The long noncoding RNA ZFAS1 promotes the progression of glioma by regulating the miR-150-5p/PLP2 axis

Numerous studies have reported that long noncoding RNA (lncRNA) dysregulation is involved in the progression of many malignant tumors, including glioma. The lncRNA ZNFX1 antisense RNA 1 (ZFAS1) plays an oncogenic role in various malignant tumors, such as gastric cancer and hepatocellular carcinoma. However, the underlying molecular mechanism of ZFAS1 in glioma has not been fully clarified. In this study, we found that the expression of ZFAS1 was upregulated in both glioma tissues and cell lines. Functional experiments revealed that ZFAS1 promoted glioma proliferation, migration and invasion, and increased resistance to temozolomide in vitro. By using online databases, RNA pull-down assays and luciferase reporter assays, ZFAS1 was demonstrated to act as a sponge of miR-150-5p. Furthermore, proteolipid protein 2 (PLP2) was shown to be the functional target of miR-150-5p. Rescue experiments revealed that ZFAS1 regulated the expression of PLP2 by sponging miR-150-5p. Finally, a xenograft tumor assay demonstrated that ZFAS1 promoted glioma growth in vivo. Our results showed that ZFAS1 promoted glioma malignant progression by regulating the miR-150-5p/PLP2 axis, which may provide a potential therapeutic target for the treatment of glioma.     

Lentiviral delivery of a shRNA sequence analogous to miR-4319/miR-125-5p induces apoptosis in NSCLC cells by arresting G2/M phase

In this study, we explored the therapeutic potential of microRNA (miR) analogs against non–small-cell lung cancer (NSCLC) using lentiviral delivery of short hairpin RNA (shRNA). By using A549 as a model cell line, we used analogs and mimics of miR-4319/miR-125-5p to target the tumorigenic RAF1 gene. Lentiviral vectors carrying shRNA of a highly efficient miRNA analog of miR-4319/miR-125-5p, Analog2, were constructed to infect A549 cells. Our results showed that, compared with the noncancerous bronchial epithelial cell line 16HBE, lentivirus delivering Analog2 shRNA induced significant G2/M arrest and subsequent apoptosis in A549 cells, but not in 16HBE cells. Western blot analysis revealed that key factors regulating cell cycle were downregulated following RAF1 inhibition. In vivo xenograft experiments showed that lentivirus carrying Analog2 shRNA markedly decreased tumor size. Therefore, lentiviral delivery of Analog2 shRNA is a valid RNA interference-based treatment against NSCLC with high potency and specificity.     

Retracted: miR-942 promotes tumor migration,invasion, and angiogenesis by regulating EMT via BARX2 in non-small-cell lung cancer

Epithelial–mesenchymal transition (EMT) has an important function in cancer. Recently, microRNAs have been reported to be involved in EMT by regulating target genes. miR-942 is considered a novel oncogene in esophageal squamous cell carcinoma. However, its role in non-small-cell lung cancer (NSCLC) has not been investigated. In this study, the expression of miR-942 in NSCLC patients tumor and paired adjacent tissues were assessed by quantitative real-time polymerase chain reaction and in situ hybridization. Transwell, wound healing, tube formation, and tail vein xenograft assays were conducted to assess miR-942′s function in NSCLC. Potential miR-942 targets were confirmed using dual-luciferase reporter assays, immunohistochemistry, immunoblot, and rescue experiments. The results showed miR-942 is relatively highly expressed in human NSCLC tissues and cells. In vitro assays demonstrated that overexpression of miR-942 promoted cell migration, invasion, and angiogenesis. Tail vein xenograft assays suggested that miR-942 contributed to NSCLC metastasis in vivo. Three bioinformatics software was searched, and BARX2 was predicted as a downstream target of miR-942. Direct interaction between them was validated by dual-luciferase assays. Rescue experiments further confirmed that BARX2 overexpression could reverse functional changes caused by miR-942. Moreover, miR-942 increased EMT-associated proteins N-cadherin and vimentin by inhibiting BARX2, while E-cadherin expression is reduced. In summary, this study reveals that miR-942 induces EMT-related metastasis by directly targeting BARX2, which may provide a potential therapeutic strategy for NSCLC.     

Knockdown of LncRNA PVT1 inhibits tumorigenesis in non-small-cell lung cancer by regulating miR-497 expression

Plasmacytoma variant translocation1 (PVT1) was reported to be upregulated in non-small-cell lung cancer (NSCLC) tissues, serve as a promising biomarker for diagnosis and prognosis of NSCLC, and promoted NSCLC cell proliferation. However, the detailed molecular mechanism of PVT1 involved in the pathogenesis and development of NSCLC remains largely unknown. In this study, the expression levels of PVT1 and miR-497 in NSCLC cells were determined by qRT-PCR. Cell viability, invasion and apoptosis were detected by MTT assay, cell invasion assay and flow cytometry analysis, respectively. RNA immunoprecipitation (RIP) and luciferase reporter assay were performed to confirm whether PVT1 directly interacts with miR-497. A xenograft mouse model was established to confirm the effect of PVT1 on tumor growth in vivo and the underlying molecular mechanism. Our findings indicated that PVT1 was significantly upregulated and miR-497 was markedly downregulated in NSCLC cell lines. si-PVT1 effectively decreased the expression of PVT1 and increased the expression of miR-497. PVT1 knockdown remarkably inhibited cell viability, invasion and promoted apoptosis in NSCLC cells. RIP and luciferase reporter assay demonstrated that PVT1 could directly interact with miR-497. Moreover, PVT1 overexpression reversed the inhibitory effect of miR-497 on cell viability, invasion and promotion effect on apoptosis of NSCLC cells. Furthermore, in vivo experiment showed that knockdown of PVT1 inhibited tumor growth in vivo and promoted miR-497 expression. In conclusion, knockdown of PVT1 inhibited cell viability, invasion and induced apoptosis in NSCLC by regulating miR-497 expression, elucidating the molecular mechanism of the oncogenic role of PVT1 in NSCLC and providing an lncRNA-directed target for NSCLC.     

Up-regulation of long noncoding RNA MINCR promotes non-small cell of lung cancer growth by negatively regulating miR-126/SLC7A5 axis

A growing body of evidence suggests that MYC induced long noncoding RNA (MINCR) is involved in the initiation and progression of various tumors. However, little is known about the biological function and clinical value of MINCR in non-small cell lung cancer (NSCLC). In the present study, results found that MINCR over expression in NSCLC tissue and cell lines was closely related to poor survival in NSCLC. Functional experiments found that decreased MINCR expression inhibits NSCLC cell proliferation and migration and promotes cells apoptosis. Tumor formation assay found that knockdown of MINCR significantly inhibited tumor growth. Results also found that MINCR functions as an oncogene in the metastasis of NSCLC, in part, by acting as a competing endogenous RNA to modulate the miR-126/SLC7A5 axis. Dysfunction of MINCR, miR-126 and SLC7A5 predicted poor prognosis of patients with NSCLC. In conclusion, results suggest that the MINCR-miR-126-SLC7A5 axis plays an important role in the progression of NSCLC and may serve as a potential target for lung cancer diagnosis and treatment.     

Knockdown of lncRNA ZFAS1-suppressed non–small cell lung cancer progression via targeting the miR-150-5p/HMGA2 signaling

Non–small cell lung cancer (NSCLC) is the main type of lung malignancy. Early diagnosis and treatments for NSCLC are far from satisfactory due to the limited knowledge of the molecular mechanisms regarding NSCLC progression. Long noncoding RNA (lncRNA) ZNFX1 antisense RNA1 (ZFAS1) has been implicated for its functional role in the progression of malignant tumors. This study aimed to determine the ZFAS1 expression from lung cancer clinical samples and to explore the molecular mechanisms underlying ZFAS1-modulated NSCLC progression. Experimental assays revealed that clinical samples and cell lines of lung malignant tumors showed an upregulation of ZFSA1. ZFAS1 expression was markedly upregulated in the lung tissues from patients with advanced stage of this malignancy. The loss-of-function assays showed that knockdown of ZFAS1-suppressed NSCLC cell proliferative, as well as invasive potentials, increased NSCLC cell apoptotic rates in vitro and also attenuated tumor growth of NSCLC cells in the nude mice. Further experimental evidence showed that ZFAS1 inversely affected miR-150-5p expression and positively affected high-mobility group AT-hook 2 (HMGA2) expression in NSCLC cell lines. MiR-150-5p inhibition or HMGA2 overexpression counteracted the effects of ZFAS1 knockdown on NSCLC cell proliferative, invasive potentials and apoptotic rates. In light of examining the clinical lung cancer samples, miR-150-5p expression was downregulated and the HMGA2 expression was highly expressed in the lung cancer tissues compared with normal ones; the ZFAS1 expression showed a negative correlation with miR-150-5p expression but a positive correlation with HMGA2 expression in lung cancer tissues. To summarize, we, for the first time, demonstrated the inhibitory effects of ZFAS1 knockdown on NSCLC cell progression, and the results from mechanistic studies indicated that ZFAS1-mediated NSCLC progression cells via targeting miR-150-5p/HMGA2 signaling.     

Circular RNA TUBA1C accelerates the progression of non-small-cell lung cancer by sponging miR-143-3p

Non-small-cell lung cancer (NSCLC) is one of the most common solid tumors and the leading cause of lung cancer-related fatality. Growing evidence has indicated that circular RNAs (circRNAs) play important roles in the progression of multiple human cancers. As a novel circRNA, very little research has focused on the function of circRNA TUBA1C (circTUBA1C) in cancer development, including NSCLC. In the present study, we found that the expression of circTUBA1C was significantly upregulated in NSCLC tissues. The loss-of function assays suggested that circTUBA1C deficiency notably hampered cell proliferation as well as accelerated cell apoptosis in NSCLC. In mechanism, we discovered that circTUBA1C could act as a sponge for miR-143-3p and then negatively regulate miR-143-3p. Moreover, rescue assays demonstrated that knockdown of miR-143-3p could reverse circTUBA1C silence-mediated effects on cell proliferation and apoptosis. Besides, we established a xenografted tumor model to investigate the function of circTUBA1C in vivo. The result illustrated that the decline of tumor growth resulted from circTUBA1C deficiency could be recovered by miR-143-3p knockdown. Taken together, these findings indicated the important role of circTUBA1C/miR-143-3p axis in NSCLC, which may provide a potential target for NSCLC therapy.     

MicroRNA-429 induces tumorigenesis of human non-small cell lung cancer cells and targets multiple tumor suppressor genes

Lung cancer is the major cause of cancer death globally. MicroRNAs are evolutionally conserved small noncoding RNAs that are critical for the regulation of gene expression. Aberrant expression of microRNA (miRNA) has been implicated in cancer initiation and progression. In this study, we demonstrated that the expression of miR-429 are often upregulated in non-small cell lung cancer (NSCLC) compared with normal lung tissues, and its expression level is also increased in NSCLC cell lines compared with normal lung cells. Overexpression of miR-429 in A549 NSCLC cells significantly promoted cell proliferation, migration and invasion, whereas inhibition of miR-429 inhibits these effects. Furthermore, we demonstrated that miR-429 down-regulates PTEN, RASSF8 and TIMP2 expression by directly targeting the 3′-untranslated region of these target genes. Taken together, our results suggest that miR-429 plays an important role in promoting the proliferation and metastasis of NSCLC cells and is a potential target for NSCLC therapy.     

TGF-β1-mediated exosomal lnc-MMP2-2 increases blood–brain barrier permeability via the miRNA-1207-5p/EPB41L5 axis to promote non-small cell lung cancer brain metastasis

Brain metastases remain a major problem in patients with advanced non-small cell lung cancer (NSCLC). The permeability of the blood–brain barrier (BBB) is highly increased during lung cancer brain metastasis; however, the underlying mechanism remains largely unknown. We previously found that lnc-MMP2-2 is highly enriched in tumor growth factor (TGF)-β1-mediated exosomes and regulates the migration of lung cancer cells. This study aimed to explore the role of exosomal lnc-MMP2-2 in the regulation of BBB and NSCLC brain metastasis. Here, using endothelial monolayers and mouse models, we found that TGF-β1-mediated NSCLC-derived exosomes efficiently destroyed tight junctions and the integrity of these natural barriers. Overexpression of lnc-MMP2-2 in human brain microvascular endothelial cells increased vascular permeability in endothelial monolayers, whereas inhibition of lnc-MMP2-2 alleviated these effects. Furthermore, lnc-MMP2-2 knockdown markedly reduced NSCLC brain metastasis in vivo. Mechanistically, through luciferase reporter assays, RNA pull-down assay, and Ago2 RNA immunoprecipitation assay, we showed that lnc-MMP2-2 served as a microRNA sponge or a competing endogenous RNA for miR-1207-5p and consequently modulated the derepression of EPB41L5. In conclusion, TGF-β1-mediated exosomal lnc-MMP2-2 increases BBB permeability to promote NSCLC brain metastasis. Thus, exosomal lnc-MMP2-2 may be a potential biomarker and therapeutic target against lung cancer brain metastasis.Subject terms: Lung cancer, Metastasis     

Circ_0025039 acts an oncogenic role in the progression of non-small cell lung cancer through miR-636-dependent regulation of CORO1C

Non-small cell lung cancer remains the leading cause of cancer-related death worldwide. Circular RNA plays vital roles in NSCLC progression. This study is designed to reveal the role of circ_0025039 in NSCLC cell malignancy. The RNA expression of circ_0025039, microRNA-636 (miR-636), and coronin 1C was detected by quantitative real-time polymerase chain reaction. Protein expression was checked by Western blot analysis or immunohistochemistry assay. Cell proliferation, migration, invasion, tube formation ability, sphere formation capacity, and apoptosis were investigated by cell counting kit-8, 5-Ethynyl-29-deoxyuridine, transwell assay, tube formation assay, sphere formation assay, and flow cytometry analysis, respectively. Mouse model assay was conducted to reveal the effect of circ_0025039 silencing on tumor formation in vivo. The interaction between miR-636 and circ_0025039 or CORO1C was identified through dual-luciferase reporter and RNA pull-down assays. The expression of circ_0025039 and CORO1C was significantly increased, while miR-636 was decreased in NSCLC tissues and cells compared with controls. Circ_0025039 depletion repressed NSCLC cell proliferation, migration, invasion, tube-forming capacity, and sphere formation ability, but induced cell apoptosis. The neoplasm formation was repressed after circ_0025039 silencing. Additionally, circ_0025039 acted as a sponge for miR-636, which was found to target CORO1C. Importantly, the contribution of circ_0025039 to NSCLC progression was mediated by miR-636/CORO1C axis. Circ_0025039 silencing repressed NSCLC malignant progression by reducing CORO1C expression through miR-636, showing the possibility of circ_0025039 as a therapeutic target for NSCLC.

     

Circular RNA hsa_circ_0004015 regulates the proliferation,invasion, and TKI drug resistance of non-small cell lung cancer by miR-1183/PDPK1 signaling pathway

Circular RNAs (circRNAs) were recently reported to be involved in the pathogenesis of Non-small cell lung cancer (NSCLC), however, the molecular mechanisms of circRNAs in cell proliferation, invasion and TKI drug resistance remain largely undetermined. Here, we identified hsa_circ_0004015 was upregulated in NSCLC tissues, and was associated with the poor overall survival rate of NSCLC patients. Knockdown of hsa_circ_0004015 significantly decreased cell viability, proliferation, and invasion, whereas overexpression exhibited opposed effects in vivo and in vitro. Furthermore, hsa_circ_0004015 could enhance the resistance of HCC827 to gefitinib. In mechanism, hsa_circ_0004015 acted as a sponge for miR-1183, and PDPK1 was revealed to be target gene of miR-1183. Subsequently, functional assays illustrated that the oncogenic effects of hsa_circ_0004015 was attributed to the regulation of miR-1183/PDPK1 axis. In conclusion, circ_0016760/miR-1183/PDPK1 signaling pathway might play vital roles in the tumorigenesis of NSCLC.     

Circular RNA circPIP5K1A promotes non-small cell lung cancer proliferation and metastasis through miR-600/HIF-1α regulation

Circular RNAs (circRNAs) have an important function in human diseases, especially in cancer. circRNA hsa_circ_0014130 (circPIP5K1A), a particularly abundant circRNA, participates in the tumorigenesis of non-small cell lung cancer (NSCLC), although the underlying regulatory mechanism remains unclear. Here, we investigated the circPIP5K1A role in NSCLC. Expression of circPIP5K1A in NSCLC cell lines was explored with quantitative real-time PCR. The effect of circPIP5K1A on NSCLC was evaluated with circPIP5K1A silencing, miR-600 mimic transfection, and hypoxia-inducible factor (HIF)-1α overexpression, followed by assessment of cell proliferation, metastasis, and tumorigenesis in nude mice. The subcellular localization of circPIP5K1A was evaluated via fluorescence in situ hybridization (FISH), and correlation between circPIP5K1A, miR-600, and HIF-1α was assessed by luciferase assay. The data demonstrated that circPIP5K1A expression was increased in NSCLC cells. FISH showed that circPIP5K1A localized to the cytoplasm. The circPIP5K1A knockdown suppressed NSCLC cell metastasis and proliferation by promoting expression of miR-600. Overexpression of miR-600 inhibited HIF-1α-mediated metastasis and proliferation of NSCLC cell by downregulating the endothelial mesenchymal transition-related proteins, Snail and vimentin, and upregulating E-cadherin. In vivo experiments illustrated that circPIP5K1A silence suppressed tumor growth and pulmonary metastasis. The circPIP5K1A may function as an miR-600 sponge to facilitate NSCLC proliferation and metastasis by promoting HIF-1α. A bifluorescein reporter experiment confirmed that miR-600 was the circPIP5K1A target, and miR-600 interacted with the 3′ untranslated region of HIF-1α. These results show that circPIP5K1A acted as a tumor promoter through a novel circPIP5K1A/miR-600/HIF-1α axis, which provides candidate markers and therapeutic targets for NSCLC.     

lncRNA HNF1A-AS1 modulates non–small cell lung cancer progression by targeting miR-149-5p/Cdk6

Growing evidence have shown the important regulation of lncRNAs (long noncoding RNAs) in non–small cell lung cancer (NSCLC). lncRNA hepatocyte nuclear factor 1 homeobox A (HNF1A)-antisense RNA 1 (AS1), an “oncogene”, was reported to regulate human tumors progression. However, the molecular mechanism of HNF1A-AS1 involved in the development of NSCLC is still under investigation. In the current study, we found that HNF1A-AS1 was relatively upregulated in both NSCLC patient tissues and cell lines. Functional studies established that overexpression of HNF1A-AS1 promoted cell proliferation, cell cycle, invasion, and migration of NSCLC cells in vitro. The promotion abilities of HNF1A-AS1 on NSCLC cell progression were suppressed via knockdown of HNF1A-AS1. miR-149-5p was then proved to be a novel target of HNF1A-AS1, whose expression was negatively correlated with HNF1A-AS1 in NSCLC patient tissues and cell lines. HNF1A-AS1 increased the expression of cyclin-dependent kinase 6 (Cdk6) via sponging with miR-149-5p. Gain- and loss-of-functional studies indicated that HNF1A-AS1 promoted NSCLC progression partially through inhibition of miR-363-3p and induction of Cdk6. Subcutaneous xenotransplanted tumor model confirmed that interference of HNF1A-AS1 suppressed the tumorigenic ability of NSCLC via upregulation of miR-149-5p and downregulation of Cdk6 in vivo. In conclusion, our findings clarified the biologic significance of the HNF1A-AS1/miR-149-5p/Cdk6 axis in NSCLC progression and provided novel evidence that HNF1A-AS1 may be a new potential therapeutic target for the treatment of NSCLC.     

Inhibitory role of bone marrow mesenchymal stem cells-derived exosome in non-small-cell lung cancer: microRNA-30b-5p,EZH2 and PI3K/AKT pathway

Exosomal microRNA (miRNA) exerts potential roles in non-small-cell lung cancer (NSCLC). The current study elucidated the role of miR-30b-5p shuttled by bone marrow mesenchymal stem cells (BMSCs)-derived exosomes in treating NSCLC. Bioinformatics analysis was performed with NSCLC-related miRNA microarray GSE169587 and mRNA data GSE74706 obtained for collection of the differentially expressed miRNAs and mRNAs. The relationship between miR-30b-5p and EZH2 was predicted and confirmed. Exosomes were isolated from BMSCs and identified. BMSCs-derived exosomes overexpressing miR-30b-5p were used to establish subcutaneous tumorigenesis models to study the effects of miR-30b-5p, EZH2 and PI3K/AKT signalling pathway on tumour growth. A total of 86 BMSC-exo-miRNAs were differentially expressed in NSCLC. Bioinfomatics analysis found that BMSC-exo-miR-30b-5p could regulate NSCLC progression by targeting EZH2, which was verified by in vitro cell experiments. Besides, the target genes of miR-30b-5p were enriched in PI3K/AKT signalling pathway. Animal experiments validated that BMSC-exo-miR-30b-5p promoted NSCLC cell apoptosis and prevented tumorigenesis in nude mice via EZH2/PI3K/AKT axis. Collectively, the inhibitory role of BMSC-derived exosomes-loaded miR-30b-5p in NSCLC was achieved through blocking the EZH2/PI3K/AKT axis.     

CircRNA fibroblast growth factor receptor 3 promotes tumor progression in non-small cell lung cancer by regulating Galectin-1-AKT/ERK1/2 signaling

The dysregulation of circular RNA (circRNA) expression is involved in the progression of several cancers, including non-small cell lung cancer (NSCLC). However, the role and underlying molecular mechanisms of circRNA FGFR3 (circFGFR3) in NSCLC progression remains unknown. Here, we used quantitative real-time polymerase chain reaction to validate that circFGFR3 expression was higher in NSCLC tissues than in the paratumor tissues. Furthermore, our study indicated that the forced circFGFR3 expression promoted NSCLC cell invasion and proliferation. Mechanistically, we found that circFGFR3 promoted NSCLC cell invasion and proliferation via competitively combining with miR-22-3p to facilitate the galectin-1 (Gal-1), p-AKT, and p-ERK1/2 expressions. Clinically, we revealed that the high circFGFR3 expression correlates with the poor clinical outcomes in patients with NSCLC. Together, these data provide mechanistic insights into the circFGFR3-mediated regulation of both the AKT and ERK1/2 signaling pathways by sponging miR-22-3p and increasing Gal-1 expression.