Let-7a-3p inhibits activity of cancer stem cells in human lung cancer A549 cells by targeting IGF1R

AIM:To study the effect of let-7a-3p on the activity of cancer stem cells in human lung cancer A549 cells and its molecular biological mechanism. METHODS:The exepression levels of let-7a-3p in lung cancer cell lines A549, NCI-H1299, SPC-A1, H1650 and HCC-827, and human normal bronchial epithilial cell line BEAS-2B were compared by RT-qPCR. The lung cancer A549 cells were transfected with let-7a-3p mimic and negative control mimic, as let-7a-3p group and negative control group, respectively, and non-transfected control group was also set up. The content of let-7a-3p in each group was detected by RT-qPCR. Tumor sphere formation assay was used to detect the tumor sphere formation ability in 3 groups of the cancer stem cells. The proportion of cancer stem cells was detected by flow cytometry. The protein levels of NANOG, OCT4 and insulin-like growth factor 1 receptor (IGF1R) were determined by Western blot. The target gene of let-7a-3p was predicted by the bioinformatic method. The relationship between let-7a-3p and IGF1R was analyzed by double luciferase assay. Western blot was used to detect whether IGF1R over-expression antagonized the inhibitory effect of let-7a-3p on the activity of cancer stem cells. A subcutaneous transplantation tumor model was also established and the effect of let-7a-3p in vivo was observed. RESULTS:The expression level of let-7a-3p in the lung cancer cell lines was significantly lower than that in the normal bronchial epithelial cell line (P<0.01). The expression level of let-7a-3p in the A549 cells of let-7a-3p group was significantly up-regulated compared with non-transfected group (P<0.01). The number of tumor spheres in let-7a-3p group was significantly lower than that in non-transfected group. The percentage of CD133⁺ cells in let-7a-3p group was significantly lower than that in non-transfected group (P<0.01). The protein expression of NANOG and OCT4 in let-7a-3p group was significantly lower than that in non-transfected group (P<0.01). Bioinformatic prediction showed that let-7a-3p complementarily matched the 3'-UTR of IGF1R, and IGF1R might be the target gene of let-7a-3p. Luciferase assay confirmed that IGF1R was the direct downstream target gene of let-7a-3p. The protein expression of IGF1R in let-7a-3p group was significantly decreased (P<0.01). Subcutaneously transplantated tumor in let-7a-3p group was significantly smaller than that in non-transfected group. CONCLUSION:Let-7a-3p may affect the expression of lung cancer stem cell-related proteins and inhibit the potential of lung cancer stem cells by down-regulating its downstream target gene IGF1R.