Invasion Of Lung Cancer Cells Research Articles

Tumor promoting effects of exosomal microRNA-210 derived from lung cancer cells on lung cancer through the RUNX3/PI3K/AKT signaling pathway axis.

Exosomes are involved in a range of processes in lung cancer such as cell proliferation, metastasis, and angiogenesis. Tumor-derived exosomes participate in the formation and progression of lung cancer by delivering functional biomolecules, including microRNAs (miRNA). The purpose of the present study was to determine the role of lung cancer cell-derived exosomal miR-210 in the proliferation and invasion of lung cancer cells and its underlying mechanism. Initially, exosomes were isolated from A549 cells and characterized by transmission electron microscopy and assessment of exosomal marker expression. RT-qPCR determined that miR-210 expression was elevated in exosomes as well as lung cancer cells. As reflected by dual-luciferase reporter assay, miR-210 negatively regulated RUNX3 expression. Following loss- and gain- function assay, it was found that miR-210 inhibition suppressed biological properties of A549 and H460 cells, which could be reversed by the silencing of RUNX3. miR-210 elevation induced the p-PI3K/PI3K and p-AKT/AKT levels, suggesting the activation of PI3K/AKT signaling pathway. Collectively, exosomal miR-210 targeted and negatively regulated RUNX3 expression to promote malignant properties of lung cancer cells by potentiating PI3K/AKT signaling pathway.

Ectodermal‑neural cortex 1 affects the biological function of lung cancer through the MAPK pathway.

Ectodermal-neural cortex 1 (ENC1), a highly expressed protein in lung cancer tissues, was identified from the Cancer Genome Atlas (TCGA) database. The objective of the present study was to examine the effects of ENC1 on the biological functions of lung cancer cells. For this purpose, the expression of ENC1 was examined by RT-qPCR to compare mRNA expression levels between 28 lung cancer tissue samples and para-cancerous tissue samples. The association between ENC1 expression and clinicopathological features was evaluated between the 2 tissue types. Using RT-qPCR and western blot analysis, the expression of ENC1 was investigated in a normal lung cell line (16HBE) and 2 lung cancer cell lines (A549 and H1299). The effect of siRNA targeting ENC1 (si-ENC1) on the proliferation of A549 and H1299 cells was detected by CCK-8 assay at the indicated time points. Transwell assay was used to measure the migration and invasion of A549 and H1299 cells following transfection with siRNA targeting ENC1 (si-ENC1). The expression levels of several proteins related to migration and invasion were examined by western blot analysis. A mouse model of subcutaneous tumor xenotransplantation was established in nude mice to examine the effects of ENC1 downregulation on cancer cells. The results revealed that the expression of ENC1 in lung cancer tissues and lung cancer cells was significantly higher than that in para-cancerous tissues and non-cancer lung cells, respectively. The knockdown of ENC1 in the A549 and H1299 cells using si-ENC1 significantly decreased cell proliferation, migration and invasion compared with the untransfected cells. The knockdown of ENC1 significantly downregulated the levels of matrix metalloproteinase (MMP)2, MMP9, N-cadherin, p-c-Jun N-terminal kinase (JNK), p-extracellular signal-regulated kinase (ERK) and p-p38. The levels of E-cadherin were upregulated. In the mouse lung tumor model, reduced levels of ENC1 inhibited the growth of lung tumors. On the whole, the present study demonstrates that ENC1 is involved in the proliferation, migration and invasion of lung cancer cells, and may thus be an effective diagnostic target for certain cancers. The inhibition or reduction of ENC1 activity may represent a breakthrough in the treatment of lung cancer.

LSR Promotes Cell Proliferation and Invasion in Lung Cancer.

The lipolysis-stimulated lipoprotein receptor (LSR) displays an important regulatory role in cancer. However, the association between LSR and lung cancer is still elusive. Here, the candidate oncogene LSR on Ch.9q was obtained and assessed by bioinformatics analysis of The Cancer Genome Atlas (TCGA) dataset of lung cancer. We conducted clinical pathology and survival analysis based on the lung cancer database. We assessed the biological effects of LSR in lung cancer cells on cell proliferation. Our data indicated that LSR was upregulated in lung cancer cells. Meanwhile, LSR was identified in this study to be a poor prognostic factor, and its high expression exhibited relations with grades, stages, and nodal metastasis status. Using in vitro analysis, our data revealed that LSR could promote lung cancer progression by regulating cell proliferation, migration, and invasion. In our study, our data demonstrated that LSR was a tumor promoter for lung cancer and was a potential biomarker and target for lung cancer prognosis and treatment.

Tumor Suppressor miR-584-5p Inhibits Migration and Invasion in Smoking Related Non-Small Cell Lung Cancer Cells by Targeting YKT6.

Simple SummaryCigarette smoke is a major carcinogen that causes lung cancer and induces DNA methylation. DNA methylation regulates the expression of microRNA (miRNAs), which are important regulators of cancer biology. However, the association between smoking and miRNAs has not been fully elucidated in smoking-related lung carcinogenesis. In this study, we found that miR-584-5p expression was downregulated with cancer progression using a lung carcinogenesis model cell line. Moreover, we demonstrated that miR-584-5p is downregulated by the methylation of its promoter region and that it suppresses migration and invasion by targeting YKT6 in smoking-related non-small cell lung cancer (NSCLC) cells. Our results provide a better understanding of the underlying changes in miRNA expression in smoking-related lung carcinogenesis and suggest that miR-584-5p is a potential molecular biomarker for smoking-related NSCLC.Cigarette smoke (CS) affects the expression of microRNAs (miRNAs), which are important regulators of gene expression by inducing DNA methylation. However, the effects of smoking on miRNA expression have not been fully elucidated in smoking-related lung carcinogenesis. Therefore, in this study, to investigate the change of miRNA expression pattern and to identify tumor suppressor miRNAs by smoking in lung carcinogenesis, we used lung carcinogenesis model cell lines that, derived from a murine xenograft model with human bronchial epithelial cells (BEAS-2B), exposed CS or not. The microarray analysis revealed that miR-584-5p expression was downregulated with cancer progression in lung carcinogenesis model cell lines. We confirmed by pyrosequencing that the methylation level of the miR-584-5p promoter increased with cancer progression. In vitro and in vivo experiments showed that miR-584-5p suppressed migration and invasion in non-small cell lung cancer (NSCLC) cells by targeting YKT6. Furthermore, we showed that high level of YKT6 was associated with a poor survival rate in NSCLC patients with a history of smoking. These results suggest that miR-584-5p acts as a tumor suppressor and is a potential molecular biomarker for smoking-related NSCLC.

P64.01 MiRNAs in Exosomes Isolated From the Blood of Non-Small Cell Lung Cancer Patients: Biomarkers for Lung Cancer Prediction and Prognosis

Non-small cell lung cancer accounts for about 80% of lung cancer, and about 75% of patients have been found in advanced stages, and their survival rate is very low. However, sensitive and simple diagnostic tests for NSCLC and highly-targeted therapeutic biomarkers are still clinically unusable. The aim of this study was to investigate the expression profiles and clinical significance of miRNAs in platelet-derived exosomes in peripheral blood as biomarkers for NSCLC. The fasting blood samples of 102 patients with NSCLC and 89 healthy people were collected. The exosomes were obtained from platelets using ExoQuick Precipitation Kit. The expression of miR-223 in exosomes from platelets were determined by QRT-PCR. Furthermore, we explored its mechanism in the tumor progress by cell migration assay. First, we found that the levels of miR-223 in platelets and P-MVs were significantly up-regulated in lung cancer patients compared with the non-cancer control group. Second, in vitro assays using qRT-PCR showed that platelet miR-223 was rapidly delivered into cancer cells by P-MVs. Third, platelet-derived exogenous miR-223 reduces the protein level of the tumor suppressor EPB41L3, thereby promoting the invasion of lung cancer cells. The effect of P-MV on the reduction and enhancement of tumor cell invasion by EPB41L3 was largely eliminated by consumption of miR-223, indicating that miR-223 plays a key role in these processes. This study provides additional evidence that functional miRNAs in MVs secreted by cells can serve as biomarkers for lung cancer prediction and prognosis.

Carbonyl Reductase 3-Antisense RNA 1 Negatively Regulates microRNA-337-3p Expression: Effects on Proliferation, Migration, and Invasion of Lung Cancer Cells

Lung cancer, a malignant tumor, is associated with high morbidity and mortality worldwide. We studied the influence and mechanism of CBR3-AS1 on lung cancer cell proliferation, migration, and infiltration. The expression of CBR3-AS1 and miRNA-337-3p were higher and lower (P< 0.05), respectively, in lung cancer tissues than in paracancerous tissues. After inhibiting the expression of CBR3-AS1, the OD value of A549 cells, cloning formation numbers, migrating and invasive numbers, N-cadherin protein expression levels were lower. The G0-G1 cell cycle periods was longer. The S cell cycle periods was shorter. The E-cadherin protein expression levels higher (P< 0.05 in all cases). CBR3-AS1 negatively regulated miRNA-337-3p expression in A549 cells (P< 0.05). After inhibiting the expression of CBR3-AS1 and miRNA-337-3p, the OD value of A549 cells was lower, cloning formation numbers, migrating and invasive numbers, N-cadherin protein expression levels were lower. The G0-G1 cell cycle periods was longer. The S cell cycle periods was shorter. The E-cadherin protein expression levels was higher (P< 0.05 in all cases). CBR3-AS1 expression was increased in lung cancer tissues, and interference with CBR3-AS1 expression could inhibit the proliferation, migration, and infiltration of lung cancer A549 cells by negatively regulating miRNA-337-3p.

Anti-Lung Cancer Targets of Radix Paeoniae Rubra and Biological Molecular Mechanism: Network Pharmacological Analyses and Experimental Validation.

ObjectiveTo systematically explore the pharmacological mechanism of Radix Paeoniae Rubra (RPR) against lung cancer (LC).MethodsA network pharmacology approach, which involves active ingredients and target forecast, network construction, gene ontology and pathway enrichment, was employed in this research. In addition, the effect of Baicalein (BAI) in RPR on A549 cells was researched in vitro and in vivo.ResultsA total of 159 targets of the 29 active components in RPR were procured by pharmacokinetic parameters. The network analysis showed that β-sitosterol, baicalein, (+)-catechin, ellagic acid, stigmasterol, (2R, 3R)-4-methoxyl-distylin were the main ingredients and JUN, VEGFA, BCL2 were the hub targets of RPR in the treatment of LC. The functional enrichment analysis showed that RPR likely was useful to LC by regulating numerous pathways including Pathways in cancer, MAPK signaling pathway and so on. MTT results showed that 100μM, 200μM, 400μM of BAI had a time and dose-dependent inhibitory effect on A549 cells proliferation; Wound healing and transwell assays showed that 100μM, 200μM, 400μM of BAI could significantly restrain the migration and invasion of A549 cells; Flow cytometry assay results showed that 100μM, 200μM, 400μM of BAI could induce apoptosis of A549 cells. In vivo, BAI (50, 100 mg/kg) significantly inhibited tumor growth and promoted apoptosis of tumor cells compared with the control group.ConclusionBAI in RPR may exert anti-tumor effects by inhibiting the proliferation, migration and invasion of LC cells, and inducing the apoptosis of LC cells.

Low expression of BTN3A3 indicates poor prognosis and promotes cell proliferation, migration and invasion in non-small cell lung cancer.

BackgroundThe butyrophilin (BTN) family has many members with diverse functions related to immunomodulation, initiation and progression of tumors. BTN3A3 belongs to the BTN family, and exploring its expression and correlation with the prognosis of non-small cell lung cancer (NSCLC) patients has great clinical significance.MethodsClinical specimens were used to detect BTN3A3 expression. Small interfering RNA (siRNA) was used to knock down BTN3A3 and analyze the proliferative, migratory and invading ability of the transfected NSCLC cells. Multiplex immunofluorescence staining was used to detect the expression of BTN3A3 protein in the tumor microenvironment (TME). We analyzed the relationship between the expression of BTN3A3 and the clinicopathological features and prognosis of NSCLC patients.ResultsThe expression of BTN3A3 in NSCLC tissues was significantly lower than in adjacent tissues, and patients with low expression of BTN3A3 had late clinical stages and lower degree of tumor differentiation. Knocking down BTN3A3 promoted the proliferation, migration and invasion of NSCLC. In the TME, the density of BTN3A3+ tumor cells positively correlated with the density of CD8+ T cells, and the patients with low expression of BTN3A3 had poor overall survival (OS).ConclusionsChanges in the BTN3A3 expression level may play a potential key role in the carcinogenesis and development of NSCLC. Patients with low expression of BTN3A3 showed a more aggressive and invasive phenotype and a lower level of CD8+ T-cell infiltration, which may be an important factor affecting the OS of NSCLC patients.

The Downregulation of Long Noncoding RNA Metastasis Associated with Colon Cancer 1 (lncRNA MACC1-AS1) Inhibits the Migration and Invasion of Non-Small Cell Lung Cancer Cells

To explore the effects of lncRNA MACC1-AS1 on the invasion and migration of non-small cell lung cancer (NSCLC), PCR was used to detect and compare lncRNA MACC1-AS1 expression in NSCLC cells to normal lung epithelial cells and assess the knockdown effect of lncRNA MACC1-AS1 in A549 and H1299 cell lines. The Transwell system was employed to compare the invasiveness of the lncRNA MACC1-AS1 knockdown cells to the control cells. The migratory ability of the lncRNA MACC1-AS1 knockdown cells was compared to the control cells via in vitro scratch assay. Western blot was used to detect the E-cadherin/N-cadherin expression. Balb/c mouse model was used to verify the metastasis ability in vivo between the control and lncRNA MACC1-AS1 knockdown groups. The level of lncRNA MACC1-AS1 was markedly higher in lung cancer cells than normal lung epithelial cells. The knockdown of lncRNA MACC1-AS1 reduced the invasiveness and migratory capacity of A549 and H1299 cells. Downregulating lncRNA MACC1-AS1 upregulated E-cadherin but down-regulated N-cadherin in A549 and H1299 cells. Besides, the silencing of lncRNA MACC1-AS1 reduced the A549 cells’ ability to metastasize in vivo. LncRNA MACC1-AS1 can enhance the invasiveness and migratory capability of NSCLC.

Red Raspberry Extracts Inhibit A549 Lung Cancer Cell Migration, Invasion, and Epithelial-Mesenchymal Transition Through the Epidermal Growth Factor Receptor/Signal Transducer and Activator of Transcription-3 Signaling Pathway

The incidence and mortality of lung cancer ranks first among all malignant tumors in the world. Because it is relatively asymptomatic at early stages, most patients do not become aware of the disease until it has progressed to an advanced stage. Advanced lung cancer metastasis results in systemic cachexia and effective treatment becomes challenging, leading to poor response and outcome. Therefore, the development of new drugs for the treatment of lung cancer is paramount. In this study, A549 cells were treated with different concentrations of red raspberry extract and the proliferation, migration, and invasion of cells were evaluated. The results indicated that red raspberry extract reduced the proliferation, migration, and invasion of A549 cells. Western blot analysis was used to detect the expression of the cyclin D1, N-cadherin, vimentin, E-cadherin, EGFR, and STAT3 proteins. Treatment with red raspberry extract reduced the expression of cyclin D1, N-cadherin, vimentin, EGFR, and STAT3, whereas the expression of E-cadherin increased. Following transfection of an EGFR overexpression vector into A549 cells, we observed a reduced inhibitory effect of the red raspberry extract on the proliferation, migration, and invasion of A549 cells. In addition, EGFR overexpression abrogated the increased expression of cyclin D1, N-cadherin, vimentin, EGFR, and STAT3 protein expression in A549 cells following extract treatment. In contrast, E-cadherin protein expression was decreased under these treatment conditions. Overall, this study suggests that red raspberry extract may reduce the proliferation, migration, invasion, and epithelialmesenchymal transition of A549 lung cancer cells by inhibiting the activation of the EGFR/STAT3 signaling pathway. These findings may lead to the development of new strategies to treat advanced lung cancer.

The Monoacylglycerol Lipase Inhibitor JZL184 Inhibits Lung Cancer Cell Invasion and Metastasis via the CB1 Cannabinoid Receptor.

A targeted modulation of the endocannabinoid system is currently discussed as a promising strategy for cancer treatment. An important enzyme for the endocannabinoid metabolism is the monoacylglycerol lipase (MAGL), which catalyzes the degradation of 2-arachidonoylglycerol (2-AG) to glycerol and free fatty acids. In this study, we investigated the influence of MAGL inhibition on lung cancer cell invasion and metastasis. Using LC-MS, significantly increased 2-AG levels were detected in A549 cells treated with the MAGL inhibitor JZL184. In athymic nude mice, JZL184 suppressed metastasis of A549 cells in a dose-dependent manner, whereby the antimetastatic effect was cancelled by the CB1 receptor antagonist AM-251. In vitro, JZL184 induced a time- and concentration-dependent reduction of A549 cell invasion through Matrigel-coated membranes, which was likewise reversed by AM-251. An MAGL inhibition-associated reduction of free fatty acids as a cause of the anti-invasive effect could be excluded by add-back experiments with palmitic acid. Both JZL184 and the MAGL substrate 2-AG led to an increased formation of the tissue inhibitor of metalloproteinase-1 (TIMP-1), whereby a TIMP-1 knockdown using siRNA significantly attenuated the anti-invasive effects of both substances. Decreased invasion and TIMP-1 upregulation was also caused by the MAGL inhibitors JW651 and MJN110 or transfection with MAGL siRNA. A CB1- and TIMP-1-dependent anti-invasive effect was further confirmed for JZL184 in H358 lung cancer cells. In conclusion, MAGL inhibition led to a CB1-dependent decrease in human lung cancer cell invasion and metastasis via inhibition of 2-AG degradation, with TIMP-1 identified as a mediator of the anti-invasive effect.

A novel circular RNA, hsa_circ_0030998 suppresses lung cancer tumorigenesis and Taxol resistance by sponging miR-558.

Circular RNAs (circRNAs) are single-stranded RNAs which form a covalently closed continuous loop. Although originally shown to be non-protein-coding, some circRNAs can give rise to micropeptides. circRNAs have also been shown to play essential regulatory roles in a variety of developmental and disease processes. In a previous study, hsa_circ_0030998 was identified as a circRNA downregulated in lung cancer, but its potential implications and mechanisms in lung cancer were not addressed. Here, we showed that overexpressing circ_0030998 decreased proliferation, migration, and invasion of lung cancer cells, while also dampening resistance to Taxol, a classical antitumor drug. Depleting circ_0030998 reversed these phenotypic effects. A high circ_0030998 expression was correlated with a high survival rate in lung cancer patients. Additionally, we found circ_0030998 could downregulate miR-558 expression, serving as a microRNA sponge. In conclusion, our data support that hsa_circ_0030998 can slow down the progression of lung cancer by targeting miR-558 and suppress malignant phenotypes such as proliferation, migration, and invasion progression of lung cancer cells. Therefore, we highlight that circ_0030998 could be a novel tumor suppressor of lung cancer.

LncRNA LINC01116 sponges miR-93-5p to promote cell invasion and migration in small cell lung cancer

BackgroundLINC01116 is a recently identified oncogenic lncRNA in glioma. Differential expression analysis using the public gene expression analysis tool GEPIA revealed the upregulation of LINC01116 in lung cancer. We studied the functions of LINC01116 in small cell lung cancer (SCLC).MethodsThe expression of LINC01116 in several types of cancer tissue and the paired non-tumor tissues was evaluated by GEPIA. The effects of the overexpression of LINC01116 and miR-93-5p on the expression of STAT3 were evaluated. The effects of the overexpression of LINC01116, miR-93-5p and STAT3 on SHP-77 cell behaviors were evaluated by Transwell assays.ResultsLINC01116 was highly expressed in SCLC and predicted poor survival. In SCLC tissues, the expression of LINC01116 was positively correlated with STAT3. Bioinformatics analysis revealed that miR-93-5p may target LINC01116. Overexpression of LINC01116 increased STAT3 but did not affect the expression of miR-93-5p. Transwell assay showed that LINC01116 and STAT3 increased cell invasion and migration rates. MiR-93-5p played an suppressed cell behaviors and suppressed the role of LINC01116.ConclusionTherefore, LINC01116 might upregulate STA3 by sponging miR-93-5p, thereby promoting cell invasion and migration in SCLC.

Impact Analysis of miR-1253 on Lung Cancer Progression Through Targeted Regulation of ANXA3.

ObjectiveThis study set out to investigate the effect of miR-1253 on lung cancer progression through targeted regulation of ANXA3.MethodsRT-PCR was employed to detect the miR-1253 expression levels in lung cancer cells and its targeted gene ANXA3 mRNA determined by biological information prediction. MTT, invasion and apoptosis rate tests were employed to detect the proliferation, invasion and apoptosis rate of lung cancer cells over-expressing miR-1253 or those with low expression of ANXA3 and the expression of related proteins.ResultsRT-qPCR results manifested that the miR-1253 level was down-regulated in lung cancer tissues and cells, and the ANXA3 expression increased. The miR-1253 and ANXA3 expression levels were negatively correlated. miR-1253 was correlated with tumor differentiation degree, TNM stage and lymph node metastasis of lung cancer patients. Cell tests confirmed that miR-1253 played a tumor-inhibiting function, including inhibiting proliferation and invasion of lung cancer cells and promoting apoptosis. Bioinformatics prediction and subsequent experiments proved that ANXA3 was the direct target of miR-1253. Moreover, after the ANXA3 expression in lung cancer cells was knocked down, proliferation and invasion of those cells were inhibited dramatically, the apoptosis rate increased markedly, and the expression levels of pro-apoptosis-related proteins Bax and caspase-3 were up-regulated, and the anti-apoptosis-related protein Bcl-2 expression was down-regulated.ConclusionmiR-1253 can inhibit the proliferation and invasion of lung cancer cells and promote their apoptosis by targeting ANXA3. It can be used as a new potential target for lung cancer treatment.

CircTMEM30A/hsa-miR-130a-3p regulates TNFα and promotes the malignant progression of COPD with primary lung cancer.

Chronic obstructive pulmonary disease (COPD) and lung cancer is the most common respiratory diseases among smokers. Lung cancer is one of the most common malignancy. COPD patients are at high risk of lung cancer, but the specific pathogenesis is still unclear. In recent years, circRNAs have become the focus of lung cancer research. This study investigated the effect of CircTMEM30A on COPD and lung cancer and its possible molecular mechanisms. QPCR was used to detect the expression of CircTMEM30A in COPD patients, lung cancer patients, and COPD patients with lung cancer. CircTMEM30A was transfected with PPF, HFL1, A549 and NCI-H446 cells, and the transfection efficiency was verified by real-time quantitative PCR. The effects of CircTMEM30A on the proliferation, migration and invasion of fibroblasts and lung cancer cells were detected by MTT, Transwell and scratch tests. The target gene of CircTMEM30A was verified by luciferase reporter assay. Real-time quantitative PCR was used to detect the expression level of CircTMEM30A target gene. CircTMEM30A was the most highly expressed in COPD patients with lung cancer. Overexpression of CircTMEM30A enhanced the effects of proliferation, migration and invasion of fibroblasts and lung cancer cells, down-regulated the expression of E-cadherin, up-regulated the expression of Vimentin, and promoted EMT. After knocking down CircTMEM30A, the migration and invasion ability of lung cancer cells was significantly reduced. Bioinformatics retrieval and luciferase reporter gene confirmed that CircTMEM30A could be bind to miR-130a, while TNFα gene was the target gene of miR-130a. Further studies showed that overexpression of miR-130a and knockdown of TNFα reversed the promoting effect of CircTMEM30A on lung cancer cells. CircTMEM30A was highly expressed in COPD patients with lung cancer, and miR-130a was low expressed in lung cancer cells. CircTMEM30A regulated the expression of TNFα through miR-130a, thereby affecting the progression of COPD and lung cancer. CircTMEM30A may be a therapeutic target for COPD patients with lung cancer.

Circular RNA circ_103820 suppresses lung cancer tumorigenesis by sponging miR-200b-3p to release LATS2 and SOCS6

A growing number of circular RNAs (circRNAs) have been identified and verified in several cancers. However, highly efficient therapeutic methods based on circRNAs in lung cancer remain largely unexplored. In the present study, we identified a novel circular RNA, hsa_circ_103820, based on Gene Expression Omnibus (GEO) data. Functionally, overexpression of hsa_circ_103820 showed significant inhibitory effects on the proliferation, migration and invasion of lung cancer cells, and knockdown of hsa_circ_103820 played promoting roles. Regarding the mechanism, we revealed that miR-200b-3p was a direct target of hsa_circ_103820 and that LATS2 and SOCS6 were the downstream target genes of miR-200b-3p. Therefore, we identified a novel potential tumor suppressive function of hsa_circ_103820 in lung cancer.

Long Non-Coding RNA CAR10 Facilitates Non-Small Cell Lung Cancer Cell Migration and Invasion by Modulating the miR-892a/GJB2 Pathway.

IntroductionNon-coding RNAs, including long non-coding (lnc)RNAs and microRNAs (miRs), play crucial roles in numerous malignant tumors, including non-small cell lung cancer (NSCLC).MethodsThe expression levels of chromatin-associated RNA Intergenic 10 (CAR10), gap junction protein beta 2 (GJB2) and miR-892a in NSCLC were evaluated by reanalyzing three Gene Expression Omnibus (GEO) datasets, and performing reverse transcription-quantitative PCR, immunohistochemistry staining and Western blot analysis, accordingly. Functionally, Transwell and Matrigel assays were performed to measure changes in the migration and invasion abilities of the A549 and H1299 cell lines. The targeted binding effects between CAR10 and miR-892a, as well as between miR-892a and GJB2 were confirmed by conducting dual-luciferase reporter and RNA pull-down assays, respectively.ResultsThe present study demonstrated that CAR10 was upregulated in patients with NSCLC, which was also associated with a poor prognosis. Functionally, CAR10 was confirmed to be oncogenic and promoted NSCLC cell migration and invasion, using overexpression and knockdown Transwell assays. Furthermore, GJB2 expression was revealed to be upregulated and was positively correlated with CAR10 expression in NSCLC. A further mechanistic study revealed that GJB2 was a downstream target of CAR10, which induced the migration and invasive potential of the A549 and H1299 cell lines. More specifically, miR-892a was found to serve as a bridge between CAR10 and GJB2, via similar miRNA response elements. The RNA pull-down and luciferase assays indicated that miR-892a directly binds both CAR10 and GJB2.ConclusionCAR10 promoted NSCLC cell migration and invasion by upregulating GJB2 and sponging miR-892a. These findings illustrated that the CAR10/miR-892a/GJB2 axis may be a novel molecular target for the treatment of NSCLC.

Circular RNA FLNA acts as a sponge of miR-486-3p in promoting lung cancer progression via regulating XRCC1 and CYP1A1

Significantly high-expressed circFLNA has been found in various cancer cell lines, but not in lung cancer. Therefore, this study aimed to explore the role of circFLNA in the progression of lung cancer. The target gene of circFLNA was determined by bioinformatics and luciferase reporter assay. Viability, proliferation, migration, and invasion of the transfected cells were detected by CCK-8, colony formation, wound-healing, and transwell assays, respectively. A mouse subcutaneous xenotransplanted tumor model was established, and the expressions of circFLNA, miR-486-3p, XRCC1, CYP1A1, and related genes in the cancer cells and tissues were detected by RT-qPCR, Western blot, or immunohistochemistry. The current study found that miR-486-3p was low-expressed in lung cancer. MiR-486-3p, which has been found to target XRCC1 and CYP1A1, was regulated by circFLNA. CircFLNA was located in the cytoplasm and had a high expression in lung cancer cells. Cancer cell viability, proliferation, migration, and invasion were promoted by overexpressed circFLNA, XRCC1, and CYP1A1 but inhibited by miR-486-3p mimic and circFLNA knockdown. The weight of the xenotransplanted tumor was increased by circFLNA overexpression yet reduced by miR-486-3p mimic. Furthermore, miR-486-3p mimic reversed the effect of circFLNA overexpression on promoting lung cancer cells and tumors and regulating the expressions of miR-486-3p, XRCC1, CYP1A1, and metastasis/apoptosis/proliferation-related factors. However, overexpressed XRCC1 and CYP1A1 reversed the inhibitory effect of miR-486-3p mimic on cancer cells and tumors. In conclusion, circFLNA acted as a sponge of miR-486-3p to promote the proliferation, migration, and invasion of lung cancer cells in vitro and in vivo by regulating XRCC1 and CYP1A1.

MiR-758-3p inhibits the proliferation and invasion of non-small cell lung cancer cells by targeting NUSAP1

Objective: To investigate the biological function of miR-758-3p and the underlying mechanism in non-small cell lung cancer (NSCLC). Methods: miR-758-3p mimics was transfected to A549 NSCLC cells, miRNA control was used as a negative control, cells transfected with nucleolar and spindle associated protein 1 (NUSAP1)-overexpression vector indicated as NUSAP1 group, cells co-transfected with miR-758-3p mimics and NUSAP1-overexpression vector indicated as miR-758-3p mimics+ NUSAP1 group. The effects of miR-758-3p on the proliferation, migration and invasion abilities of A549 cells were detected by cell counting kit-8 (CCK-8) and Transwell assays, respectively. Bioinformatics and dual luciferase reporter assay were used to predict and verify the target gene of miR-758-3p. Results: The expressions of miR-758-3p and NUSAP1 in A549 cells were significantly up-regulated at 24 hours after transfection with miR-758-3p mimics (P<0.05). Compared with the miRNA control group (1.15±0.06), the OD value of miR-758-3p mimic group (0.78±0.06) was significantly decreased at 72 hours after transfection (P<0.05). The number of migrated cells of miR-758-3p mimic group (119.04±11.49) was significantly lower than that of the control group (271.38±19.05) (P<0.05). The number of invaded cells of miR-758-3p mimic group (71.33±5.36) was significantly lower than the control group (164.30±8.11) (P<0.05). The result of dual-luciferase reporter assay showed that NUSAP1 was a direct target of miR-758-3p. Moreover, up-regulation of NUSAP1 abolished the inhibitory effects of miR-758-3p on the proliferation, migration and invasion abilities of A549 cells. Conclusion: miR-758-3p inhibits the proliferation, migration and invasion of NSCLC cells by targeting NUSAP1.

Rhophilin-2 Upregulates Glutamine Synthetase by Stabilizing c-Myc Protein and Confers Resistance to Glutamine Deprivation in Lung Cancer.

IntroductionRHPN2, a member of rhophilin family of rho-binding proteins, regulates actin cytoskeleton and vesicular trafficking, and promotes mesenchymal transformation in cancer. We have found that RHPN2 was significantly mutated in lung adenocarcinoma (LUAD). However, the role of RHPN2 in lung cancer is not fully understood.MethodsIn the present study, we investigated the expression of RHPN2 in 125 patients with LUAD by qRT-PCR and correlated its expression with clinical characteristics. The effects of RHPN2 on the proliferation and invasion of lung cancer cells were determined by CCK-8 and in vitro transwell assays, clonogenic assay, and xenograft mouse model. The RhoA pull down assay and Western blotting were performed to elucidate the mechanism of RNPN2 in tumorigenesis of lung cancer.ResultsRHPN2 was overexpressed in tumors from LUAD, and high levels of RHPN2 were associated with poor prognosis of LUAD patients. RHPN2 was required for proliferation and invasion of lung cancer cells. Intriguingly, overexpression of RHPN2 conferred the resistance to glutamine depletion in lung cancer cells. Mechanistic studies revealed that ectopic overexpression of RHPN2 promoted the stability of c-Myc protein via phosphorylation at Ser62 and increased c-Myc target glutamine synthetase (GS). Analysis of GS expression in clinical sample showed that the expression of GS was elevated in tumor cells. Kaplan-Meier analysis revealed that high levels of GS were significantly associated with worse overall survival time of the patients with LUAD.ConclusionsTaken together, this study suggested that RHPN2 was involved in tumorigenesis of lung cancer via modulating c-Myc stability and the expression of its target GS in lung adenocarcinoma, which links RHPN2 and glutamine metabolism.