Mutational Landscape of Secondary Glioblastoma Guides MET-Targeted Trial in Brain Tumor.

Low-grade gliomas almost invariably progress into secondary glioblastoma (sGBM) with limited therapeutic option and poorly understood mechanism. By studying the mutational landscape of 188 sGBMs, we find significant enrichment of TP53 mutations, somatic hypermutation, MET-exon-14-skipping (METex14), PTPRZ1-MET (ZM) fusions, and MET amplification. Strikingly, METex14 frequently co-occurs with ZM fusion and is present in ∼14% of cases with significantly worse prognosis. Subsequent studies show that METex14 promotes glioma progression by prolonging MET activity. Furthermore, we describe a MET kinase inhibitor, PLB-1001, that demonstrates remarkable potency in selectively inhibiting MET-altered tumor cells in preclinical models. Importantly, this compound also shows blood-brain barrier permeability and is subsequently applied in a phase I clinical trial that enrolls MET-altered chemo-resistant glioma patients. Encouragingly, PLB-1001 achieves partial response in at least two advanced sGBM patients with rarely significant side effects, underscoring the clinical potential for precisely treating gliomas using this therapy.

Glial-Cell-Line-Derived Neurotrophic Factor Promotes Glioblastoma Cell Migration and Invasion via the SMAD2/3-SERPINE1-Signaling Axis.

Glial-cell-line-derived neurotrophic factor (GDNF) is highly expressed and is involved in the malignant phenotype in glioblastomas (GBMs). However, uncovering its underlying mechanism for promoting GBM progression is still a challenging work. In this study, we found that serine protease inhibitor family E member 1 (SERPINE1) was a potential downstream gene of GDNF. Further experiments confirmed that SERPINE1 was highly expressed in GBM tissues and cells, and its levels of expression and secretion were enhanced by exogenous GDNF. SERPINE1 knockdown inhibited the migration and invasion of GBM cells promoted by GDNF. Mechanistically, GDNF increased SERPINE1 by promoting the phosphorylation of SMAD2/3. In vivo experiments demonstrated that GDNF facilitated GBM growth and the expressions of proteins related to migration and invasion via SERPINE1. Collectively, our findings revealed that GDNF upregulated SERPINE1 via the SMAD2/3-signaling pathway, thereby accelerating GBM cell migration and invasion. The present work presents a new mechanism of GDNF, supporting GBM development.

Yes-Associated Protein Nuclear Translocation Is Regulated by Epidermal Growth Factor Receptor Activation Through Phosphatase and Tensin Homolog/AKT Axis in Glioblastomas.

Gliomas are the most common and lethal primary brain tumors in adults. Glioblastomas, the most frequent and aggressive form of gliomas, represent a therapeutic challenge as no curative treatment exists to date, and the prognosis remains extremely poor. Recently, the transcriptional cofactors Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) belonging to the Hippo pathway have emerged as a major determinant of malignancy in solid tumors, including gliomas. However, the mechanisms involved in its regulation, particularly in brain tumors, remain ill-defined. In glioblastomas, EGFR represents one of the most altered oncogenes affected by chromosomal rearrangements, mutations, amplifications, and overexpression. In this study, we investigated the potential link between epidermal growth factor receptor (EGFR) and the transcriptional cofactors YAP and TAZ by in situ and in vitro approaches. We first studied their activation on tissue microarray, including 137 patients from different glioma molecular subtypes. We observed that YAP and TAZ nuclear location was highly associated with isocitrate dehydrogenase 1/2 (IDH1/2) wild-type glioblastomas and poor patient outcomes. Interestingly, we found an association between EGFR activation and YAP nuclear location in glioblastoma clinical samples, suggesting a link between these 2 markers contrary to its ortholog TAZ. We tested this hypothesis in patient-derived glioblastoma cultures by pharmacologic inhibition of EGFR using gefinitib. We showed an increase of S397-YAP phosphorylation associated with decreased AKT phosphorylation after EGFR inhibition in phosphatase and tensin homolog (PTEN) wild-type cultures, unlike PTEN-mutated cell lines. Finally, we used bpV(HOpic), a potent PTEN inhibitor, to mimic the effect of PTEN mutations. We found that the inhibition of PTEN was sufficient to revert back the effect induced by Gefitinib in PTEN-wild-type cultures. Altogether, to our knowledge, these results show for the first time the regulation of pS397-YAP by the EGFR-AKT axis in a PTEN-dependent manner.

CDKN2A/B Homozygous Deletions in Astrocytomas: A Literature Review.

Genomic alterations of CDKN2A and CDKN2B in astrocytomas have been an evolving area of study for decades. Most recently, there has been considerable interest in the effect of CDKN2A and/or CDKN2B (CDKN2A/B) homozygous deletions (HD) on the prognosis of isocitrate dehydrogenase (IDH)-mutant astrocytomas. This is highlighted by the adoption of CDKN2A/B HD as an essential criterion for astrocytoma and IDH-mutant central nervous system (CNS) WHO grade 4 in the fifth edition of the World Health Organisation (WHO) Classification of Central Nervous System Tumours (2021). The CDKN2A and CDKN2B genes are located on the short arm of chromosome 9. CDKN2A encodes for two proteins, p14 and p16, and CDKN2B encodes for p15. These proteins regulate cell growth and angiogenesis. Interpreting the impact of CDKN2A/B alterations on astrocytoma prognosis is complicated by recent changes in tumour classification and a lack of uniform standards for testing CDKN2A/B. While the prognostic impact of CDKN2A/B HD is established, the role of different CDKN2A/B alterations-heterozygous deletions (HeD), point mutations, and promoter methylation-is less clear. Consequently, how these alternations should be incorporated into patient management remains controversial. To this end, we reviewed the literature on different CDKN2A/B alterations in IDH-mutant astrocytomas and their impact on diagnosis and management. We also provided a historical review of the changing impact of CDKN2A/B alterations as glioma classification has evolved over time. Through this historical context, we demonstrate that CDKN2A/B HD is an important negative prognostic marker in IDH-mutant astrocytomas; however, the historical data is challenging to interpret given changes in tumour classification over time, variation in the quality of evidence, and variations in the techniques used to identify CDKN2A/B deletions. Therefore, future prospective studies using uniform classification and detection techniques are required to improve the clinical interpretation of this molecular marker.

Borax regulates iron chaperone- and autophagy-mediated ferroptosis pathway in glioblastoma cells.

Glioblastoma (GBM) is classified as a stage-IV glioma. Unfortunately, there are currently no curative treatments for GBM. Poly(rC)-binding protein 1 (PCBP1) is a cytosolic iron chaperone with diverse functions. PCBP1 is also known to regulate autophagy, but the role of PCBP1 in ferroptosis, iron-dependent cell death pathway, remains unrevealed in GBM cells. Here, we investigated the effects of borax, a boron compound, on the ferroptosis signaling pathway mediated by PCBP1 and autophagy. The study analyzed cell viability, proliferation, and cell cycle on U87-MG and HMC3 cells to investigate the effects of borax. After determining the cytotoxic concentrations of borax, morphological analyzes and measurement of PCBP1, Beclin1, malondialdehyde (MDA), glutathione (GSH), glutathione peroxidase 4 (GPx4) and acyl-CoA synthetase long chain family member 4 (ACSL4) levels were performed. Finally, expression levels of PCBP1, Beclin1, GPx4 and ACSL4, and caspase-3/7 activity were determined. We found that borax reduced U87-MG cell viability in a concentration- and time-dependent manner. Additionally, borax altered cell proliferation and remarkably reduced S phase in the U87-MG cells and exhibited selectivity by having an opposite effect on normal cells (HMC3). According to DAPI staining, borax caused nuclear deficits in U87-MG cells. The result showed that borax in U87-MG cells induced reduction of the PCBP1, GSH, and GPx4 and enhancement of Beclin1, MDA, and ACSL4. Furthermore, borax triggered apoptosis by activating caspase 3/7 in U87-MG cells. Our study indicated that the borax has potential as an anticancer treatment for GBM via regulating PCBP1/Beclin1/GPx4/ACSL4 signaling pathways.

Molecular landscape of IDH-wild-type, H3-wild-type glioblastomas of adolescents and young adults.

OBJECTIVE: We aimed to characterise glioblastomas of adolescents and young adults (AYAs) that were isocitrate dehydrogenase (IDH) wild type (wt) and H3wt. MATERIALS AND METHODS: Fifty such patients (aged 16-32) were studied by methylation profiling, targeted sequencing and targeted RNA-seq. RESULTS: Tumours predominantly clustered into three methylation classes according to the terminology of Capper et al. (2018): (anaplastic) pleomorphic xanthoastrocytoma (PXA) (21 cases), GBM_midline (15 cases) and glioblastoma RTK/mesenchymal (seven cases). Two cases clustered with ANA_PA, four cases with LGG classes and one with GBM_MYCN. Only fifteen cases reached a calibrated score >0.84 when the cases were uploaded to DKFZ Classifier. GBM_midline-clustered tumours had a poorer overall survival (OS) compared with the PXA-clustered tumours (p = 0.030). LGG-clustered cases had a significantly better survival than GBM_midline-clustered tumours and glioblastoma RTK/mesenchymal-clustered tumours. Only 13/21 (62%) of PXA-clustered cases were BRAF V600E mutated. Most GBM_midline-clustered cases were not located in the midline. GBM_midline-clustered cases were characterised by PDGFRA amplification/mutation (73.3%), mutations of mismatch repair genes (40.0%), and all showed H3K27me3 and EZH1P loss, and an unmethylated MGMT promoter. Across the whole cohort, MGMT promoter methylation and wt TERT promoter were favourable prognosticators. Mismatch repair gene mutations were poor prognosticators and together with methylation class and MGMT methylation, maintained their significance in multivariate analyses. BRAF mutation was a good prognosticator in the PXA-clustered tumours. CONCLUSION: Methylation profiling is a useful tool in the diagnosis and prognostication of AYA glioblastomas, and the methylation classes have distinct molecular characteristics. The usual molecular diagnostic criteria for adult IDHwt glioblastoma should be applied with caution within the AYA age group.

Phenotypic characterization of human peripheral γδT-cell subsets in glioblastoma.

The antitumoral contribution of γδT cells depends on their activation and differentiation into effectors. This depends on different molecules and membrane receptors, which conditions their physiology. This study aimed to determine the phenotypic characteristics of γδT cells in glioblastoma (GBM) according to five layers of membrane receptors. Among ten GBM cases initially enrolled, five of them who had been confirmed by pathological examination and ten healthy controls underwent phenotyping of peripheral γδT cells by flow cytometry, using the following staining: αßTCR, γδTCR, CD3, CD4, CD8, CD16, CD25, CD27, CD28, CD45, CD45RA, CD56, NKG2D, CD272(BTLA), and CD279(PD-1). Compared with the controls, the results showed no significant change in the number of γδT cells. However, there was a decrease of double-negative (CD4- CD8- ) Tγδ cells and an increase of naive γδT cells, a lack of CD25 expression, a decrease of the expression of CD279, and a remarkable, but not significant, increase in the expression of the CD27 and CD28 costimulation markers. Among the γδT cell subsets, the number of Vδ2 decreased in glioblastoma and showed no significant difference in the expression of CD16, CD56, and NKG2D. In contrast, the number of Vδ1 increased in glioblastoma with overexpression of CD16, CD56, and NKG2D. Our results showed that γδT cells are prone to adopt a pro-inflammatory profile in the glioblastoma context, which suggests that they might be a potential tool to consider in T cell-based immunotherapy in glioblastoma. However, this requires additional investigation on a larger sample size.

Extracellular Vesicles Secreted by Glioma Stem Cells Are Involved in Radiation Resistance and Glioma Progression.

Glioblastoma is the most aggressive brain tumour with short survival, partly due to resistance to conventional therapy. Glioma stem cells (GSC) are likely to be involved in treatment resistance, by releasing extracellular vesicles (EVs) containing specific molecular cargoes. Here, we studied the EVs secreted by glioma stem cells (GSC-EVs) and their effects on radiation resistance and glioma progression. EVs were isolated from 3 GSCs by serial centrifugation. NanoSight measurement, cryo-electron microscopy and live imaging were used to study the EVs size, morphology and uptake, respectively. The non-GSC glioma cell lines LN229 and U118 were utilised as a recipient cell model. Wound healing assays were performed to detect cell migration. Colony formation, cell viability and invadopodium assays were conducted to detect cell survival of irradiated recipient cells and cell invasion post GSC-EV treatment. NanoString miRNA global profiling was used to select for the GSC-EVs' specific miRNAs. All three GSC cell lines secreted different amounts of EVs, and all expressed consistent levels of CD9 but different level of Alix, TSG101 and CD81. EVs were taken up by both LN229 and U118 recipient cells. In the presence of GSC-EVs, these recipient cells survived radiation exposure and initiated colony formation. After GSC-EVs exposure, LN229 and U118 cells exhibited an invasive phenotype, as indicated by an increase in cell migration. We also identified 25 highly expressed miRNAs in the GSC-EVs examined, and 8 of these miRNAs can target PTEN. It is likely that GSC-EVs and their specific miRNAs induced the phenotypic changes in the recipient cells due to the activation of the PTEN/Akt pathway. This study demonstrated that GSC-EVs have the potential to induce radiation resistance and modulate the tumour microenvironment to promote glioma progression. Future therapeutic studies should be designed to interfere with these GSC-EVs and their specific miRNAs.

Primary cerebellar glioblastomas in children: clinical presentation and management.

Pediatric cerebellar glioblastomas (pcGBMs) are rare and their characteristics remain ill-defined. We conducted a retrospective analysis of pediatric cerebellar glioblastomas who underwent surgery from 2008 to 2019 in our department. Besides, we performed a literature review of the literature data on pcGBMs. Ten children with mean age of 9.4 years were included. During the follow-up, six patients died with mean survival time of 11.7 months, four patients survived with mean follow-up of 28 months. Seven patients underwent molecular analysis, no patients detected IDH1 mutations, four patients (57.1%) had H3K27M mutations, and two patients (28.6%) had MGMT promoter methylation. The literature review identified 38 pcGBMs cases (including ours), with mean age of 8.84 ± 4.20 years (range, 1-16 years). Increased ICP was the commonest sign. Eighteen (47.4%) patients underwent GTR and fifteen (45.5%) patients received STR. Postoperative radiation (RT) was conducted in 28 patients (75.7%) and 23 patients (65.7%) received chemotherapy. During the follow-up, 25 patients died with mean survival time of 12.21 months and 11 patients survived with average follow-up of 29.3 months. Kaplan-Meier survival depicted chemotherapy (P < 0.001) or radiation (P < 0.001) had positive impact on overall survival. Multivariate analysis revealed chemotherapy was a significant predictor of survival with a hazard ratio of 3.264 (P = 0.038). Our study found mean overall survival time for pcGBMs patients was 12.21 months. PcGBMs may have distinct molecular features, with higher incidence of H3K27M mutation and were always IDH1 wild-type. We recommend the routine postoperative radiotherapy and chemotherapy in pcGBMs.

[Radiosurgery for recurrent glioblastoma]. / Radiokhirurgicheskoe lechenie retsidivov glioblastom.

BACKGROUND: Despite the combined treatment in accordance with modern standards, recurrent glioblastoma usually occurs within several months after resection and causes low relapse-free and overall survival. One of the most effective methods for malignant glioma progression is repeated radiotherapy. Indications for this approach have expanded after introduction of stereotactic irradiation into routine clinical practice. OBJECTIVE: To evaluate the results of radiosurgery in patients with recurrent glioblastoma and to identify the factors determining its effectiveness. MATERIAL AND METHODS: Radiosurgery has been carried out in 168 patients with relapses of glioblastoma between 2005 and 2021. This study enrolled 88 patients with 180 foci of local and distant progression. Mean age of patients was 42.8±2.1 years (range 4-73). Mean period between diagnosis and repeated irradiation was 12.7 months. Mean volume of focus was 2.4 cm3, mean dose - 20 Gy. Median follow-up period after radiosurgery was 11.2 months. RESULTS: Repeated irradiation with correction of systemic therapy improved progression-free survival and overall survival with insignificant radiation-induced toxicity. Annual overall survival was 62.2%, median of overall survival after radiosurgery - 15.1 months. Significant factors of local control were marginal dose of at least 18 Gy and distant relapse. Median of progression-free survival in the group of distant progression of glioblastoma was only 3.6 months vs. 9.1 months in patients with local recurrence. CONCLUSION: Repeated irradiation in radiosurgery mode with a dose of 18 Gy and higher is an effective option for local treatment increasing progression-free and overall survival in patients with progression of glioblastoma.