Identification of Implications of Angiogenesis and m6A Modification on Immunosuppression and Therapeutic Sensitivity in Low-Grade Glioma by Network Computational Analysis of Subtypes and Signatures.

Angiogenesis is a complex process in the immunosuppressed low-grade gliomas (LGG) microenvironment and is regulated by multiple factors. N6-methyladenosine (m6A), modified by the m6A modification regulators ("writers" "readers" and "erasers"), can drive LGG formation. In the hypoxic environment of intracranial tumor immune microenvironment (TIME), m6A modifications in glioma stem cells are predominantly distributed around neovascularization and synergize with complex perivascular pathological ecology to mediate the immunosuppressive phenotype of TIME. The exact mechanism of this phenomenon remains unknown. Herein, we elucidated the relevance of the angiogenesis-related genes (ARGs) and m6A regulators (MAGs) and their influencing mechanism from a macro perspective. Based on the expression pattern of MAGs, we divided patients with LGG into two robust categories via consensus clustering, and further annotated the malignant related mechanisms and corresponding targeted agents. The two subgroups (CL1, CL2) demonstrated a significant correlation with prognosis and clinical-pathology features. Moreover, WGCNA has also uncovered the hub genes and related mechanisms of MAGs affecting clinical characters. Clustering analysis revealed a synergistic promoting effect of M6A and angiogenesis on immunosuppression. Based on the expression patterns of MAGs, we established a high-performance gene-signature (MASig). MASig revealed somatic mutational mechanisms by which MAGs affect the sensitivity to treatment in LGG patients. In conclusion, the MAGs were critical participants in the malignant process of LGG, with a vital potential in the prognosis stratification, prediction of outcome, and therapeutic sensitivity of LGG. Findings based on these strategies may facilitate the development of objective diagnosis and treatment systems to quantify patient survival and other outcomes, and in some cases, to identify potential unexplored targeted therapies.

Construction of Interferon-Gamma-Related Gene Signature to Characterize the Immune-Inflamed Phenotype of Glioblastoma and Predict Prognosis, Efficacy of Immunotherapy and Radiotherapy.

Interferon-gamma (IFNG) has profound impacts on tumor-immune interaction and is of great clinical significance for multiple cancers. Exploring the role of IFNG in glioblastoma (GBM) may optimize the current treatment paradigm of this disease. Here, multi-dimensional data of 429 GBM samples were collected. Various bioinformatics algorithms were employed to establish a gene signature that characterizes immunological features, genomic alterations, and clinical characteristics associated with the IFNG response. In this way, a novel IFNG-related gene signature (IFNGrGS, including TGFBI, IL4I1, ACP5, and LUM) has been constructed and validated. Samples with increased IFNGrGS scores were characterized by increased neutrophil and macrophage infiltration and exuberant innate immune responses, while the activated adaptive immune response may be frustrated by multiple immunosuppressive mechanisms. Notably, the IFNG pathway as well as its antagonistic pathways including IL4, IL10, TGF-beta, and VEGF converged on the expression of immune checkpoints. Besides, gene mutations involved in the microenvironment were associated with the IFNGrGS-based stratification, where the heterogeneous prognostic significance of EGFR mutation may be related to the different degrees of IFNG response. Moreover, the IFNGrGS score had solid prognostic value and the potential to screen ICB and radiotherapy sensitive populations. Collectively, our study provided insights into the role of IFNG on the GBM immune microenvironment and offered feasible information for optimizing the treatment of GBM.

Dihydroartemisinin prompts amplification of photodynamic therapy-induced reactive oxygen species to exhaust Na/H exchanger 1-mediated glioma cells invasion and migration.

Photodynamic therapy (PDT) is a promising glioma therapy; however, its efficacy is compromised due to the PDT-induced reactive oxygen species (ROS) production being limited by the local hypoxic tumor microenvironment. Furthermore, Hypoxia activates sodium/hydrogen exchanger 1 (NHE1), an essential component for tumor progression and metastasis, enables glioma cells (GC) to escape PDT-mediated phototoxicity via increased H+ extrusion. However, interactions between NHE1 expression with ROS level involving response of GC remain unclear. Dihydroartemisinin (DHA), a ROS generator, has extensive anti-tumor effects. This study aimed to explore whether PDT along with DHA could amplify the total ROS levels and diminish GC invasion and migration by inhibiting NHE1 expression. Proliferation and invasion of U251 and LN229 cells were evaluated under different treatments using cell counting Kit-8 (CCK-8), transwell, and wound healing assays. ROS levels were measured using fluorescence probes and flow cytometry. NHE1 levels were detected by immunofluorescence and western blotting. Co-treatment effects and molecular events were further confirmed in a bilateral tumor-bearing nude mouse model. PDT with synergistic DHA significantly increased the total abundance of ROS to further suppress the invasion and migration of GC by reducing NHE1 levels in vitro. Using a bilateral glioma xenograft mouse model with primary and recurrent gliomas, we found that PDT markedly suppressed primary tumor growth, while PDT in synergy with DHA also suppressed recurrent tumors, and improved overall survival by regulating the ROS-NHE1 axis. No evident side effects were observed. Our results suggest that PDT with DHA can amplify the total ROS levels to weaken GC invasion and migration by suppressing NHE1 expression in vitro and in vivo, thus abolishing the resistance of GC to PDT. The synergistic therapy of PDT and DHA therefore represents a more efficient and safe strategy for comprehensive glioma treatment.

m6A regulator-mediated methylation modification patterns and characteristics of immunity and stemness in low-grade glioma.

m6A RNA methylation is an emerging epigenetic modification, and its potential role in immunity and stemness remains unknown. Based on 17 widely recognized m6A regulators, the m6A modification patterns and corresponding characteristics of immune infiltration and stemness of 1152 low-grade glioma samples were comprehensively analyzed. Machine-learning strategies for constructing m6AScores were trained to quantify the m6A modification patterns of individual samples. Here, we reveal a significant correlation between the multi-omics data of regulators and clinicopathological parameters. We identified two distinct m6A modification patterns (an immune-activated differentiation pattern and an immune-desert dedifferentiation pattern) and four regulatory patterns of m6A methylation on immunity and stemness. We show that the m6AScores can predict the molecular subtype of low-grade glioma, the abundance of immune infiltration, the enrichment of signaling pathways, gene variation and prognosis. The concentration of high immunogenicity and clinical benefits in the low-m6AScore group confirmed the sensitive response to radio-chemotherapy and immunotherapy in patients with high-m6AScore. The results of the pan-cancer analyses illustrate the significant correlation between m6AScore and clinical outcome, the burden of neoepitope, immune infiltration and stemness. The assessment of individual tumor m6A modification patterns will guide us in improving treatment strategies and developing objective diagnostic tools.

Identification of Prognostic Model and Biomarkers for Cancer Stem Cell Characteristics in Glioblastoma by Network Analysis of Multi-Omics Data and Stemness Indices.

The progression of most human cancers mainly involves the gradual accumulation of the loss of differentiated phenotypes and the sequential acquisition of progenitor and stem cell-like features. Glioblastoma multiforme (GBM) stem cells (GSCs), characterized by self-renewal and therapeutic resistance, play vital roles in GBM. However, a comprehensive understanding of GBM stemness remains elusive. Two stemness indices, mRNAsi and EREG-mRNAsi, were employed to comprehensively analyze GBM stemness. We observed that mRNAsi was significantly related to multi-omics parameters (such as mutant status, sample type, transcriptomics, and molecular subtype). Moreover, potential mechanisms and candidate compounds targeting the GBM stemness signature were illuminated. By combining weighted gene co-expression network analysis with differential analysis, we obtained 18 stemness-related genes, 10 of which were significantly related to survival. Moreover, we obtained a prediction model from both two independent cancer databases that was not only an independent clinical outcome predictor but could also accurately predict the clinical parameters of GBM. Survival analysis and experimental data confirmed that the five hub genes (CHI3L2, FSTL3, RPA3, RRM2, and YTHDF2) could be used as markers for poor prognosis of GBM. Mechanistically, the effect of inhibiting the proliferation of GSCs was attributed to the reduction of the ratio of CD133 and the suppression of the invasiveness of GSCs. The results based on an in vivo xenograft model are consistent with the finding that knockdown of the hub gene inhibits the growth of GSCs in vitro. Our approach could be applied to facilitate the development of objective diagnostic and targeted treatment tools to quantify cancer stemness in clinical tumors, and perhaps lead considerable benefits that could predict tumor prognosis, identify new stemness-related targets and targeted therapies, or improve targeted therapy sensitivity. The five genes identified in this study are expected to be the targets of GBM stem cell therapy.

FtMt promotes glioma tumorigenesis and angiogenesis via lncRNA SNHG1/miR-9-5p axis.

OBJECTIVE: This study is to investigate the effects and the mechanisms of mitochondrial ferritin (FtMt) on the glioma tumorigenesis and angiogenesis. METHODS: FtMt expression was detected in glioma tissues and cells as well as in nude mouse tissues. Cell proliferation and apoptosis rate were observed following transfection of LV-FtMt or sh-FtMt in glioma cell line. Moreover, glioma cells with FtMt over-expression/knockdown were co-cultured with human umbilical vein endothelial cells (HUVECs) to observe its function on HUVEC proliferation, angiogenic ability and the vascular endothelial growth factor (VEGF) content. Gain and loss of function of small nucleolar RNA host gene 1 (SNHG1) and miR-9-5p were performed in glioma cells and GBM nude mice to observe its effect on glioma cell proliferation and HUVEC angiogenic ability. Luciferase reporter gene and RIP assay were employed to inspect the interactions among SNHG1, FtMt and miR-9-5p. Additionally, a xenograft mouse model was applied to determine the role of FtMt in glioma. RESULTS: In this work, FtMt was strongly expressed in glioma tissues and cells as well as in nude mouse tumor tissues. The employment of the loss-of and gain-of functions assays illustrated that FtMt enhanced glioma tumorigenesis and angiogenesis. Mechanistically, our findings showed that FtMt positively related to SNHG1 while negatively correlated with miR-9-5p, and both SNHG1 and FtMt can competitively bind with miR-9-5p. Besides, the inhibition effects of sh-FtMt on glioma were surveyed in vivo experiments. CONCLUSION: Evidence in this study suggested that FtMt promotes glioma tumorigenesis and angiogenesis via SNHG1 mediated miR-9-5p expression, which may provide a theoretical basis for glioma treatment.

Malignant Evaluation and Clinical Prognostic Values of m6A RNA Methylation Regulators in Glioblastoma.

N6-methyladenosine (m6A) RNA methylation, the most common form of mRNA modification and regulated by the m6A RNA methylation regulators ("writers," "erasers," and "readers"), has been reported to be associated with the progression of the malignant tumor. However, its role in glioblastoma (GBM) has been poorly known. This study aimed to identify the expression, potential functions, and prognostic values of m6A RNA methylation regulators in GBM. Here, we revealed that the 13 central m6A RNA methylation regulators were firmly related to the clinical and molecular phenotype of GBM. Taking advantage of consensus cluster analysis, we obtained two categories of GBM samples and found malignancy-related processes of m6A methylation regulators and compounds that specifically targeted the malignant processes. Besides, we also obtained a list of genes with poor prognosis in GBM. Finally, we derived a risk-gene signature with three selected m6A RNA methylation regulators, which allowed us to extend the in-depth study and dichotomized the OS of patients with GBM into high- and low-risk subgroups. Notably, this risk-gene signature could be used as independent prognostic markers and accurate clinicopathological parameter predictors. In conclusion, m6A RNA methylation regulators are a type of vital participant in the malignant progression of GBM, with a critical potential in the prognostic stratification and treatment strategies of GBM.

Overexpression of circular RNA circ-CDC45 facilitates glioma cell progression by sponging miR-516b and miR-527 and predicts an adverse prognosis.

Glioma (GM) is a highly lethal human cancer. Circular RNAs (circRNAs) act as an imperative factor in oncogenesis. We aimed to investigate the biological functions and mechanisms of circ-CDC45 in GM. circ-CDC45 expression in GM specimens and cell lines was measured by real-time quantitative reverse transcription polymerase chain reaction. Fisher's exact test and Kaplan-Meier curves further analyzed its clinical implications. A gain/loss-of-function study was conducted to investigate the role of circ-CDC45 in GM. Additionally, luciferase reporter and rescue assays were performed to unravel the mechanisms of circ-CDC45. High circ-CDC45 expression was found in GM specimens and cells, which was tightly related to a larger tumor size, higher world health organization (WHO) stages, and worse survival for patients with GM. Functionally, manipulation of circ-CDC45 expression strongly affected cell growth, apoptosis, migration and invasion, which suggests the oncogenic function of circ-CDC45 in GM oncogenesis. Stepwise mechanism studies indicated that circ-CDC45 sponged and regulated the expression of miR-516b and miR-527 to promote cell growth and invasion. Briefly, the regulatory network of circ-CDC45/miR-516b/miR-527 plays a pivotal role in GM tumorigenesis and may act as a potential target for GM treatment.