GINS2 promotes the progression of human HNSCC by altering RRM2 expression.

INTRODUCTION: GINS2 exerts a carcinogenic effect in multiple human malignancies, while it is still unclear that the potential roles and underlying mechanisms of GINS2 in HNSCC. METHODS: TCGA database was used to screen out genes with significant differences in expression in HNSCC. Immunohistochemistry and qRT-PCR were used to measure the expression of GINS2 in HNSCC tissues and cells. GINS2 was detected by qRT-PCR or western blot after knockdown or overexpression. Celigo cell counting, MTT, colony formation, and flow cytometric assay were used to check the ability of proliferation and apoptosis. Bioinformatics and microarray were used to screen out the downstream genes of GINS2. RESULTS: GINS2 in HNSCC tissues and cells was up-regulated, which was correlated with poor prognosis. GINS2 gene expression was successfully inhibited and overexpressed in HNSCC cells. Knockdown of GINS2 could inhibit proliferation and increase apoptosis of cells. Meanwhile, overexpression of GINS2 could enhance cell proliferation and colony formation. Knockdown of RRM2 may inhibit HNSCC cell proliferation, while overexpression of RRM2 rescued the effect of reducing GINS2 expression. CONCLUSION: Our study reported the role of GINS2 in HNSCC for the first time. The results demonstrated that in HNSCC cells, GINS2 promoted proliferation and inhibited apoptosis via altering RRM2 expression. Therefore, GINS2 might play a carcinogen in HNSCC, and become a specific promising therapeutic target.

Therapeutic potential of the novel Bcl-2/Bcl-XL dual inhibitor, APG1252, alone or in combination against non-small cell lung cancer.

Targeting the induction of apoptosis is a promising cancer therapeutic strategy with some clinical success. This study focused on evaluating the therapeutic efficacy of the novel Bcl-2/Bcl-XL dual inhibitor, APG1252-M1 (also named APG-1244; an in vivo active metabolite of APG1252 or pelcitoclax), as a single agent or in combination, against non-small cell lung cancer (NSCLC) cells. APG1252-M1 effectively decreased the survival of some NSCLC cell lines expressing low levels of Mcl-1 and induced apoptosis. Overexpression of ectopic Mcl-1 in the sensitive cells substantially compromised APG1252-M1's cell-killing effects, whereas inhibition of Mcl-1 greatly sensitized insensitive cell lines to APG1252-M1, indicating the critical role of Mcl-1 levels in impacting cell response to APG1252-M1. Moreover, APG1252-M1, when combined with the third generation epidermal growth factor receptor (EGFR) inhibitor, osimertinib, synergistically decreased the survival of EGFR-mutant NSCLC cell lines including those resistant to osimertinib with enhanced induction of apoptosis and abrogated emergence of acquired resistance to osimertinib. Importantly, the combination was effective in inhibiting the growth of osimertinib-resistant tumors in vivo. Collectively, these results demonstrate the efficacy of APG1252 alone or in combination against human NSCLC cells.

The lncRNA ZNF667-AS1 Inhibits Propagation, Invasion, and Angiogenesis of Gastric Cancer by Silencing the Expression of N-Cadherin and VEGFA.

Purpose: Gastric cancer is one of the most common malignancies with high mortality worldwide. It is known that long noncoding RNAs (lncRNAs) play important roles in the pathogenesis of gastric cancer. This study investigates the role of lncRNA ZNF667-AS1 in gastric cancer cells. Methods: We have applied real-time quantitative PCR (qPCR) to study the levels of ZNF667-AS1 in gastric cancer biopsies and cell lines. The effects of ZNF667-AS1 on the propagation, clonogenicity, metastasis, and angiogenesis of gastric cancer cells were evaluated by calorimetry, colony formation, cell migration, and angiogenesis assays. Western blotting was used to identify the levels of proteins involved in cancer invasion and angiogenesis signaling pathways. Result: It was found that lncRNA ZNF667-AS1 was downregulated in gastric cancer biopsies. Overexpression of ZNF667-AS1 reduced the propagation, migration, and angiogenesis of gastric cancer cells. Molecular mechanism studies displayed that the high level of lncRNA ZNF667-AS1 promoted the expression of E-cadherin and inhibited the expression of N-cadherin and VEGFA, leading to the inhibition of the proliferation, migration, and angiogenesis of gastric cancer cells. Conclusion: As a tumor suppressor gene, lncRNA ZNF667-AS1 significantly hinders the propagation, metastasis, and angiogenesis of gastric cancer cells by promoting the expression of E-cadherin and inhibiting the expression of N-cadherin and VEGFA. Therefore, lncRNA ZNF667-AS1 could play a synergistic therapeutic role by targeting tumor cells and vascular endothelial cells, which represents a new therapeutic scheme for novel therapeutics of gastric cancer.

The novel BET degrader, QCA570, is highly active against the growth of human NSCLC cells and synergizes with osimertinib in suppressing osimertinib-resistant EGFR-mutant NSCLC cells.

Lung cancer remains the leading cause of cancer deaths worldwide despite advances in knowledge in cancer biology and options of various targeted therapies. Efforts in identifying innovative and effective therapies are still highly appreciated. Targeting bromodomain and extra terminal (BET) proteins that function as epigenetic readers and master transcription coactivators is now a potential cancer therapeutic strategy. The current study evaluates the therapeutic efficacies of the novel BET degrader, QCA570, in lung cancer and explores its underlying mechanisms. QCA570 at low nanomolar ranges effectively decreased the survival of a panel of human lung cancer cell lines with induction of apoptosis in vitro. As expected, it potently induced degradation of BET proteins including BRD4, BRD3 and BRD2. Moreover, it potently decreased Mcl-1 levels due to transcriptional suppression and protein degradation; this event is critical for mediating apoptosis induced by QCA570. Moreover, QCA570 synergized with osimertinib in suppressing the growth of osimertinib-resistant cells in vitro and in vivo, suggesting potential in overcoming acquired resistance to osimertinib. These preclinical findings support the potential of QCA570 in treatment of lung cancer either as a single agent or in combination with others.

Overcoming acquired resistance to third-generation EGFR inhibitors by targeting activation of intrinsic apoptotic pathway through Mcl-1 inhibition, Bax activation, or both.

Treatment of EGFR-mutant non-small cell lung cancer (NSCLC) with mutation-selective third-generation EGFR-tyrosine kinase inhibitors (EGFR-TKIs) such as osimertinib has achieved remarkable success in the clinic. However, the immediate challenge is the emergence of acquired resistance, limiting the long-term remission of patients. This study suggests a novel strategy to overcome acquired resistance to osimertinib and other third-generation EGFR-TKIs through directly targeting the intrinsic apoptotic pathway. We found that osimertinib, when combined with Mcl-1 inhibition or Bax activation, synergistically decreased the survival of different osimertinib-resistant cell lines, enhanced the induction of intrinsic apoptosis, and inhibited the growth of osimertinib-resistant tumor in vivo. Interestingly, the triple-combination of osimertinib with Mcl-1 inhibition and Bax activation exhibited the most potent activity in decreasing the survival and inducing apoptosis of osimertinib-resistant cells and in suppressing the growth of osimertinib-resistant tumors. These effects were associated with increased activation of the intrinsic apoptotic pathway evidenced by augmented mitochondrial cytochrome C and Smac release. Hence, this study convincingly demonstrates a novel strategy for overcoming acquired resistance to osimertinib and other 3rd generation EGFR-TKIs by targeting activation of the intrinsic apoptotic pathway through Mcl-1 inhibition, Bax activation or both, warranting further clinical validation of this strategy.

LncRNA EPB41L4A-AS2 represses Nasopharyngeal Carcinoma Metastasis by binding to YBX1 in the Nucleus and Sponging MiR-107 in the Cytoplasm.

Nasopharyngeal carcinoma (NPC) is known for its potential to progress to the lymph nodes and distant metastases at an early stage. As an important regulator in tumorigenesis biological processes, the functions of lncRNA in NPC tumor development remain largely unclear. In this research, the expression of EPB41L4A-AS2 in NPC tissues and cells was analyzed via real-time quantitative polymerase chain reaction (qRT-PCR). CCK8, colony formation, and EDU experiments were used to determine the viability of NPC cells. Transwell and wound healing assays were performed to test NPC cell migration and invasion. RNA pull-down and mass spectrometry analysis were used to identify potential binding proteins. Then, a popliteal lymph node metastasis model was established to test NPC metastasis. EPB41L4A-AS2 is repressed by transforming growth factor-beta, which is downregulated in NPC cells and tissue. It is associated with the presence of distant metastasis and adverse outcomes. The univariate and multivariate survival assays confirmed that EPB41L4A-AS2 expression was an independent predictor of progression-free survival (PFS) in patients with NPC. Biological analyses showed that overexpression of EPB41L4A-AS2 reduced the metastasis and invasion of NPC in vitro and in vivo, but had no significant effect on cell proliferation. Mechanistically, in the nucleus we identified that EPB41L4A-AS2 relies on binding to YBX1 to reduce the stability of Snail mRNA to enhance the expression of E-cadherin and reverse the progression of epithelial-to-mesenchymal transition (EMT). In the cytoplasm, we found that EPB41L4A-AS2 blocked the invasion and migration of NPC cells by promoting LATS2 expression via sponging miR-107. In a whole, the findings of this study help to further understand the metastasis mechanism of NPC and could help in the prevention and treatment of NPC metastasis.

KIF18A knockdown reduces proliferation, migration, invasion and enhances radiosensitivity of esophageal cancer.

Kinesin family member 18A (KIF18A) is significantly overexpressed and is related to the poor prognosis of human cancers. However, the function of KIF18A in esophageal cancer (EC) is still unclear. Human EC cell lines were used in this study. KIF18A expression in human tissues was assessed using Gene Expression Profiling Interactive Analysis 2.0 (GEPIA2). The expressions of KIF18A or IGF2BP3 in EC cells were detected using qRT-PCR or WB. Cells were transfected using si-KIF18A, si-IGF2BP3, and plasmid IGF2BP3. The abilities of proliferation, migration, and invasion were detected by EdU, wound-healing, and transwell assay. The interaction between KIF18A and IGF2BP3 was predicted by starBase v3.0 and studied by RIP and RNA stability assay. Colony formation assay was used to reflect the changes of radiosensitivity in EC cells. KIF18A was upregulated in EC, and KIF18A knockdown inhibited EC cell proliferation, migration, invasion, and radioresistance. The prediction in starBase and RIP assay results showed that KIF18A mRNA could bind to IGF2BP3 protein in EC cells. RNA stability assay was performed to confirm that IGF2BP3 affects mRNA stability of KIF18A. Further studies also showed that IGF2BP3 could positively regulate KIF18A on proliferation, migration, invasion, and radioresistance. Our findings first revealed an oncogenic effect of KIF18A in human EC progression. KIF18A expression was associated with radioresistance of EC cells. The binding relationship between KIF18A and IGF2BP3 might influence the mRNA stability of KIF18A in EC cell lines.

Inhibition of ACK1 delays and overcomes acquired resistance of EGFR mutant NSCLC cells to the third generation EGFR inhibitor, osimertinib.

OBJECTIVES: The emergence of acquired resistance to the third generation EGFR inhibitor, osimertinib (AZD9291 or TAGRISSO™), is an unavoidable huge clinical challenge. The involvement of ACK1, a non-receptor tyrosine kinase with an oncogenic function, in regulating cell response to osimertinib has not been investigated and thus is the focus of this study. MATERIAL AND METHODS: Drug effects on cell growth were evaluated by measuring cell numbers and colony formation. Apoptosis was monitored with flow cytometry for annexin V-positive cells and Western blotting for protein cleavage. Intracellular protein and mRNA alterations were detected with Western blotting and qRT-PCR, respectively. Drug effects on delaying osimertinib acquired resistance were determined using colony formation in vitro and xenografts in nude mice in vivo, respectively. Cell senescence was assayed by ß-galactosidase staining. RESULTS: Inhibition of ACK1 with the novel ACK1 inhibitor, (R)-9b synergized with osimertinib in inhibiting the growth of EGFR mutant NSCLC cell lines. Similar results were also generated with ACK1 gene knockdown. The combination of osimertinib and (R)-9b enhanced induction of apoptosis. In both in vitro and in vivo long-term resistance delay assays, the combination of (R)-9b and osimertinib clearly delayed the emergence of osimertinib-resistance. Further, the (R)-9b and osimertinib combination was also effective in inhibiting the growth of EGFR mutant NSCLC cell lines with acquired resistance to osimertinib, which possess elevated levels of ACK1, and the growth of osimertinib-resistant tumors in vivo. In some resistant cell lines, the combinations induced senescence in addition to induction of apoptosis. CONCLUSIONS: These novel findings suggest that ACK1 inhibition might be a potential and innovative strategy for delaying and overcoming osimertinb acquired resistance.

The Utility of Diffusion and Perfusion Magnetic Resonance Imaging in Target Delineation of High-Grade Gliomas.

BACKGROUND: The tumor volume of high-grade glioma (HGG) after surgery is usually determined by contrast-enhanced MRI (CE-MRI), but the clinical target volume remains controversial. Functional magnetic resonance imaging (multimodality MRI) techniques such as magnetic resonance perfusion-weighted imaging (PWI) and diffusion-tensor imaging (DTI) can make up for CE-MRI. This study explored the survival outcomes and failure patterns of patients with HGG by comparing the combination of multimodality MRI and CE-MRI imaging with CE-MRI alone. METHODS: 102 patients with postoperative HGG between 2012 and 2016 were included. 50 were delineated based on multimodality MRI (PWI, DTI) and CE-MRI (enhanced T1), and the other 52 were delineated based on CE-MRI as control. RESULTS: The median survival benefit was 6 months. The 2-year overall survival, progression-free survival, and local-regional control rates were 48% vs. 25%, 42% vs. 13.46%, and 40% vs. 13.46% for the multimodality MRI and CE-MRI cohorts, respectively. The two cohorts had similar rates of disease progression and recurrence but different proportions of failure patterns. The univariate analysis shows that characteristics of patients such as combined with epilepsy, the dose of radiotherapy, the selection of MRI were significant influence factors for 2-year overall survival. However, in multivariate analyses, only the selection of MRI was an independent significant predictor of overall survival. CONCLUSIONS: This study was the first to explore the clinical value of multimodality MRI in the delineation of radiotherapy target volume for HGG. The conclusions of the study have positive reference significance to the combination of multimodality MRI and CE-MRI in guiding the delineation of the radiotherapy target area for HGG patients.

MiR-154-5p Suppresses Cell Invasion and Migration Through Inhibiting KIF14 in Nasopharyngeal Carcinoma.

BACKGROUND: Mounting evidence has reported that microRNA-154-5p (miR-154-5p) is involved in the development of multiple cancers, but its function in nasopharyngeal carcinoma (NPC) remains not well investigated. METHODS: Real-time quantitative PCR (qRT-PCR) was used to detect miR-154-5p expression in NPC tissues and cells. CCK8, colony formation, wound healing and transwell assays were performed to assess cell proliferation, migration and invasion. Dual-luciferase reporter assays and Western blots were performed to confirm the target gene of miR-154-5p. Rescue experiments were conducted to explore the influence of target gene KIF14 on the functions of miR-154-5p. Xenograft tumor model was conducted to detect the effect of miR-154-5p in vivo. RESULTS: qRT-PCR results revealed that the expression of miR-154-5p was down-regulated in NPC tissues and cell lines compared to normal nasopharyngeal tissues and cell line. Overexpression of miR-154-5p inhibited cell migration and invasion. However, miR-154-5p had no influence on the proliferation of NPC cells. MiR-154-5p overexpression suppressed xenograft tumor metastasis in vivo. Dual-luciferase reporter analysis identified KIF14 as a target gene of miR-154-5p. Rescue experiments showed that knockdown of KIF14 reversed the effect of inhibiting miR-154-5p expression on NPC cell migration and invasion. CONCLUSION: Taken together, miR-154-5p suppresses tumor migration and invasion by targeting KIF14 in NPC. The newly identified miR-154-5p/KIF14 interaction offers further insights into the progression of NPC, which may represent a novel target for NPC diagnosis and treatment.

Feedback loop in miR-449b-3p/ADAM17/NF-κB promotes metastasis in nasopharyngeal carcinoma.

An emerging body of evidence has promoted the understanding of the role of microRNAs (miRNAs) in tumorigenesis and progression, but the mediating function of miRNAs in nasopharyngeal carcinoma (NPC) development remains poorly elucidated. In this study, miR-449b-3p was downregulated in NPC specimens (P < .001) and cells (P < .05). Cytological and animal experiments provided evidence that miR-449b-3p inhibited NPC metastasis in vitro and in vivo. Disintegrin and metalloproteinase 17 (ADAM17) was revealed as a direct target of miR-449b-3p. Rescue experiments suggested that the downregulation of ADAM17 in the miR-449b-3p knockdown cells partially reversed the inhibition of cell invasion and migration. Luciferase reporter assay, chromatin immunoprecipitation assay, and Western blot analysis showed that ADAM17 could suppress the promoter activity and expression of miR-449b-3p by inducing NF-κB transcriptional activity. In conclusion, our study provided new insights into the underlying mechanism of the invasion and metastasis of NPC. The novel miR-449b-3p/ADAM17/NF-κB feedback loop could be a target for the clinical treatment of NPC.

Long non-coding RNA LINC01133 mediates nasopharyngeal carcinoma tumorigenesis by binding to YBX1.

Recently, long non-coding RNAs (lncRNAs) have been reported as the vital regulators of various cancers including nasopharyngeal carcinoma (NPC). An increasing number of studies have suggested that lncRNA LINC01133 is dysregulated and involved in human carcinogenesis. However, the roles of LINC01133 in NPC remain largely unknown. In this work, we demonstrated that LINC01133 was significantly downregulated in NPC tissues and cell lines. Loss and gain of function experiments provided evidence that LINC01133 inhibited NPC cell proliferation, invasion and migration both in vitro and in vivo. Besides, Fluorescence in situ hybridization (FISH) assay was performed to determine the localization of LINC01133 and LINC01133 was observed mainly distributed in the nucleus. Importantly, RNA pull-down and RIP assays showed that LINC01133 directly combined with YBX1, and YBX1 can partly reverse the repression of NPC cell proliferation, migration, and invasion caused by LINC01133. Collectively, our exploration indicate that LINC01133 inhibits the malignant-biological behavior of NPC cells by binding to YBX1, thereby suggesting a novel biomarker for the NPC prognosis and treatment.

Identification of key pathways and genes in nasopharyngeal carcinoma using bioinformatics analysis.

Nasopharyngeal carcinoma (NPC) is one of the most common malignancies in the head and neck. The aim of the current study was to identify the key pathways and genes involved in NPC through bioinformatics analysis and to identify potential molecular mechanisms underlying NPC proliferation and progression. Three gene expression profiles (GSE12452, GSE34573 and GSE64634) were downloaded from the Gene Expression Omnibus database. A total of 76 samples were analyzed, of which 59 were NPC samples and 17 were normal samples. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were subsequently conducted. The protein-protein interaction (PPI) network of the differentially expressed genes (DEGs) was constructed using Cytoscape software. Analysis of GSE12452, GSE34573 and GSE64634 datasets identified 1,301 (553 upregulated and 748 downregulated), 1,232 (348 upregulated and 884 downregulated) and 1,218 (555 upregulated and 663 downregulated) DEGs, respectively. Using Venn diagram analysis, 268 DEGs (59 upregulated and 209 downregulated) that intersected all three datasets, were selected for further analysis. The results of GO analysis revealed that upregulated DEGs were significantly enriched in biological processes, including 'cell adhesion', 'cell division', 'mitosis' and 'mitotic cell cycle'. The downregulated DEGs were mainly enriched in 'microtubule-based movement', 'cilium movement', 'cilium axoneme assembly' and 'epithelial cell differentiation'. The KEGG pathway analysis results revealed that the upregulated DEGs were highly associated with several pathways, including 'extracellular matrix-receptor interaction', 'human papillomavirus infection', 'arrhythmogenic right ventricular cardiomyopathy' and 'focal adhesion', whereas the downregulated DEGs were enriched in 'metabolic pathways', 'Huntington's disease', 'fluid shear stress and atherosclerosis' and 'chemical carcinogenesis'. On the basis of the PPI network of the DEGs, the following top 10 hub genes were identified: Dynein axonemal light intermediate chain 1, dynein axonemal intermediate chain 2, calmodulin 1, coiled-coil domain containing 114, dynein axonemal heavy chain 5, radial spoke head 9 homolog, radial spoke head component 4A, NDC80 kinetochore complex component, thymidylate synthetase and coiled-coil domain containing 39. In conclusion, by performing a comprehensive bioinformatics analysis of DEGs, putative targets that could be used to elucidate the molecular mechanisms underlying NPC were identified.

miR-130a-3p regulated TGF-ß1-induced epithelial-mesenchymal transition depends on SMAD4 in EC-1 cells.

Metastasis and invasion are the primary causes of malignant progression in esophageal squamous cell carcinoma (ESCC). Epithelial-mesenchymal transition (EMT) is crucial step of acquisition of "stemness" properties in tumor cells. However, the mechanism of esophageal cancer metastasis remains unclear. This research was designed to explore the role and mechanism of SMAD4 and miR-130a-3p in the progression of transforming growth factor-ß (TGF-ß)-induced EMT in vivo and in vitro. The expression of miR-130a-3p in ESCC cell line and normal esophageal epithelial cell was determined by RT-qPCR. The protein expression levels of TGF-ß-induced changes in EMT were analyzed by western blotting and immunofluorescence. Dual-luciferase report assays were used to validate the regulation of miR-130a-3p-SMAD4 axis. The effect of miR-130a-3p and SMAD4 in TGF-ß-induced migration, invasion in the ESCC cell line EC-1 was investigated by wound healing assays and Transwell assays. Here we found that knocked down SMAD4 could partially reverse TGF-ß-induced migration, invasion, and EMT progression in the ESCC cell line EC-1. miR-130a-3p, which directly targets SMAD4, is down-regulated in ESCC. miR-130a-3p inhibits the migration and invasion of EC-1 cells both in vitro and in vivo. Finally, miR-130a-3p inhibits TGF-ß-induced EC-1 cell migration, invasion, and EMT progression in a SMAD4-dependent way. In conclusion, this study provides new insights into the mechanism underlying ESCC metastasis. The TGF-ß/miR-130a-3p/SMAD4 pathway could be potential targets for clinical treatment of ESCC.

miR-184 Inhibits Tumor Invasion, Migration and Metastasis in Nasopharyngeal Carcinoma by Targeting Notch2.

BACKGROUND/AIMS: A recent study found that dysregulated microRNA-184 (miR-184) is involved in the proliferation and survival of nasopharyngeal carcinoma (NPC). This study aimed to evaluate the detailed mechanisms of invasion, migration and metastasis of NPC cells. METHODS: Quantitative reverse-transcription PCR (qRT-PCR) and Western blot were used to confirm the expression levels of miR-184 and Notch2. NPC cell invasion and migration were subsequently examined using in vitro cell invasion and wound-healing assays, respectively. MicroRNA (miRNA) target gene prediction databases and dual-luciferase reporter assay were adopted to validate the target genes of miR-184. RESULTS: MiR-184 was downregulated in the NPC cell lines. The miR-184 inhibitor increased the number of invading NPC cells, whereas miR-184 mimics inhibited the invasive ability of such cells. The protein level of E-cadherin decreased, whereas those of N-cadherin and vimentin increased in the anti-miR-184 group. This result showed that miR-184 inhibited NPC cell invasion and metastasis by regulating EMT progression. MiRNA target gene prediction databases indicated the potential of Notch2 as a direct target gene of miR-184. Such a notion was then validated by results of dual-luciferase reporter assay. Notably, shRNANotch2 restrained the EMT and partially abrogated the inhibitory effects of miR-184 on EMT progression in NPC cells. CONCLUSION: MiR-184 functions as a tumour-suppressive miRNA targeting Notch2 and inhibits the invasion, migration and metastasis of NPC.

MiRNA-34a reversed TGF-ß-induced epithelial-mesenchymal transition via suppression of SMAD4 in NPC cells.

Epithelial-mesenchymal transition (EMT) is considered a prerequisite for tumor invasion and metastasis in many cancers. However, the mechanisms underlying EMT in nasopharyngeal carcinoma (NPC) is largely unknown. In this study, we found that transforming growth factor-ß (TGF-ß), which reportedly promotes EMT in multiple cancers, can trigger EMT and increase the invasive and migratory capacities of NPC cells. Conversely, the downregulation of SMAD4, a vital member of the canonical TGF-ß pathway, reversed the TGF-ß-induced EMT, invasion, and migration. Further experiments revealed that SMAD4 was the target of miRNA-34a, which was downregulated in NPC tissues and suppressed NPC cell metastasis in vivo. miRNA-34a overexpression also antagonized the TGF-ß-induced EMT progression, invasion, and migration through SMAD4 inhibition. However, the restoration of SMAD4 expression rescued the inhibitory effects of miRNA-34a on tumorigenesis. All these results confirmed that miRNA-34a suppressed the TGF-ß-induced EMT, invasion, and migration of NPC cells by directly targeting SMAD4, which indicated the potential of miR-34a as a therapeutic target against NPC.

MIIP gene expression is associated with radiosensitivity in human nasopharyngeal carcinoma cells.

The present study aims to investigate the radiosensitization effect of the migration and invasion inhibitory protein (MIIP) gene on nasopharyngeal carcinoma (NPC) cells. The MIIP gene was transfected into NPC 5-8F and CNE2 cells. The level of MIIP was analyzed by quantitative reverse transcription-polymerase chain reaction analysis and western blot. The changes in radiosensitivity of the cells were analyzed by colony formation assay. The changes in cell apoptosis and cycle distribution following irradiation were detected by flow cytometry. The expression of BCL2 associated X, apoptosis regulator/B-cell lymphoma 2 was evaluated using western blot. DNA damage was analyzed by counting γ-H2AX foci. The expression levels of γ-H2AX were evaluated by immunofluorescence and western blot. In a previous study by the authors, the results indicated that the expression of MIIP gene evidently increased in MIIP-transfected 5-8F (5-8F OE) and MIIP-transfected CNE2 (CNE2 OE) cells compared with the parental or negative control cells. In the present study, the survival rate of 5-8F OE and CNE2 OE cells markedly decreased following irradiation (0, 2, 4, 6 and 8 Gy) compared with the negative control (5-8F NC and CNE2 NC) and the untreated (5-8F and CNE2) groups. The expression of MIIP was able to increase apoptosis, which resulted in G2/M cell cycle arrest and DNA damage repair was attenuated in 5-8F and CNE2 cells following irradiation as measured by the accumulation of γ-H2AX. It was indicated that MIIP expression is associated with the radiosensitivity of NPC cells and has a significant role in regulating cell radiosensitivity.

TGF-? regulates the ERK/MAPK pathway independent of the SMAD pathway by repressing miRNA-124 to increase MALAT1 expression in nasopharyngeal carcinoma.

Transforming growth factor beta (TGF-?), a pleiotropic cytokine, promotes cell proliferation and migration in multiple cancers, including nasopharyngeal carcinoma (NPC). microRNA-124 (miR-124) becomes downregulated in NPC and inhibits the tumorigenesis of this disease. However, the role of miR-124 in TGF-?-induced NPC development remains unknown. In this study, constant TGF-? stimulation repressed miR-124 expression, whereas miR-124 overexpression antagonized TGF-?-promoted NPC cell growth and migration. miR-124 overexpression decreased p-SMAD2/3, SMAD4, and p-ERK levels, indicating that ectopic miR-124 overexpression inhibited SMAD and non-SMAD pathways. Pro-oncogenic lncRNA MALAT1 was targeted by miR-124 that regulated ERK/MAPK by targeting MALAT1 independent of the SMAD signaling pathway. In conclusion, our work clarified the significant role of miR-124 in TGF-? signaling pathways independent of the SMAD signaling pathway and showed the potential of miR-124 as a new therapeutic target against NPC.

MiR-152 functioning as a tumor suppressor that interacts with DNMT1 in nasopharyngeal carcinoma.

BACKGROUND: In recent years, miR-152 has been dysregulated in a variety of tumors and used as a tumor suppressor. Nevertheless, its role in nasopharyngeal carcinoma (NPC) remains unidentified. MATERIALS AND METHODS: Real-time quantitative PCR (polymerase chain reaction) was performed to analyze the expression of miR-152 in NPC cell lines. MiR-152 expression profiles in NPC tissues were obtained from Gene Expression Omnibus (GEO GSE36682). The effect of miR-152 on the invasion and proliferation of NPC cells was determined through cell invasion, wound healing, and cell viability assays. Apoptosis was examined by flow cytometry, and Western blot was performed to measure expression of the target gene. Pyrosequencing was used to detect the methylation level of NPC cells. RESULTS: In this study, miR-152 was downregulated in the NPC tissues and cell lines. When miR-152 was enhanced, the invasion and migration of NPC cells were inhibited. However, miR-152 had no effect on the proliferation of NPC cells. Luciferase reporter gene analysis was performed, and the results showed that DNMT1 (DNA (cytosine-5)-methyltransferase 1) is a direct target of miR-152 in NPC cells. DNMT1 downregulation and miR-152 overexpression both reversed the effects of miR-152 inhibition on the NPC cells. In addition, miR-152 expression increased as a result of the inhibition of the methylation level of miR-152 when DNMT1 expression was downregulated. CONCLUSION: The overexpression of miR-152 inhibited the migration and invasion of NPC cells by targeting DNMT1. Furthermore, DNMT1 regulated miR-152 expression through DNA methylation. Overall, the novel miR-152-DNMT1 regulatory circuit may provide better understanding of the pathogenesis of NPC and new epigenetic therapeutic target in NPC.

Long non-coding RNA n326322 promotes the proliferation and invasion in nasopharyngeal carcinoma.

Long non-coding RNAs (lncRNAs) have been reported to perform significant roles in cancer development and progression. Our research has found that a novel lncRNA n326322 was higher in nasopharyngeal carcinoma (NPC) cells. Moreover, the gain and loss of functional approaches revealed that the overexpression of lncRNA-n326322 promoted NPC cell proliferation and invasion, whereas the downregulation of lncRNA-n326322 suppressed cell proliferation and invasion. Further experiments demonstrated that potential mechanism may be associated with the activation of PI3K/AKT and ERK/MAPK pathways. Taken together, these results indicate that lncRNA-n326322 is associated with tumorigenesis of NPC.