Objective To investigate the effect and mechanism of circular RNA (circRNA)_0128846 on the radioresistance of human non-small cell lung cancer (NSCLC) cells.

Methods The circRNA with the most significant differential expression in the human NSCLC cell line A549 and its radioresistant cell line (A549R) was considered as the target circRNA. It was screened by using the GEO database and detected through fluorescence real-time quantitative polymerase chain reaction (qRT-PCR). qRT-PCR was utilized to detect the expression of the target circRNA in the human bronchial epithelial cell strain Beas-2B and NSCLC cells A549, H460, H1299, and H1975. A549 cells were transfected with a silencing vector, and A549R cells were transfected with an overexpression vector. Clone formation assay was used to detected the clone formation ability of the cells after 8 Gy X-ray irradiation. The downstream miRNA of the target circRNA was predicted with the human circular RNA database and circinteractome database. qRT-PCR was applied to detect the miRNA with the expression level that had increased most significantly after the target circRNA was silenced in the A549R cells, and regarded it as the target miRNA. The expression of the target miRNA after the overexpression of the target circRNA in A549 cells was detected with qRT-PCR. A dual-luciferase reporter gene system was employed to analyze the expression of the target circRNA after the target miRNA was overexpressed in human embryonic kidney cells 293T. The expression level of the target miRNA in A549 and A549R cells was detected through qRT-PCR. The silencing and overexpression vectors of the target miRNA were transfected into A549 and A549R cells, respectively. Then, clone formation ability was detected after 8 Gy X-ray irradiation. The A549 cells overexpressing the target circRNA were irradiated with 8 Gy X-ray, and the target miRNA was overexpressed in the cells to detect their clone formation ability. Data were compared between groups by using independent sample t test.

Results qRT-PCR results revealed that circRNA_0128846 was the target circRNA, and its relative expression level in human NSCLC cells A549, H460, H1299, and H1975 was higher than that in the human bronchial epithelial cell line Beas-2B (t=6.200, 7.903, 6.010, 6.132; all P<0.01). After circRNA_0128846 was overexpressed, the clone formation ability of irradiated A549 cells was enhanced (0.22% vs. 0.45%, t=4.427, P<0.05). After circRNA_0128846 was silenced, the clone formation ability of irradiated A549R cells decreased (0.23% vs. 0.10%, t=3.780, P<0.05). qRT-PCR results showed that miR-1183 was the target miRNA, and the overexpression of circRNA_0128846 in A549 cells significantly down-regulated the expression of miR-1183 (t=6.002, P<0.01). Dual-luciferase reporter gene analysis confirmed that the overexpression of miR-1183 could significantly reduce the expression of circRNA_0128846 (t=4.562, P<0.05). The relative expression level of miR-1183 in A549R cells was significantly lower than that in parental A549 cells (t=6.025, P<0.01). The overexpression of miR-1183 significantly reduced the clone formation ability of A549R cells after irradiation; the silencing of miR-1183 significantly enhanced the clone formation ability of A549 cells after irradiation (0.26% vs. 0.15%, 0.21% vs. 0.31%; t=3.671, 3.293; both P<0.05); and the overexpression of miR-1183 significantly reduced the promoting effect of the overexpression of circRNA_0128846 on the clone formation ability in A549 cells (1.90% vs. 1.20%, t=6.325, P<0.01).

Conclusion In human NSCLC cells, circRNA_0128846 acts as a miR-1183 adsorption sponge that can inhibit the radiosensitization effect of miR-1183 by reducing miR-1183 expression, thereby promoting NSCLC radioresistance.