IDH-mutant Gliomas Research Articles

Multidisciplinary quantitative and qualitative assessment of IDH-mutant gliomas with full diagnostic deep learning image reconstruction

Rationale and Objectives: Diagnostic accuracy and therapeutic decision-making for IDH-mutant gliomas in tumor board reviews are based on MRI and multidisciplinary interactions. Materialsand Methods: This study explores the feasibility of deep learning-based reconstruction (DLR) in MRI for IDH-mutant gliomas. The research utilizes a multidisciplinary approach, engaging neuroradiologists, neurosurgeons, neuro-oncologists, and radiotherapists to evaluate qualitative aspects of DLR and conventional reconstructed (CR) sequences. Furthermore, quantitative image quality and tumor volumes according to Response Assessment in Neuro-Oncology (RANO) 2.0standards were assessed. ResultsAll DLR sequences consistently outperformed CR sequences (median of 4 for all) in qualitative image quality across all raters (p < 0.001 for all) and revealed higher SNR and CNR values (p < 0.001 for all). Preference for all DLR over CR was overwhelming, with ratings of 84% from the neuroradiologist, 100% from the neurosurgeon, 92% from the neuro-oncologist, and 84% from the radiation oncologist. The RANO 2.0 compliant measurements showed no significant difference between the CR and DRL sequences (p = 0.142). ConclusionThis study demonstrates the clinical feasibility of DLR in MR imaging of IDH-mutant gliomas, with significant time savings of 29.6% on average and non-inferior image quality to CR. DLR sequences received strong multidisciplinary preference, underscoring their potential for enhancing neuro-oncological decision-making and suitability for clinical implementation.

Homozygous CDKN2A/B deletions in low- and high-grade glioma: a meta-analysis of individual patient data and predictive values of p16 immunohistochemistry testing.

CDKN2A/B deletions are prognostically relevant in low- and high-grade gliomas. Data on this is derived from heterogeneous series, an accurate estimation of survival risk from homozygous CDKN2A/B deletion is missing. Besides genetic testing, p16-immunohistochemistry (IHC) as a less cost intensive means for indirect detection of CDKN2A/B alterations is possible but not validated in larger datasets. The present meta-analysis aimed to (1) reconstruct individual patient data (IPD) and estimate overall survival (OS) stratified by CDKN2A/B status from all literature and to (2) determine accuracy of p16 testing for CDKNA2/B detection from published studies. For survival analysis according to CDKN2A/B status 460 records were screened, four articles with 714 participants were included. In IDH-wildtype (IDH-wt) gliomas, 57.07% harbored the deletion compared to 9.76% in IDH-mutant (IDH-mut) gliomas. Median OS of patients with IDH-wt gliomas and homozygous CDKN2A/B deletion was 13.0 months compared to 18.0 months with non-deleted CDKN2A/B (p = 0.014, Log-Rank). With homozygous deletion of CDKN2A/B the risk of death was increased by 1.5 (95%-CI 1.1-2.1). Median OS in patients with IDH-mut gliomas without CDKN2A/B deletion was 92.0 months compared to 40.0 months with CDKN2A/B deletion (p < 0.001, Log-Rank). CDKN2A/B deletions were associated with a significantly shorter OS (HR = 3.2; 95%-CI 2.2-5.5). For p16 IHC analysis, 10 eligible studies with 1087 examined samples were included. The cut-off for retention differed between the studies. In 588/662 p16 retained cases CDKN2A/B deletions was not detected, implying a negative predictive value (NPV) of p16 staining of 88.8%. Conversely, 279/425 p16 absent cases showed a CDKN2A/B deletion resulting in a positive predictive value (PPV) of 65.6%. Sensitivity of p16 staining for CDKN2A/B detection was 79.0%, specificity 80.1%. Highest diagnostic accuracy of p16 IHC was reached with a cut-off of > 5% and within IDH-mut glioma.

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.

The Correlation Patterns of miRNA Expression with Targeted mRNA Transcripts in Glioma Patients with Wild-Type and Mutated Isocitrate Dehydrogenase (IDH) Genotypes.

Low-grade gliomas are divided into two main genetic phenotypes based on the presence or absence of mutations in the isocitrate dehydrogenase (IDH) genes. The mutated IDH phenotype (IDHmut), in contrast to the wild-type phenotype (IDHwt), is characterized by a more positive response to pharmacological intervention and a significantly longer survival time. In this study, we analyzed the differential co-expression of 225,000 microRNA-mRNA pairs at the level of correlations between microRNA levels and their potential mRNA targets. Analysis of the associative relationships of individual representatives of the selected pairs revealed that the level of mRNAs encoded by the ELN, ARL4C, C9orf64, PLAT, and FKBP9 genes associated with aggressive progression of glioma was increased in the IDHwt group. Meanwhile, the levels of miRNA-182, miRNA-455, and miRNA-891a associated with the negative prognosis in glioma were generally increased in the IDHmut group. Most (16/21) of the detected 21 microRNA-mRNA pairs with significant difference in regulation between IDHwt and IDHmut glioma samples had a weak or moderate positive correlation in IDHmut samples and a negative correlation in IDHwt samples. Therefore, our findings indicate that glioma samples from the IDHmut group with a positive prognosis potentially have a significantly less pronounced ability to microRNA-mediated regulation. We further suggest that such physiological disorders can lead to reduced tumor viability, resulting in an increased ability of the host to resist the spread of a malignant transformation of this genetic phenotype.

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.

DDDR-08. PIM1 INHIBITION: A NOVEL THERAPEUTIC APPROACH FOR IDH-MUTANT GLIOMAS

Abstract BACKGROUND PIM1, a constitutively active serine/threonine kinase has emerged as a relevant target in cancer treatment. Previously, we discovered that IDH-mutant glioma exhibited high kinase activity of PIM1, suggesting an increased sensitivity to PIM1 inhibition. Here we sought to investigate PIM1 inhibition as a therapeutic approach for IDH-mutant gliomas. METHODS Patient-derived IDH-mutant and wildtype glioma cell lines were used to evaluate the impact of PIM1inhibition on cell viability, proliferation, and colony formation. mRNA and protein expression of PIM1 were determined by qRT-PCR, Western-blot, and immunohistochemistry. m6A dot-blot assay was conducted to detect m6A RNA levels in glioma cells. m6A modification status and its location in PIM1 transcript were evaluated by m6A RNA immunoprecipitation array. RNA-binding protein immunoprecipitation was performed using anti-YTHDF1 antibody to identify PIM1 RNA-YTHDF1 protein interactions. RESULTS An upregulation of PIM1 protein is demonstrated in IDH-mutant patient tumor tissues and cells. A pan-PIM inhibitor, AZD1208 significantly suppressed cell viability, cell proliferation, and colony formation in IDH-mutant cells but had a lesser effect in IDH-wildtype cells. Interestingly, a lower mRNA expression of PIM1 is observed, while the protein level is significantly elevated in IDH-mutant cells compared to wildtype cells, suggesting post-transcriptional and translational regulation of PIM1 associated with IDH mutation. Mechanistically, 2-hydroxyglutarate, unique to IDH-mutant gliomas, competitively inhibits α-KG-dependent RNA demethylase FTO. We demonstrated suppressed FTO activity and an increased m6A RNA level in IDH-mutant glioma cells. An enrichment of m6A modification at motif “GG (m6A) CU” located in the 3’UTR is found in the PIM1 transcript. Furthermore, we demonstrated that the m6A modification site is recognized by the m6A reader YTHDF1 to promote protein translation, leading to an elevated PIM1 protein expression. CONCLUSION m6A RNA modification contributes to an enhanced PIM1 activity and thereby a selective sensitivity to PIM1 inhibition in IDH-mutant gliomas.

EXTH-70. PRECLINICAL TRIALS OF THE ANTIBODY-DRUG CONJUGATE TISOTUMAB VEDOTIN IN ADULT-TYPE DIFFUSE GLIOMAS

Abstract The gene F3, encoding Tissue Factor (TF), is expressed in many cancers and contributes to their malignancy. Our previous work showed that, among adult-type diffuse gliomas, IDH wild-type (IDHwt) glioblastomas (GBM) express more TF than IDH mutant (IDHmut) gliomas (PMID: 27664011, 30266764, 31222125). Tisotumab vedotin (TisVed), an anti-TF antibody conjugated to monomethyl auristatin E, is a therapeutic designed to target cells expressing TF. We therefore sought to determine the therapeutic potential of TisVed in IDHwt vs. IDHmut gliomas. TisVed was more active against cultured IDHwt GBM cells than IDHmut glioma cells (P&amp;lt;0.0001). This activity was increased by 55% with daily washout of the soluble TF secreted by IDHwt GBM cells. TisVed extended the survival of mice intracranially engrafted with IDHwt GBM by 34% (37.5 days vs. 28 days for IgG control, P=0.006), but not mice with IDHmut glioma (30.5 days, P=0.88). TisVed also reduced the growth of IDHwt GBM flank xenografts by 58% (P=0.045). Unconjugated Tis had no antitumor effect intracranially or in the flank. All 12 flank-engrafted mice treated with Tis, and 9/12 mice treated with TisVed, developed large hematomas in and around the flank xenografts, compared to 0/12 IgG-treated mice (P&amp;lt;0.001). Similarly, a semiquantitative analysis showed that intracerebral hemorrhage was more extensive in Tis/TisVed- treated GBM12 mice than in IgG-treated GBM12 mice (P=0.046). These data indicate that TisVed targets high TF-expressing IDHwt GBM, but not low TF-expressing IDHmut glioma, that its activity is predominately through the vedotin conjugate rather than inhibition of TF signaling, and that it may have some effect against TF-expressing IDHwt GBM, but intracerebral hemorrhage may be a concern.

EXTH-45. PRECLINICAL EVALUATION OF THE ANTIBODY-DRUG CONJUGATE TISOTUMAB VEDOTIN IN ADULT-TYPE DIFFUSE GLIOMAS

Abstract The gene F3, encoding Tissue Factor (TF), is expressed in many cancers and contributes to their malignancy. Our previous work showed that, among adult-type diffuse gliomas, IDH wild-type (IDHwt) glioblastomas (GBM) express more TF than IDH mutant (IDHmut) gliomas (PMID: 27664011, 30266764, 31222125). Tisotumab vedotin (TV), an anti-TF antibody conjugated to monomethyl auristatin E, is a therapeutic designed to target cells expressing TF. We therefore sought to determine the therapeutic potential of TV in IDHwt vs. IDHmut gliomas. TV was more active against cultured IDHwt GBM cells than IDHmut glioma cells (P&amp;lt;0.0001). This activity was increased by 55% with daily washout of the soluble TF secreted by IDHwt GBM cells. TV extended the survival of mice intracranially engrafted with IDHwt GBM by 34% (37.5 days vs. 28 days for IgG control, P=0.006), but not mice with IDHmut glioma (30.5 days, P=0.88). TV also reduced the growth of IDHwt GBM flank xenografts by 58% (P=0.045). Unconjugated tisotumab had no antitumor effect intracranially or in the flank. All 12 flank-engrafted mice treated with tisotumab, and 9/12 mice treated with TV, developed large hematomas in and around the flank xenografts, compared to 0/12 IgG-treated mice (P&amp;lt;0.001). Similarly, a semiquantitative analysis showed that intracerebral hemorrhage was more extensive in tisotumab/TV- treated GBM12 mice than in IgG-treated GBM12 mice (P=0.046). These data indicate that TV targets high TF-expressing IDHwt GBM, but not low TF-expressing IDHmut glioma, that its activity is predominately through the vedotin conjugate rather than inhibition of TF signaling, and that it may have some effect against TF-expressing IDHwt GBM, but intracerebral hemorrhage may be a concern.

DDDR-28. SPHINGOSINE AND N,N-DIMETHYLSPHINGOSINE MODULATE CHOLESTEROL HOMEOSTASIS IN IDH1MUT GLIOMAS, CAUSING PRO-APOPTOTIC MEMBRANE DEGRADATION

Abstract BACKGROUND Previous work in our team revealed a therapeutic potential for metabolic reprogramming along the sphingolipid pathway within IDH1mut gliomas. The dosing of various IDH1mut glioma subtypes with sphingosine and sphingosine kinase inhibitor (SphKi), N,N-dimethylsphingosine (NDMS) led to the enrichment of pro-apoptotic ceramides and sphingosines while suspending the proliferation of the IDH1mut neurospheres. However, the mechanism of drug action (MOA) was not well understood. We postulated that suppressing the production of S1P production with SphKi treatment combined with the global accumulation of sphingosine and certain ceramides triggers apoptotic membrane stress in highly susceptible IDH1mut gliomas. METHODS We cultured and treated IDH1mut (TS603, BT142 &amp; NCH1681) and IDH1wt (GSC923) glioma cell lines with a combination of NDMS and C17-sphingosine. We used a comprehensive multi-omics approach involving LC-MS lipidomic, RNA sequencing, immunoblotting, flow cytometry, and fluorescent microscopy. RESULTS Following combination treatment, a global increase in sphingosines, ceramides, and their derivatives was accompanied by a decline in cholesterol levels. The transcriptomic expression was elevated for NR1H3, MYLIP, ABCA1, and ABCG1, which regulate the trafficking and biosynthesis of membrane-associated cholesterol. In contrast, total gene expression for catalytic enzymes involved in cholesterol (and isoprenoid) biosynthesis was negatively impacted. In addition, early onset changes included an elevation in the expression of ROCK (a marker for apoptotic membrane blebbing). Moreover, fluorescent microscopy revealed that sphingosine and cholesterol tend to become associated within the cellular membranes. This interaction then saturates the plasma membrane (PM), causing apparent vesiculation, permeation, and disruption of membrane integrity. CONCLUSION The MOA involved concomitant attenuation of cholesterol metabolism and influx in response to membrane saturation with sphingosine-associated cholesterol. As a result, PM stress was incurred, leading to PM disintegration in a blebbing fashion. Our study revealed how reprogramming sphingolipid metabolism increases the susceptibility of IDHmut gliomas to sphingosine-associated cholesterol-induced blebbing and resulting cell death.

TMOD-36. NANOGLIOS: A MINIATURIZED PATIENT AVATAR ENABLING RAPID AND HIGH-THROUGHPUT PHENOTYPIC SCREENING FOR GLIOMAS

Abstract BACKGROUND Establishing pre-clinical patient derived orthtopical xenograft models for IDH-mutant and H3K27M-mutant gliomas has presented significant histological challenges. There is increasing interest in developing faithful patient-derived ex vivo 3D models to replicate these type of gliomas, aiming to accelerate drug screening efforts. METHOD We developed a rapid approach for establishing nanoliter-sized miniature glioma organoids, termed NanoGlios, by leveraging emulsion microfluidics. We tested the robustness of NanoGlios for neurosphere culture, glioma xenografts, and primary patient samples, focusing on H3K27M-mutant diffuse midline gliomas (DMGs) and IDH-mutant gliomas. We compared the morphological features of NanoGlios to their counterparts using high-content imaging, H&amp;E staining, and histological analysis. Additionally, we evaluated the mutation and gene expression profiles of NanoGlios relative to their counterparts. Furthermore, we conducted proof-of-concept drug screening of NanoGlios with various inhibitors and established an AI-enhanced high-throughput phenotypic imaging-based screening platform. RESULTS We successfully established NanoGlios from various sample sources, including neurospheres, xenografts, and primary tissues harboring H3K27M or IDH1/2 mutations. NanoGlios derived from xenografts or primary tissues captured more heterogeneous patterns as compared to the neuropsheres. We were able to generate thousands of NanoGlios and perform drug assays for hundreds of conditions at passage 0 within two weeks from both resected glioma tissues and xenografts. The AI phenotypic screening enabled us to identify differential responses of NanoGlios to ONC-201 treatment and drug responses to mutant IDH inhibitors Vorasidenib and Ivosidenib, which was not able to be detected by conventional cell viability bulk assays.

CTIM-07. A PHASE II, OPEN LABEL, SINGLE ARM STUDY OF NIVOLUMAB FOR RECURRENT OR PROGRESSIVE IDH MUTANT GLIOMAS WITH PRIOR EXPOSURE TO ALKYLATING AGENTS

Abstract BACKGROUND Somatic mutations in isocitrate dehydrogenase 1 (IDH1) are an important prognostic factor in patients with gliomas. IDH-mutant gliomas are prone to develop hypermutation after exposure to alkylating agents; in other solid tumors, hypermutation may be a predictive biomarker for PD-1 inhibitor response. In this study, we hypothesize that this specific subgroup of patients would clinically benefit from nivolumab as measured by overall response rate (ORR), duration of response (DOR), median progression-free survival (PFS), and overall survival (OS). METHODS Patients aged 18 years and older with recurrent/progressive IDH-mutant (WHO Grades 2-4) gliomas and prior exposure to alkylating agents were included. Nivolumab was given 240 mg q2w for 8 cycles, then 480 mg q4w until progression, toxicity, study withdrawal, or completion at 2 years. Efficacy was evaluated by ORR of partial (PR) and complete responses (CR) per RANO criteria; low-grade glioma RANO criteria was used for patients with non-enhancing disease. Secondary endpoints included median PFS, median OS, DOR. Toxicity and survival were followed every 3 months until death/study completion. RESULTS Thirty-three patients (66.7% male) were evaluable at study completion. Histology consisted of oligodendroglioma (N=11; 55% Grade 3) and astrocytoma (N=22; 36% Grade 3, 32% Grade 4). Median number of prior lines of systemic therapy was 1 (range 1-6). Median dexamethasone dose at screening was 0 mg (range 0-4). ORR was 9% with two PR and one CR, median DOR 33 months. 11 (32%) patients had stable disease (7 stable for 6+ months) with median PFS 2.2 months and median OS 29.1 months. There were two treatment-related Grade 3 adverse events (lymphopenia and hypotension). One patient discontinued treatment for Grade 2 transaminitis. CONCLUSIONS Though the primary endpoint was not met, three patients have ongoing CR and PR at 54+, 33+ and 24+ months. Future combination therapies with PD-1 inhibitors may increase efficacy in recurrent IDH-mutant gliomas.

PATH-07. CLINICAL AND RADIOGRAPHIC FEATURES OF DIFFUSE GLIOMA MOLECULAR SUBTYPES IN ACCORDANCE WITH THE 2021 CNS5 WHO CLASSIFICATION

Abstract BACKGROUND The incorporation of molecular parameters into WHO CNS5 has led to reclassification of many gliomas. We undertook to describe presenting clinical and radiographic features of diffuse adult gliomas according to CNS5 category. METHODS We reviewed pathology reports, clinical and MRI data at presentation of 972 adult patients with glioma diagnosed between 1/2010-2/2022. Continuous variables were presented as median; categorical variables as numbers and percentages, and comparison were performed using chi-square test. RESULTS 736 patients had sufficient data for CNS5 re-classification and were included in the analysis: Grade 2 IDH-mutant (IDHmut) astrocytoma (A2) n=69, grade 3 IDHmut astrocytoma (A3) n=37, grade 4 IDHmut astrocytoma (A4), n=32, grade 2 oligodendroglioma (O2) n=60, grade 3 oligodendroglioma (O3) n=23, and grade 4 IDH-wildtype glioblastoma (GBM) n=515. Age at presentation significantly differed between grade 2&amp;3 gliomas and grade 4 (mean age:39 versus 61.7; p&amp;lt;0.001); A4 were also younger than GBM (39.8 versus 63.1). A majority in all categories except grade 2 gliomas were male. Seizure was more common in IDHmut gliomas than GBM (58.4% versus 31.8%;p&amp;lt;0.001), while cognitive impairment was more common in GBM than in IDHmut gliomas (64.4% versus 34.4%;p&amp;lt;0.001) and in A4 (64% versus 40.6%;p=0.013). Focal deficits at presentation were more frequent in GBM than in IDHmut gliomas (74.2% versus 30.5%;p&amp;lt;0.001), but not significantly different compared to A4 (74.2% versus 58.1%;p=0.06). Contrast enhancement on MRI was more frequent in GBM compared to IDHmut gliomas (93% versus 48%;p&amp;lt;0.001), but similar frequency with A4 (93% versus 96%;p=1.00). The minimum ADC was significantly higher in IDHmut glioma including A4 than GBM(p&amp;lt;0.001). Calcification was more common in oligodendroglioma than astrocytoma and GBM (p&amp;lt;0.001), and more frequent in A4 than GBM(p=0.016). CONCLUSION Significant differences in the clinical presentation and radiologic features exist among CNS5 glioma subtypes, informing potential diagnosis, management and prognosis at initial presentation.

IMMU-13. TRANS-SPECIES STUDY OF IDH-MUTANT REPLICATION-REPAIR DEFICIENT HIGH-GRADE GLIOMAS (RRD-HGG) AND RESPONSE TO COMBINED TARGETED AND IMMUNOTHERAPY: AN IRRDC STUDY

Abstract BACKGROUND AND AIMS IDH-mutant gliomas comprise &amp;lt;10% of HGG in children. RRD-HGG comprise 5-10% of childhood HGG, demonstrate high mutation burden (TMB) and respond to immune-checkpoint inhibition (ICI). The impact of RRD within childhood IDH-mutant gliomas, and effective therapeutic options for these patients are not well-established. METHODS Clinical and multi-omic analyses were performed on IDH mut -RRD-HGG registered to the IRRDC and the Toronto glioma taskforce. Tumor development and genomic data were studied from a novel IDH mut -RRD-HGG immunocompetent mouse model. Outcome following ICI and targeted therapy were evaluated. RESULTS RRD was detected in &amp;gt;60% of childhood IDH mut -HGG. IDH1-mutations were detected in 20% of RRD-HGG. All patients with IDH mut -RRD-HGG (n=49) harboured germline variants in MMR genes (CMMRD: 65%, Lynch: 35%). Contrary to sporadic IDH mut -gliomas, &amp;gt;91% were WHO-grades 3/4. Diffuse/multifocal disease involving the frontal lobe was frequent. TMB (median: 28 mutations/Mb) was lower than IDH-wildtype RRD-HGG (p&amp;lt;0.05). POLE/POLD1 mutations were absent. TP53 and ATRX were frequent somatic hits. Copy number changes, particularly CDKN2A/2B loss, were common. IDH-mutation contributed to an immune-suppressed microenvironment, with lower CD8-T-cell infiltration (immunohistochemistry) and tumor-inflammation scores (transcriptome; p&amp;lt;0.05). A novel mouse model (Olig2-Cre+/Msh2LoxP/LoxP/LSL-Idh1R132H/+) demonstrated similar TMB, diffuse cerebral involvement, slower growth, and lesser immune infiltrates as compared with IDH-wildtype RRD-HGG models, and provided opportunity for therapeutic testing. RRD contributed to poor survival in IDHmut-gliomas (p&amp;lt;0.001). Despite hypermutation, ICI monotherapy resulted in inferior survival in IDH mut -RRD-HGG vs IDH-wildtype RRD-HGG (p&amp;lt;0.05). Five patients developed metachronous IDH mut -RRD-HGG while on anti-PD1 treatment for IDH-wildtype RRD-HGG, suggesting intrinsic ICI-resistance. Ivosidenib (IDH-inhibitor) demonstrated objective response in 4/8 IDH mut -RRD-HGG. Furthermore, for IDH mut -RRD-HGG on ICI treatment, the addition of ivosidenib prolonged survival at 12-months in comparison to those without IDH-inhibition (p=0.01) CONCLUSIONS Hypermutant RRD-HGG with IDH1;p.R132H harbour unique immuno-biology and do not respond to anti-PD1 monotherapy. The addition of IDH-inhibition demonstrated favourable responses, supporting need for evaluation of the combination in clinical trials.

CTNI-65. DETERMINANTS OF LONG-TERM SURVIVAL IN PATIENTS WITH IDH-MUTANT GLIOMAS

Abstract BACKGROUND Survival times of patients with IDH-mutant gliomas is variable and can extend to decades. Many studies only provide progression-free rather than overall survival times and prognostic factors remain ill-defined. Here we explored clinical disease characteristics of long-term survivors within a large cohort of patients with extended follow-up. METHODS This single-center, retrospective analysis included 114 patients with IDH-mutant glioma that either died due to tumor-related causes or survived at least 15 years after first diagnosis. Patient characteristics, outcomes, and prognostic factors were stratified by short- (&amp;lt; 15 years) versus long-term (≥ 15 years) survival. Initial diagnosis was (re-)classified according to the WHO 2016 classification. Uni- and multivariate analyses were performed. RESULTS Overall, 66 patients (58%) were diagnosed with astrocytoma and 48 patients (42%) with oligodendroglioma. Median follow-up of the survivors was 16.6 years (range 15-28.9). 62 tumor-related deaths (54%) had been reported at database closure. Patients that died before reaching 15 years survival time had been diagnosed with astrocytoma in 41 (72%) and oligodendroglioma in 16 cases (28%). The long-term survivor cohort comprised 25 patients (44%) with astrocytoma and 32 patients (56%) with oligodendroglioma. Long-term survival was associated with grade 2 histology (p &amp;lt; 0.01), smaller tumor volumes (p &amp;lt; 0.01), and lack of contrast enhancement (p = 0.02). 41 long-term survivors (72%) were initially monitored by a wait-and-scan strategy as opposed to 16 patients (28%) in the cohort of short-term survival (p&amp;lt;0.01). In patients with oligodendroglioma, a higher KPS was associated with longer survival (HR 0.25; 95% CI 0.02-0.67; p = 0.05). Tumor resection (HR 0.43; 95% CI 0.22-0.86; p = 0.02) and wait-and-scan strategies (HR 0.34; 95% CI 0.14-0.79; p = 0.01) were associated with longer survival in patients with astrocytoma. CONCLUSION Tumor resection followed by a wait-and-scan strategy may yield excellent survival times, especially in patients with IDH-mutant grade 2 astrocytoma. Age appears not be an important predictor for survival in patients with IDH-mutant gliomas.

CTNI-57. REAL-WORLD EVIDENCE OF THE FREQUENCY OF POST-SURGICAL INITIAL OBSERVATION IN PATIENTS WITH IDH-MUTANT GLIOMA

Abstract BACKGROUND After initial resection of IDH-mutant gliomas, initial observation is an option for low-risk patients, to postpone potential therapy-related toxicities without impacting survival. Since NCCN guidelines for risk stratification are considered controversial, we aimed to assess the frequency of initial observation in IDH-mutant gliomas using real-world evidence in the US. METHODS We evaluated Optum claims from 2016-2021 of patients age ≥12 with ICD-10 codes for incident glioma. Since ICD-10 does not distinguish IDH-mutant from IDH-wild type (GBM) gliomas, we excluded patients with factors indicative of GBM (age&amp;gt;60/ hypofractionated RT/ carmustine wafer/ tumor-treating fields) to create a baseline cohort. We then evaluated the subgroup age &amp;lt;40 (&amp;lt;40 cohort), who have higher IDH-mutant incidence. To distinguish initial observation from surgical recovery, we assessed time from index surgery to first intervention. When 90% of first interventions had occurred those still without intervention were considered to undergo initial observation. RESULTS The baseline and &amp;lt;40 cohorts comprised 5404 and 646 patients, respectively. In the baseline cohort, 56% had intervention by day 90 (most commonly concurrent chemoradiation); peak intervention rate was 60%. Initial observation rate was 44%. In the &amp;lt;40 cohort, 47% had an intervention by day 90; peak intervention rate was 52%. Initial observation rate was 53%. CONCLUSIONS In both cohorts ≥90% of patients with likely IDH-mutant gliomas not initiating treatment within 3 months post-surgery continued in active observation. Initial observation rates were 44-53%, well above the levels we had expected based on NCCN guidelines. The cohorts likely include some GBM patients, for whom observation is unaccepted, and therefore these rates may understate actual initial observation rates for IDH-mutant glioma patients. These findings suggest that clinicians and patients have chosen initial observation over existing treatments, evincing a need for additional treatment options for this younger group of patients with considerably long life expectancy.

NCOG-30. FACTORS ASSOCIATED WITH COGNITIVE IMPAIRMENT IN LONG-TERM IDH-MUTANT GLIOMA SURVIVORS

Abstract BACKGROUND Cognitive deficits are less frequent in patients with IDH mutant (IDHm) than wild-type glioma at time of diagnosis, but less is known about factors that influence cognition for long-term survivors. We evaluated long-term neuropsychological outcomes of patients with IDHm glioma. METHODS Patients with IDHm glioma who underwent clinical neuropsychological assessment at Massachusetts General Hospital 3+ years after diagnosis were included and compared to a previously published cohort of IDHm patients tested prior to surgery/diagnosis (Wefel et al., 2016). RESULTS Sixty-three patients were included for analysis (54% male; median age 45 [range 21-75], 16 years of education, and 7 years from diagnosis [range 3-28]). Thirty-eight patients (60%) had astrocytoma and 24 (40%) had oligodendroglioma; 28 (44%) were grade 2, 31 (49%) grade 3, and 4 (6%) grade 4. Fifty-five patients (87%) received radiation and 51 (81%) received chemotherapy. Cognitive impairment (defined as Z score ≤-1.5 on at least two tests) was present in 30 patients (48%) and was associated with lower scores in neurologic aspects of quality of life (QoL; p&amp;lt;0.01), grade 3/4 relative to grade 2 (p=0.01), older age (p=0.03), left-sided lateralization (p=0.02), and less education (p=0.01). Prior radiation (p=0.07) and photon relative to proton radiation (p=0.06) trended toward poorer function. Tumor type, disease duration, and extent of resection were not significant factors (p&amp;gt;0.05). Compared to the pre-diagnosis IDHm cohort, impairment was significantly more common on tests of processing speed and language. Learning/memory, executive, and visuospatial function trended toward more frequent impairment but were not significant. Attention had comparable frequency of impairment. CONCLUSIONS In this cohort of IDHm glioma survivors, neurocognitive deficits were common, related to QoL, and associated with some expected clinical factors, but not length of survival. These findings emphasize the need for future research on interventions aimed at supporting patients’ neurocognitive abilities and QoL.

EPCO-26. LONGITUDINAL SINGLE-CELL ANALYSES IDENTIFY DRIVERS OF GENETIC, EPIGENOMIC, AND CELLULAR EVOLUTION IN IDH-MUTANT GLIOMA

Abstract Despite standard treatment, IDH-mutant gliomas inevitably recur. Previous studies suggest that therapeutic resistance may result from a combination of intratumoral cellular heterogeneity, epigenetic evolution, and acquired genetic alterations. However, how these multilayered molecular features interact to influence the evolutionary paths of IDH-mutant gliomas remains incompletely understood. To chart this evolution, 75 glioma samples were longitudinally collected from 35 patients (n = 13 oligodendroglioma, n = 22 astrocytoma) and profiled using single nucleus RNA sequencing, ATAC sequencing, and bulk DNA sequencing. Analysis of 331,016 nuclei (snRNA) revealed eight tumor microenvironment cell types and five pan-IDH-mutant malignant cellular states. The malignant states were distributed along a cellular hierarchy of stem-like populations (neural progenitor cell-like, oligodendrocyte progenitor cell-like, and undifferentiated) and more differentiated populations (mesenchymal-like and astrocyte-like) with cycling cells enriched in the stem-like populations. Joint open chromatin accessibility data (snATAC) was available for a subset of malignant cells (71,088 nuclei) and state-specific differentially accessible peaks supported that these states are epigenetically encoded. Across both subtypes, higher tumor grade was associated with reduced malignant cell differentiation and increased cycling populations. Between the two time points, we identified that there was a longitudinal increase in the cycling, undifferentiated, and mesenchymal-like populations with a corresponding decrease in the astrocyte-like population. Longitudinal genetic analysis revealed that 19 of 35 tumors acquired at least one of the following genetic alterations: treatment-associated hypermutation, cell cycle alteration, or large changes in copy number alterations. Importantly, tumors that acquired these key genetic alterations demonstrated significant shifts towards reduced differentiation and increased cycling populations while those tumors that did not had more stable malignant profiles. Collectively, our results suggest a common cellular hierarchy across IDH-mutant gliomas with a shift towards reduced differentiation, increased cycling populations during disease progression that is driven by acquired genetic alterations.

TMET-12. TARGETING AMINO ACID METABOLIC VULNERABILITIES IN GLIOMAS

Abstract Isocitrate dehydrogenase (IDH) wild-type gliomas and IDH mutant gliomas are similar in histologic appearance but differ in their genetic and metabolic backgrounds. This study aimed to reveal the difference in metabolites between IDH-wildtype glioma and IDH-mutant glioma, and to find the effective treatment according to the status of IDH in gliomas. Two artificial cell lines made from human astrocyte were used: NHAE6E7hTERTRas (IDH-wildtype) and NHAE6E7hTERTIDHmut (IDH-mutant). Using capillary electrophoresis- and ion chromatography–coupled mass spectrometry, the amount of asparagine was lower in NHAE6E7hTERTRas cells compared with NHAE6E7hTERTIDHmut cells, whereas the amount of glutamine, glutamate and 2-oxoglutarate in NHAE6E7hTERTIDHmut cells was lower than NHAE6E7hTERTRas cells. L-asparaginase, which converts asparagine into aspartate, was more effective in NHAE6E7hTERTRas cells than NHAE6E7hTERTIDHmut cells. L-asparaginase induced autophagy and inhibition of autophagy by 3-MA suppressed L-asparaginase-induced antitumor effect, which meant that the antitumor effect was at least partially due to the L-asparaginase-induced autophagy. Pharmacological or genetical inhibition of GLUD1 which converts glutamate to 2-oxoglutarate, suppressed proliferation of the cells by inducing ROS and apoptosis in NHAE6E7hTERTIDHmut cells. ROS inhibitor, NAC suppressed GLUD1 inhibitor-induced ROS, apoptosis, and cytotoxicity in NHAE6E7hTERTIDHmut cells, revealing that cytotoxicity by GLUD1 inhibitor was at least partially due to the inhibitor-induced ROS. The experiments using mutant IDH1 overexpressed glioma cell line showed similar sensitivity to GLUD1 inhibitor to NHAE6E7hTERTIDHmut, which suggested that the difference of sensitivity to GLUD1 inhibitor was due to the status of mutant IDH. In vivo experiments, L-asparaginase suppressed the growth of xenografted glioma cells without mutant IDH, whereas GLUD1 inhibitor inhibited the proliferation of xenografted glioma cells with overexpressed mutant IDH1. In conclusion, L-asparaginase and Glud1 inhibitor will be new therapeutic option for IDH-wildtype glioma and IDH-mutant glioma, respectively.

QOL-09. “I’VE DEFINITELY CHANGED WHO I AM”: A THEMATIC ANALYSIS OF CHALLENGES FACED BY PATIENTS WITH IDH-MUTANT GLIOMAS

Abstract BACKGROUND Despite their more favorable prognosis, patients with isocitrate dehydrogenase (IDH)-mutant gliomas experience significant psychological distress across their disease trajectory. The specific drivers of distress in this population are poorly understood, and interventions to support these patients are lacking. We sought to better characterize drivers of distress and ways to enhance support for patients with IDH-mutant gliomas as they confront potentially prolonged but ultimately unpredictable survival with an incurable, progressive brain tumor. METHODS Eligible individuals were English-speaking adults ages 65 and younger diagnosed with an IDH-mutant glioma at a single academic medical center. We conducted semi-structured interviews (N=20) to better understand the distress patients experience as they progress along their illness trajectories. We audio-recorded and transcribed interviews and analyzed interview content using reflexive thematic analysis. RESULTS Distress among participating patients (female = 7/20; age &amp;lt;40 years = 10/20) was linked to their struggles to maintain a coherent sense of identity while adapting to 1) neurocognitive changes and 2) altered life plans amidst prognostic uncertainty. Patients described feeling self-conscious and distrustful of their cognitive capacities, frequently comparing their current performance to their premorbid functioning (e.g. by checking their symptoms and indexing perceived deficits against scan results). Patients felt overwhelmed by the need to make decisions and set priorities (e.g., regarding family planning, retirement) due to the uncertain timing of their tumor’s progression and its unclear impact on their future functioning. Patients identified the transition from treatment to surveillance as a time of heightened worry, as they shifted their focus from the logistics of cancer care to the task of planning – financially, occupationally, socially – for an uncertain future. CONCLUSION These findings highlight the need for tailored supportive care to help patients with IDH-mutant gliomas cope with the distress caused by significant changes to identity and ongoing uncertainties regarding their future.

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.