Circ_0005015 upregulates BACH1 to promote aggressive behaviors in glioblastoma by sponging microRNA-382-5p.

To investigate the potential role and molecular mechanism of circ_0005015 in GBM progression. Circ_0005015, microRNA-382-5p (miR-382-5p), and BTB domain and CNC homolog 1 (BACH1) levels were measured by real-time quantitative polymerase chain reaction (RT-qPCR). Cell proliferation was determined by MTT, colony formation, and EdU assays. Cell apoptosis was analyzed using flow cytometry. Cell migration and invasion were assessed using wound healing and transwell assays. Glucose accumulation and lactate levels were examined by the corresponding kit. RNA pull-down and dual-luciferase reporter assays were performed to confirm the interaction between miR-382-5p and circ_0005015 or BACH1. Protein levels of MMP9, PCNA, and BACH1 were examined using western blot assay. Role of circ_0005015 on tumor growth in vivo was analyzed using a xenograft tumor model. Circ_0005015 content was up-regulated in GBM patients and cells, its knockdown restrained GBM cell proliferation, migration, invasion, glycolysis, and triggered apoptosis. Mechanistically, we found that circ_0005015 could directly interact with miR-382-5p and serve as a miRNA sponge to regulate BACH1 expression. In addition, circ_0005015 knockdown might repress tumor growth in vivo. Circ_0005015 boosted GBM progression via binding to miR-382-5p to up-regulate BACH1, which may offer new effective targets for GBM treatment.

circ_0030018 promotes glioma proliferation and metastasis.

BACKGROUND: Circular RNA (circRNA) plays an essential role in tumor progression, including glioma. circ_0030018 is a newly discovered circRNA that is highly expressed in glioma. However, its role and mechanism in glioma need to be further elucidated. METHODS: The expression of circ_0030018, microRNA (miR)-194-5p, and tripartite motif containing 44 (TRIM44) was examined using quantitative real-time PCR. Cell proliferation, migration, invasion, and apoptosis were determined using MTT assay, colony formation assay, transwell assay, and flow cytometry. Moreover, dual-luciferase reporter assay and RNA pull-down assay were used to verify the interactions among circ_0030018, miR-194-5p, and TRIM44. The protein expression of TRIM44 was assessed by western blot analysis. Animal experiments were conducted to explore the role of circ_0030018 in glioma tumor growth in vivo. RESULTS: circ_0030018 was overexpressed in glioma tissues and cells, and its silencing could inhibit glioma cell proliferation, migration, invasion, and accelerate apoptosis. miR-194-5p could be sponged by circ_0030018, and its overexpression could hinder the progression of glioma cells. Further experiments revealed that miR-194-5p inhibitor reversed the negative regulation of circ_0030018 knockdown on glioma cell progression. In addition, TRIM44 was a target of miR-194-5p, and its downregulation could repress glioma cell progression. Overexpressed TRIM44 reversed the inhibition effect of miR-194-5p on glioma cell progression. Animal experiments suggested that circ_0030018 knockdown could reduce glioma tumor growth through regulating miR-194-5p and TRIM44. CONCLUSION: Our 8data showed that circ_0030018 enhanced glioma progression by sponging miR-194-5p to regulate TRIM44, indicating that circ_0030018 might be a potential treatment target for glioma.

Overexpression of Paxillin Correlates with Tumor Progression and Predicts Poor Survival in Glioblastoma.

AIMS: To explore the prognostic and clinicopathological features of glioma with Paxillin (PXN) expression based on a large number of samples. METHODS: RNA sequencing data of 325 glioma samples from Chinese Glioma Genome Atlas (CGGA) database were obtained as discovery set. Three additional datasets were further obtained as validation sets. The protein expression pattern of PXN in glioma was measured by IHC. Kaplan-Meier survival and multivariate Cox analysis were used to estimate the survival distributions. Moreover, the functional annotation of PXN was also analyzed. RESULTS: In the discovery set, PXN overexpression was significantly associated with high-grade glioma as well as the higher mortality in survival analysis (log-rank test, P < 0.01). The results of the other validation datasets showed similar findings. PXN also served as an independent prognostic biomarker in glioblastoma patients. Functional assays showed that PXN contributed to glioma cell proliferation and invasion. CONCLUSION: PXN plays as an oncogene in glioma progression and suggests a new potential biotarget for therapy.

KIF23 is an independent prognostic biomarker in glioma, transcriptionally regulated by TCF-4.

Kinesin family member 23 (KIF23), a nuclear protein and a key regulator of cellular cytokinesis, has been found to be overexpressed as an oncogene in glioma. However, the prognostic and clinicopathological features of glioma with KIF23 expression was not clear yet. Here, we analyzed KIF23 expression pattern by using whole genome mRNA expression microarray data from Chinese Glioma Genome Atlas (CGGA) database (http://www.cgga.org.cn), and found that KIF23 overexpression was significantly associated with high grade glioma as well as the higher mortality in survival analysis (log-rank test, p<0.01). The results of the three other validation datasets showed similar findings. Furthermore, KIF23 also served as an independent prognostic biomarker in glioma patients. Finally, functional assay showed that reduction of KIF23 suppressed glioma cell proliferation both in vivo and vitro. Additionally, we found that KIF23 was regulated by TCF-4 at transcriptionally level. Therefore, this evidence indicates KIF23 over-expression is associated with glioma malignancy and conferred a worse survival time in glioma, which suggests KIF23 is a new novel prognostic biomarker with potential therapeutic implications in glioma.

High expression of CXCR3 is an independent prognostic factor in glioblastoma patients that promotes an invasive phenotype.

Chemokines are a superfamily of small heparin-binding cytokines that induce leukocytes to migrate to sites of inflammation or injury through interacting with specific transmembrane G protein-coupled receptors. Currently, attention is focused on chemokine/chemokine receptor pairs and their ability to promote tumor cell migration and angiogenesis. The chemokine receptor CXCR3 is involved in tumor metastasis and is used as a prognostic biomarker. However, its relationship with the clinicopathological features of primary glioblastoma multiforme (pGBM) and its potential prognostic value have yet to be investigated. Here, we report that high CXCR3 expression conferred poor survival in pGBM patients. Further analysis showed that CXCR3 served as an independent prognostic biomarker for pGBM patients. In addition, functional assays indicated that CXCR3 induced glioma cell invasion. Therefore, this evidence indicates CXCR3 is an independent prognostic factor for pGBM patients and promotes an invasive phenotype, which suggests a new potential biotarget for glioblastoma multiforme therapy.

MiR-328 expression is decreased in high-grade gliomas and is associated with worse survival in primary glioblastoma.

MicroRNAs, a group of small endogenous, noncoding RNAs, are aberrantly expressed in many human cancers and can act as oncogene or anti-oncogene. Recent evidence suggests that some miRNAs have prognostic value for tumors. MiR-328 is known as a tumor suppressor; however, its relationship with the clinicopathological features of glioblastoma (GBM) and its prognostic value has yet not been investigated. We found that expression of miR-328 was significantly decreased both in anaplastic and GBM cohorts and that low miR-328 expression also conferred poor survival in primary GBM (PGBM) patients. MiR-328 might, therefore, serve as an independent prognostic marker. Furthermore, expression profiles of miR-328-associated mRNAs were established via microarrays for 60 GBM samples. The ontology of the miR-328-associated genes was then analyzed, which identified gene sets tightly related to cell mitosis. In addition, ectopic expression of miR-328 inhibited U87 cell proliferation and induced U87 cell cycle arrest. In conclusion, this is the first report showing that miR-328 is associated with patient's survival time and that miR-328 might serve as an independent prognostic biomarker for GBM.

Methylation mediated silencing of miR-23b expression and its role in glioma stem cells.

MicroRNAs (miRNAs) are a class of small non-coding RNAs that negatively regulate gene expression at a post-transcriptional level. Some miRNAs harboring CGIs undergo methylation mediated silencing, a characteristic of many tumor suppressor genes. To identify such miRNAs in glioma stem cells (GSCs), we first showed that miR-23b is frequently methylated in GSCs but not in parallel U87 cells. Meanwhile, miR-23b expression was also markedly reduced in GSCs compared with matching U87 cells. Furthermore, treatment with 5-aza can increase miR-23b expression in GSCs. In addition, ectopic expression of miR-23b in GSCs induces cell cycle arrest and proliferation inhibition. Further analysis showed that miR-23b could enhance the sensitivity of U87 GSCs to TMZ. Our results suggest that miR-23b is epigenetically down-regulated and restoration of miR-23b can effectively suppress cell growth in GSCs.

MicroRNA-34a suppresses cell proliferation and induces apoptosis in U87 glioma stem cells.

Tumor stem cells (TSCs) have been identified in many malignant tumors and are unique in their self-renewal, multi-differentiation and tumor growth maintenance capabilities. MicroRNAs (miRNAs) are small endogenous, non-coding RNAs that predominately modulate target gene expression at the post-transcriptional level. Accumulating evidence suggests that some miRNAs have an essential role in TSC proliferation, growth, apoptosis, and differentiation. In particular, miR-34a has been found to inhibit proliferation and induce apoptosis in stem cells. The insensitivity of glioma to standard therapies combined with the hypothesis that glioma stem cells (GSCs) are responsible for this chemorefractory nature suggests that investigations exploring the function and mechanism of miR-34a in GSCs would be of value. In our study, we found that miR-34a is down-regulated in CD133-positive U87 cells. Furthermore, miR-34a could significantly suppress cell proliferation and induce apoptosis in GSCs. These findings suggest that miR-34a acts as a tumor suppressor in U87 GSCs. Therefore, this observation highlights a new potential therapeutic agent for GSC-based glioma treatment.

Bone morphogenetic protein 4 inhibits cell proliferation and induces apoptosis in glioma stem cells.

Glioma stem cells (GSCs), which are originated from transformed neural stem cells, are tumor-initiating cells of glioma, the most common primary malignant neoplasm of the central nervous system. Extensive studies have shown that bone morphogenetic protein 4 (BMP4) plays an important role in the differentiation and proliferation of neural stem cells. To seek the functions and mechanisms of BMP4 in GSCs, GSCs isolated from U87 human glioma cells by using vincristine were exposed to BMP4 protein. This study shows that BMP4 inhibited U87 GSC proliferation (p < 0.01) via downregulation of cyclin D1 level and promoted GSC apoptosis through induction of Bax expression and inhibition of Bcl-2 and Bcl-xL levels. Thus, these results indicate a new approach of GSC-based glioma treatment.

MicroRNA-10b induces glioma cell invasion by modulating MMP-14 and uPAR expression via HOXD10.

MicroRNAs are small endogenous noncoding RNAs, which modulate target gene expression by binding with target mRNA sequences in the 3'untranslated region (UTR) with an imperfect complementarity that inhibits the mRNA translation. Many microRNAs have been reported to function as tumor oncogenes or anti-oncogenes. Recently, more and more microRNAs have been reported to contribute to a tumor's invasive potential. Here, we show that microRNA-10b (miR-10b) was over-expressed in glioma samples and directly associated with the glioma's pathological grade and malignancy. We also found that miR-10b induced glioma cell invasion by modulating tumor invasion factors MMP-14 and uPAR expression via the direct target HOXD10. The miR-10b/HOXD10/MMP-14/uPAR signaling pathway might contribute to the invasion of glioma. Accordingly, glioma cells lost their invasive ability when treated with specific antisense oligonucleotides (miR-10b inhibitors), suggesting that miR-10b could be used as a new bio-target to cure glioma.

Combination of tamoxifen and antisense human telomerase RNA inhibits glioma cell proliferation and anti-apoptosis via suppression of telomerase activity.

Accumulating evidence indicates that telomerase activity and human telomerase RNA (hTR) play a potentially crucial role in maintaining the malignant progression of human gliomas. Tamoxifen (TAM) induces cell growth inhibition by modulating several cellular activities, including signaling pathways and the cell cycle. In this study, we aimed to evaluate the effect of combination therapy with TAM and antisense hTR on malignant glioma growth in vitro. TAM treatment and the down-regulation of hTR expression by antisense hTR resulted in a significant suppression of cell growth and the induction of cell apoptosis through the inhibition of telomerase activity. The antitumor effect of treatment with TAM alone was found to be mediated in part by the down-regulation of telomerase activity and of PKC and IGF-1 expression. Our results indicate that TAM and antisense hTR may serve as a novel strategy for glioma therapy.