated EGFR-TKIs resistance may help us to improve clinical response rates.MethodTo investigate the expression of growth arrest-specific 5 (GAS5) in lung adenocarcinoma, we performed real-time reverse-transcriptase polymerase chain reaction. The correlation between GAS5 expression levels and the samples’ clinicopathological features was also analyzed. Primary resistance to EGFR-TKIs was identified in the human lung adenocarcinoma cell line A549. Plasmid vectors were used to overexpress GAS5 in A549 cells. MTT (3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide) colony formation assays and EdU (5-ethynyl-2’-deoxyuridine) assays were used to assess cell proliferation, and flow-cytometric analysis was used to evaluate the apoptosis rate. The expression levels of our target proteins, namely, EGFR, p-EGFR, ERK, p-ERK, Akt, p-Akt, IGF-1R (insulin-like growth factor 1 receptor), and p-IGF-1R, were analyzed by western blotting. A549 cells transfected with pcDNA-GAS5 were injected into nude mice. The transplanted mice were treated with gefitinib to study the effect of GAS5 on the resistance to EGFR-TKIs in vivo.ResultsOur results showed that GAS5 was significantly downregulated in lung adenocarcinoma tissues compared with the paired adjacent non-tumorous tissue samples. Furthermore, lower GAS5 expression levels were associated with larger tumor sizes, poor tumor differentiation, and advanced pathological stages. However, GAS5 was almost equally expressed between benign tumors compared with the adjacent normal tissues. GAS5 was also overexpressed in EGFR-TKI sensitive cell lines compared with the resistant cell line. Using MTT, EdU incorporation, and colony formation assays, we showed that GAS5-expressing A549 cells displayed an elevated level of cell death. In addition to its pro-apoptotic effect in the A549 cell line, GAS5 overexpression also suppressed the growth of A549-derived tumors in nude mice treated with gefitinib. GAS5 overexpression was inversely correlated with the expression of the EGFR pathway and IGF-1R proteins.ConclusionsCollectively, our results indicated that GAS5 LncRNA may represent a potential biomarker for the diagnosis of lung adenocarcinoma and that GAS5 might play a novel role in the development of the resistance to gefitinib, which could be reversed by overexpressing GAS5.

Repression of PDK1- and LncRNA HOTAIR-Mediated EZH2 Gene Expression Contributes to the Enhancement of Atractylenolide 1 and Erlotinib in the Inhibition of Human Lung Cancer Cells

“…Disregulation of lncRNAs other than HOTAIR plays critical roles in EGFR-TKIs resistant NSCLC cells. The lncRNA-mediated EGFR-TKIs resistance might help us to improve clinical response rates and could be used as novel potential targets to reverse the EGFR-TKI resistance for NSCLC patients [45,46]. Our findings implied that targeting In addition, we demonstrated an important role of EZH2 in this process.…”

Genome-wide profiling of long non-coding RNA expression patterns in the EGFR-TKI resistance of lung adenocarcinoma by microarray

“…lncRNAs also take part in the regulation of EGFR-TKI resistance. GAS5, one lncRNA, was found to be overexpressed in EGFR-TKI-sensitive cells compared with its expression in resistant cells, and was found to enhance gefitinib-induced cell death in innate EGFR-TKIresistant LAC cells with wild-type EGFR via downregulation of IGF-1R expression (10). These findings indicate that lncRNAs may be promising biomarkers as diagnostic and therapeutic targets in the resistance of EGFR-TKIs.…”

Long Non-Coding RNA in Non-Small Cell Lung Cancers

“…GAS5 could inhibit autophagy and therefore enhance cisplatin sensitivity in NSCLC cells [86]. Another study found that GAS5 overexpression was inversely correlated with EGFR pathway and the expression of IGF-1R proteins in human ADC cell line, indicating its role in reversing EGFR-TKIs resistance [87]. These indings indicate the tumour suppressor lncRNA GAS5 may represent a potential biomarker for diagnosis and therapy target for NSCLC intervention.…”