Correlation of LLT-1 and NLRC4 inflammasome and its effect on glioblastoma prognosis.

PURPOSE: LLT-1 is a well-known ligand for the natural killer (NK) cell inhibitory receptor NKRP1A. Here, we examined NLRC4 inflammasome components and LLT-1 expression in glioblastoma (GBM) tissues to elucidate potential associations and interactions between these factors. METHODS: GBM tissues were collected for RNA sequencing (RNA-seq) and Immunofluorescent experiments. Colocalization of LLT-1 and other proteins was assessed by immunofluorescence. Computational analyses utilized RNA-seq data from 296 to 52 patients from the Chinese Glioma Genome Atlas and CHA medical records, respectively. These data were subjected to survival, non-negative matrix factorization clustering, Gene Ontology enrichment, and protein-protein interaction analyses. Receptor-ligand interactions between tumor and immune cells were confirmed by single-cell RNA-seq analysis. RESULTS: In GBM tissues, LLT-1 was predominantly colocalized with glial fibrillary acidic protein (GFAP)-expressing astrocytes, but not with microglial markers like Iba-1. Additionally, LLT-1 and activated NLRC4 inflammasomes were mainly co-expressed in intratumoral astrocytes, suggesting an association between LLT-1, NLRC4, and glioma malignancy. High LLT-1 expression correlates with poor prognosis, particularly in the mesenchymal subtype, and is associated with TNF and NOD-like receptor signaling pathway enrichment, indicating a potential role in tumor inflammation and progression. At the single-cell level, mesenchymal-like malignant cells showed high NF, NLR, and IL-1 signaling pathway enrichment compared to other malignant cell types. CONCLUSION: We revealed an association between NLRC4 inflammasome activity and LLT-1 expression, suggesting a novel regulatory pathway involving TNF, inflammasomes, and IL-1, potentially offering new NK-cell-mediated anti-glioma approaches.

Dysregulation of inflammasome activation in glioma.

Gliomas are the most common brain tumors characterized by complicated heterogeneity. The genetic, molecular, and histological pathology of gliomas is characterized by high neuro-inflammation. The inflammatory microenvironment in the central nervous system (CNS) has been closely linked with inflammasomes that control the inflammatory response and coordinate innate host defenses. Dysregulation of the inflammasome causes an abnormal inflammatory response, leading to carcinogenesis in glioma. Because of the clinical importance of the various physiological properties of the inflammasome in glioma, the inflammasome has been suggested as a promising treatment target for glioma management. Here, we summarize the current knowledge on the contribution of the inflammasomes in glioma and therapeutic insights. Video Abstract.

Comparing the Expression of Canonical and Non-Canonical Inflammasomes Across Glioma Grades: Evaluating Their Potential as an Aggressiveness Marker.

BACKGROUND: Inflammasomes are key in the initiation of inflammatory responses and serve to defend the organism. However, when the immune system is imbalanced, these complexes contribute to tumor progression. The purpose of this study was to investigate the effect of non-canonical inflammasomes on glioma malignancy. METHODS: We performed bioinformatics analysis to confirm the expression of canonical and non-canonical inflammasome-related molecules according to the degree of malignancy through immunohistochemical examination of glioma tissues obtained with patient consent from our institution. RESULTS: Bioinformatics analysis confirmed that the expression levels of non-canonical inflammasome-related molecules were significantly higher in tumor tissues than in normal tissues, and they also increased according to malignancy, which adversely affected the survival rate. Furthermore, in gliomas, positive correlations were found between N-form gasdermin-D, a key molecule associated with the non-canonical inflammasome, and other related molecules, including NLRP3, caspase-1, caspase-4, and caspase-5. These results were verified by immunohistochemical examination of glioma tissues, and the expression levels of these molecules also increased significantly with increasing grade. In addition, the features of pyroptosis were confirmed. CONCLUSION: This study identified the potential of non-canonical inflammasomes as aggressiveness markers for gliomas and presented a perspective for improving glioma treatment.

Non-canonical NLRC4 inflammasomes in astrocytes contribute to glioma malignancy.

BACKGROUND: The present study was designed to explore the pathological role of non-canonical NLRC4 inflammasome in glioma. METHODS: This retrospective study included bioinformatical analysis, including survival, gene ontology, ssGSEA, cox regression, IPA and drug repositioning with TCGA and DepMap database. Experimental validations were conducted in glioma patient's sample and evaluated with histological or cellular functional analysis. RESULT: Clinical dataset analysis revealed that non-canonical NLRC4 inflammasomes significantly contribute to glioma progression and poor survival rates. Experimental validation was revealed that the expression of non-canonical NLRC4 inflammasomes were co-localized with astrocytes in malignant gliomas, with a sustained clinical correlation observed between astrocytes and inflammasome signatures. Indeed, the formation of an inflammatory microenvironment increased in malignant gliomas, leading to pyroptosis, known as inflammatory cell death. Molecular interaction analysis revealed that NF-κB pathways potentially serve as the connecting point between the canonical and noncanonical pathways of the NLRC4 inflammasome. Finally, drug repositioning analysis of non-canonical NLRC4 inflammasome-associated molecules revealed that MK-5108, PF4981517, and CTEP may represent effective options for glioma therapy. CONCLUSION: The findings of this study suggest that non-canonical NLRC4 inflammasomes contribute to poor prognosis in patients with glioma and induce an inflammatory microenvironment. We propose the pathological phenomenon of non-canonical NLRC4 inflammasomes and several therapeutic strategies based on the modulation of the inflammatory tumor microenvironment.

Molecular characteristics of incidental lower-grade glioma for treatment decision-making.

OBJECTIVE: Several limitations are associated with the early diagnosis and treatment of incidental lower-grade glioma (iLGG), and due to its unknown molecular features, its management is categorized as either the "wait-and-see" strategy or immediate treatment. Therefore, in this study the authors explored iLGG's clinical and molecular landscape to improve its management. METHODS: The authors retrospectively assessed the differences between the molecular and clinical characteristics of iLGG and symptomatic lower-grade glioma (sLGG) samples filtered based on symptom data corresponding to The Cancer Genome Atlas cohort with mutations. Thereafter, genomic and transcriptomic analysis was performed. RESULTS: There was no significant difference between iLGG and sLGG with respect to mutation status; however, there was an increase in the interaction between major mutations in sLGG, depending on the histological subtype and the IDH1 mutation status. Furthermore, the IDH1 mutation characteristics corresponding to wild-type glioma were much more obvious in sLGG than in iLGG. Additionally, in sLGG, genes associated with malignancy, including cell proliferation-related, cell migration-related, epithelial-to-mesenchymal transition-related, and negative regulation of cell death-related genes, were significantly upregulated, and groups showing higher expression levels of these genes were associated with worse prognosis. Also, 8 of the 75 identified upregulated genes showed positive correlation with resistance to the drugs that are normally used for glioma treatment, including procarbazine, carmustine, vincristine, and temozolomide. CONCLUSIONS: The new insights regarding the different molecular features of iLGG and sLGG indicated that the immediate management of iLGG could result in better prognosis than the wait-and-see strategy.

Association of Tim-3/Gal-9 Axis with NLRC4 Inflammasome in Glioma Malignancy: Tim-3/Gal-9 Induce the NLRC4 Inflammasome.

Tim-3/Gal-9 and the NLRC4 inflammasome contribute to glioma progression. However, the underlying mechanisms involved are unclear. Here, we observed that Tim-3/Gal-9 expression increased with glioma malignancy and found that Tim-3/Gal-9 regulate NLRC4 inflammasome formation and activation. Tim-3/Gal-9 and NLRC4 inflammasome-related molecule expression levels increased with WHO glioma grade, and this association was correlated with low survival. We investigated NLRC4 inflammasome formation by genetically regulating Tim-3 and its ligand Gal-9. Tim-3/Gal-9 regulation was positively correlated with the NLRC4 inflammasome, NLRC4, and caspase-1 expression. Tim-3/Gal-9 did not trigger IL-1ß secretion but were strongly positively correlated with caspase-1 activity as they induced programmed cell death in glioma cells. A protein-protein interaction analysis revealed that the FYN-JAK1-ZNF384 pathways are bridges in NLRC4 inflammasome regulation by Tim-3/Gal-9. The present study showed that Tim-3/Gal-9 are associated with poor prognosis in glioma patients and induce NLRC4 inflammasome formation and activation. We proposed that a Tim-3/Gal-9 blockade could be beneficial in glioma therapy as it would reduce the inflammatory microenvironment by downregulating the NLRC4 inflammasome.

NOX2-Induced High Glycolytic Activity Contributes to the Gain of COL5A1-Mediated Mesenchymal Phenotype in GBM.

The alteration of the cellular metabolism is a hallmark of glioma. The high glycolytic phenotype is a critical factor in the pathogenesis of high-grade glioma, including glioblastoma multiforme (GBM). GBM has been stratified into three subtypes as the proneural, mesenchymal, and classical subtypes. High glycolytic activity was found in mesenchymal GBM relative to proneural GBM. NADPH oxidase 2 (NOX2) has been linked to cellular metabolism and epithelial-mesenchymal transition (EMT) in tumors. The role of NOX2 in the regulation of the high glycolytic phenotype and the gain of the mesenchymal subtype in glioma remain unclear. Here, our results show that the levels of NOX2 were elevated in patients with GBM. NOX2 induces hexokinase 2 (HK2)-dependent high glycolytic activity in U87MG glioma cells. High levels of NOX2 are correlated with high levels of HK2 and glucose uptake in patients with GBM relative to benign glioma. Moreover, NOX2 increases the expression of mesenchymal-subtype-related genes, including COL5A1 and FN1 in U87MG glioma cells. High levels of NOX2 are correlated with high levels of COL5A1 and the accumulation of extracellular matrix (ECM) in patients with GBM relative to benign glioma. Furthermore, high levels of HK2 are correlated with high levels of COL5A1 in patients with GBM relative to benign glioma. Our results suggest that NOX2-induced high glycolytic activity contributes to the gain of the COL5A1-mediated mesenchymal phenotype in GBM.

Autologous adoptive immune-cell therapy elicited a durable response with enhanced immune reaction signatures in patients with recurrent glioblastoma: An open label, phase I/IIa trial.

Glioblastoma multiforme (GBM) is an aggressive malignancy classified by the World Health Organization as a grade IV glioma. Despite the availability of aggressive standard therapies, most patients experience recurrence, for which there are currently no effective treatments. We aimed to conduct a phase I/IIa clinical trial to investigate the safety and efficacy of adoptive, ex-vivo-expanded, and activated natural killer cells and T lymphocytes from peripheral blood mononuclear cells of patients with recurrent GBM. This study was a single-arm, open-label, investigator-initiated trial on 14 patients recruited between 2013 and 2017. The immune cells were administered via intravenous injection 24 times at 2-week intervals after surgical resection or biopsy. The safety and clinical efficacy of this therapy was examined by assessing adverse events and comparing 2-year overall survival (OS). Transcriptomic analysis of tumor tissues was performed using NanoString to identify the mechanism of therapeutic efficacy. No grade 4 or 5 severe adverse events were observed. The most common treatment-related adverse events were grade 1 or 2 in severity. The most severe adverse event was grade 3 fever. Median OS was 22.5 months, and the median progression-free survival was 10 months. Five patients were alive for over 2 years and showed durable response with enhanced immune reaction transcriptomic signatures without clinical decline until the last follow-up after completion of the therapy. In conclusion, autologous adoptive immune-cell therapy was safe and showed durable response in patients with enhanced immune reaction signatures. This therapy may be effective for recurrent GBM patients with high immune response in their tumor microenvironments. Trial registration: The Korea Clinical Research Information Service database: KCT0003815, Registered 18 April 2019, retrospectively registered.