IDH-wildtype Astrocytomas Research Articles

BIOM-23. IDENTIFICATION OF NOVEL BIOMARKERS FOR BRAIN TUMOR USING PRECISION MEDICINE APPROACHES: ORC6 PLAYS A CRITICAL ROLE IN GLIOBLASTOMA THERAPEUTICS

Abstract INTRODUCTION Brain tumors remain challenging to treat despite diagnostic advancements. Current approaches involving surgery, radiotherapy, and chemotherapy tend to fail due to drug resistance. Our team identified potential biomarkers associated with aggressive brain tumors, including ORC6 and GPC4. ORC6, implicated in DNA replication and overexpressed in many cancers, presents a promising therapeutic target. In this study, we leverage advanced bioinformatics and experimental validation to further elucidate ORC6's potential for cancer treatment. Material and methods: This study uses R programming and packages to identify biomarkers from genomic databases like TCGA, CGGA, and GLASS. Protein expression is also assessed via tissue microarray and immunohistochemistry. Changes in the ORC6 gene are studied using GO, KEGG, and gene set enrichment analysis techniques. The tumor microenvironment is predicted using CIBERSORT. RESULTS are presented with 3D CLARITY images, and a deep learning algorithm is introduced for potential clinical application optimization. RESULTS Utilizing bioinformatics analyses, we identified ORC6 and GPC4 as potential biomarkers impacting tumor growth. This study focuses on ORC6, which demonstrated significantly increased expression in IDH-wildtype astrocytoma versus normal brain tissue. This heightened ORC6 expression correlated with tumor grading, poor prognosis, and increased KI67 expression in all glioma subgroups. ATAC sequencing and gene set enrichment analysis revealed E2F factors as potential ORC6 regulators and showed that increased ORC6 expression affected cell proliferation signaling pathways. Single-cell sequencing data further indicated ORC6 expression correlation with tumor cells, proliferative stem cells, and M0, M1, and M2 macrophages. Additionally, high ORC6 expression was observed in several brain tumor cell lines (U87MG, U118MG, LN229, GBM8401) that could be down-regulated using siRNA, paving the way for more complex gene editing and functional modeling experiments. CONCLUSION This study employs sophisticated bioinformatics and experimental validation techniques to explore the potential of targeting ORC6 for cancer therapy. Keywords: ORC6, Glioblastoma, TCGA, CGGA, GLASS, biomarker

P06.09.B TRIAL IN PROGRESS: PHASE 1 DOSE-FINDING STUDY TO EVALUATE SAFETY AND TOLERABILITY OF CVGBM IN PATIENTS WITH NEWLY DIAGNOSED AND SURGICALLY RESECTED MGMT-UNMETHYLATED GLIOBLASTOMA

Abstract BACKGROUND Glioblastoma (GBM) is characterized by a low mutational burden limiting the number of neoantigen targets for cancer vaccines. Vaccination against tumor-associated antigens over-presented via MHC class I and II in GBM is an alternative that has shown promise in previous trials. One cancer vaccine containing peptides presented by HLA-A*02-01 (class I) and some DR alleles (class II), showed evidence of peripheral vaccine-induced immune response against these peptides in patients with GBM. Our study will assess the safety and immunogenicity of the messenger ribonucleic acid (mRNA)-based multiepitope vaccine CV-GBM, an investigational therapeutic mRNA vaccine, encoding eight of these peptides that have demonstrated immunogenicity as peptide vaccines. CV-GBM consists of a mRNA with unmodified nucleotides formulated with lipid nanoparticles. MATERIAL AND METHODS CV-GBLM-001 is a first-in-human, open-label, international, dose-escalation phase 1 trial. HLA-A*02:01-positive patients with newly diagnosed MGMT-unmethylated GBM (CNS WHO Grade 4), including IDH-wildtype astrocytoma with a molecular signature of unmethylated MGMT-GBM who have had a gross total or partial resection and who completed post-surgery radiotherapy with or without chemotherapy, are eligible to receive CVGBM. The trial comprises a dose-escalation part (Part A) followed by a dose-expansion part (Part B). In Part A, the starting dose is 12 µg mRNA, which may be escalated to 25 µg, 50 µg and 100 µg, with 3 to 6 patients per dose level. Dose escalation will be guided by a Bayesian logistic regression model. Patients will receive 7 doses of CVGBM by intramuscular injection on Days 1, 8, 15, 29, 43, 57, and 71, followed by 6 optional doses at 6-week intervals, up to 1 year after the first dose or until disease progression or intolerable toxicity. An independent Data and Safety Monitoring Board will recommend the dose for expansion in Part B in which about 20 patients will be enrolled. Biomarkers and vaccine-induced innate and adaptive immunogenicity, including, but not limited to, systemically induced cytokines and chemokines and antigen-specific CD4+ and CD8+ T cells, will be monitored in the blood, and optionally in vaccine-draining lymph nodes and relapsed tumor. In addition to the primary safety objectives, secondary objectives of efficacy (progression-free survival, overall survival) and patient-reported quality of life outcomes will be assessed. Trial sponsor: CureVac SE, Germany.

Epidemiology and survival of adult-type diffuse glioma in Belgium during the molecular era.

Survival data of diffuse adult-type glioma is mostly based on prospective clinical trials or small retrospective cohort studies. Real-world data with large patient cohorts is currently lacking. Using the nationwide, population-based Belgian Cancer Registry, all known histological reports of patients diagnosed with an adult-type diffuse glioma in Belgium between 2017 and 2019 were reviewed. The ICD-O-3 morphology codes were matched with the histological diagnosis. The gathered data were transformed into the 2021 World Health Organization classification of CNS tumors using the IDH- and 1p/19q-mutation status. Between 2017 and 2019, 2233 diffuse adult-type gliomas were diagnosed in Belgium. Full molecular status was available in 67.1% of identified cases. The age-standardized incidence rate of diffuse adult-type glioma in Belgium was estimated at 8.55 per 100 000 person-years and 6.72 per 100 000 person-years for grade 4 lesions. Median overall survival time in IDH-wild-type glioblastoma was 9.3 months, significantly shorter compared to grade 4 IDH-mutant astrocytoma (median survival time: 25.9 months). The 3-year survival probability was 86.0% and 75.7% for grades 2 and 3 IDH-mutated astrocytoma. IDH-wild-type astrocytoma has a worse prognosis with a 3-year survival probability of 31.6% for grade 2 and 5.7% for grade 3 lesions. This registry-based study presents a large cohort of adult-type diffuse glioma with known molecular status and uses real-world survival data. It adds to the current literature which is mainly based on historical landmark trials and smaller retrospective cohort studies.

Distress in Neuro-Oncology Patients and Its Implications for Communication.

Psychoemotional distress affects patients with cancer, including patients with a diagnosis of a malignant brain tumor. Empathy, professional expertise, and conversational skills are required to ensure successful communication with patients. The purpose of this study was to assess whether knowing the communication needs of patients would be helpful to neuro-oncologists before meeting with them. Patients in our neuro-oncology center were asked to complete the National Comprehensive Cancer Network Distress Thermometer (DT) and a study-specific questionnaire on patients' expectations for communication with the treating physician. The questions targeted issues such as attention/caring and awareness of their disease and prognosis. Importance ratings were compared between patients, with high vs. low distress scores to analyze the impact of distress on the patient's needs in physician-patient communication. A total of 81 patients completed the DT and questionnaire. One third (n = 27) had IDH wild-type astrocytoma, and 42 patients (51.9%) were undergoing therapy for primary or recurrent disease. Mean distress was 4.88 (standard deviation ± 2.64) in the whole cohort, and 56.8% of patients had a high distress score (≥ 5 on a 10-point scale). All issues were assessed as important or very important for communication by the majority of patients, and importance ratings increased in patients with high distress levels for most items. Mean importance ratings correlated significantly with distress scores (p < .001). Distress was increased in neuro-oncology patients. Patients with higher distress levels considered issues of both attention/caring and medical information about the disease as more important than patients with lower distress levels. Using distress assessment may help physicians and advanced practitioners to tailor the contents of their discussion for successful communication with patients.

Ancient ubiquitous protein 1 (AUP1) is a prognostic biomarker connected with TP53 mutation and the inflamed microenvironments in glioma

IntroductionGlioblastoma (GBM) is the most common and lethal brain tumor. The current treatment is surgical removal combined with radiotherapy and chemotherapy, Temozolomide (TMZ). However, tumors tend to develop TMZ resistance which leads to therapeutic failure. Ancient ubiquitous protein 1 (AUP1) is a protein associated with lipid metabolism, which is widely expressed on the surface of ER and Lipid droplets, involved in the degradation of misfolded proteins through autophagy. It has recently been described as a prognostic marker in renal tumors. Here, we aim to use sophisticated bioinformatics and experimental validation to characterize the AUP1's role in glioma.Material and methodsWe collected the mRNA, proteomics, and Whole-Exon-Sequencing from The Cancer Genome Atlas (TCGA) for bioinformatics analyses. The analyses included the expression difference, Kaplan–Meier-survival, COX-survival, and correlation to the clinical factors (tumor mutation burden, microsatellite instability, and driven mutant genes). Next, we validated the AUP1 protein expression using immunohistochemical staining on the 78 clinical cases and correlated them with P53 and KI67. Then, we applied GSEA analyses to identify the altered signalings and set functional experiments (including Western Blot, qPCR, BrdU, migration, cell-cycle, and RNAseq) on cell lines when supplemented with small interfering RNA targeting the AUP1 gene (siAUP1) for further validation. We integrated the single-cell sequencing and CIBERSORT analyses at the Chinese Glioma Genome Atlas (CGGA) and Glioma Longitudinal AnalySiS (GLASS) dataset to rationale the role of AUP1 in glioma.ResultsFirstly, the AUP1 is a prognostic marker, increased in the tumor component, and correlated with tumor grade in both transcriptomes and protein levels. Secondly, we found higher AUP1 associated with TP53 status, Tumor mutation burden, and increased proliferation. In the function validation, downregulated AUP1 expression merely impacted the U87MG cells' proliferation instead of altering the lipophagy activity. From the single-cell sequencing and CIBERSORT analyses at CGGA and GLASS data, we understood the AUP1 expression was affected by the tumor proliferation, stromal, and inflammation compositions, particularly the myeloid and T cells. In the longitudinal data, the AUP1 significantly dropped in the recurrent IDH wildtype astrocytoma, which might result from increased AUP1-cold components, including oligodendrocytes, endothelial cells, and pericytes.ConclusionAccording to the literature, AUP1 regulates lipophagy by stabilizing the ubiquitination of lipid droplets. However, we found no direct link between AUP1 suppression and altered autophagy activity in the functional validation. Instead, we noticed AUP1 expression associated with tumor proliferation and inflammatory status, contributed by myeloid cells and T cells. In addition, the TP53 mutations seem to play an important role here and initiate inflamed microenvironments. At the same time, EGFR amplification and Chromosome 7 gain combined 10 loss are associated with increased tumor growth related to AUP1 levels. This study taught us that AUP1 is a poorer predictive biomarker associated with tumor proliferation and could report inflamed status, potentially impacting the clinical application.

Tumor volume and calcifications as indicators for preoperative differentiation of grade II/III diffuse gliomas.

To retrospectively evaluate preoperative clinical factors for their ability to preoperatively differentiate malignancy grades in patients with incipient supratentorial grade II/III diffuse gliomas. This retrospective study included 206 adult patients with incipient supratentorial grade II/III diffuse gliomas according to the 2016 World Health Organization classification of tumors of the central nervous system. The cohort included 136 men and 70 women, with a median age of 41 years. Preoperative factors included age, sex, presence of calcifications on computed tomography scans, and preoperative tumor volume measured using preoperative magnetic resonance imaging. In patients with oligodendrogliomas (IDH-mutant and 1p/19q-codeleted), calcifications were significantly more frequent (p = 0.0034) and tumor volume was significantly larger (p < 0.001) in patients with grade III tumors than in those with grade II tumors. Moreover, in patients with IDH-mutant astrocytomas, preoperative tumor volume was significantly larger (p = 0.0042) in patients with grade III tumors than in those with grade II tumors. In contrast, none of the evaluated preoperative clinical factors were significantly different between the patients with grade II and III IDH-wildtype astrocytomas. In adult patients with suspicison incipient supratentorial grade II/III diffuse gliomas, presence of calcifications and larger preoperative tumor volume might be used as preoperative indices to differentiate between malignancy grades II and III in oligodendrogliomas (IDH-mutant and 1p/19q-codeleted) and larger preoperative tumor volume might have similar utility in IDH-mutant astrocytomas.

SURG-14. LOWER-GRADE GLIOMA IN ELDERLY PATIENTS: TREATMENT AND OUTCOMES IN A MOLECULARLY CHARACTERIZED CONTEMPORARY COHORT

Abstract Lower-grade glioma (LGG) is rare among patients above the age of 60. Previous series reported poor outcomes, likely due to a high proportion of missed IDH wildtype astrocytomas which today would be diagnosed as glioblastoma, and thus proposed defensive treatment in elderly patients. We questioned these findings in our contemporary cohort of “true” LGGs, i.e. IDH mutant astrocytoma and oligodendroglioma WHO grade 2 and 3. Patients aged 60 years and older (“elderly”) treated for hemispheric LGG were retrospectively analyzed for demographic, tumor- and treatment-related factors and progression-free survival (PFS) and were compared to a cohort of patients younger than 60 years. Inclusion criteria, amongst others, were availability of molecular data and volumetric assessment of pre- and postoperative tumor burden. 212 patients were analyzed, among those 21 elderlies (9.9%). Elderly patients did not differ from younger ones regarding preoperative tumor volumes, eloquent location, presence of contrast enhancement and clinical presentation (seizures, focal deficits). In both groups (elderly vs. younger), most patients underwent tumor resection (81% vs. 90.6%; p=0.25) with comparable median residual tumor volumes (5.12 cm3 vs. 3.28 cm3; p=0.66). There was a trend towards more aggressive surgical approaches in younger patients, e.g., use of IOM and awake surgery. However, rates of functional deterioration (p=0.2) and revision surgery (p=0.98) were comparable. Oligodendroglioma, in relation to astrocytoma, was more common in the elderly (76.2% vs. 46.1%; p=0.011). Adjuvant radio- and/or chemotherapy was administered in 76.2% of elderly and 59.8% of younger patients (p=0.163). Median PFS was comparable (50 vs. 56 months; p=0.62), regardless of WHO grade (p=0.43) and tumor subtype (p=0.93). In conclusion, favorable surgical and survival outcomes were achieved in our series that were comparable to those of younger patients. Thus, intensified treatment including maximal safe resection should be advocated in elderly patients whenever feasible.

BIOM-27. PROGNOSTIC SIGNIFICANCE OF EXTENT OF RESECTION IN GLIOMA ACCORDING TO THE 2021 WHO CLASSIFICATION

Abstract BACKGROUND In the updated 2021 WHO classification, diffuse gliomas were strictly reclassified by molecular statuses. In glioblastoma (GBM), the prognostic significance of the resection has been shown, but the excision rate to be achieved in the new classification has not been defined. Moreover, the correlation between the extent of resection (EOR) and the prognosis is still controversial for other gliomas. METHODS IDH 1/2 mutations and TERT promoter mutations were analyzed by Sanger sequencing, 1p/19q co-deletion by microsatellite or MLPA, and EGFR amplification and CDKN2A deletions by MLPA. The correlation between the resection rate and prognosis of each tumor type in the 2021/2016 WHO classification was analyzed retrospectively. RESULTS According to the 2021 classification, 293 patients with GBM, IDH-wild type, 68 with astrocytoma, IDH-mutant (2/3/4 22/26/20), and 58 with oligodendroglioma, IDH-mutant and 1p/19q co-deleted were identified. Based on the 2016 classification, they were diagnosed as 314 GBM, 106 astrocytoma (Gd II/III 37/69), and 59 oligodendroglioma. For GBM, IDH-wild type (2021 classification), a prolonged OS benefit was observed with &amp;gt;25% removal (p=0.042), whereas, more than 20% of tumor removal for patients with GBM (2016 classification) resulted in OS prolongation. EOR was a significant favorable prognostic factor for grade II/III astrocytoma by the 2016 classification, but not for astrocytoma, IDH-mutant (grade 2/3) in the 2021 classification. When analyzed only in patients with astrocytoma, IDH-mutant by the 2021 classification, EOR was not significantly prognostic, suggesting that poorer prognosis of patients with unresectable IDH-wild type astrocytomas (2016 classification) might have a negative impact on the outcome. CONCLUSION There was a strong correlation between EOR and prognosis in patients with GBM, and &amp;gt;25% removal was considered significant according to the 2021 classification. For astrocytoma in the 2021 classification, EOR did not show a prognostic significance, but further validation including grading effect is warranted.

Expression of CD133 Cancer Stem Cell Marker in IDH-Mutant and IDH-wildtype (Isocitrate Dehydrogenase) Astrocytoma.

This study evaluated the differences between IDH1-R132H and CD133 expression in different categories of astrocytoma. This study used a cross-sectional design. Sixty-seven paraffin embedded block of Diffuse Astrocytoma (DA), Anaplastic Astrocytoma (AA) and Glioblastoma (GB) were assessed using using the monoclonal antibody IDH1-R132H and Rabbit polyclonal antibody CD133. It was found that there was a significant relationship between the expression of IDH1-R132H and CD133 in DA, AA and GB (p<0.001). Astrocytoma with IDH-mutant molecular status will express more markers of cancer stem cell CD133 than IDH-wildtype. The IDH1-R132H and CD133 can provide predictive value on treatment success, disease prognosis, recurrence and can be considered as target combination therapy with chemotherapy.

Glioblastoma Extracellular Vesicle-Specific Peptides Inhibit EV-Induced Neuronal Cytotoxicity.

WHO Grade 4 IDH-wild type astrocytoma (GBM) is the deadliest brain tumor with a poor prognosis. Meningioma (MMA) is a more common “benign” central nervous system tumor but with significant recurrence rates. There is an urgent need for brain tumor biomarkers for early diagnosis and effective treatment options. Extracellular vesicles (EVs) are tiny membrane-enclosed vesicles that play essential functions in cell-to-cell communications among tumor cells. We aimed to identify epitopes of brain tumor EVs by phage peptide libraries. EVs from GBM plasma, MMA plasma, or brain tumor cell lines were used to screen phage-displayed random peptide libraries to identify high-affinity peptides. We purified EVs from three GBM plasma pools (23 patients), one MMA pool (10 patients), and four brain tumor cell lines. We identified a total of 21 high-affinity phage peptides (12 unique) specific to brain tumor EVs. The peptides shared high sequence homologies among those selected by the same EVs. Dose–response ELISA demonstrated that phage peptides were specific to brain tumor EVs compared to controls. Peptide affinity purification identified unique brain tumor EV subpopulations. Significantly, GBM EV peptides inhibit brain tumor EV-induced complement-dependent cytotoxicity (necrosis) in neurons. We conclude that phage display technology could identify specific peptides to isolate and characterize tumor EVs.

Abstract 5264: Glioblastoma degraded to low-grade astrocytoma two-years after surgical resection: A rare case report

Abstract Background: Glioblastoma (GBM) is the most common and malignant brain tumor. The prognosis of patients with GBM is poor, with a five-year survival rate of 5% and a median overall survival of 12-16 months. In addition, there is a high risk for the recurrence of GBM patient, with the median time of 9.5 months. However, little was reported about the degradation of GBM recurrence. Methods: Immunohistochemistry (IHC) staining and next-generation sequencing (NGS) were conducted to address the histopathological and molecular features of the patient. Results: This patient was firstly diagnosed with GBM in April 2019, followed by surgical resection to completely remove tumor occupancy. IHC staining of the tumor sample showed positive expression of P53, S-100, GFAP, EGFR, MGMT, and a 60% expression of Ki67. Thus, this tumor was clinically classified as WHO grade 4 according to its histopathological features. Then we performed NGS and found that the tumor contained four SNVs (EGFR c.865G&amp;gt;A, EPHA5 c.2152C&amp;gt;T, RB1 c.1615G&amp;gt;T, TP53 c.742C&amp;gt;T), EGFR gene amplification, and without IDH1 or IDH2 mutation. These molecular features of the tumor were classified as WHO grade 4 and were consistent with the histopathological grading. In May 2021, two recurrent tumors were found in this patient and were surgical resected again. However, IHC staining of the tumor sample showed positive expression of P53, S-100, and GFAP, while EGFR and MGMT were undetectable, and expression of Ki67 was only 5%. Thus, these recurrent tumors were diagnosed as low-grade astrocytoma (WHO grade 2) according to their histopathological features. Furthermore, NGS was performed with one of those recurrent tumors and showed that it contained 38 SNVs including those four variants in the primary tumor, EGFR gene amplification, and without IDH1 or IDH2 mutation. According to the fifth edition of the WHO Classification of Tumors of the Central Nervous System (CNS), IDH-wildtype astrocytoma combined with EGFR gene amplification should be classified as IDH-wildtype GBM (WHO grade 4), which was different from the histopathological grading. From our points of view, though these recurrent tumors are actually high-grade glioma from their ‘molecular identity’, maybe they have not developed to high-grade tumors at the time of diagnosis because of our timely follow-up examination. Conclusion: In summary, we reported a rare GBM patient survived for two-years post surgical resection and recurred with a low-grade astrocytoma in histopathological diagnosis. However, the molecular features revealed that the recurrent tumor should be diagnosed as a high-grade glioma, suggesting the clinical importance of combining the molecular and conventional histopathological diagnosis. Moreover, we provided a clinical evidence of histological degradation of GBM, and showed a rare clinical case of inconsistence of histopathological and molecular diagnosis in recurrent GBM. Citation Format: Zhipeng Su, Xiang Zhang, Bella Guo, Ximeng Zhao, Qun Li, Tonghui Ma. Glioblastoma degraded to low-grade astrocytoma two-years after surgical resection: A rare case report [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5264.

Utility of targeted next-generation sequencing assay to detect 1p/19q co-deletion in formalin-fixed paraffin-embedded glioma specimens

Molecular classification of brain neoplasms is important for diagnosis, prognosis, and treatment outcome of histologically similar tumors. Oligodendroglioma is a glioma subtype characterized by 1p/19q co-deletion and IDH1/IDH2 mutations, which predict a good prognosis, responsiveness to therapy, and an improved overall survival compared to other adult gliomas. In a routine clinical setting, 1p/19q co-deletion is detected by interphase-FISH and SNP microarray, and somatic mutations are detected by targeted next-generation sequencing (NGS). The aim of this proof-of-principle study was to investigate the feasibility of using targeted NGS to simultaneously detect both 1p/19q co-deletion and somatic mutations. Among 247 consecutive patients with formalin-fixed paraffin-embedded brain tumors with various subtypes, NGS revealed 1p/19q co-deletion in 26 oligodendrogliomas and an IDH-wildtype astrocytoma, and partial loss across chromosomes 1p and 19q/whole-arm loss of 1p or 19q/copy neutral loss of heterozygosity in 11 nonoligodendrogliomas. For this 247 brain-tumor cohort, the overall sensitivity, specificity, and accuracy of detecting 1p/19q co-deletion by NGS in oligodendrogliomas were 96.2%, 99.6%, and 99.2%, respectively. The oligodendroglioma cohort had more mutations in IDH1/IDH2, CIC, FUBP1, and TERT, and fewer mutations in ATRX and TP53 than the nonoligodendroglioma cohort. This proof-of-concept study demonstrated that targeted NGS can simultaneously detect both 1p/19q co-deletion and somatic mutations, which can provide a more comprehensive genetic profiling for patients with gliomas using a single assay in a clinical setting.

Deciphering of Adult Glioma Vulnerabilities through Expression Pattern Analysis of GABA, Glutamate and Calcium Neurotransmitter Genes.

Adult infiltrating gliomas are highly aggressive tumors of the central nervous system with a dismal prognosis despite intensive multimodal therapy (chemotherapy and/or radiotherapy). In this study, we studied the expression, methylation and interacting miRNA profiles of GABA-, glutamate- and calcium-related genes in 661 adult infiltrating gliomas available through the TCGA database. Neurotransmitter-based unsupervised clustering identified three established glioma molecular subgroups that parallel major World Health Organization glioma subclasses (IDH-wildtype astrocytomas, IDH-mutant astrocytomas, IDH-mutant oligodendroglioma). In addition, this analysis also defined a novel, neurotransmitter-related glioma subgroup (NT-1), mostly comprised of IDH-mutated gliomas and characterized by the overexpression of neurotransmitter-related genes. Lower expression of neurotransmission-related genes was correlated with increased aggressivity in hypomethylated IDH-wildtype tumors. There were also significant differences in the composition of the tumor inflammatory microenvironment between neurotransmission-based tumor categories, with lower estimated pools of M2-phenotype macrophages in NT-1 gliomas. This multi-omics analysis of the neurotransmission expression landscape of TCGA gliomas—which highlights the existence of neurotransmission-based glioma categories with different expression, epigenetic and inflammatory profiles—supports the existence of operational neurotransmitter signaling pathways in adult gliomas. These findings could shed new light on potential vulnerabilities to exploit in future glioma-targeting drug therapies.

Radiomics and Qualitative Features From Multiparametric MRI Predict Molecular Subtypes in Patients With Lower-Grade Glioma.

BackgroundIsocitrate dehydrogenase (IDH) mutation and 1p19q codeletion status have been identified as significant markers for therapy and prognosis in lower-grade glioma (LGG). The current study aimed to construct a combined machine learning-based model for predicting the molecular subtypes of LGG, including (1) IDH wild-type astrocytoma (IDHwt), (2) IDH mutant and 1p19q non-codeleted astrocytoma (IDHmut-noncodel), and (3) IDH-mutant and 1p19q codeleted oligodendroglioma (IDHmut-codel), based on multiparametric magnetic resonance imaging (MRI) radiomics, qualitative features, and clinical factors.MethodsA total of 335 patients with LGG (WHO grade II/III) were retrospectively enrolled. The sum of 5,929 radiomics features were extracted from multiparametric MRI. Selected robust, non-redundant, and relevant features were used to construct a random forest model based on a training cohort (n = 269) and evaluated on a testing cohort (n = 66). Meanwhile, preoperative MRIs of all patients were scored in accordance with Visually Accessible Rembrandt Images (VASARI) annotations and T2-fluid attenuated inversion recovery (T2-FLAIR) mismatch sign. By combining radiomics features, qualitative features (VASARI annotations and T2-FLAIR mismatch signs), and clinical factors, a combined prediction model for the molecular subtypes of LGG was built.ResultsThe 17-feature radiomics model achieved area under the curve (AUC) values of 0.6557, 0.6830, and 0.7579 for IDHwt, IDHmut-noncodel, and IDHmut-codel, respectively, in the testing cohort. Incorporating qualitative features and clinical factors into the radiomics model resulted in improved AUCs of 0.8623, 0.8056, and 0.8036 for IDHwt, IDHmut-noncodel, and IDHmut-codel, with balanced accuracies of 0.8924, 0.8066, and 0.8095, respectively.ConclusionThe combined machine learning algorithm can provide a method to non-invasively predict the molecular subtypes of LGG preoperatively with excellent predictive performance.

Local detection of microvessels in IDH-wildtype glioblastoma using relative cerebral blood volume: an imaging marker useful for astrocytoma grade 4 classification

BackgroundThe microvessels area (MVA), derived from microvascular proliferation, is a biomarker useful for high-grade glioma classification. Nevertheless, its measurement is costly, labor-intense, and invasive. Finding radiologic correlations with MVA could provide a complementary non-invasive approach without an extra cost and labor intensity and from the first stage. This study aims to correlate imaging markers, such as relative cerebral blood volume (rCBV), and local MVA in IDH-wildtype glioblastoma, and to propose this imaging marker as useful for astrocytoma grade 4 classification.MethodsData from 73 tissue blocks belonging to 17 IDH-wildtype glioblastomas and 7 blocks from 2 IDH-mutant astrocytomas were compiled from the Ivy GAP database. MRI processing and rCBV quantification were carried out using ONCOhabitats methodology. Histologic and MRI co-registration was done manually with experts’ supervision, achieving an accuracy of 88.8% of overlay. Spearman’s correlation was used to analyze the association between rCBV and microvessel area. Mann-Whitney test was used to study differences of rCBV between blocks with presence or absence of microvessels in IDH-wildtype glioblastoma, as well as to find differences with IDH-mutant astrocytoma samples.ResultsSignificant positive correlations were found between rCBV and microvessel area in the IDH-wildtype blocks (p < 0.001), as well as significant differences in rCBV were found between blocks with microvascular proliferation and blocks without it (p < 0.0001). In addition, significant differences in rCBV were found between IDH-wildtype glioblastoma and IDH-mutant astrocytoma samples, being 2–2.5 times higher rCBV values in IDH-wildtype glioblastoma samples.ConclusionsThe proposed rCBV marker, calculated from diagnostic MRIs, can detect in IDH-wildtype glioblastoma those regions with microvessels from those without it, and it is significantly correlated with local microvessels area. In addition, the proposed rCBV marker can differentiate the IDH mutation status, providing a complementary non-invasive method for high-grade glioma classification.

Comprehensive bioinformatic analysis of key genes and signaling pathways in glioma

The identification of specific survival-related differentially expressed genes (DEGs) is a method for uncovering therapeutic approaches for various cancers, including glioma. However, the key target genes associated with the occurrence and development of gliomas remain unknown. In this study, we performed bioinformatics analysis on 17 GSE datasets and identified DEGs correlated with glioma. A total of 74 mutual-DEGs with downregulated expression in gliomas compared with that in normal brain tissues were found in 17 datasets. These DEGs were related to GABAergic synaptic transmission, chloride transmembrane transport, glutamate secretion, and gamma-aminobutyric acid signaling pathway. Gamma-aminobutyric acid type A receptor subunit gamma 2 (GABRG2) was identified as a hub gene in the protein-protein interaction network. GABRG2 exhibited lower expression in IDH wild-type astrocytoma than that in IDH mutant astrocytoma and indicated poor prognosis in glioma patients. GABRG2 may contribute to the progression of glioma by affecting GABA receptor-related pathways and is a potential biomarker for the diagnosis and treatment of glioma.

Extent, pattern, and prognostic value of MGMT promotor methylation: does it differ between glioblastoma and IDH-wildtype/TERT-mutated astrocytoma?

IntroductionThe cIMPACT-NOW update 6 first introduced glioblastoma diagnosis based on the combination of IDH-wildtype (IDHwt) status and TERT promotor mutation (pTERTmut). In glioblastoma as defined by histopathology according to the WHO 2016 classification, MGMT promotor status is associated with outcome. Whether this is also true in glioblastoma defined by molecular markers is yet unclear.MethodsWe searched the institutional database for patients with: (1) glioblastoma defined by histopathology; and (2) IDHwt astrocytoma with pTERTmut. MGMT promotor methylation was analysed using methylation-specific PCR and Sanger sequencing of CpG sites within the MGMT promotor region.ResultsWe identified 224 patients with glioblastoma diagnosed based on histopathology, and 54 patients with IDHwt astrocytoma with pTERTmut (19 astrocytomas WHO grade II and 38 astrocytomas WHO grade III). There was no difference in the number of MGMT methylated tumors between the two cohorts as determined per PCR, and also neither the number nor the pattern of methylated CpG sites differed as determined per Sanger sequencing. Progression-free (PFS) and overall survival (OS) was similar between the two cohorts when treated with radio- or chemotherapy. In both cohorts, higher numbers of methylated CpG sites were associated with favourable outcome.ConclusionsExtent and pattern of methylated CpG sites are similar in glioblastoma and IDHwt astrocytoma with pTERTmut. In both tumor entities, higher numbers of methylated CpG sites appear associated with more favourable outcome. Evaluation in larger prospective cohorts is warranted.

The dominant TP53 hotspot mutation in IDH -mutant astrocytoma, R273C, has distinctive pathologic features and sex-specific prognostic implications.

BackgroundInfiltrative astrocytic tumors with and without isocitrate dehydrogenase (IDH) mutation frequently contain mutations in the TP53 tumor suppressor gene. Disruption of normal p53 protein activity confers neoplastic cells with a number of oncogenic properties and is a common feature of aggressive malignancies. However, the high prevalence of TP53 mutation and its pathogenic role in IDH-mutant (IDHmut) astrocytoma is not well understood.MethodsWe performed a retrospective analysis of molecular and clinical data from patients with IDHmut astrocytoma at the University of Pittsburgh Medical Center between 2015 and 2019 as our initial cohort. We validated and expanded our findings using molecular and clinical data from The Cancer Genome Atlas.ResultsWe show that the TP53 mutational spectrum in IDHmut astrocytomas is dominated by a single hotspot mutation that codes for the R273C amino acid change. This mutation is not enriched in IDH-wildtype astrocytomas. The high prevalence of TP53R273C mutation is not readily explained by known mutagenic mechanisms, and TP53R273C mutant tumors have lower transcriptional levels of proliferation-related genes compared to IDHmut astrocytomas harboring other forms of mutant p53. Despite lower proliferation, TP53R273C mutant tumors tend to progress more quickly and have a shorter overall survival than those with other TP53 mutations, particularly in male patients.ConclusionsOur findings suggest that compared to other TP53 mutations, IDHmut astrocytomas may select for TP53R273C mutations during tumorigenesis. The genotype, sex, and mutation-specific findings are clinically relevant and should prompt further investigation of TP53R273C.

MPC-17 2021 WHO Classification of Tumors of the CNS, 5th ed

The grading of gliomas based on histological features has been a subject of debate for several decades. While the traditional grading system has failed to stratify the risk of IDH-mutant astrocytoma, canonical histological and proliferative markers may be applicable to the risk stratification of IDH-wildtype astrocytoma. Numerous studies have examined molecular markers to obtain more clinically relevant information that will improve the risk stratification of gliomas. The CDKN2A/B homozygous deletion for IDH-mutant astrocytoma and the following three criteria for IDH-wildtype astrocytoma: the concurrent gain of whole chromosome 7 and loss of whole chromosome 10, TERT promoter mutations, and EGFR amplification, were identified as independent molecular markers of the worst clinical outcomes. Therefore, the 2021 World Health Organization (WHO) Classification of Tumors of the Central Nervous System adopted these molecular markers into the revised grading criteria of IDH-mutant and -wildtype astrocytoma respectively, as a grading system within tumor types. For diffuse gliomas in children, molecular alteration-based classification was adopted, dividing low-grade and high-grade subcategories. New tumor types and subtypes were introduced, some based on DNA methylation profiling. To achieve this novel classification in a resource-limited setting, an integrated diagnosis combining clinical, histological, and molecular information became more important.

MPC-1 DNA methylome analysis suggested the presence of “true” IDH-wildtype lower-grade gliomas

Background: There will be significant changes in the diagnosis of IDH-wildtype adult-type gliomas in the upcoming 5th edition of the WHO Classification of Central Nervous System Tumours. IDH-wildtype lower grade gliomas (IDHwt LGGs) that harbor molecular features of glioblastoma (EGFR amplification, the combination of whole chromosome 7 gain and whole chromosome 10 loss (7+/10-), or TERT promoter mutations) will be diagnosed as glioblastomas (GBMs), while IDH-wildtype astrocytomas will not be included as a separate tumor type. However, IDHwt LGGs are a very heterogeneous group of tumors, and further investigation is warranted particularly in those without molecular features of glioblastoma. To elucidate the biology of IDHwt LGGs, we analyzed DNA methylation profile and survival time. Materials and Methods: Of the 724 adult-type diffuse glioma samples from a multi-institutional study, 64 IDHwt LGG, including 54 without any of molecular features of GBM and 10 with PDGFRA amplification or TERT promoter mutation, were examined using Infinium MethylationEPIC BeadChip. The raw data files (IDAT files) were analyzed by the web-based DNA methylation classifier provided by DKFZ (MolecularNeuropathology.org) or by R (Version 4.0.4) using the minfi (1.34.0) and Rtsne (0.15) packages. [Result] Twenty-three out of 54 IDHwt LGGs matched known methylation classes using the DKFZ methylation classifier. In t-Distributed Stochastic Neighbor Embedding clustering analysis, 20 cases formed a cluster within the methylation class family glioblastoma, IDH-wildtype, mainly subclass RTK I (“GBM” cluster). Another 29 IDHwt LGGs formed an independent cluster (“LGG” cluster) separate from any of the existing reference groups near but not overlapping with several subtypes of pediatric-type lower grade gliomas. The “LGG” cluster cases had significantly longer overall survival than the “GBM” cluster cases. Discussion: Methylation profiling showed that IDHwt LGGs without molecular features of GBM were heterogeneous group of tumors. Our data suggested the presence of “true” IDHwt LGGs with intermediate prognosis.