Secondary Glioblastomas Research Articles

Targeted alpha therapy for glioblastoma.

According to the 2021 World Health Organization Classification of Tumors of the Central Nervous System, glioblastoma (GB) is a primary brain tumor and presents with the worst prognosis. Due to its infiltrating characteristic, molecular heterogeneity, and only partly preserved function of the blood-brain barrier, the median overall survival time is short (9-15 months), regardless of comprehensive treatment including surgery, radiotherapy, and chemotherapy. Several novel treatment strategies are under investigation. Unfortunately, none of them produced successful results; 90% of patients have a recurrence of the disease within 6 months. Local administration of the drug could be a promising approach to delivering treatment with minimized side effects, due to the recurrence of 95% glioblastomas in a margin of 2 cm at the primary site. Several ligand-receptor systems have been evaluated, such as targeting tenascin, the extracellular matrix protein, or radiolabeled somatostatin analogs, as it is overexpressed with the SSTR-2 receptor system in around 80% of gliomas. Moreover, this study revealed that the NK-1 receptor is overexpressed in GB, suggesting that substance P (SP) may serve as a ligand. A variety of radioisotopes, beta- (131I, 90Y, or 177 Lu) and alpha emitters (213Bi, 225Ac, or 211At), with different physical properties were tested for treatment. Alpha particles have many advantages over beta radiation such as short range with higher linear energy transfer. According to that characteristic, it is extremely dose delivered to the targeted cells, while reducing harm to nearby healthy tissue. Additionally, the biological effect of alpha radiation is independent of the cell cycle phase, cell oxygenation and O-6-methylguanine-DNA methyltransferase (MGMT) gene promoter methylation status. In this article, we summarize the experience with local treatment of primary and secondary GBs with locally used radioisotopes such as [213Bi]Bi-DOTA-SP or [225Ac]Ac-DOTA-SP.

Detection of Isocitrate Dehydrogenase (IDH-1), Epidermal Growth Factor Receptor (EGFR), P53 and C-erbB2/HER2 Mutation in Glial Tumour

Introduction: In the last decade, several molecular pathways in gliomagenesis have been discovered, with each involving a unique set of molecular alterations. IDH1 has become a diagnostic tool in the latest 2016 WHO Classification. The tumour protein, p53, is involved in the IDH-mutant arm, observed in astrocytoma and oligodendroglioma (grades II and III), and secondary glioblastoma. Meanwhile, EGFR and c-erbB2/HER2 were postulated to be expressed in higher-grade glioma as the disease progresses. Methods: A retrospective cross-sectional study was conducted to evaluate the association of IDH1, EGFR, p53 and c-erbB2/HER2 protein expression in astrocytic and oligodendroglial tumours with clinicopathological data in HUSM, Kota Bharu, Kelantan, Malaysia. This study examined 61 archived formalin-fixed paraffin-embedded (FFPE) tissue blocks of patients diagnosed with glioma. The immunohistochemistry (IHC) test was performed using antibodies, IDH1, EGFR, p53 and c-erbB2/HER2, and the protein expressions were evaluated microscopically. Finally, the association between IDH1, p53, EGFR and c-erbB2/HER with the clinicopathology variables were statistically analysed. Results: A total of 61 glioma cases consisting of 36 (59%) males and 25 (41%) females were included in this study. The IDH1 protein was positively expressed in 14 cases (23%), P53 was highly expressed in 26 cases (42.6%), and EGFR was substantially observed in 34 cases (55.7%). For glioblastoma cases, IDH1 was expressed in two cases (11.1%), EGFR in 14 cases (77.7%), p53 in 12 cases (66.7%) and c-erbB2 in 1 case (5.6%). Significant associations exist between IDH1, p53 and EGFR expressions in astrocytoma and oligodendroglial tumours with the histological types and WHO tumour grades. Conclusion: Recently, our knowledge regarding the genetics of central nervous system (CNS) tumors has expanded; hence, newer antibodies or molecular markers, which can be used in IHC, are continuously being developed. These antibodies, IDH1, p53 and EGFR markers are useful for diagnostic, prognostication and therapeutic. In addition, help to clarify the nature of cellular maturation, tissue differentiation, and tumor progression to be considered as an integral part of WHO classification of CNS tumours.

MET fusions and splicing variants convergently define a subgroup of glioma sensitive to MET inhibitors

PurposeOur previous study has shown that PTPRZ1-MET (ZM) fusion is a viable target for MET inhibitors in gliomas. However, the diversity and prevalence of somatic MET alterations in diffuse gliomas are still elusive and need to be extensively characterized for identifying novel therapeutic targets.MethodsTotally, 1,350 glioma patients and 31 patient-derived cells were collected from the Chinese Glioma Genome Atlas (CGGA) and published data. All kinds of MET fusions and/or splicing variants (MET F/SVs) were identified by bioinformatical methods. Single-cell RNA sequencing (scRNA-seq) were used for validation. In vitro experiments of drug resistance were conducted for the possibility of MET-targeted treatment.ResultsMET F/SVs but not genomic amplification, were highly enriched in the secondary glioblastomas (sGBM) and marked worse prognosis. Further molecular and scRNA-seq analysis revealed that MET F/SVs were induced in the course of glioma evolution and highly associated with MET overexpression. Subsequent in vitro and the clinical study showed that cells and patients harboring MET F/SVs have better response to MET inhibitors.ConclusionOur findings expanded the percentage of gliomas with abnormal MET alterations and suggested that a subgroup of gliomas harboring MET F/SVs may benefit from MET-targeted therapy.

Development of a sensitive, specific and cost-effective T-ARMS PCR assay for the genotyping of R132H of IDH1 gene in glioma patients

The discovery of isocitrate dehydrogenase isoform 1 (IDH1) mutation as a key molecular marker has resulted in a change in glial tumour classification [1]. IDH1 mutations are commonly in gliomas, particularly in low-grade gliomas and secondary glioblastoma [2]. IDH1 p.R132H (c.395G>A) accounted for more than 90% of the mutation in IDH1/2 mutation and had a significant association with clinical outcomes. IDH1/2 mutations cause gain-of-function resulting in the formation of an oncometabolite, R-2-hydroxyglutarate instead of α-ketoglutarate implying a disruption of oxidative decarboxylation of Kreb’s cycle and other cellular mechanisms [3,4,5]. Immunohistochemistry (IHC) and Sanger sequencing is the most common approach used in molecular diagnostics. Nevertheless, both IHC and Sanger sequencing methods have shortcomings. IHC is prone to miss out on the mutation if the tumour samples are of poor quality, and it has also been reported to have low sensitivity when compared to sequencing-based techniques [6,7].
 
 This study aimed to develop a sensitive, specific and cost-effective assay for genotyping IDH1 p.R132H mutations in glioma patients in order to shorten the time required for confirmatory diagnosis to be made. The tetra primer amplification refractory mutation system polymerase chain reaction (T-ARMS PCR) was used in this study to develop and validate the clinical applicability of the assay. A total of 61 glial specimens were collected and genomic DNA was isolated from all of them. All the samples were subjected to endpoint PCR and Sanger sequencing for mutation detection. T-ARMS PCR was developed and optimized prior to the screening of all the samples, and comparative mutation analysis was carried out.
 Overall, IDH1 p.R132H mutation was found to be 45.90% (n=28/61) prevalent and was found to be significantly associated with gender, tumour subtypes and grading, and location of the lesions (p=<0.05). With an F1 score of 0.966, we reported T-ARMS PCR with sensitivity, specificity and accuracy of 100% (95% CI: 87.94-100.00%), 93.94% (95% CI: 80.39-98.92%) and 96.72%, respectively. To compare with published studies, a meta-analysis is T-ARMS PCR detected one case with IDH1 p.R132C (c.394C>T), howbeit the case could have a double mutation of p.R132C (c.394C>T) and p.R132H (c.395G>A) or single mutation of p.R132Y (c.394_395CG>TA). However, this study was able to detect IDH1 p.R132G (c.394C>G) mutation via PCR by Sanger sequencing whereas T-ARMS PCR excluded the mutation, suggesting the assay is very specific to IDH1 p.R132HH (c.395G>A) only.
 Hereby, the performance of the T-ARMS PCR assay sheds a light that can be adapted for preoperative or intraoperative diagnosis.

Effects of cancer-associated point mutations on the structure, function, and stability of isocitrate dehydrogenase 2

Mutations in isocitrate dehydrogenase (IDH) are frequently found in low-grade gliomas, secondary glioblastoma, chondrosarcoma, acute myeloid leukemias, and intrahepatic cholangiocarcinoma. However, the molecular mechanisms of how IDH2 mutations induce carcinogenesis remain unclear. Using overlapping PCR, transfection, immunoblotting, immunoprecipitation, measurements of enzyme activity, glucose, lactic acid, ATP, and reactive oxygen species (ROS), cell viability, protein degradation assays post-inhibition of the 26S proteasome (bortezomib) or HSP90 (17-AAG), and a homology model, we demonstrated that the properties of ten cancer-associated IDH2 variants (R140G/Q/W and R172S/K/M/W/G/C/P) arising from point mutations are closely related to their structure and stability. Compared with wild-type IDH2, the R172 and R140 point mutations resulted in a decrease in IDH2 activity, ROS, and lactate levels and an increase in glucose and ATP levels under normal and hypoxic conditions, indicating that mutant IDH2 increases cell dependency on mitochondrial oxidative phosphorylation, and reduces glycolysis under hypoxia. Overexpression of most of IDH2 point mutants showed anti-proliferative effects in the 293T and BV2 cell lines by inhibition of PI3K/AKT signaling and cyclin D1 expression and/or induced the expression of TNF-α and IL-6. Furthermore, bortezomib treatment resulted in dramatic degradation of IDH2 mutants, including R140G, R140Q, R140W, R172S and R172K, whereas it had little impact on the expression of WT and other mutants (R172M, R172W, R172G, R172C and R172P). In addition, targeting HSP90 minimally affected the expression of mutated IDH2 due to a lack of interaction between HSP90 and IDH2. The homology model further revealed that changes in conformation and IDH2 protein stability appeared to be associated with these point mutations. Taken together, our findings provide information important for understanding the molecular mechanisms of IDH2 mutations in tumors.

Meningeosis Neoplastica in Patients with Glioblastoma: Analysis of 36 Cases

Meningeosis neoplastica is a rare manifestation of high-grade gliomas and is usually associated with a devastating outcome. The aim of this bicenter series was to investigate the clinical course and outcome of patients with meningiosis neoplastica. This case series included patients in whom surgery was performed for World Health Organization grade IV primary and secondary glioblastoma (GBM) at the University Medical Center Göttingen, Göttingen, Germany between 2009 and 2021 and Burdenko Institute of Neurosurgery, Moscow, Russia between 2012 and 2018. Inclusion criteria were manifestation of clinical and neuroradiologic signs of leptomeningeal, ependymal, or spinal dissemination of GBM at various time points during the course of the disease. Meningeosis neoplastica was found in 36 patients. Nine patients developed spinal metastases and 12 ependymal dissemination and 15 patients had a leptomeningeal manifestation of high-grade glioma. The median age of patients at first diagnosis of primary tumor was 56 years. Typical symptoms were headache, nausea, vomiting, and acute paraplegia. The median overall survival was 11 months and progression-free survival was 8 months. Meningeosis neoplastica developed a median 2 months after the initial tumor diagnosis. Salvage therapies included ventriculoperitoneal shunting, decompression of spinal metastases, and spinal radiation therapy. The median time between meningeosis manifestation and death was 3 months. Meningeosis neoplastica is a rare manifestation of GBM. It has a poor prognosis. The overall survival after the manifestation of meningeosis was barely longer than 3 months. Salvage therapies did not improve the outcome in our patient cohort.

Recent findings on molecular alterations in IDH1, TP53, and CASP9 in gliomagenesis

Isocitrate dehydrogenase (IDH) mutations have been the focus of glioma-related neuroscience research since the discovery of the gene in 2008. IDH1 has been identified as a key enzyme in cellular metabolism, epigenetic regulation, redox regulation, and DNA repair and is thought to be one of the most important factors in gliomagenesis. IDH1 mutations cause neomorphic activity, resulting in an increase in 2-hydroxyglutarate production and a decrease in NADPH production. Emerging research has identified IDH1 mutations in the vast majority of low-grade gliomas and secondary glioblastomas, but these mutations are extremely uncommon in primary glioblastomas. Other genetic defects appear to play a significant role in glioma initiation and progression. In this study, we review recent findings on oncogenic alterations in IDH1, TP53, and CASP9 to identify any potential molecular correlations and interrelationships that lead to gliomagenesis. The roles and molecular interactions of these glioma-associated genes in gliomagenesis are elucidated. In addition, we highlight studies on stem cell modeling in glioma-associated genetic alterations that have been conducted over the past several decades.

P10.22.A Prognostic role ofTCF7L2 in glioblastoma

Abstract Background Glioblastoma is the most common and devastating brain cancer and despite maximal surgical resection, chemotherapy and radiotherapy, the five-year survival rate is less than 7%. The TCF7L2 gene encodes transcription factor 7-like 2, a main effector of Wnt/β-Catenin signaling pathway involved in cancer development and progression. Aberrant activation of the TCF7L2 gene is one of the primary events in tumorigenesis by maintaining stem-cell characteristics, however its role in glioblastoma is poorly studied. Material and Methods The expression of TCF7L2 was analyzed by qRT-PCR in a series of consecutive glioblastoma samples from patients that underwent surgical resection. Secondary or recurrent glioblastomas were excluded from the analysis. All patients underwent radio- and chemotherapy according to the protocol. Housekeeping gene RPL13 was used as reference gene. The 2-ΔCT method was used to calculate the relative expression of TCF7L2 to the reference gene. Results A total of 49 glioblastoma samples were included in the study. The mean survival period was 58.2 weeks. The median value of TCF7L2 expression was used as a cut-off point for the survival analysis. There were no statistically significant differences regarding the clinical and histopathological factors between the groups with low and high expression of TCF7L2. By using the Kaplan-Meier analysis, we observed that patients with overexpression of TCF7L2 had a decreased survival rate compared to patients with low levels of TCF7L2 (38.8 weeks vs. 77.34 weeks, p = 0.001). Conclusion These results demonstrate that glioblastoma patients with high levels of TCF7L2 have a poorer survival rate and TCF7L2 might be a useful prognostic factor for these patients. However, further studies are needed to elucidate the mechanisms through which TCF7L2 affects the prognosis and its implication as a possible therapeutic target. This work was supported by an ERA PerMed grant - ERAPERMED2018-270 (PerProGlio).

CNPY4 is a potential promising prognostic-related biomarker and correlated with immune infiltrates in gliomas.

Glioblastomas are classified into primary and secondary; primary glioblastomas develop rapidly and aggressively, whereas secondary glioblastomas are more common in grade II and III gliomas. Here, we aimed to demonstrate the role of the CNPY4 gene as a potential biomarker in immune infiltration in gliomas. Based on gene expression profile interaction analysis (GEPIA), we studied the survival model of CNPY4 and evaluated its effect on patients with glioma. The glioma dataset was downloaded from The Cancer Genome Atlas (TCGA) database. Logistic regression was used to analyze the relationship between clinical data and CNPY4 expression. Univariate and multivariate Cox proportional-hazards models were used to compare clinical features and patient survival. The relationship between CNPY4 and immune infiltration in glioma was studied using GEPIA and CIBERSORT online tools. TCGA data were analyzed using gene set enrichment analysis (GSEA). Finally, TIMER was used to analyze the expression and immune infiltration of CNPY4 in glioma to study the cumulative survival rate. Univariate logistic regression analysis showed that increased CNPY4 expression was associated with tumor age, grade, IDH status, and 1p/19q codeletion. Multivariate analysis showed that that downregulation of CNPY4 expression was an independent and satisfactory prognostic factor. CNPY4 expression was correlated with the infiltration level of dendritic cells in glioblastoma. In contrast, in low-grade gliomas, the infiltration level of B cells, dendritic cells, macrophages, neutrophils, and CD4+ T cells was significantly correlated with CNPY4 expression. The GSEA results showed that CNPY4 played an immunoregulatory role in immune-related phenotypic pathways between lymphoid and nonlymphoid cells. The intestinal immune networks for IgA production, rabbit thyroid disease, primary immunodeficiencies, and cancer immunotherapy were enriched by PD-1 blockade. High CNPY4 expression is a biomarker of glioma prognosis and is associated with the immune invasion of glioma.

Looking Beyond the Glioblastoma Mask: Is Genomics the Right Path?

Glioblastomas are the most frequent and malignant brain tumor hallmarked by an invariably poor prognosis. They have been classically differentiated into primary isocitrate dehydrogenase 1 or 2 (IDH1 -2) wild-type (wt) glioblastoma (GBM) and secondary IDH mutant GBM, with IDH wt GBMs being commonly associated with older age and poor prognosis. Recently, genetic analyses have been integrated with epigenetic investigations, strongly implementing typing and subtyping of brain tumors, including GBMs, and leading to the new WHO 2021 classification. GBM genomic and epigenomic profile influences evolution, resistance, and therapeutic responses. However, differently from other tumors, there is a wide gap between the refined GBM profiling and the limited therapeutic opportunities. In addition, the different oncogenes and tumor suppressor genes involved in glial cell transformation, the heterogeneous nature of cancer, and the restricted access of drugs due to the blood–brain barrier have limited clinical advancements. This review will summarize the more relevant genetic alterations found in GBMs and highlight their potential role as potential therapeutic targets.

Diagnostic algorithm for pathological evaluation of gliomas in a resource-constrained setting.

Gliomas are the most common primary intracranial tumors. The current World Health Organization (WHO) classification of central nervous system tumors recommends integrated histo-molecular diagnosis of gliomas. However, molecular testing is not available in even most of the advanced centers of our country, and histopathology aided with immunohistochemistry (IHC) is still widely used for diagnosis. Immunohistochemical markers such as iso-citrate dehydrogenase1 (IDH1) and Alpha Thalassemia/Mental Retardation Syndrome X-linked (ATRX) can be reliably used for the correct diagnosis, prognosis, and treatment of gliomas. We aimed to develop a diagnostic algorithm by integrating morphology, IDH1, and ATRX status of gliomas seen in our institute for 1 year. Analytical cross-sectional study. This study included 60 histopathologically confirmed cases of astrocytic (n = 51) and oligodendroglial tumors (n = 9). Clinical, radiological, and histopathological features were noted and tumor grades assigned according to the WHO recommendations. IDH1 and ATRX mutation status was evaluated using IHC. The tumors were divided into three molecular groups on the basis of their IDH1 and ATRX mutation status: (1) Group 1: IDH1 negative and ATRX positive, (2) Group 2: IDH1 positive and ATRX positive, (3) Group 3: IDH1 positive and ATRX negative. The mean age of presentation was 45.0 ± 15.8 years with a male-to-female ratio of 2:1. Seizures, headache, and hemiparesis were the most common modes of presentation. The tumor subtypes studied were glioblastoma (n = 32), anaplastic astrocytoma (n = 7), diffuse astrocytoma (n = 6), oligodendroglioma (n = 6), pilocytic astrocytoma (n = 6), and anaplastic oligodendroglioma (n = 3). IDH1 mutation was present in 26 cases including anaplastic astrocytoma (n = 7), diffuse astrocytoma (n = 6), oligodendroglioma (n = 5), secondary glioblastoma (n = 5), and anaplastic oligodendroglioma (n = 3). ATRX mutation, i. e., loss of ATRX was observed in 17 cases including diffuse astrocytoma (n = 5), anaplastic astocytoma (n = 5), anaplastic oligodendroglioma (n = 3), oligodendroglioma (n = 3), and secondary glioblastoma (n = 1). All six cases of pilocytic astrocytoma were negative for IDH1 and ATRX mutation. There were 34 patients in Group 1 (IDH1- and ATRX +), nine cases in Group 2 (IDH1 + and ATRX +), and 17 patients in Group 3 (IDH1 + and ATRX-). Diagnosis of gliomas should be based on a detailed clinicoradiological and histopathological assessment, followed by genotypic characterization. Evaluation for IDH1and ATRX status has both diagnostic and prognostic value as it helps in differentiating gliomas from reactive gliosis, primary glioblastoma from secondary glioblastoma, and pilocytic astrocytoma (WHO grade I) from diffuse astrocytoma (WHO grade II). Tumors with IDH1 mutations have a better outcome than those with wild-type IDH. IHC can serve as a useful surrogate to conventional molecular tests in resource-constrained settings. By devising an algorithm based on morphological and IHC features, we were able to stratify gliomas into three prognostic subgroups.

Recurrence- and Malignant Progression-Associated Biomarkers in Low-Grade Gliomas and Their Roles in Immunotherapy.

Despite a generally better prognosis than high-grade glioma (HGG), recurrence and malignant progression are the main causes for the poor prognosis and difficulties in the treatment of low-grade glioma (LGG). It is of great importance to learn about the risk factors and underlying mechanisms of LGG recurrence and progression. In this study, the transcriptome characteristics of four groups, namely, normal brain tissue and recurrent LGG (rLGG), normal brain tissue and secondary glioblastoma (sGBM), primary LGG (pLGG) and rLGG, and pLGG and sGBM, were compared using Chinese Glioma Genome Atlas (CGGA) and Genotype-Tissue Expression Project (GTEx) databases. In this study, 296 downregulated and 396 upregulated differentially expressed genes (DEGs) with high consensus were screened out. Univariate Cox regression analysis of data from The Cancer Genome Atlas (TCGA) yielded 86 prognostically relevant DEGs; a prognostic prediction model based on five key genes (HOXA1, KIF18A, FAM133A, HGF, and MN1) was established using the least absolute shrinkage and selection operator (LASSO) regression dimensionality reduction and multivariate Cox regression analysis. LGG was divided into high- and low-risk groups using this prediction model. Gene Set Enrichment Analysis (GSEA) revealed that signaling pathway differences in the high- and low-risk groups were mainly seen in tumor immune regulation and DNA damage-related cell cycle checkpoints. Furthermore, the infiltration of immune cells in the high- and low-risk groups was analyzed, which indicated a stronger infiltration of immune cells in the high-risk group than that in the low-risk group, suggesting that an immune microenvironment more conducive to tumor growth emerged due to the interaction between tumor and immune cells. The tumor mutational burden and tumor methylation burden in the high- and low-risk groups were also analyzed, which indicated higher gene mutation burden and lower DNA methylation level in the high-risk group, suggesting that with the accumulation of genomic mutations and epigenetic changes, tumor cells continued to evolve and led to the progression of LGG to HGG. Finally, the value of potential therapeutic targets for the five key genes was analyzed, and findings demonstrated that KIF18A was the gene most likely to be a potential therapeutic target. In conclusion, the prediction model based on these five key genes can better identify the high- and low-risk groups of LGG and lay a solid foundation for evaluating the risk of LGG recurrence and malignant progression.

The Impact of Resistance Exercise on Muscle Mass in Glioblastoma in Survivors (RESIST): Protocol for a Randomized Controlled Trial.

BackgroundGlioblastoma is the most common primary brain malignancy in adults, accounting for approximately 48% of all brain tumors. Standard treatment includes radiation and temozolomide chemotherapy. Glioblastomas are highly vascular and can cause vasogenic brain edema and mass effect, which can worsen the neurologic symptoms associated with the disease. The steroid dexamethasone (DEX) is the treatment of choice to reduce vasogenic edema and intracranial pressure associated with glioblastoma. However high-dose DEX or long-term use can result in muscle myopathy in 10%-60% of glioblastoma patients, significantly reducing functional fitness and quality of life (QOL). There is a wealth of evidence to support the use of exercise as an adjuvant therapy to improve functional ability as well as help manage treatment-related symptoms. Specifically, resistance training has been shown to increase muscle mass, strength, and functional fitness in aging adults and several cancer populations. Although studies are limited, research has shown that exercise is safe and feasible in glioblastoma populations. However, it is not clear whether resistance training can be successfully used in glioblastoma to prevent or mitigate steroid-induced muscle myopathy and associated loss of function.ObjectiveThe primary purpose of this study is to establish whether an individualized circuit-based program will reduce steroid-induced muscle myopathy, as indicated by maintained or improved functional fitness for patients on active treatment and receiving steroids.MethodsThis is a 2-armed, randomized controlled trial with repeated measures. We will recruit 38 adult (≥18 years) patients diagnosed with either primary or secondary glioblastoma who are scheduled to receive standard radiation and concurrent and adjuvant temozolomide chemotherapy postsurgical debulking and received any dose of DEX through the neurooncology clinic and the Nova Scotia Health Cancer Center. Patients will be randomly allocated to a standard of care waitlist control group or standard of care + circuit-based resistance training exercise group. The exercise group will receive a 12-week individualized, group and home-based exercise program. The control group will be advised to maintain an active lifestyle. The primary outcome, muscle myopathy (functional fitness), will be assessed using the Short Physical Performance Battery and hand grip strength. Secondary outcome measures will include body composition, cardiorespiratory fitness, physical activity, QOL, fatigue, and cognitive function. All measures will be assessed pre- and postintervention. Participant accrual, exercise adherence, and safety will be assessed throughout the study.ResultsThis study has been funded by the Canadian Cancer Society Atlantic Cancer Research Grant and the J.D. Irving Limited–Excellence in Cancer Research Fund (grant number 707182). The protocol was approved by the Nova Scotia Health and Acadia University’s Research Ethics Boards. Enrollment is anticipated to begin in March 2022.ConclusionsThis study will inform how individualized circuit-based resistance training may improve functional independence and overall QOL of glioblastoma patients.Trial RegistrationClinicalTrails.gov NCT05116137; https://www.clinicaltrials.gov/ct2/show/NCT05116137International Registered Report Identifier (IRRID)DERR1-10.2196/37709

Synergistic anti-tumor efficacy of mutant isocitrate dehydrogenase 1 inhibitor SYC-435 with standard therapy in patient-derived xenograft mouse models of glioma

Clinical outcomes in patients with WHO grade II/III astrocytoma, oligodendroglioma or secondary glioblastoma remain poor. Isocitrate dehydrogenase 1 (IDH1) is mutated in > 70% of these tumors, making it an attractive therapeutic target. To determine the efficacy of our newly developed mutant IDH1 inhibitor, SYC-435 (1-hydroxypyridin-2-one), we treated orthotopic glioma xenograft model (IC-BT142AOA) carrying R132H mutation and our newly established orthotopic patient-derived xenograft (PDX) model of recurrent anaplastic oligoastrocytoma (IC-V0914AOA) bearing R132C mutation. In addition to suppressing IDH1 mutant cell proliferation in vitro, SYC-435 (15 mg/kg, daily x 28 days) synergistically prolonged animal survival times with standard therapies (Temozolomide + fractionated radiation) mediated by reduction of H3K4/H3K9 methylation and expression of mitochondrial DNA (mtDNA)-encoded molecules. Furthermore, RNA-seq of the remnant tumors identified genes (MYO1F, CTC1 and BCL9) and pathways (base excision repair, TCA cycle II, sirtuin signaling, protein kinase A, eukaryotic initiation factor 2 and α-adrenergic signaling) as mediators of therapy resistance. Our data demonstrated the efficacy SYC-435 in targeting IDH1 mutant gliomas when combined with standard therapy and identified a novel set of genes that should be prioritized for future studies to overcome SYC-435 resistance.

In vivo efficacy of decitabine as a natural killer cell-mediated immunotherapy against isocitrate dehydrogenase mutant gliomas.

Isocitrate dehydrogenase (IDH) mutations are found in more than 80% of low-grade gliomas and in the majority of secondary glioblastomas. IDH mutation (IDHmut) leads to aberrant production of an oncogenic metabolite that promotes epigenetic dysregulation by inducing hypermethylation to suppress transcription of various tumor suppressor genes. Hypermethylation in IDHmut gliomas leads to transcriptional repression of NKG2D ligands, especially UL16-binding protein (ULBP)-1 and ULBP-3, and subsequent evasion of natural killer (NK) cell-mediated lysis. The demethylating agent 5-aza-2'deoxycytodine (decitabine [DAC]) is a DNA methyltransferase 1 inhibitor that prevents hypermethylation and is capable of restoring NKG2D ligand expression in IDHmut gliomas to resensitize them to NK cells. Given its capacity for sustained epigenetic reprogramming, the authors hypothesized that DCA would be an effective immunotherapeutic agent in treating IDHmut gliomas in an NK cell-dependent manner by upregulating epigenetically repressed activating NKG2D ligands in IDHmut tumors. In this study, the authors sought to use a glioma stem cell, preclinical animal model to determine the efficacy of DAC in IDHmut and IDH wild-type (IDHwt) tumors, and to characterize whether the activity of DAC in gliomas is dependent on NK cell function. Xenograft models of IDHwt and IDHmut gliomas were established in athymic-nude mice. When tumors were grossly visible and palpable, mice were treated with either DCA or dimethylsulfoxide intraperitoneally every 7 days. Tumor sizes were measured every 2 to 3 days. After the animals were euthanized, xenografts were harvested and analyzed for the following: tumor expression of NKG2D ligands, tumor susceptibility to human and murine NK cells, immunohistochemistry for NK infiltration, and tumor-infiltrating lymphocyte characterization. DAC significantly inhibited the growth of IDHmut xenografts in the athymic nude mice. This effect was abrogated with NK cell depletion. Ex vivo analysis of tumor cells from harvested xenografts confirmed that DAC increased NKG2D ligand ULBP-1 and ULBP-3 expressions, and enhanced susceptibility to lysis of both human and murine IDHmut glial cells with corresponding NK cells. Immunohistochemical analysis of the xenografts indicated that DCA-treated IDHmut gliomas had a greater level of NK infiltration into the tumor compared with the negative control. Finally, DCA radically altered the tumor-infiltrating lymphocyte landscape of IDHmut glioma xenografts by increasing NK cells, dendritic cells, and M1 macrophages, while decreasing suppressive monocyte infiltration. DCA displayed novel immunotherapeutic functions in IDHmut gliomas. This effect was critically dependent on NK cells. Additionally, DCA significantly altered the tumor immune landscape in IDHmut gliomas from suppressive to proinflammatory.

Decreased Expression of Metastasis Suppressor 1 Correlates with Poor Prognosis of Patients with Rectal Adenocarcinoma

Background: Venous thromboembolic events (VTE) are common causes of morbidity and mortality in glioblastoma patients. Mutation in the isocitrate dehydrogenase 1 enzyme (IDH1) is frequent in secondary glioblastoma and results in altered metabolomics. Objectives: This study evaluates whether IDH-1 status correlates with incidence of VTE in glioblastoma patients. Methods: Observational study of 398 cases of patients with glioblastoma, who all underwent surgery in a regional Neurosurgical centre between April 2012 and December 2014. IDH -1 status and Tissue factor (F3) protein expression were assessed by immunohistochemistry. Deep venous thrombosis (DVT) and pulmonary embolism (PE) were diagnosed by Doppler ultrasound and pulmonary CT angiogram respectively. Results: 336 cases were wild type (WT) IDH-1 (94.1%) and 21 cases were IDH-1 mutated (R132H) (5.9%). 51 patients had a thromboembolic event (15.3%), with all cases of VTE in WT IDH-1 tumors, a rate of 21.8% within this group. IDH-1 status had a significant correlation with VTE (p=0.033 Fisher exact test). As expected, mutant IDH was associated with prolonged patient survival (p=0.024 Log rank). The mean expression in IDH-1 wild type GBM was 7.14 and in R132h mutant GBM was 4.87 (log2 scale). This was highly statistically significant with a corrected P value of less than 0.0001. Conclusion: A significant association exists between IDH1 status in glioblastoma patients and the risk of VTE. Patients with wild type IDH-1 appear at high risk of VTE and appropriate precautions should be considered.

TMOD-18. DIRECT IN VIVO CRISPR SCREEN IDENTIFIES COOPERATING TUMOR SUPPRESSORS THAT DRIVE PROGRESSION OF IDH1-MUTANT LOW-GRADE GLIOMA TO AGGRESSIVE GLIOBLASTOMA

Abstract Low-grade glioma (LGG) are generally slowly growing brain cancers, that frequently undergo malignant progression to aggressive, secondary glioblastoma with a dismal prognosis. By combining genetically engineered Idh1-mutant mice with in vivo CRISPR gene editing we generated a mouse model faithfully recapitulating the founder mutations of LGG. Clonal activation of the neomorphic Idh1 R132H mutation cooperates with Trp53 and Atrx mutations to trigger development of brain tumors but only with ~30% penetrance and very long latency. To elucidate the molecular mechanisms underlying the malignant progression of IDH1-mutant LGG, we devised and deployed a direct in vivo CRISPR screen targeting genes commonly mutated in human IDH-mutant secondary glioblastoma. Stereotaxic delivery of a lentiviral sgRNA library targeting the mouse orthologs of these genes into the brain of Idh1 R132H ;Trp53;Atrx;Cas9 and control Idh1 wt ;Trp53;Atrx;Cas9 compound mutant mice resulted in rapid formation of tumors that recapitulate human Idh1-mutant glioblastoma. Deconvoluting the screen showed that PI3K pathway members Pten and Pik3ca as well as Notch1, Smarca4 and Fat1 are preferentially enriched in Idh1 R132H-tumors, while Rb1 and NF2 were enriched in Idh1 wt tumors. Co-mutation analysis further identified additional co-occurring driver combinations such as Bcor-Met, Olig2-Met, Olig2-Med12 or Bcor-Olig2. We validated the tumor suppressive function of Notch1 and Pten using conventional floxed knock-out alleles and found that Notch1 functions in a haploinsufficient manner. Interestingly, Idh1 R132H did not alter tumor latency or pathology in a high grade p53;Pten;Rb1 mutant background, indicating that the neomorphic IDH-mutations can drive low but not high grade glioma development. Our study provides a functional landscape of gliomagenesis suppressors in vivo.

BIOM-17. DIFFERENCES IN THE IMMUNE MICROENVIRONMENT OF GLIOMAS HARBORING IDH2 VERSUS IDH1 MUTATIONS

Abstract INTRODUCTION IDH mutations are a defining feature of lower-grade glioma and secondary glioblastoma. Approximately 95% of glioma-associated IDH mutations are in codon 132 of IDH1, but a small proportion are in IDH2. IDH mutations produce the oncometabolite 2-hydroxyglutarate, which induces global DNA hypermethylation and is associated with an immunosuppressive tumor microenvironment. IDH1 is localized in the cytosol while IDH2 is found in the mitochondrial matrix, and mutations in these genes may have differing effects on the tumor microenvironment. METHODS Formalin-fixed, paraffin-embedded tissue from 633 IDH-mutant gliomas (615 IDH1-mutant, 18 IDH2-mutant) underwent whole-exome and whole-transcriptome sequencing at Caris Life Sciences (236 grade 2/3 astrocytoma, 158 grade 2/3 oligodendroglioma, 202 IDH-mutant glioblastoma, 37 glioma, NOS). QuantiSEQ was used to infer tumor-infiltrating immune cell populations from RNAseq data, and gene-set enrichment analyses (GSEA) were performed using Wikipathway. RESULTS IDH1-mutant gliomas had higher levels of pro-inflammatory M1 macrophages (P=0.04), modestly higher levels of monocytes (P=0.08), and lower levels of neutrophils (P=0.04) – typically considered immunosuppressive – compared with IDH2-mutant gliomas. No differences were observed in levels of B cells, dendritic cells, NK cells, or T cell subsets (Treg, CD4+, CD8+). IDH2-mutant gliomas were enriched for hallmark oligodendroglioma mutations (TERT promoter, CIC, FUBP1), while IDH1-mutant gliomas were enriched for hallmark astrocytoma mutations (ATRX, TP53). However, associations with tumor-infiltrating immune cells persisted after excluding 1p/19q co-deleted oligodendroglioma from analyses. GSEA revealed upregulation of the microglial TYROBP signaling pathway, the microglial phagocytic pathway, and of Type II Interferon signaling in IDH1-mutant gliomas versus IDH2-mutant gliomas. CONCLUSIONS Although IDH2 mutations are generally thought to function similarly to IDH1 mutations, we observe differences in tumor-infiltrating immune cells across groups. IDH2-mutant gliomas appeared to have a more immunosuppressive tumor microenvironment than their IDH1-mutant counterparts. Early-phase immunotherapy trials should consider covariate-adaptive randomization approaches to equally allocate IDH2-mutant gliomas across treatment arms.

Cerebral Glioblastoma: A Review on Genetic Alterations, Signaling Pathways, and Clinical Managements

Context: Glioblastoma, previously known as glioblastoma multiforme (GBM), is a grade IV astrocytoma that frequently affects patients with a mean age of 45 and above. It is generally categorized into primary and secondary subtypes, based on research conducted by Hans Joachim Scherer. Evidence Acquisition: This review concentrates on cellular and genetic drawbacks that can lead to the appearance of glioblastoma. National Center for Biotechnology Information (NCBI) was the main source used for writing this review article, followed by Google scholar. The following keywords were used to retrieve articles: 'glioblastoma', 'brain tumors', 'glioma', 'LOH', and 'cellular and signaling pathways in glioblastoma'. Results: Several genetic alterations and cellular pathways are involved in the appearance and progression of glioblastoma, including loss of heterozygosity (LoH), TP53 mutation, isocitrate dehydrogenase 1 (IDH1) mutation, P16INK4/RB1 pathway, and EGFR/PTEN/Akt/mTOR pathway. The majority (70%) of primary glioblastomas are caused by (LoH), and it mostly occurs in older people. Secondary glioblastoma is mainly manifested by TP53 mutation and usually affects younger people. Understanding the alterations and cellular mechanisms involved in glioblastoma is important to develope new therapeutic regimes. Surgery, radiation therapy, temozolomide, and TTFields are the four most important therapeutic options available for treating patients. Conclusions: In this review, the genetic alterations and cellular pathways which could lead to the appearance of this tumor were highlighted, and the latest options for treating patients dealing with glioblastoma were discussed.

Targeting gene fusions in glioma.

Glioma represents of variety of brain malignancies, the majority of which confer a poor prognosis despite treatment. With the widespread use of next-generation sequencing, gene fusions are being found in greater numbers. Gene fusions in glioma represent an opportunity to deliver targeted therapies to those with limited options for treatment. Extensive studies on these gene fusions have shown that they can exhibit distinct phenotypes, such as PTPRZ1-MET fusions in secondary glioblastoma or FGFR3-TACC3 fusions in IDH wildtype gliomas. Responses have been observed with the use of targeted therapies but some have been short lived because of the development of treatment resistance. Increasing detection of gene fusions in glioma along with basket trials have helped define different fusion phenotypes and paved the way for targeted kinase inhibitor-based therapies. Targeting NTRK fusions has been the most successful fusion-guided therapy to date and evaluating all patients for these fusions may be warranted.