Enhanced invasion and migration of non-small cell lung cancer (NSCLC) cells is the major cause of metastasis and poor prognosis in NSCLC. This study was conducted to investigate the role and mechanism of lncRNA KCNQ1OT1 in the proliferation, invasion, and migration of NSCLC cells. The expression of KCNQ1OT1 in NSCLC was analyzed in the StarBase database, and the target miRNA of KCNQ1OT1 as well as the target genes of the miRNA was predicted. Then, the mRNA expression levels of KCNQ1OT1, miR-496, and HMGB1 were detected in clinical tissue samples and cells by qRT-PCR assay. Besides, the protein levels of HMGB1 were detected by Western blot. MTT assay, transwell assay, and scratch assay were used to determine the proliferation, invasion, and migration ability of NSCLC cells, respectively. Correlation analysis was performed to assess the correlation between the expression of KCNQ1OT1, miR-496, and HMGB1 in clinical NSCLC samples. Dual-luciferase reporter gene assay was conducted to analyze the interaction between KCNQ1OT1 and miR-496 and between miR-496 and HMGB1. The database results showed that KCNQ1OT1 was highly expressed in NSCLC. Similarly, we found that the expression level of KCNQ1OT1 was significantly higher in NSCLC tissues and cells than that in the corresponding normal tissues and cells. The results of MTT assay, transwell assay, and scratch assay demonstrated that KCNQ1OT1 significantly enhanced the proliferation, invasion, and migration of NSCLC cells. Further mechanism exploration revealed that KCNQ1OT1 could sponge miR-496, and miR-496 directly targeted and regulated the expression of HMGB1. The expression of miR-496 and either KCNQ1OT1 or HMGB1 were negatively correlated in NSCLC, while the expression of KCNQ1OT1 and HMGB1 were positively correlated. Compared with normal paracancer tissues, miR-496 was much lower and HMGB1 was much higher expressed in NSCLC tissues. The results of cotransfection also further demonstrated that miR-496 inhibitor or sh-HMGB1 cotransfected with sh-KCNQ1OT1 could significantly decrease or increase the ability of sh-KCNQ1OT1 to inhibit the proliferation, invasion, and migration of H1299 cells, respectively. In conclusion, lncRNA KCNQ1OT1 promotes the invasion and migration of NSCLC cells through miR-496/HMGB1 signaling axis.

Background Non-small cell lung cancer (NSCLC) is the most deadly cancer worldwide. LncRNA KCNQ1OT1 has been reported to be involved in the progression of various tumors, including NSCLC. However, the precise mechanism of KCNQ1OT1 in NSCLC requires further investigation. Methods The expression levels of KCNQ1OT1, miR-129-5p and JAG1 were detected by qRT-PCR or western blot. Kaplan-Meier survival analysis was used to assess the correlation between KCNQ1OT1 expression and the overall survival of NSCLC patients. CCK-8 assay was used to measure cell viability. Cell migration and invasion were detected by transwell assay. The targets of KCNQ1OT1 and miR-129-5p were predicted by bioinformatics, which was confirmed by dual-luciferase reporter assay or pull-down assay. Results KCNQ1OT1 expression was significantly enhanced, while miR-129-5p expression was dramatically reduced in NSCLC tissues and cells. Higher KCNQ1OT1 shortened overall survival and was positively associated with tumor stage and lymph node metastasis. KCNQ1OT1 knockdown inhibited proliferation, migration and invasion of NSCLC cells. Inhibition of miR-129-5p attenuated the inhibition of NSCLC cell viability, migration and invasion induced by KCNQ1OT1 knockdown. In addition, JAG1 was confirmed as a target of miR-129-5p. Knockdown of JAG1 reversed the effects of miR-129-5p knockdown on NSCLC progression. KCNQ1OT1 regulated JAG1 expression by sponging miR-129-5p in NSCLC cells. Conclusion KCNQ1OT1 induced proliferation, migration and invasion of NSCLC cells by sponging miR-129-5p and regulating JAG1 expression, indicating that KCNQ1OT1 was a therapeutic target for NSCLC.

Objective The purpose of this study was to investigate whether CDH13 gene expression could be restored by demethylation to reverse the resistance of A549/cisplatin (DDP) cells to cisplatin.Methods The expression of the CDH13 in A549 and A549/DDP cells was detected by reverse transcription-polymerase chain reaction,and the CDH13 expression in the A549/DDP cells that interfered with the methyltransferase inhibitor 5-Aza-2'-deoxy-citydine (5-Aza-dc) was also detected by reverse transcription-polymerase chain reaction.MSP was used to detect the methylation of CDH13 in groups.Results The expression of CDH13 mRNA in A549 cells was higher than the A549/DDP cells.The median inhibitory concentration value of DDP for A549 and A549/DDP cells was (7.053±0.366) μmol/L and (36.351±2.648) μmol/L (t =8.153,P ＜0.05),respectively,and the resistance index was 5.15.MSP showed that the CDH13 gene promoter region was partially methylated.DNA demethylation occurred in the CDH13 gene promoter region with 5-Aza-dc at non-toxic and low-toxic doses after 48 h for treating A549/DDP cells.With increasing concentration of 5-Aza-dc up to 40 μmol/L,the median inhibitory concentration value of DDP for A549/DDP cells is (9.840± 1.337) μmol/L,and the resistance ratio of reversed DDP was about 3.41.Conclusions CDH13 methylation may effect the process of A549/DDP cells on DDP resistance.

Reversing cisplatin resistance of lung cancer cell line A549/DDP through recovering cadherin 13 (CDH13) expression by demethylation was investigated in the current study. RT-PCR was used to measure CDH13 expression in lung cancer A549 and A549/DDP cells with or without 5-Aza-CdR intervention. Methylation-specific PCR was used to detect CDH13 methylation. MTT assay and flow cytometry were used to measure the effects of cisplatin on inhibiting cell proliferation, apoptosis, and the reversal of cisplatin resistance. The IC50 value of cisplatin for A549 and A549/DDP cells was 3.278±0.532 and 28.341±1.435 µmol/l, respectively (P<0.05). The cisplatin-resistance index of A549/DDP cells was up to 8.65. After 2.5, 10, or 40 µmol/l 5-Aza-CdR treatment, the apoptotic rates of A549/DDP cells were 9.4±0.86, 18.1±1.42 and 42±2.01%, respectively, which were significantly different to those of the control group (P<0.05). Methylation-specific PCR detected both methylation (M) and unmethylation (U) bands at CDH13 promoter region before 5-Aza-CdR intervention while it only detected an unmethylation band after the treatment with a higher concentration of 5-Aza-CdR, which indicates the transformation to unmethylation state. When 10 µmol/l 5-Aza-CdR was added, the IC50 of cisplatin to A549/DDP cells was 8.472±0.415 µmol/l, and cisplatin resistance was reversed by 3.35-fold. CDH13 methylation is related to the cisplatin resistance of A549/DDP cells. 5-Aza-CdR can inhibit CDH13 methylation and recover CDH13 expression. With the increase in 5-Aza-CdR concentration, the unmethylation state of CDH13 is enhanced, which can strengthen the function of cisplatin inhibiting proliferation and apoptosis in A549/DDP cells.