IDH-mutant Tumors Research Articles

Evolving cell states and oncogenic drivers during the progression of IDH-mutant gliomas.

Isocitrate dehydrogenase (IDH) mutants define a class of gliomas that are initially slow-growing but inevitably progress to fatal disease. To characterize their malignant cell hierarchy, we profiled chromatin accessibility and gene expression across single cells from low-grade and high-grade IDH-mutant gliomas and ascertained their developmental states through a comparison to normal brain cells. We provide evidence that these tumors are initially fueled by slow-cycling oligodendrocyte progenitor cell-like cells. During progression, a more proliferative neural progenitor cell-like population expands, potentially through partial reprogramming of 'permissive' chromatin in progenitors. This transition is accompanied by a switch from methylation-based drivers to genetic ones. In low-grade IDH-mutant tumors or organoids, DNA hypermethylation appears to suppress interferon (IFN) signaling, which is induced by IDH or DNA methyltransferase 1 inhibitors. High-grade tumors frequently lose this hypermethylation and instead acquire genetic alterations that disrupt IFN and other tumor-suppressive programs. Our findings explain how these slow-growing tumors may progress to lethal malignancies and have implications for therapies that target their epigenetic underpinnings.

NCOG-42. SPEECH-SPECIFIC ERRORS IN LEXICAL RETRIEVAL PREDICT GLIOMA MOLECULAR FEATURES

Abstract Recent work confirms that low and high-grade gliomas, remodel neural circuits which influence speech production. The extent to which speech kinetics and category of speech error during lexical retrieval may predict glioma molecular features remains unknown. Here, we extracted audiovisual speech production from 550 patients totaling over 26,400 individual stimuli to identify correlations with glioma molecular features including IDH, TERT, 1p19q co-deletion, and CDK2AB. Nonparametric statistics were used for analysis. IDH wild-type tumors were more likely to be temporoparietal, while IDH mutant tumors were more likely to be insular (p=0.02). Patients with IDH wild-type glioblastoma exhibited more errors (8.23 vs 3.79, p=0.02), with a higher occurrence of anomia (p=0.004), circumlocution (p=0.034) and general delay (p=0.04) when compared with IDH mutant WHO grade 4 astrocytoma. Binomial logistic regression identified lexical retrieval anomia, circumlocution, and initiation delays as an independent predictor of having an IDH wild-type tumor. Within the IDH mutant group, anomia (p=0.03) and articulation (p=0.014) errors were associated with malignant molecular features (after controlling for prognostic variables such as tumor volume). Binomial logistic regression identified articulation or anomia for picture naming as an independent risk predictor for WHO grade 2 gliomas with greater risk molecular features (based on Ki67 and CDK2AB). Distinct patterns of language errors were observed in patients with IDH wild-type and IDH mutant tumors. Further work is required to elucidate the functional and biological correlates of these findings.

QOL-31. RESULTS OF A TELEHEALTH-BASED PSYCHOLOGICAL INTERVENTION FOR CAREGIVERS OF PATIENTS WITH PRIMARY MALIGNANT BRAIN TUMORS: A MODERATION ANALYSIS OF A RANDOMIZED CONTROLLED TRIAL

Abstract BACKGROUND In a recent randomized controlled trial, caregivers of patients with primary malignant brain tumors (PMBT) who received NeuroCARE—a six-session, telehealth-based psychological intervention—demonstrated improved anxiety and depression symptoms, coping, and self-efficacy at 11-weeks compared to usual care (UC) participants. We now explore how sociodemographic variables, disease-specific factors, and baseline mood symptoms moderate intervention effects on anxiety and depression symptoms. METHODS We examined data from 120 enrolled caregivers (Mage=53 years; 83% female, 92% White, 70% spouses/partners) of PMBT patients (84% glioblastoma). Eligible caregivers had elevated anxiety (Generalized Anxiety Disorder-7 ≥ 5). Participants were randomized 1:1 to NeuroCARE or UC and completed baseline sociodemographic questionnaires and the Hospital Anxiety and Depression Scale at baseline and 11-weeks. We used linear regression to evaluate whether selected sociodemographic characteristics (gender, age, relationship to patient), tumor characteristics (tumor location/laterality, IDH mutation status), and baseline mood symptoms moderated intervention effects on anxiety and depression symptoms. We report the group-by-moderator (b) effects, using a 0.20 significance level. RESULTS The beneficial effects of NeuroCARE on reducing 11-week anxiety symptoms were greater in women compared to men (b=-2.63, CI: -6.17, 0.91, p=0.14) and greater in caregivers who reported higher baseline anxiety symptoms (b=-0.43, CI: -0.78, -0.08, p=0.02) and depression symptoms (b=-0.56, CI: -0.96, -0.17, p=0.01). The beneficial effects of NeuroCARE on reducing 11-week depression symptoms were greater in caregivers of patients with IDH-wildtype tumors compared to caregivers of patients with IDH-mutant tumors (b=2.49, CI: -0.26, 5.24, p=0.08). Caregiver age, relationship to patient, and tumor location/laterality did not significantly moderate NeuroCARE effects on mood. CONCLUSIONS The positive effects of NeuroCARE were particularly salient for caregivers with elevated anxiety and/or depression symptoms and those caring for patients with IDH-wildtype tumors. Content may be further tailored to account for the unique needs of men and caregivers of patients with IDH-mutant tumors.

TMET-30. ONCOLYTIC HSV REWIRES METABOLISM TO PROMOTE ANTITUMOR IMMUNITY THROUGH REDUCTIVE CARBOXYLATION AND FERROPTOSIS

Abstract Cancer cells autonomously rewire metabolism to meet increased bioenergetic demands. Oncolytic HSVs (oHSVs) are attenuated Herpes simplex type 1 derived viruses that lyse tumor cells releasing damage-associated molecules triggering antitumor immunity. Currently one oHSV-1 derived virus is approved for melanoma treatment in the USA and another for recurrent GBM in Japan. While several other viruses are being evaluated in clinical trials for safety and efficacy, an understanding of the impact of oHSV therapy on cancer cell metabolism is largely unexplored. A detailed understanding of how oHSV impact tumor cell metabolism and oxidative stress is important to understand the unique susceptibilities that can further be exploited to improve the therapeutic index. Here, we analyzed transcriptomic changes in tumor biopsies of recurrent high-grade glioma patients treated with oHSV. RNA-seq data obtained from pre- and post-G207 treated biopsies from phase-Ib study patients (NCT00028158) revealed enrichment in pathways related to citric acid cycle, oxidative phosphorylation, and glutamate metabolism. We confirmed these changes by RNA sequencing of oHSV-infected patient derived GBM cells. Mechanistically, oHSV hijacked cellular glucose metabolism to increase oxidative phosphorylation and modulated cellular glutamine metabolism towards reductive carboxylation and reduced free glutathione, collectively contributing to increased ROS generation. Kinome profiling revealed PKC activation upon treatment with oHSV. Increased ROS combined with reduced glutathione, and PKC activation promoted ferroptotic cell death characterized by increased lipid peroxidation and smaller, ruptured mitochondria, and contributed to immunotherapy by this treatment. IDH mutant tumors that block reductive carboxylation were resistant to oHSV-induced ferroptotic cell death and did not benefit from oHSV-induced antitumor benefit. Together, this study presents avenues to enhance antitumor immunity by oHSV therapy.

CTIM-09. INB-200: FULLY ENROLLED PHASE 1 STUDY OF GENE-MODIFIED AUTOLOGOUS GAMMA-DELTA (ΓΔ) T CELLS IN NEWLY DIAGNOSED GLIOBLASTOMA MULTIFORME (GBM) PATIENTS RECEIVING MAINTENANCE TEMOZOLOMIDE (TMZ)

Abstract IN8bio’s DeltEx drug resistant immunotherapy (DRI), comprising TMZ-resistant γδ T cells engineered to express methylguanine-DNA methyltransferase (MGMT), enables γδ T cells to target upregulated NKG2D ligands on tumor cells in real time during alkylating chemotherapy exposure. Updated results from the fully enrolled Phase 1 trial evaluating DRI in adult newly diagnosed GBM subjects with IDH wild type and mutant tumors, adequate organ function and KPS ≥ 70%. Cohorts (C) 1, 2 and 3 received 1, 3 or 6 doses (1 x 107 DRI cells/dose) into the resection cavity with 150 mg/m2 of IV TMZ on Day 1 of each of 6 Stupp maintenance cycles. The primary endpoint is safety and secondary endpoints include survival; immunologic correlative analyses are included. Dose limiting toxicities (DLTs) are defined as treatment related ≥ grade (G) 3 cardiopulmonary or hepatic toxicity, G4 toxicity exceeding 72 hours or neurologic deterioration that exceeds 2 weeks. A total of 23 subjects were enrolled and 13 dosed (61% male; median age 68 (range: 21-74); 92% IDH-WT, 54% MGMT unmethylated). No DLTs, cytokine release syndrome (CRS), neurotoxicity (ICANS) or treatment related deaths were reported. The most common adverse events were decreased WBC/platelet count, asthenia, fatigue, hydrocephalus, headache, decreased appetite, urinary tract infection, thrombosis and balance disorder with ≥G3 platelet count decreased in 23% and WBC/ neutrophil/ lymphocyte count decreased in 7.7% each. Peripheral TMZ-based lymphodepletion was maintained for up to 1 year evidenced by near or below normal range T, B, and NK subsets. With median follow-up of 10.8 months, 83% of subjects exceeded the median 7 month Progression Free Survival (PFS) of the Stupp regimen alone, had manageable toxicity with a continued encouraging trend in PFS. Subject with IDH mutant tumor remains progression free for > 35 months. Long-term follow-up for durability of PFS and OS continues.

TMIC-53. USE OF CEREBRAL OPEN-FLOW MICROPERFUSION TO IDENTIFY POTENTIAL ONCOMETABOLITE, L-2-HYDROXYGLUTARATE, IN GLIOBLASTOMA CHEMOTHERAPY ADAPTATION

Abstract Glioblastoma (GBM) is among the most prevalent and lethal brain tumors, comprising nearly half of all central nervous system malignancies and imparting a median overall survival of only 18-20 months. One major challenge in curing this disease is the limited understanding of how GBM cells biologically respond to treatments like radiotherapy and chemotherapy, including temozolomide (TMZ), in real-time. A significant challenge is observing changes within the tumor interstitial fluid (ISF), which mediates adaptive intercellular signaling between tumor cells and their surrounding tumoral and stromal neighbors. Microfluidic sampling methods, such as cerebral microdialysis and microperfusion, have started to provide valuable insights into the ISF signaling dynamics. These methods enable real-time biomolecular monitoring of this often-neglected component of the tumor microenvironment (TME). We have performed in vivo cerebral open-flow microperfusion (cOFM) pilot studies to identify metabolomic and proteomic changes within the tumor ISF resulting from differential treatment with either TMZ or vehicle control. Among these metabolite and protein changes, we identified an increase in 2-hydroxyglutarate (2-HG). While D-2-HG is a hallmark metabolite of isocitrate dehydrogenase mutant (IDH-mut) tumors, our model utilized the IDH wild-type (IDH-WT) CT2A mouse-syngeneic glioma model. Subsequent analysis of enantiomeric confirmation characterized this to be predominantly L-2-HG. Follow-up in vitro studies have shown that elevated L-2-HG plays a functional role in IDH-WT patient-derived glioma cells via non-canonical HIF1α stabilization and subsequent expression of HRE genes. This suggested that L-2-HG may play a similar role in IDH-WT GBM as D-2-HG plays in IDH-mutant tumors. This study demonstrates the potential of L-2-HG to function as a chemo-responsive oncometabolite in the TME of highly aggressive IDH-WT glioblastoma. By targeting the metabolic and signaling properties of L-2-HG, it may be possible to prevent chemo-resistant adaptations mounted by GBM cells when treated with TMZ.

CNSC-54. CENTRAL AND BOUNDARY-DRIVEN GROWTH PATTERNS DOMINATE RESPECTIVELY IDH WILD-TYPE AND MUTANT GLIOMAS

Abstract Diffuse gliomas are the most prevalent primary malignant brain tumors in adults and are characterized by invasive growth and poor prognosis. Tumor cells infiltrate the brain parenchyma and spread to distant sites. The aim of this study is to characterize the spatial heterogeneity, mode, and direction of tumor development to identify key areas for localized treatment. Neuronavigation was employed to collect n=134 image-guided samples from n=16 adult patients with diffuse gliomas, including n=7 IDH wild-type (IDHwt) and n=9 IDH mutant (IDHmut) tumors from regions with and without MRI abnormalities. Collected samples were subjected to targeted and shallow whole-genome sequencing. Somatic variants were used to determine the evolutionary processes governing heterogeneity. Examination of the dN/dS ratio, Tajima’s D score, and MOBSTER model suggested that all tumors are consistent with neutral evolution, as previously reported. Considering that genetic heterogeneity was a result of stochastic mutations, we used phylogeographic relationships between samples to dissect the spatiotemporal development of these tumors. Evolutionary (patristic) and cartesian (Euclidean) distances between sample pairs from the same patient were correlated in n=2/9 IDHmut and n=2/7 IDHwt patients, suggesting that migratory or punctuated evolutionary processes play a major role in tumor diffusion. An examination of spatial diffusion revealed that the latest descendant appeared peripherally in n=5/9 IDHmut patients, suggesting boundary-driven growth. In the remaining cases, three patients exhibited centrally-driven growth, whereas a fourth patient displayed a mixed pattern. In contrast, in n=4/7 IDHwt patients, descendants appeared closer to the tumor center, suggesting centrally-driven growth. In the remaining three IDHwt patients, one patient showed a diffusion that was consistent with boundary-driven growth, while the other two patients showed a mixed pattern. Taken together our data suggest that IDHwt tumors tend to develop centrally, whereas IDHmut tumors preferably develop outward. Our data could aid in tailoring localized treatment based on projected growth patterns.

EPID-02. BLOOD-BASED CLASSIFICATION OF IDH-STATUS OF NON-ENHANCING GLIOMAS: A MACHINE LEARNING APPROACH

Abstract BACKGROUND Non-invasive determination of IDH mutational status in patients with glioma could offer significant therapeutic opportunities. While IDH wildtype (WT) tumors typically show enhancement on MRI, IDH mutant (MUT) tumors often lack this enhancement. However, relying solely on anatomic radiology may lead to misclassification, and currently, tissue acquisition is the primary method for assessing IDH-status in gliomas. These limitations hamper the development of neoadjuvant or intraoperative therapeutic strategies based on IDH-status; thus, more minimally invasive methods to determine IDH-status are needed. In this study, we assessed peripheral immune cell proportions, which vary by glioma subtype, from pre-surgery peripheral blood samples as an alternative method of classifying the IDH-status in gliomas without enhancement. MATERIALS AND METHODS We employed a highly accurate large language model (GPT-4-Turbo-128k) with over 99% accuracy to read radiology notes and exclude patients with enhancing gliomas. Then, we identified 12 immune cell subtypes from whole blood using deconvolution algorithms based on DNA methylation data. These immune cell subtypes, along with patient age, were integrated into a machine learning model (random forest) to predict IDH-status (WT vs. MUT), leveraging conditional Generative Adversarial Networks to generate synthetic data and mitigate bias in the datasets. Two independent datasets were included for training and validating the model. RESULTS The random forest model had an AUC of 0.90 and accurately identified IDH-status in 81% of a training set of 287 gliomas (65 WT and 222 MUT at varied time points after diagnosis). In an independent validation data set of 99 gliomas (6 WT and 93 MUT) from pre-surgery blood samples, the AUC was 0.93, and accurately predicted IDH-status in 92% of cases with a sensitivity of 93% and specificity of 83%. CONCLUSIONS We predicted the IDH-status of non-enhancing gliomas using patients’ age and immunomethylomic data from pre-surgical peripheral blood samples. This minimally invasive approach is a promising step toward reducing risk and enabling earlier therapies based on IDH-status.

TMIC-26. IMMUNE CELL DYNAMICS WITHIN GLIOMAS: THE INFLUENCE OF GRADE AND IDH STATUS

Abstract Mutations in isocitrate dehydrogenase (IDH) are known to significantly impact the prognosis and biology of diffuse gliomas. A number of studies have compared the immune microenvironment of IDH-wildtype glioma versus IDH-mutant glioma, and have also characterized the impact of mutant-IDH on lymphocytes. However, few studies have explored how mutant-IDH may affect tumors as they progress and increase in grade, or have delved into the myeloid immune cell compartment. Thus, we sought to understand whether tumor progression affected intratumoral immune cell composition within IDH-mutant glioma, and aimed to compare IDH-mutant versus IDH-wildtype glioma in both humans and mice. Using bulk RNA sequencing, we compared paired samples from patients with IDH-mutant glioma before and after progression, focusing on differentially-expressed immunological genes and relative immune cell proportions. We also compared grade 4 IDH-mutant astrocytoma cases to IDH-wildtype glioblastoma, performing both transcriptomic and cellular analyses. Finally, we engineered murine glioma cells to test the effect of mutant-IDH alone in the tumor microenvironment. We found that multiple immune-related genes distinguish low- and high-grade IDH-mutant glioma, and that high-grade tumors contain greater proportions of suppressive myeloid cells. When comparing IDH-wildtype to IDH-mutant glioma, we found enrichment of suppressive macrophages and myeloid-derived suppressor cells (MDSC) in IDH-wildtype tumors. Finally, our murine model of IDH-wildtype versus IDH-mutant glioma mirrored many of the myeloid cell shifts observed in human tumors. Our data show that progression of IDH-mutant tumors is a significant immunological phenomenon, associated with marked alterations in the tumor immune microenvironment. We also show that IDH-mutant glioma harbors greater inflammatory signatures and fewer suppressive myeloid cells than IDH-wildtype glioma, and that engineering mutant IDH into murine tumors is sufficient to model these differences. These findings improve our understanding of key intratumoral cell populations that may be harnessed by future immunotherapeutic strategies.

PATH-39. AI-BASED IDENTIFICATION OF GLIOMA IDH MUTATIONAL STATUS FROM H&E-STAINED WHOLE SLIDE IMAGES

Abstract INTRODUCTION Isocitrate-dehydrogenase (IDH) mutational status is diagnostically critical in adult gliomas, with prognostic and therapeutic implications. IDH status cannot be determined by sole histologic assessment and requires molecular testing. We hypothesize that AI-based analysis can accurately predict IDH mutational status in glioma, by retrieving sub-visual cues in H&E-stained digitized tissue sections. MATERIALS AND METHODS A retrospective discovery cohort of 1,534 cases (756/778 IDH-wildtype/IDH-mutant, including all available grades) from the TCGA-LGG/TCGA-GBM collections was used for model development with 10-fold cross-validation (CV). A replication cohort of 114 cases (82/32 IDH-wildtype/IDH-mutant) from the University of Pennsylvania Health System (UPHS) was used for independent hold-out evaluation. Each 20X magnification whole slide image (WSI) underwent comprehensive curation for elimination of artifactual content, followed by partitioning into 256x256 patches, and feature extraction using pre-trained i) ImageNet weights (baseline), and ii) self-supervised vision transformer (ViT). A weakly-supervised attention-based multiple-instance-learning framework distinguished cases as IDH-wildtype or IDH-mutant, while generating visually interpretable attention heatmaps. RESULTS Evaluation on discovery and replication cohorts with the ViT yield accuracy of 90.84% (AUCDiscovery=97.16%) and 93.94% (AUCReplication=97.36%), respectively, with high sensitivity (i.e., high confidence in correctly predicting IDH-wildtype glioma). Comparison of the ViT with the baseline model on the replication cohort indicates the ViT superiority by improvements of 8% and 9.5% in accuracy and AUC, respectively. Histologic assessment of heatmaps indicates IDH-wildtype tumors exhibiting distinct regions of significant pleomorphism and microvascular proliferation, while IDH-mutant tumors exhibit dense nodular cell concentrations, microcystic architecture, and gemistocytic cells. CONCLUSION Our accurate H&E-based computational determination of glioma IDH status, with interpretations aligned with human-identifiable features, can contribute to obviating the need for molecular analyses and enable expedited diagnosis even in community healthcare settings. This study robustly identifies morpho-IDH correlates and could enable expedited patient prognostic stratification.

PATH-52. DIFFERENCES IN GENE EXPRESSION, GENOME-WIDE METHYLATION AND METABOLITES IN ADULT DIFFUSE GLIOMAS

Abstract Metabolic reprogramming is a hallmark of cancer, including diffuse gliomas. IDH1 and IDH2 are key enzymes linking cellular metabolism to epigenetic regulation and redox states. IDH1/IDH2 enzymes catalyze the conversion of isocitrate to α-ketoglutarate, while the mutant enzymes increase the production of D-2-hydroxyglutarate, an oncometabolite that induces epigenetic changes, leading to gene expression alterations. Understanding how epigenetic alterations affect metabolic gene expression and metabolite levels in diffuse gliomas could help identify diagnostic biomarkers and therapeutic targets. We studied the expression of metabolism-related genes, metabolites, and genome-wide methylation status, which will facilitate an integral understanding of the metabolic changes in IDH-mutant and IDH-wildtype gliomas. Study design included 47 fresh frozen tissue samples. RNA was extracted and used for gene expression analysis using the NanoString platform (nCounter® Metabolic Pathways Panel), while extracted DNA was used for targeted sequencing with a panel interrogating 600 genes and for genome-wide methylation analysis using the Illumina Epic v2. In addition, unbiased metabolomic analysis by mass spectrometry was performed in 25 tissue and CSF samples out of 47 samples. Methylation status, gene expression and metabolite levels were compared between IDH-mutant and IDH-wildtype gliomas. Results showed significant differences in DNA methylation, gene expression and metabolites between IDH-mutant and IDH-wildtype gliomas. UPP1, an enzyme involved in uracil metabolism, was studied in depth to understand the interplay between DNA methylation, gene expression, and metabolic reprogramming in diffuse glioma. Epigenetic analysis revealed that the CpG sites within the UPP1 gene displayed reduced methylation in IDH wild-type compared to IDH-mutant tumors. IDH wild-type tumors showed higher UPP1 expression compared to their IDH-mutant counterparts. Consequently, we observed higher levels of uracil in tissue and CSF samples from patients with IDH wild-type gliomas. These data provide a framework to study the relationship between mutations, methylation, gene expression and metabolism in diffuse gliomas.

P10.09.A BLOOD-BASED CLASSIFICATION OF IDH-STATUS OF NON-ENHANCING GLIOMAS: A MACHINE LEARNING APPROACH

Abstract BACKGROUND Non-invasive determination of IDH mutational status in patients with glioma could offer significant therapeutic opportunities. While IDH wildtype tumors (e.g., GBM) typically show enhancement on MRI, IDH mutant tumors often lack this enhancement. However, relying solely on anatomic radiology may lead to misclassification, and currently, tissue acquisition is the primary method for assessing IDH status in gliomas. This limitation hampers the development of neoadjuvant or intraoperative therapeutic strategies based on IDH status; thus, more minimally invasive methods to determine IDH status are needed. In this study, we assessed immune cell proportions, which vary by glioma subtype, from pre-surgery peripheral blood samples as an alternative method of classifying the IDH status in gliomas without enhancement. MATERIAL AND METHODS First, we employed a highly accurate large language model (GPT-4-Turbo-128k) with over 99% accuracy to read radiology notes and exclude patients with enhancing gliomas. Then, we identified twelve immune cell subtypes from whole blood using deconvolution algorithms based on DNA methylation data. These immune cell subtypes, along with patient age, were integrated into a machine learning model (random forest) to predict IDH status (wildtype vs. mutant), leveraging conditional Generative Adversarial Networks to generate synthetic data and mitigate bias in the datasets. Two independent datasets were included for training and validating the model. RESULTS The random forest model had an area under the ROC curve (AUC) of 0.90 and accurately identified IDH status in 81% of a training set of 287 gliomas (65 wildtype and 222 mutant tumors at varied time points after diagnosis). In an independent validation data set of 99 gliomas (6 wildtype and 93 mutant) from pre-surgery blood samples, the AUC was 0.93, and accuracy predicted IDH status in 92% of cases with a sensitivity of 93% and specificity of 83%. CONCLUSION We could accurately predict IDH status in pre-surgical peripheral blood samples from patients with non-enhancing gliomas. This minimally invasive approach is a promising step toward reducing risk and exploring opportunities for earlier therapies based on IDH status.

Sex differences in the molecular profile of adult diffuse glioma are shaped by IDH status and tumor microenvironment.

Sex differences in adult diffuse glioma (ADG) are well-established clinically, yet the underlying molecular mechanisms remain inadequately understood. Here, we aim to reveal molecular features and cellular compositions unique to each sex in ADG to comprehend the role of sex in disease etiology. We quantified sex differences in transcriptome of ADG using multiple independent glioma patient datasets. Next, we delved into the single-cell landscape to examine sex differences in gene expression and cellular composition. To explore how sex influences disease progression, we analyzed paired samples from primary and recurrent ADG cases, aiming to identify sex-specific differences in molecular and cellular features. Our analysis revealed that mutations in isocitrate dehydrogenase (IDH) genes and the tumor microenvironment emerged as primary influencers of sex-differential molecular enrichments. In IDHwt tumors, genes in neuronal signaling pathway are found to be enriched in male tumors, while genes in hypoxia and inflammatory response pathways are enriched in female tumors. This pattern was reversed in IDHmut gliomas. We hypothesized that these distinctions could be attributed to heterogeneous cellular composition between sexes. Using single-cell data, we observed distinctive patterns of sex differences in cell states, cell composition and cell-cell interaction in IDHwt and IDHmut tumors separately. Further, by comparing molecular changes in paired primary and recurrent ADG samples, we identified sex-specific differences in molecular characteristics and cellular compositions of recurrent tumors. Our results provide a comprehensive multi-level characterization of sex differences in ADG, such findings provide novel insights into glioma disease progression in each sex.

Integrated electrophysiological and genomic profiles of single cells reveal spiking tumor cells in human glioma

Prior studies have described the complex interplay that exists between glioma cells and neurons; however, the electrophysiological properties endogenous to glioma cells remain obscure. To address this, we employed Patch-sequencing (Patch-seq) on human glioma specimens and found that one-third of patched cells in IDH mutant (IDHmut) tumors demonstrate properties of both neurons and glia. To define these hybrid cells (HCs), which fire single, short action potentials, and discern if they are of tumoral origin, we developed the single cell rule association mining (SCRAM) computational tool to annotate each cell individually. SCRAM revealed that HCs possess select features of GABAergic neurons and oligodendrocyte precursor cells, and include both tumor and non-tumor cells. These studies characterize the combined electrophysiological and molecular properties of human glioma cells and describe a cell type in human glioma with unique electrophysiological and transcriptomic properties that may also exist in the non-tumor brain.

Mutant IDH Modulates Suppressive Myeloid Populations in Malignant Glioma.

Mutations in the isocitrate dehydrogenase (IDH) genes IDH1 and IDH2 have critical diagnostic and prognostic significance in diffuse gliomas. Neomorphic mutant IDH activity has been previously implicated in T-cell suppression; however, the effects of IDH mutations on intratumoral myeloid populations remain underexplored. In this study, we investigate the influence of IDH status on the myeloid compartment using human glioma specimens and preclinical models. We performed RNA sequencing and quantitative immunofluorescence on newly diagnosed, treatment-naive IDH-mutant grade 4 astrocytoma and IDH-wild-type (IDH-WT) glioblastoma (GBM) specimens. We also generated a syngeneic murine model, comparing transcriptomic and cell-level changes in paired isogenic glioma lines that differ only in IDH mutational status. Among patient samples, IDH-mutant tumors displayed an underrepresentation of suppressive myeloid transcriptional signatures, which was confirmed at the cellular level with decreased numbers of intratumoral M2-like macrophages and myeloid-derived suppressor cells. Introduction of the mutant IDH enzyme into murine glioma was sufficient to recapitulate the transcriptomic and cellular shifts observed in patient samples. We provide transcriptomic and cellular evidence that mutant IDH is associated with a quantitative reduction of suppressive myeloid cells in gliomas and that introduction of the mutant enzyme is sufficient to result in corresponding cellular changes using an in vivo preclinical model. These data advance our understanding of high-grade gliomas by identifying key myeloid cell populations that are reprogrammed by mutant IDH and may be targetable through therapeutic approaches.

Valproic acid targets IDH1 mutants through alteration of lipid metabolism

Histone deacetylases (HDACs) have a wide range of targets and can rewire both the chromatin and lipidome of cancer cells. In this study, we show that valproic acid (VPA), a brain penetrant anti-seizure medication and histone deacetylase inhibitor, inhibits the growth of IDH1 mutant tumors in vivo and in vitro, with at least some selectivity over IDH1 wild-type tumors. Surprisingly, genes upregulated by VPA showed no enhanced chromatin accessibility at the promoter, but there was a correlation between VPA-downregulated genes and diminished promoter chromatin accessibility. VPA inhibited the transcription of lipogenic genes and these lipogenic genes showed significant decreases in promoter chromatin accessibility only in the IDH1 MT glioma cell lines tested. VPA inhibited the mTOR pathway and a key lipogenic gene, fatty acid synthase (FASN). Both VPA and a selective FASN inhibitor TVB-2640 rewired the lipidome and promoted apoptosis in an IDH1 MT but not in an IDH1 WT glioma cell line. We further find that HDACs are involved in the regulation of lipogenic genes and HDAC6 is particularly important for the regulation of FASN in IDH1 MT glioma. Finally, we show that FASN knockdown alone and VPA in combination with FASN knockdown significantly improved the survival of mice in an IDH1 MT primary orthotopic xenograft model in vivo. We conclude that targeting fatty acid metabolism through HDAC inhibition and/or FASN inhibition may be a novel therapeutic opportunity in IDH1 mutant gliomas.

Targeting Isocitrate Dehydrogenase (IDH) in Solid Tumors: Current Evidence and Future Perspectives.

The isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) enzymes are involved in key metabolic processes in human cells, regulating differentiation, proliferation, and oxidative damage response. IDH mutations have been associated with tumor development and progression in various solid tumors such as glioma, cholangiocarcinoma, chondrosarcoma, and other tumor types and have become crucial markers in molecular classification and prognostic assessment. The intratumoral and serum levels of D-2-hydroxyglutarate (D-2-HG) could serve as diagnostic biomarkers for identifying IDH mutant (IDHmut) tumors. As a result, an increasing number of clinical trials are evaluating targeted treatments for IDH1/IDH2 mutations. Recent studies have shown that the focus of these new therapeutic strategies is not only the neomorphic activity of the IDHmut enzymes but also the epigenetic shift induced by IDH mutations and the potential role of combination treatments. Here, we provide an overview of the current knowledge about IDH mutations in solid tumors, with a particular focus on available IDH-targeted treatments and emerging results from clinical trials aiming to explore IDHmut tumor-specific features and to identify the clinical benefit of IDH-targeted therapies and their combination strategies. An insight into future perspectives and the emerging roles of circulating biomarkers and radiomic features is also included.

Mutant IDH1 inhibition induces dsDNA sensing to activate tumor immunity.

Isocitrate dehydrogenase 1 (IDH1) is the most commonly mutated metabolic gene across human cancers. Mutant IDH1 (mIDH1) generates the oncometabolite (R)-2-hydroxyglutarate, disrupting enzymes involved in epigenetics and other processes. A hallmark of IDH1-mutant solid tumors is T cell exclusion, whereas mIDH1 inhibition in preclinical models restores antitumor immunity. Here, we define a cell-autonomous mechanism of mIDH1-driven immune evasion. IDH1-mutant solid tumors show selective hypermethylation and silencing of the cytoplasmic double-stranded DNA (dsDNA) sensor CGAS, compromising innate immune signaling. mIDH1 inhibition restores DNA demethylation, derepressing CGAS and transposable element (TE) subclasses. dsDNA produced by TE-reverse transcriptase (TE-RT) activates cGAS, triggering viral mimicry and stimulating antitumor immunity. In summary, we demonstrate that mIDH1 epigenetically suppresses innate immunity and link endogenous RT activity to the mechanism of action of a US Food and Drug Administration-approved oncology drug.

The IDH paradox: Meta-analysis of alkylating chemotherapy in IDH-wild type and -mutant lower grade gliomas.

IDH-wild type (-wt) status is a prerequisite for the diagnosis of glioblastoma (GBM); however, IDH-wt gliomas with low-grade or anaplastic morphology have historically been excluded from GBM trials and may represent a distinct prognostic entity. While alkylating agent chemotherapy improves overall survival (OS) and progression-free survival (PFS) for IDH-wt GBM and also IDH-mutant gliomas, irrespective of grade, the benefit for IDH-wt diffuse histologic lower-grade gliomas is unclear. We performed a meta-analysis of randomized clinical trials for World Health Organization (WHO) grades 2-3 gliomas (2009 to present) to determine the effect of alkylating chemotherapy on IDH-wt and -mutant gliomas using a random-effects model with inverse-variance pooling. We identified 6 trials with 1204 patients (430 IDH-wt, 774 IDH-mutant) that evaluated alkylating chemoradiotherapy versus radiotherapy alone, allowing us to perform an analysis focused on the value of adding alkylating chemotherapy to radiotherapy. For patients with IDH-wt tumors, alkylating chemotherapy added to radiotherapy was associated with improved PFS (HR:0.77 [95% CI: 0.62-0.97], P = .03) but not OS (HR:0.87 [95% CI: 0.64-1.18], P = .17). For patients with IDH-mutant tumors, alkylating chemotherapy added to radiotherapy improved both OS (HR:0.52 [95% CI: 0.42-0.64], P < .001) and PFS (HR = 0.47 [95% CI: 0.39-0.57], P < .001) compared to radiotherapy alone. The magnitude of benefit was similar for IDH-mutant gliomas with or without 1p19q-codeletion. Alkylating chemotherapy reduces mortality by 48% and progression by 53% for patients with IDH-mutant gliomas. Optimal management of IDH-wt diffuse histologic lower-grade gliomas remains to be determined, as there is little evidence supporting an OS benefit from alkylating chemotherapy.

The biological significance of tumor grade, age, enhancement, and extent of resection in IDH-mutant gliomas: How should they inform treatment decisions in the era of IDH inhibitors?

The 2016 and 2021 World Health Organization 2021 Classification of central nervous system tumors have resulted in a major improvement in the classification of isocitrate dehydrogenase (IDH)-mutant gliomas. With more effective treatments many patients experience prolonged survival. However, treatment guidelines are often still based on information from historical series comprising both patients with IDH wild-type and IDH-mutant tumors. They provide recommendations for radiotherapy and chemotherapy for so-called high-risk patients, usually based on residual tumor after surgery and age over 40. More up-to-date studies give a better insight into clinical, radiological, and molecular factors associated with the outcome of patients with IDH-mutant glioma. These insights should be used today for risk stratification and for treatment decisions. In many patients with IDH-mutant grades 2 and 3 glioma, if carefully monitored postponing radiotherapy and chemotherapy is safe, and will not jeopardize the overall outcome of patients. With the INDIGO trial showing patient benefit from the IDH inhibitor vorasidenib, there is a sizable population in which it seems reasonable to try this class of agents before recommending radio-chemotherapy with its delayed adverse event profile affecting quality of survival. Ongoing trials should help to further identify the patients that are benefiting from this treatment.