Malignant Progression Of Glioma Research Articles

Knockdown of LncRNA SCAMP1 suppressed malignant biological behaviours of glioma cells via modulating miR-499a-5p/LMX1A/NLRC5 pathway.

Dysregulation of long non‐coding RNAs (lncRNAs) confirm that it plays a crucial role in tumourigenesis and malignant progression of glioma. The present study demonstrated that LncRNA secretory carrier membrane protein 1 (SCAMP1) was up‐regulated and functioned as an oncogene in glioma cells. In addition, miR‐499a‐5p was down‐regulated meanwhile exerted tumour‐suppressive function in glioma cells. Subsequently, inhibition of SCAMP1 significantly restrained the cell proliferation, migration and invasion, as well as promoted apoptosis by acting as a molecular sponge of miR‐499a‐5p. Transcription factor LIM homeobox transcription factor 1, alpha (LMX1A) was overexpressed in glioma tissues and cells. Moreover, miR‐499a‐5p targeted LMX1A 3′‐UTR in a sequence‐specific manner. Hence, down‐regulation of SCAMP1 remarkably reduced the expression level of LMX1A, indicating that LMX1A participated in miR‐499a‐5p‐induced tumour‐suppressive effects on glioma cells. Furthermore, knockdown of LMX1A decreased NLR family, CARD domain containing 5 (NLRC5) mRNA and protein expression levels through directly binding to the NLRC5 promoter region. Down‐regulation of NLRC5 obviously inhibited malignant biological behaviours of glioma cells through attenuating the activity of Wnt/β‐catenin signalling pathway. In conclusion, our study clarifies that SCAMP1/miR‐499a‐5p/LMX1A/NLRC5 axis plays a critical role in modulating malignant progression of glioma cells, which provide a novel therapeutic strategy for glioma treatment.

Long Noncoding RNA linc00665 Stabilized by TAF15 Impeded the Malignant Biological Behaviors of Glioma Cells via STAU1 Mediated mRNA Degradation

BACKGROUND: Glioma is a poor prognostic brain cancer with strong invasiveness and limited treatment options. Moreover, lncRNAs dysregulation has emerged as important component in tumorigenesis, and participated in diverse biomolecular regulatory networks of glioma therapies. METHODS: RNA FISH assays and lncRNA microarrays were used to determine the expression of linc00665. RNA-IP, RNA pull-down, ChIP and luciferase reporter gene assays were used to testify the association among TAF15/linc00665/MTF1(YY2)/GTSE1 axis in glioma. RESULTS: In this study, we demonstrated that TAF15 and linc00665 were identified as tumor-suppressors in glioma. Moreover, TAF15 overexpression enhanced the stability of linc00665 and inhibited malignant progression of glioma cells. Both MTF1 and YY2 showed high expression level, while knockdown of either MTF1 or YY2 inhibited malignant progression of glioma. Mechanistically, overexpression of linc00665 was cofirmed to destabilize MTF1 and YY2 mRNAs respectively by interacting with STAU1. Moreover, knockdown of MTF1 or YY2 decreased the expression levels of GTSE1 through direct binding to GTSE1 promoter region. CONCLUSIONS: Our study aimed to clarify important roles of TAF15/linc00665/MTF1(YY2)/GTSE1 axis in modulating malignant progression of glioma cells, and might propose novel mechanisms in which lncRNA functions via affecting STAU1-mediated mRNA stability in glioma, thus contribute to molecular therapies for glioma. FUNDING STATEMENT: This work is supported by grants from the Nature Science Foundation of China (81573010, 81672511 and 81502179), Liaoning Science and Technology Plan Project (No. 2017225020, 2015225007), Project of Key Laboratory of Neuro-oncology in Liaoning Province (112-2400017005), special developmental project guided by central government of Liaoning Province (No. 2017011553-301). DECLARATION OF INTERESTS: The authors declare no competing interests. ETHICS APPROVAL STATEMENT: Normal brain tissues and four grade of human brain tissues obtained from patients who have signed the Informed consent hospitalized in the Department of Neurosurgery of Shengjing Hospital of China Medical University, moreover our study was approved by the Ethics Committee of Shengjing Hospital of China Medical University. All assays on nude mice were operated strictly according to the protocol approved by the Administrative Panel on Laboratory Animal Care of the Shengjing Hospital.

CD48 is a key molecule of immunomodulation affecting prognosis in glioma.

Purpose: Glioma is a refractory disease associated with immune cell infiltration, and the effectiveness of checkpoint blockade remains suboptimal. As an adhesion and costimulatory molecule, CD48 plays a significant role in immunomodulation. As such, studying CD48 may provide additional understanding of the immune and inflammation response of glioma.Methods: Using R language and GraphPad Prism 7, RNA sequencing data of 946 patients from Chinese Glioma Genome Atlas and The Cancer Genome Atlas cohorts were analyzed.Results: CD48 was highly expressed in the malignant progression of glioma. As an independent risk factor, high-CD48 patients were associated with poor prognosis. CD48 influenced glioma purity and the local immune cell subpopulation. CD48 was closely related to immune function in glioma. Patients with an enhanced immune phenotype, high CD48, were associated with immune suppressive molecules and checkpoints. In addition, CD48 correlated with the immune and inflammatory response. A checkpoint risk score including CD48, SLAMF8 and PD-L1 was used to assess the role of checkpoints. Risk score was particularly high in a malignant subtype of glioma and was an independent predictive indicator of unfavorable outcome. Additionally, age, IDH subtype and MGMT promoter status influenced the predictive significance of checkpoint risk score.Conclusion: CD48 exhibits a crucial role in reduced survival and immunomodulation in glioma. In addition, we found that checkpoints play a greater role in patients older than 40 years old with IDH wild-type and MGMT methylated status. These findings suggest that combining CD48 blockade with PD-L1 may be a promising approach to glioma immunotherapy for specific subpopulations of patients.

MicroRNA-374b inhibits migration and invasion of glioma cells by targeting EGFR.

To investigate the expression level of microRNA-374b in glioma tissues and its influence on the invasive ability of glioma cells. Meanwhile, the regulatory mechanism of microRNA-374b in glioma was also explored. The expression level of microRNA-374b in 32 glioma tissues and para-cancerous tissues were detected by quantitative Real-time polymerase chain reaction (qRT-PCR). The relationship between microRNA-374b expression and clinical indicators of glioma was analyzed. Meanwhile, the expression of microRNA-374b in glioma cells was verified by qRT-PCR as well. Subsequently, microRNA-374b over-expression model was constructed in glioma cell lines, including U251 and U87. Next, the effect of microRNA-374b on cellular biological functions was analyzed using cell counting kit-8 (CCK-8) assay, Wound healing test and transwell invasion assay, respectively. Finally, the relationship between miRNA and epidermal growth factor receptor (EGFR) was explored. QRT-PCR results showed that the expression level of microRNA-374b in glioma was significantly lower than that of adjacent tissues, and the difference was statistically significant. Compared with patients with higher expression of microRNA-374b, the occurrence rate of lymph node or distant metastasis was significantly higher in those with lower microRNA-374b expression. In addition, compared with NC group, the proliferation, invasion and migration abilities of cells in microRNA-374b mimics group was significantly decreased. Subsequently, results demonstrated that the expression of EGFR was significantly increased in glioma cells and tissues, which was negatively correlated with microRNA-374b expression. Subsequent cell recovery experiment indicated that microRNA-374b and EGFR had a mutual regulation and could affect the malignant progression of glioma all together. MicroRNA-374b could inhibit the invasion and migration of glioma by regulating EGFR. Moreover, the expression of microRNA-374b was significantly associated with lymph node metastasis, distant metastasis and poor prognosis.

Circular RNA circ_001350 regulates glioma cell proliferation, apoptosis, and metastatic properties by acting as a miRNA sponge.

Glioma is an aggressive malignancy with increasing incidence and threatens people's health worldwide. Accumulating evidence revealed that circular RNAs (circRNAs) play important functions in cancers. A previous study demonstrated that circ_001350 was elevated in glioma tissue samples than nontumorous tissue specimens screened by high-throughput microarray. The level of circ_001350 in glioma tissue specimens and cell lines was detected by quantitative real-time polymerase chain reaction. The Fisher exact test was carried out to estimate the correlation of circ_001350 level with clinical characteristics. Cell proliferation, apoptosis, and motility abilities were detected using cell counting kit-8, clonogenic, flow cytometry, and transwell experiments, respectively. The potential target of circ_001350 was identified by the luciferase assay. circ_001350 level was significantly enhanced in glioma tissue specimens and cells. Further, elevated expression of circ_001350 was closely linked to patients' clinical severity. Knockdown of circ_001350 could inhibit cell proliferation and metastatic properties and increase apoptotic cells. circ_001350 could directly bind to miR-1236 and regulate its expression to exert oncogenic functions. Collectively, circ_001350 directly sponges miR-1236, thus contributing to malignant progression of glioma.

Increased expression of NAF1 contributes to malignant phenotypes of glioma cells through promoting protein synthesis and associates with poor patient survival

The H/ACA ribonucleoprotein (RNP) complex noncore subunit NAF1 is an indispensable factor during H/ACA RNP maturation, and one of the widely known functions of H/ACA RNP is modulating ribosome biosynthesis. However, the specific biological role and exact mechanism of NAF1 in human cancers including glioma remain largely unclear. In this study, we found that NAF1 was highly expressed in gliomas relative to normal brain tissues, and demonstrated that increased expression of NAF1 was strongly correlated with poor patient survival. Further studies revealed that NAF1 was transcriptionally regulated by c-Myc, NRF2, and telomerase reverse transcriptase (TERT), which are the key molecules associated with malignant progression of gliomas. Moreover, we demonstrated that NAF1 was a functional oncogene in glioma cells through promoting cell growth in vitro and in vivo, survival, migration, and invasion. Mechanistically, NAF1 acted as a rate-limiting controller of cell growth and invasiveness through enhancing 40S subunit assembly and protein synthesis including c-Myc, NRF2, TERT, POLR1A, and POLR2A. These molecules in turn enhanced the transcription and translation of NAF1, thereby forming positive feedback loops between them to promote malignant phenotypes of glioma cells. In addition, our data also showed that NAF1 depletion could trigger ribosome stress, not only impairing ribosomal biosynthesis but also reactivating p53 signaling via blocking MDM2. Taken together, we demonstrated that NAF1 promotes the tumorigenesis and progression of glioma through modulating ribosome assembly and protein synthesis, and predicted that NAF1 may be a potential therapeutic target and valuable prognostic biomarker in gliomas.

Tumor growth under rhGM-CSF application in an orthotopic rodent glioma model.

Regulation of the host immune response serves a pivotal role in the persistence and progression of malignant glioma. To date, cytotoxic cluster of differentiation (CD)-8+ T and natural killer cells are considered the main cellular components of host tumor control. The influence of macrophages in an orthotropic C6 tumor implantation model was investigated and the aim of the present study was to characterize the effects of systemic macrophage-activation on glioma growth by using the granulocyte macrophage colony stimulating factor (rhGM-CSF). A total of 20 male Sprague-Dawley rats were orthotopically implanted with C6 glioma spheroids and treated subcutaneously with 10 µg/kg rhGM-CSF every other day; 9 animals served as controls. Serial magnetic resonance imaging was performed on days 7, 14, 21, 28, 32 and 42 post-implantation to monitor tumor volume. Histological work-up included hematoxylin and eosin, CD68/ED-1 macrophage, CD8 T-cell and Ki-67 MIB1 proliferation staining in gliomas and spleen. Experimental C6-gliomas developed in 15/20 (75%) animals. In rhGM-CSF treated rats, tumors developed significantly later and reached a smaller size (median, 134 mm3) compared with the controls (median, 262 mm3). On day 14, solid tumors presented in 11/17 (65%) rhGM-CSF-treated animals; in control animals tumor growth was detected in 3/9 animals on day 7 and in all animals on day 14. The mean survival time was 35 days in the rhGM-CSF group and significantly longer when compared with the control group (24 days). Immunohistochemistry exhibited significantly more macrophages in tumors, particularly in the perivascular zone of the rhGM-CSF group when compared with untreated animals; intratumoral CD8+ counts were equal in both groups. A systemic stimulation of macrophages by rhGM-CSF resulted in significantly reduced and delayed tumor growth in the rodent C6 glioma model. The present data suggested a significant role of macrophages in host control of experimental gliomas on the innate immune response. Until now, the role of macrophages may have been underestimated in host glioma control.

M6A RNA methylation regulators contribute to malignant progression and have clinical prognostic impact in gliomas.

N6-methyladenosine (m6A) RNA methylation, associated with cancer initiation and progression, is dynamically regulated by the m6A RNA methylation regulators (“writers”, “erasers” and “readers”). Here, we demonstrate that most of the thirteen main m6A RNA methylation regulators are differentially expressed among gliomas stratified by different clinicopathological features in 904 gliomas. We identified two subgroups of gliomas (RM1/2) by applying consensus clustering to m6A RNA methylation regulators. Compared with the RM1 subgroup, the RM2 subgroup correlates with a poorer prognosis, higher WHO grade, and lower frequency of IDH mutation. Moreover, the hallmarks of epithelial-mesenchymal transition and TNFα signaling via NF-κB are also significantly enriched in the RM2 subgroup. This finding indicates that m6A RNA methylation regulators are closely associated with glioma malignancy. Based on this finding, we derived a risk signature, using seven m6A RNA methylation regulators, that is not only an independent prognostic marker but can also predict the clinicopathological features of gliomas. Moreover, m6A regulators are associated with the mesenchymal subtype and TMZ sensitivity in GBM. In conclusion, m6A RNA methylation regulators are crucial participants in the malignant progression of gliomas and are potentially useful for prognostic stratification and treatment strategy development.

LATS2 Inhibits Malignant Behaviors of Glioma Cells via Inactivating YAP.

We previously reported that LATS2, the upstream serine/threonine kinase of Yes-associated protein (YAP), is downregulated in gliomas and exhibits negative correlation with the prognosis of glioma patients. In this work, we aimed to explore the role and mechanism of large tumor suppressor kinase (LATS2) in the progression of malignant gliomas. We found that over-expression of LATS2 inhibited glioma cell proliferation and migration/invasion, while LATS2 downregulation promoted them. Mechanistically, LATS2 promoted the phosphorylation of YAP without affecting YAP mRNA expression. Inconsistent with some previous reports, both YAPWT and YAP5SA did not affect the level of LATS2, suggesting that LATS2 did not form a feedback loop with YAP in gliomas. Furthermore, mammalian Ste20-like kinases (MST1) over-expression did not affect the phosphorylation of YAP, suggesting that MST1 may not be the essential upstream serine/threonine kinase of Hippo/YAP pathway in gliomas. Together, LATS2 inhibits glioma cell proliferation and migration/invasion by inactivating YAP.

DNA demethylation is associated with malignant progression of lower-grade gliomas

To elucidate the mechanisms of malignant progression of lower-grade glioma, molecular profiling using methylation array, whole-exome sequencing, and RNA sequencing was performed for 122, 36 and 31 gliomas, respectively. This cohort included 24 matched pairs of initial lower-grade gliomas and recurrent tumors, most of which showed malignant progression. Nearly half of IDH-mutant glioblastomas that had progressed from lower-grade gliomas exhibited characteristic partial DNA demethylation in previously methylated genomic regions of their corresponding initial tumors, which had the glioma CpG island methylator phenotype (G-CIMP). In these glioblastomas, cell cycle-related genes, RB and PI3K-AKT pathway genes were frequently altered. Notably, late-replicating domain was significantly enriched in the demethylated regions that were mostly located in non-regulatory regions, suggesting that the loss of DNA methylation during malignant transformation may involve mainly passive demethylation due to a delay in maintenance of methylation during accelerated cell division. Nonetheless, a limited number of genes including IGF2BP3, which potentially drives cell proliferation, were presumed to be upregulated due to demethylation of their promoter. Our data indicated that demethylation of the G-CIMP profile found in a subset of recurrent gliomas reflects accelerated cell divisions accompanied by malignant transformation. Oncogenic genes activated by such epigenetic change represent potential therapeutic targets.

ZNF326 promotes malignant phenotype of glioma by up-regulating HDAC7 expression and activating Wnt pathway

BackgroundZinc-finger protein-326 (ZNF326) was initially found in the NIH3T3 cell line to regulate cell growth, however, the expression and underlying role of ZNF326 in human tumours, especially in glioma, is not fully understood.MethodsImmunohistochemistry was applied to detect the expression of ZNF326 in glioma tissues, and statistical analysis was used to analyse the relationship between ZNF326 expression and clinicopathological factors. The effect of ZNF326 on glioma cells proliferation and invasion was conducted by functional experiments both in vivo and in vitro. Chromatin immunoprecipitation and dual-luciferase assays were performed to demonstrate that histone deacetylase enzyme-7 (HDAC7) is the target gene of ZNF326. Immunoblotting, real-time PCR, GST-pulldown and co-immunoprecipitation assays were used to clarify the underlying role of ZNF326 on Wnt pathway activation.ResultsHigh nuclear expression of ZNF326 was observed in glioma cell lines and tissues, and closely related with advanced tumour grade in the patients. Moreover, ectopic ZNF326 expression promoted the proliferation and invasiveness of glioma cells. Mechanistically, ZNF326 could activate HDAC7 transcription by binding to a specific promoter region via its transcriptional activation domain and zinc-finger structures. The interaction of the up-regulated HDAC7 with β-catenin led to a decrease in β-catenin acetylation level at Lys-49, followed by a decrease in β-catenin phosphorylation level at Ser-45. These changes in β-catenin posttranscriptional modification levels promoted its redistribution and import into the nucleus. Additionally, ZNF326 directly associated with β-catenin in the nucleus, and enhanced the binding of β-catenin to TCF-4, serving as a co-activator in stimulating Wnt pathway.ConclusionsOur findings elucidated ZNF326 promotes the malignant phenotype of human glioma via ZNF326-HDAC7-β-catenin signalling. This study reveals the vital role and mechanism of ZNF326 in the malignant progression of glioma, and provides the reference for finding biomarkers and therapeutic targets for glioma.

DHHC protein family targets different subsets of glioma stem cells in specific niches

BackgroundGlioblastomas (GBM) comprise different subsets that exhibit marked heterogeneity and plasticity, leading to a lack of success of genomic profiling in guiding the development of precision medicine approaches against these tumors. Accordingly, there is an urgent need to investigate the regulatory mechanisms for different GBM subsets and identify novel biomarkers and therapeutic targets relevant in the context of GBM-specific niches. The DHHC family of proteins is associated tightly with the malignant development and progression of gliomas. However, the role of these proteins in the plasticity of GBM subsets remains unclear.MethodsThis study utilized human glioma proneural or mesenchymal stem cells as indicated. The effects of DHHC proteins on different GBM subsets were investigated through in vitro and in vivo assays (i.e., colony formation assay, flow cytometry assay, double immunofluorescence, western blot, and xenograft model). Western blot, co-immunoprecipitation, and liquid chromatograph mass spectrometer-mass spectrometry assays were used to detect the protein complexes of ZDHHC18 and ZDHHC23 in various GBM subtypes, and explore the mechanism of DHHC proteins in targeting different subsets of GSCs in specific niches.ResultsZDHHC18 and ZDHHC23 could target the glioma stem cells of different GBM subsets in the context of their specific niches and regulate the cellular plasticity of these subtypes. Moreover, mechanistic investigations revealed that ZDHHC18 and ZDHHC23 competitively interact with a BMI1 E3 ligase, RNF144A, to regulate the polyubiquitination and accumulation of BMI1. These events contributed to the transition of glioma stem cells in GBM and cell survival under the stressful tumor microenvironment.ConclusionsOur work highlights the role of DHHC proteins in the plasticity of GBM subsets and reveals that BMI1 represents a potential therapeutic target for human gliomas.

ACYP2 Contributes to Malignant Progression of Glioma Through Promoting Ca <sup>2+</sup> Efflux and Subsequently Activating c-Myc and STAT3 Signals

Background: Acylphosphatase 2 (ACYP2) is involved in cell differentiation, energy metabolism and hydrolysis of intracellular ion pump. It has been reported as a negative regulator in leukemia and a positive regulator in colon cancer, respectively. However, its biological role in glioma remains unclear. The aim of this study was to clarify the biological functions and related molecular mechanism of ACYP2 in glioma tumorigenesis. Methods: We performed quantitative RT-PCR (qRT-PCR), immunohistochemistry (IHC) and western blot assays to evaluate expression of ACYP2. The functions of ACYP2 in glioma cells were determined by a series of in vitro and in vivo experiments, including cell proliferation, colony formation, cell cycle, apoptosis, migration, invasion and nude mouse tumorigenicity assays. Moreover, western blot and co-immunoprecipitation (co-IP) were used to identify its downstream targets. Findings: Knocking down ACYP2 in glioma cells significantly inhibited cell proliferation, colony formation, migration, invasion and tumorigenic potential in nude mice, and induced cell cycle arrest and apoptosis. Conversely, ectopic expression of ACYP2 in glioma cells dramatically promoted malignant phenotypes of glioma cells. Mechanistically, ACYP2 promoted malignant progression of glioma cells through regulating intracellular Ca2+ homeostasis via its interaction with PMCA4, thereby activating c-Myc and PTP1B/STAT3 signals. This could be effectively reversed by Ca2+ chelator BAPTA-AM or calpain inhibitor calpeptin. Interpretation: Our data demonstrate that ACYP2 functions as an oncogene in glioma through activating c-Myc and STAT3 signals via the regulation of intracellular Ca2+ homeostasis, and indicate that ACYP2 may be a potential therapeutic target and prognostic biomarker in gliomas. Funding Statement: This work was supported by the National Natural Science Foundation of China (No. 81572627 and 81672645), Innovation Talent Promotion Plan in Shaanxi Province (No. 2018TD-006) and The Natural Science Foundation Research Project in Shaanxi Province (No. 2019JQ-230). Declaration of Interests: The authors declare that they have no competing interest. Ethics Approval Statement: All animal procedures were conducted under the approval of the Animal Care and Use Committee of Xi’an Jiaotong University. All patients signed an informed consent before the surgery. The study protocol was approved by the Institutional Review Board and Human Ethics Committee of the First Affiliated Hospital of Xi’an Jiaotong University.

ALDH1A3 induces mesenchymal differentiation and serves as a predictor for survival in glioblastoma

As aldehyde dehydrogenase (ALDH) is a novel stem cell marker, increasing studies have confirmed that high ALDH activity promotes tumorigenesis and progression in cancers. Some preliminary studies have found that ALDH1A3 may play an important role in glioma malignant progression, but so far there was no conclusive conclusion. The purpose of our study was to elucidate the mechanisms by which ALDH1A3 regulated in glioma and to provide practical tools for clinical application. Aldefluor, flow cytometry sorting and qRT-PCR were performed to verify the role of ALDH1A3 in ALDH activity maintenance. Transwell, immunofluorescence, glycolytic assays, and orthotopic xenograft models were used to explore ALDH1A3 bio-functions in GBM. LASSO-COX, COX survival analysis and Kaplan–Meier analysis were used to establish the prognostic evaluation system and predict postoperative chemotherapy sensitivity of GBMs. Our integrated study found that (1) ALDH1A3 associates with mesenchymal differentiation of GBM in Eastern and Western world patients. (2) ALDH1A3 plays a critical role in ALDH activity maintenance. (3) ALDH1A3 is an activator of mesenchymal transformation in GBM. (4) ALDH1A3-derived PMT markers’ molecular signature can predict 1-, 2-, and 3-year survival rates of GBMs precisely. In conclusion, ALDH1A3 was a major contributor to ALDH activity and a key driver in triggering mesenchymal transformation in GBM. ALDH1A3-based molecular classification scheme can help to improve guidance for prognosis forecasting and individualized treatment decision making for GBM patients.

MiRNA-139–3p inhibits the proliferation, invasion, and migration of human glioma cells by targeting MDA-9/syntenin

Gliomas are the most common primary malignant brain tumor in adults. Although these tumors are aggressive and frequently lethal, there are currently few therapeutic approaches available to prolong patient survival. MicroRNAs play important roles in regulating the expression of genes that control diverse cellular processes. Here, we investigated the expression and function of miR-139–3p in gliomas using clinical specimens, cultured cells, and a mouse xenograft tumor model. We found that miR-139–3p expression is markedly lower in human glioma tissues than in normal brain tissues. We identified melanoma differentiation-associated gene-9 (MDA-9)/syntenin, an adaptor protein implicated in tumor metastasis, as a novel direct target of miR-139–3p and showed that syntenin mRNA and miR-139–3p levels were inversely correlated in clinical specimens (r = −0.6817, P = 0.0002). Overexpression of miR-139–3p in human glioma cell lines inhibited cell proliferation, migration, and invasion, and these effects were rescued by co-transfection with syntenin. Our results indicate that miR-139–3p plays a significant role in controlling behaviors associated with the malignant progression of gliomas, and we identify the miR-139-3p–syntenin axis as a potential therapeutic target for glioma.

Knockdown of NEAT1 repressed the malignant progression of glioma through sponging miR-107 and inhibiting CDK14.

Aberrant expressions of long noncoding RNAs (lncRNAs) contribute to carcinogenesis via regulating tumor suppressors or oncogenes. LncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) has been recognized as an oncogene to promote tumor progression of many cancers. However, the function of NEAT1 in glioma remains poorly discovered. Currently, we focused on the role of NEAT1 in glioma. Here, we found that NEAT1 was greatly upregulated in glioma cells compared with normal human astrocytes (NHAs). Meanwhile, miR-107 was significantly downregulated in glioma cell lines. Then, we observed that knockdown of NEAT1 suppressed the growth and invasion of glioma cells including U251 and SW1783 cells. Reversely, overexpression of NEAT1 dramatically induced glioma cell survival, increased cell colony formation, and promoted cell invasion ability. Subsequently, bioinformatics analysis was performed to predict the correlation between NEAT1 and miR-107. Moreover, it was revealed that NEAT1 could modulate miR-107 via serving as an endogenous sponge of miR-107. The direct binding correlation between NEAT1 and miR-107 was validated in our study. In addition, cyclin dependent kinase 14 (CDK14) was predicted as an messenger RNA target of miR-107 and the association between them was confirmed in our research. Moreover, we implied that NEAT1 demonstrated its biological functions via regulating miR-107 and CDK14 in vivo. In summary, our findings indicated that NEAT1/miR-107/CDK14 axis participated in glioma development. NEAT1 could act as a significant prognostic biomarker in glioma progression.

Regulation of human glioma cell migration, tumor growth, and stemness gene expression using a Lck targeted inhibitor

Migration of human glioma cells (hGCs) within the brain parenchyma makes glioblastoma one of the most aggressive and lethal tumors. Studies of the cellular and molecular mechanisms underlying hGC migration are hindered by the limitations of existing glioma models. Here we developed a dorsal root ganglion axon-oligodendrocyte-hGC co-culture to study in real time the migration and interaction of hGCs with their microenvironment. hGCs interact with myelinated and non-myelinated axons through the formation of pseudopodia. Isolation of pseudopodia-localized polysome-bound RNA reveals transcripts of Lck, Paxillin, Crk-II, and Rac1 that undergo local translation. Inhibition of Lck phosphorylation using a small-molecule inhibitor (Lck-I), blocks the phosphorylation of Paxillin and Crk-II, the formation of pseudopodia and the migration of hGCs. In vivo intraventricular administration of the Lck-I using an orthotopic xenograft glioma model, results in statistically significant inhibition of tumor size and significant down-regulation of Nanog-targeted genes, which are associated with glioblastoma patient survival. Moreover, treatment of human glioma stem cells (hGSCs) with Lck-I, results in significant inhibition of self-renewal and tumor-sphere formation. The involvement of Lck in different levels of glioma malignant progression, such as migration, tumor growth, and regulation of cancer stemness, makes Lck a potentially important therapeutic target for human glioblastomas.

TRIM28 is overexpressed in glioma and associated with tumor progression.

BackgroundTripartite motif containing 28 (TRIM28) is a transcriptional co-factor targeting many genes with pleiotropic biological activities, but the study on the role of TRIM28 in glioma is rare.MethodsTo explore the function of TRIM28 in glioma, we first detected the expression levels of TRIM28 in glioma tissues and analyzed the correlations of TRIM28 expression with clinicopathological variables of patients in 85 cases of glioma. Meanwhile, we used shRNA to knockdown TRIM28 in glioma cell lines to detect the biological functions of TRIM28 in cell and animal experiments.ResultsWe found that TRIM28 was expressed at significantly higher level in glioma tissues than in non-tumor brain, and TRIM28 expression correlated significantly with tumor malignancy. Furthermore, TRIM28 higher expression was also correlated with poor survival of glioma patients (P<0.01). Functionally, knockdown of TRIM28 could significantly inhibit cell proliferation and migration in glioma cells. Additionally, we found that TRIM28 could inhibit the expression of E-cadherin significantly by reducing its mRNA stability at the post-transcriptional level.ConclusionOur results suggest that TRIM28 overexpression is correlated with glioma malignant progression and patients’ poor survival, so targeting TRIM28 could be an efficacious strategy in glioma.

MiR-126 Suppresses Invasion and Migration of Malignant Glioma by Targeting Mature T Cell Proliferation 1 (MTCP1).

BackgroundThe aim of this study was to assess the utility of miR-126 in promoting malignant glioma progression and determine if miR-126 might be a target for malignant glioma treatment.Material/MethodsThe expression of miR-126 in malignant glioma tissues and cells was detected by reverse transcription polymerase chain reaction (RT-PCR). Western blot analysis was used to detect changes in protein levels. Transwell assay was applied to assess the migration and invasion in vitro. Luciferase reporter assay was used to confirm the binding of miR-126 and mature T cell proliferation 1 (MTCP1). A nude mouse tumor model was used to assess the molecular mechanism in vivo.ResultsThe expression level of miR-126 in patients with stage III~IV malignant glioma was significant lower than that in patients with stage I~II. In different malignant glioma cell lines, the expression was significantly reduced in U87MG. Compared with the control mimics group, the expression of MTCP1 was significantly decreased. The results of Transwell assay showed that the invasiveness and migration in the miR-126 mimics group was significantly lower than in the control mimics groups. miR-126 mimics did not affect luciferase activity in the Mut-miR-126/MTCP1 group, while miR-126 mimics reduced luciferase activity by 54% in the Wt-miR-126/MTCP1 group. The results of invasion showed that the invasion ability in the miR-126 inhibitor group was significantly increased compared with that in the normal control (NC) group, while the invasion and migration abilities in the MTCP1 siRNA group were significantly increased. After 6 weeks, the tumor volume in the miR-126 inhibitor group was significantly increased, while that in the MTCP1 siRNA group was significantly decreased.ConclusionsmiR-126 inhibits the migration of malignant glioma cells by inhibiting MTCP1.

RUNX2 Promotes Malignant Progression in Glioma.

Glioblastoma (GBM) is the most aggressive and lethal form of brain tumor. However, therapeutic strategies against malignant gliomas have not been completely established. Runt-related transcription factor 2 (Runx2) is an essential gene for skeletal development but its regulatory role in the malignant progression of glioma remains unclear. Here we investigated expression levels of RUNX2 in glioma tissues and its regulatory effects on aberrant growth of glioma cells. RUNX2 mRNA levels were higher in GBM tissues than that of normal brains or low-grade gliomas. RUNX2 protein was detected in five out of seven human GBM cell lines and its level was positively correlated with proliferative capacity. Stable transduction of dominant-negative Runx2 in rat-derived C6 glioma cells not only inhibited the promoter activity containing Runx2 response element, but also decreased mRNA expression levels of Runx2 target genes, such as Mmp13 and Spp1, as well as the proliferative capacity. Furthermore, transient introduction of Runx2-targeted siRNAs into C6 glioma cells significantly decreased mRNA expression levels of Mmp13 and Spp1 and the proliferative capacity. Furthermore, Runx2 knockdown suppressed both Ccnd1 mRNA expression and activation of the Ccnd1 promoter by forskolin, a PKA-activating reagent, in C6 glioma cells. Our results demonstrate that cross-talk between cAMP/PKA signaling and RUNX2 promotes a malignant phenotype of glioma cells.