Abstract Background Atypical teratoid/rhabdoid (AT/RT) is an aggressive pediatric brain tumor, prevalent in children < 3 years(Y) of age; methylation profiles subclassify AT/RTs into AT/RT-MYC (MYC), AT/RT-SHH (SHH), and AT/RT-TYR (TYR). At the molecular level, AT/RT shows alterations in SMARCB1 and SMARCA4. Despite treatment with surgery, radiation (RT), and chemotherapy, the cure rate remains suboptimal, and predictors of clinical outcome are unknown. Methods We studied the association between minimal residual disease (MRD), as measured by circulating tumor DNA (ctDNA) in CSF over multiple longitudinal time points on therapy, and outcomes in newly diagnosed, non-metastatic patients enrolled in phase 2 clinical trial SJATRT, stratum B1 (age < 36 months). Whole-genome cell-free DNA methylomes were generated for copy number variation profiling and tumor classification by an integrative computational pipeline (M-PACT). Treatment consisted of postoperative induction chemotherapy till >12 months of age, and then focal RT followed by consolidation chemotherapy and maintenance with alisertib. Results There were 30 evaluable patients (SHH: 18, TYR: 7, MYC: 3, NC: 2); the 3Y progression free survival (PFS) and overall survival (OS) were 30.0% and 56.7%. Methylation class was not associated with outcome. A total of 94 CSF samples from 24 patients (SHH:12, TYR:7, MYC:3, NC: 2) were analyzed. The median number of CSF samples per patient was 4 (range: 1-8). Positive MRD as a time-dependent covariate was associated with worse PFS and OS (p = 0.041, 0.037). Furthermore, based on landmark analyses, positive MRD at the end of induction chemotherapy (n = 7/18) and focal RT (n = 8/14) were associated with worse PFS (p = 0.012, 0.047). Conclusion PFS and OS were not improved for patients enrolled in stratum B1 in SJATRT. Positive ctDNA in CSF, as a time-dependent covariate, was associated with worse PFS and OS in non-metastatic ATRT. Positive MRD at early time points may predict early relapse and serve as a biomarker for therapeutic intervention in future trials.

Abstract Background Pediatric high-grade gliomas (pHGGs), including diffuse midline glioma (DMG) and diffuse intrinsic pontine glioma (DIPG), are the leading cause of central nervous system tumor-related morbidity and mortality in children. Neuronal activity promotes pHGG growth; one key mechanism is neuronal activity-regulated shedding of neuroligin-3 into the glioma microenvironment, mediated by protease ADAM (A Disintegrin and Metalloprotease) 10. INCB007839, an ADAM 10 inhibitor, was used in this trial to block neuroligin-3 shedding and thus strategically target the tumor microenvironment that promotes glioma progression. Study objectives were to determine safety, tolerability and recommended phase 2 dose. Methods Patients aged 3-21 years old with recurrent/progressive pHGGs, including DMG and DIPG, were eligible. One dose level (120mg/m2/dose twice daily [BID]) was tested, and the trial was amended to require prophylactic anticoagulation with enoxaparin due to a thrombotic toxicity. Results Twelve of 13 subjects were eligible, and 10 were evaluable for dose-confirming. Median age was 14.5 years (4.9-20.7 years). Diagnoses included: DIPG (n = 7, 58.3%), glioblastoma multiforme (n = 4, 33.3%), and anaplastic astrocytoma (n = 1, 8.3%). All patients were treated at DL1 and remained on study for 1-4 courses. The most common toxicities were lymphopenia, transaminitis, and fatigue. Nine patients progressed/relapsed during active treatment, 1 died on treatment, and 2 withdrew (1 prior to therapy). Conclusions INCB007839 was tolerated in the majority of patients at 120mg/m2/dose BID with concurrent prophylactic anti-coagulation, although MTD was not declared. These data, combined with foundational preclinical studies targeting neuron-cancer interactions, open the door to possible cancer neuroscience strategies for pediatric high-grade gliomas.

Previous studies have demonstrated poor outcomes in pediatric patients with H3 G34-mutant diffuse hemispheric glioma (DHG). However, the biological basis for this therapeutic resistance remains poorly understood. Furthermore, the effectiveness of temozolomide (TMZ) and the role of surgery in pediatric patients remain uncertain. Therefore, we performed a multi-institutional retrospective analysis of the clinical, imaging, and molecular characteristics of 36 pediatric (≤ 18 years) patients with newly diagnosed H3 G34-mutant DHG. The median age of the cohort was 14 years (8–18 years). The median progression-free survival (PFS) was 0.7 years (95

Abstract Background Aberrant activation of mesenchymal epithelial transition receptor (MET) contributes to tumorigenesis in pediatric CNS tumors making MET inhibition a potential therapeutic target in this patient population. Methods We conducted a first-in-children multicenter phase 1 trial of the MET inhibitor savolitinib in children with recurrent or refractory MB, HGG and DIPG. The main objectives were to determine the maximum tolerated dose (MTD)/recommended phase 2 dose (RP2D) of once-daily oral savolitinib and to characterize its toxicity and pharmacokinetics. Following MTD/RP2D determination, an efficacy expansion cohort limited to participants with MET-altered CNS tumors (including MET mutations, fusions, MET/HGF amplification) was activated. Results 41 participants were enrolled (median age: 12.5 years, range 5.3-21.5); 1 was deemed ineligible. The MTD/RP2D was 350 mg/m2 (dose level 3). Two dose-limiting toxicities (grade 3 fatigue, grade 3 ALT elevation) occurred. The most common grade 3/4 adverse events at least possibly related to savolitinib were neutropenia and lymphopenia (each 3/39, 7.7%), and leukopenia (2/39, 5.1%). In the phase 1 cohort, 1 objective response was observed in a participant with recurrent HGG on dose level 1 (150 mg/m2); they completed 39 treatment courses. Three participants with progressive DIPG remained on treatment for a median of 9 courses (range 4-24); One other participant with recurrent HGG remained on treatment for 7 courses.The median (range) apparent oral savolitinib clearance was 30.4 L/h/m2 (3.8–220.1) and half-life 3.2 hr (1.63–22.2). Among 7 participants with tumors harboring MET aberrations, 2 demonstrated sustained stable disease for 6 courses, while none achieved an objective response. Correlative analyses are underway. Conclusions Savolitinib was well-tolerated in children with recurrent CNS tumors with an MTD/RP2D of 350 mg/m2. Although preliminary antitumor activity was observed in the phase 1 cohort, no objective responses were seen in biomarker-selected participants.

GD2-CAR T cell therapy has demonstrated clinical benefit in patients with H3K27M + diffuse midline glioma (DMG), but the durability of response has been limited in many patients 1,2 . To identify mechanisms of therapeutic resistance, we conducted longitudinal single-cell RNA and TCR sequencing of cerebrospinal fluid (CSF) lymphocytes from DMG patients receiving intravenous followed by sequential intracerebral GD2-CAR therapy, with lymphodepleting chemotherapy administered once prior to the start of CAR T cell therapy ( NCT04196413 ). CSF GD2-CAR T cells manifested limited persistence and clonal expansion, while non-engineered CSF lymphocytes underwent significant clonal expansion and repertoire stabilization, ultimately dominating the CSF immune compartment. Concurrently, peripheral blood CD4 + and CD8 + T cells manifested anti-CAR immune reactivity targeting epitopes enriched within murine-derived and engineered junctional regions of the CAR construct. This was associated with appearance of circulating Human Anti-CAR Antibodies (HACAs) that bound cells expressing the GD2-CAR, as well as clonal expansion of CSF B cells which produced HACA which impeded the cytotoxic activity of GD2-CAR T cells. In several cases, appearance of circulating HACA temporally correlated with disease progression and across the patient population, and levels of circulating HACA inversely correlated with circulating CAR T cell persistence. These findings reveal robust induction of systemic and CNS adaptive T cell and B cell responses to GD2-CAR T cells following intravenous then sequential intracerebroventricular GD2-CAR therapy and provide strong evidence that anti-CAR immunity is a significant contributor to therapeutic resistance in this setting.

Lay Summary The Pediatric Brain Tumor Consortium Study, PBTC-N13, sampled 100 archived patient samples from four different pediatric, malignant brain tumor categories. Each sampled was tested for PD-1 and PD-L1 expression and then had more extensive immune profiling. This enables learning how the immune landscape differs among tumors in order to better understand the potential for individual tumor immune profiling to be considered in patient-directed care.

Abstract Introduction The PBTC conducted a re-treatment study evaluating selumetinib, a MEK I/II inhibitor, in children with recurrent/progressive pLGG. Eligible patients were previously enrolled on the phase 1/2 PBTC-029 trial, had stable disease (SD) for a minimum of 12 courses or had response during initial selumetinib exposure, and then progressed after coming off selumetinib. Methods Dual primary endpoints were objective response and 12-month disease stabilization. The secondary endpoint was PFS. The design followed a bivariate binomial two-stage approach. Results Thirty-five patients who progressed at a median of 7.3 months (0.3-40.8) post-initial therapy were enrolled. 19/35 (54%) had optic pathway/hypothalamic location, and 21/35 (60%) had pilocytic astrocytoma histology. Success criteria were > 7 objective responses or > 23 patients with SD after 12 cycles. The overall response rate was 5/35 (14.3%), 4 within 12 courses, all partial responses. Thirty patients (86%) had stable disease for ≥ 12 cycles. Median treatment duration was 27.6 months (1.9-86.9), and median response duration was 22.4 months (5.8-65.6). The 1-year/5-year PFS were 88.6%/55.6%, respectively. There was no association between re-treatment PFS to BRAF aberration, initial response to re-treatment response, or initial PFS to re-treatment PFS. 17/35 (49%) patients progressed, 9 on re-treatment and 8 after stopping re-treatment. The most common attributable toxicities were grade 1 and similar to initial treatment. The most common grade 3/4 toxicity was elevated CPK among 5/35 (14.2%) patients. There were no notable differences in toxicities between initial therapy and re-treatment. Conclusion Per the statistical design, re-treatment with selumetinib was effective in pLGG patients who had recurred/progressed after stopping initial treatment with selumetinib. The observed benefit was primarily in prolongation of PFS with 86% of patients being progression-free at 1-year, and over 50% being progression free at 5 years. Selumetinib re-treatment should be considered in patients who benefited from initial treatment.

Abstract GD2-CAR T cell therapy has demonstrated clinical benefit in a high fraction of patients with H3K27M-mutant diffuse midline glioma (DMG), but the durability of response is limited in some patients. To define mechanisms underlying loss of response and immune adaptation during repeated CAR T infusions, we performed longitudinal single-cell RNA and TCR sequencing of over 1.4 million cells from apheresis, CAR T products, and serial cerebrospinal fluid (CSF) samples collected from 13 DMG patients enrolled in the Arm A of Phase I trial of GD2-CAR T cell therapy (NCT04196413). Our analysis showed that GD2-CAR T cells exhibited poor persistence in vivo, with minimal detection after six infusion cycles. Endogenous lymphocytes, rather than CAR T cells, dominated the CSF immune compartment over time. Regulatory T cells were significantly enriched in non-responders, suggesting a suppressive immune milieu that limits CAR T activity. Among four patients who initially achieved sustained responses for over 12 months, three eventually relapsed after repeated infusions. Flow cytometry detected presence of anti-CAR antibodies in plasma and CSF these three patients, accompanied by an increase in CSF B cells at the time of relapse, indicating a humoral anti-CAR immune response. Complementing this, single-cell TCR-seq revealed that endogenous T cell repertoires stabilized toward the end of treatment in these relapsed responders, with hyper-expanded clonotypes exhibiting activated, cytotoxic, and exhausted transcriptional programs. In contrast, two durable responders maintained a dominant tissue-resident memory-like T cell program, which is associated with sustained disease control. Altogether, we generated a comprehensive longitudinal collection of single-cell sequencing and other omics data, and identified co-existing humoral and cellular mechanisms of anti-CAR immunity that emerge during prolonged GD2-CAR T cell therapy. These findings illuminate correlates of response and resistance and underscore the immunogenic potential of CAR constructs. Future approaches to reduce host immunoreactivity could improve CAR T cell efficacy and durability in solid tumors. Citation Format: Yiyun Chen, Kun-Wei Song, Moksha H. Desai, Ying-Wen Huang, Nadya Iswari, Zachary J. Ehlinger, Hossein Daghagh, Maximilian R. Koch, Kevin Reynolds, Kelvin C. Mo, Kristin C. Tsui, Skyler Rietberg, Mark P. Hamilton, Snehit Prabhu, Sonia Partap, Jasia Mahdi, Emily Egeler, Steven A. Feldman, Sabine Heitzeneder, Sean A. Yamada-Hunter, Elena Sotillo, Bita Sahaf, Zinaida Good, Michelle Monje, Sneha Ramakrishna, Crystal L. Mackall. Longitudinal single-cell atlas of GD2-CAR T cell therapy in H3K27M-mutant diffuse midline glioma identifies humoral and cellular anti-CAR immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 6463.

Abstract Background Phosphoproteomics studies have identified a critical role for the protein kinase CK2 in SHH MB growth by affecting the terminal-most components of the pathway. CX-4945 is an oral selective inhibitor of CK2 with promising preclinical data in medulloblastoma models. Methods PBTC conducted the first multicenter prospective clinical trial of CX-4945 in children and adults with recurrent SHH MB. There were 3 components: phase 1 for skeletally immature patients; phase 2 for skeletally mature patients/adults; and a surgical arm for patients undergoing tumor resection. Stratification by skeletal maturity addressed potential growth plate toxicities with SHH pathway inhibition. SHH subgroup was confirmed by methylation profiling. The phase 1 component followed the rolling-6 design, phase 2 used a Simon two-stage minimax design to assess objective response rate. Results Six subjects (median age: 10.8 years, range 8.3-13.9) were enrolled to the phase 1 component. No dose-limiting toxicities occurred at dose level 2 (800 mg/m2 twice a day given continuously); growth plate toxicities were not observed. Twelve participants (of 16 planned for interim efficacy analysis, median age 24.7 years [range 13.3-42]) enrolled in the phase 2 arm, with no objective responses observed. Three of the planned 6 surgical arm subjects had tumors analyzed for CX-4945 brain tissue concentrations, and 2/3 had measurable concentrations (54 and ∼131 ng/ml). Grade 3/4 adverse related events were limited to hypokalemia and nausea experienced by one subject. Two participants achieved sustained stable disease and remained on treatment for 6 and 9 cycles. Trial enrollment was terminated early due to discontinuation of consortium funding. Conclusions CX-4945 was well tolerated in patients with recurrent SHH MB at doses reaching 800 mg/m2 twice daily, a final maximum tolerated dose was not determined. Early study closure restricts definitive conclusions; however the lack of an efficacy signal suggests limited activity in this patient population.

H3K27M-mutant diffuse midline gliomas (DMG) are aggressive pediatric brain tumors with dismal prognoses and no effective treatments. GD2, a disialoganglioside overexpressed on DMG cells, is a promising target for chimeric antigen receptor (CAR) T cell therapy. In Arm A of the Phase I clinical trial (NCT04196413), 12 patients (11 on study) received GD2-CAR T cells through an initial intravenous (IV) infusion, and if patients had clinical or radiographic improvement, they subsequently received intracerebroventricular (ICV) administrations. This regimen achieved clinical benefit in ten patients, with >50% tumor volumetric reductions in 5 patients and a durable ongoing complete response in one patient (Majzner, Ramakrishna et al. Nature 2022; Monje et al. Nature 2024). However, disease progression ultimately occurred in 11 patients underscoring challenges in therapeutic optimization. To elucidate mechanisms underlying treatment responses and resistance, we performed a comprehensive multi-omic analysis, integrating single-cell RNA and TCR sequencing, mass cytometry, and clinical and molecular datasets. Sequencing 1, 062, 398 cells from patient apheresis, CAR T products, and cerebrospinal fluid (CSF) samples collected longitudinally at pre-defined trigger events, we reconstructed the dynamic landscape of immune and tumor cell populations in 12 DMG patients. Our analysis revealed profound CAR T cell infiltration into the CSF after treatment, accompanied by increases in endogenous T cell, mast cell, and monocyte populations and a relative decrease in microglia evident in CSF. This works sought to deeply characterize the lymphocyte profiles in these samples. Inflammatory and proliferation pathways were elevated in CAR T cells at early post-ICV time points, transitioning into an NK-like state and TGF-β receptor signaling at later stages. TCR clonal tracking identified persistent CAR T clones, alongside endogenous T cell activation and expansion. Moreover, non-responders to the therapy showed elevated Treg signatures in the CSF. Using CellChat-based cell-cell communication analysis, we uncovered differential cytokine-receptor interactions in responders versus non-responders, offering novel insights into immune-tumor interactions in the context of CAR T cell therapy. By contextualizing single-cell data within multi-omic frameworks and clinical outcomes, this study advances our understanding of the GD2-CAR T cell therapy, the tumor microenvironment, and biomarkers of response and resistance. These findings provide critical insights for optimizing CAR T cell therapies in CNS malignancies and inform the development of next-generation immunotherapies. Yiyun Chen, Moksha H. Desai, Zachary J. Ehlinger, Kun Wei Song, Skyler Rietberg, Hossein Daghagh, Mark P. Hamilton, Snehit Prabhu, Sonia Partap, Jasia Mahdi, Elena Sotillo, Bita Sahaf, Sneha Ramakrishna, Zinaida Good, Michelle Monje, Crystal Mackall. Unravelling temporal dynamics of GD2-CAR T cell therapy in H3K27M-mutant diffuse midline glioma through comprehensive single-cell and multi-omic profiling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7123.

H3 G34-mutant diffuse hemispheric glioma (DHG) is a newly recognized tumor type in WHO CNS5. Previous studies have demonstrated inferior outcomes in pediatric patients, but the underlying mechanisms remain unknown. Additionally, the effectiveness of temozolomide (TMZ) and the role of surgery in pediatric G34 tumors remain uncertain. Multi-institutional retrospective review of clinical, imaging, and molecular characteristics of 36 pediatric (≤ 18 years) patients with newly diagnosed H3 G34-mutant DHG. The cohort’s median MGMT expression values were used as the cut-off for MGMT-low vs high. The median age was 14.2 years (8-18 years). The median progression-free survival (PFS) was 0.7 years (95% CI 0.4-1.2 years), and the median overall survival (OS) was 1.8 years (95% CI 1.1-3.2 years). Gross total resection (GTR) was associated with improved PFS (p=0.0046) compared to non-GTR. Out of 31 patients with evaluable baseline imaging, seven patients (22.6%) presented with involvement of more than 3 cerebral lobes. Twenty-one patients (58.3%) received frontline TMZ and had improved PFS compared to those without (p=0.0049). Low MGMT expression level was associated with better PFS (p=0.0039), better OS (p<0.0001), and increased TMZ efficacy. In pediatric DHG, gene body/intronic CpG methylation rather than promoter methylation correlated better with MGMT expression (p<1x105); MGMT promoter methylation was not significantly associated with PFS/OS or TMZ efficacy. PDGFRA amplification/mutation (n=13) was associated with inferior OS (p=0.0035), and CDKN2A homozygous deletion (n=14) was associated with inferior PFS (p=0.0352). Of the evaluable patients with progressive disease, 81% progressed within the high-dose RT field. Our findings reaffirm the dismal outcomes of pediatric H3 G34-mutant DHG, which exhibits radiation resistance, a tendency toward widespread disease, and a novel mechanism of MGMT regulation. Our data support frontline TMZ use in pediatric patients and underscore the importance of GTR when feasible.

Responses to immunotherapy in pediatric brain cancers have thus far been disappointing. We, therefore, sought to understand the immunophenotype/immune environment in these tumors. The Pediatric Brain Tumor Consortium studied 100 patient samples from ependymomas, high-grade gliomas (HGGs), medulloblastomas, and historically-diagnosed supratentorial primitive neuroectodermal tumor [sPNET]). PD-1/PD-L1 expression were determined by immunohistochemistry as previously presented, and an updated study of immune-related gene expression was analyzed via a Nanostring immunologic codeset. Immune scores were calculated with a GSVA ssGSEA algorithm, and hierarchical clustering visualized by pheatmap and ComplexHeatmap. Tumor-type specific subgrouping was applied using the scores generated from Consensus Non-negative Matrix factorization (cNMF). Approximately half of the tumors expressed PD-1 (52/100) or PD-L1 (44/100) at an intensity score of at least 1/5, with higher scores in HGG. Ependymoma and HGG demonstrated more robust immune cell signatures, while medulloblastoma and sPNET demonstrated decreased monocytic and myeloid cell signatures. Reclustering delineated three immune-related subgroups in each tumor with unique expression profiles. Ependymoma group E1 was enriched for Th17 cell differentiation, and E3 with EGFR tyrosine kinase inhibitor resistance. HGG G1 pathways related to IL-17 signaling, NOD-like receptor signaling, and cytokine signaling, while G3 was enriched for PI3K-Akt signaling and ECM-receptor interaction. Medulloblastoma M1 and M2 was enriched for pathways related to pro-inflammatory genes, and M3 enriched with PI3K-Akt signaling and NK cell mediated cytotoxicity. sPNET P1 and P2 was also enriched for pathways related to immune system genes, while P3 showed upregulated HSC lineage and antigen presentation pathways. The addition of immune expression profiling in our study to PD-1/PD-L1 IHC scores enables better understanding of relevant immune-specific genes, with better characterization of noteworthy immune alterations that may aid in designing more effective immunotherapy for pediatric brain cancers.

Pediatric high-grade gliomas (HGGs) are the leading cause of brain cancer-related death in children. HGGs include distinct subtypes defined by anatomical location as well as molecular characteristics that stratify into H3K27M-altered diffuse midline gliomas (DMG) and hemispheric HGGs such as isocitrate dehydrogenase-wild type (IDH-WT) glioblastoma. Neuronal activity drives HGG progression both through paracrine signaling and direct neuron-to-glioma synapses. Using whole-cell patch clamp electrophysiology, in vivo optogenetics, and patient-derived glioma xenograft mouse models, we identify functional, tumor-promoting GABAergic neuron-to-glioma synapses mediated by GABAA receptors in DMGs. GABAergic input has a depolarizing effect on DMG cells due to chloride uptake by the Na-K-2Cl cotransporter NKCC1 and consequent elevated intracellular chloride concentration. In contrast, depolarizing GABAergic currents were not detected in IDH-WT HGG. Membrane depolarization is known to increase glioma proliferation and accordingly, we find that the activity of GABAergic interneurons promotes DMG proliferation in vivo. Increasing GABA signaling with the benzodiazepine lorazepam increases glioma proliferation and reduces survival in xenograft models of DMG but not IDH-WT HGG. Conversely, we find that the anti-seizure medication levetiracetam attenuates low-frequency depolarizing GABAergic synaptic currents in a glioma-specific manner, reducing those in DMG but not in healthy neurons. The effect in DMG is independent of action on synaptic vesicle glycoprotein 2A (SV2A), the chief mechanism by which levetiracetam suppresses seizures. Levetiracetam reduces glioma proliferation and extends survival of mice bearing DMG xenografts, but not IDH-WT HGG xenografts. Retrospective real-world clinical data demonstrate longer overall survival for children with DMG who were taking levetiracetam, which was not evident in pediatric hemispheric HGG. These findings uncover growth-promoting GABAergic synaptic communication between GABAergic neurons and DMG cells, underscoring a tumor subtype-specific mechanism of brain cancer neurophysiology with potentially important implications for commonly used drugs in this disease context, which should be further studied in future prospective clinical studies.

The SJMB12 (NCT01878617) clinical trial introduced a molecular subgroup-based approach to the treatment of newly diagnosed medulloblastoma by stratifying 649 evaluable patients into WNT, SHH, and non-WNT/non-SHH strata. While the genomic landscape of medulloblastoma has been extensively characterized, most prior studies analyzed heterogenous or non-trial-associated cohorts, which may not capture the true distribution of oncogenic drivers within a clinically homogeneous population. To address this limitation, we used 1) Illumina EPIC DNA methylation array for subgroup/subtype classification and detection of copy number alterations; 2) whole exome sequencing for detection of single-/multi-nucleotide variants and small insertions/deletions; 3) RNA sequencing for detection of gene overexpression and fusions. In the WNT-stratum (n=90), 96% of profiled tumors harbored canonical exon 3 hotspot mutations in CTNNB1, while the remaining 4% exhibited inactivation of APC, consistent with aberrant WNT/β-catenin pathway activation. Additionally, we identified mutually exclusive loss-of-function mutations affecting several genes on chromosome 6 - LATS1 (8%), FOXO3 (8%), and CSNK2B (9%), co-occurring exclusively with monosomy 6. In the SHH-stratum (n=107), 51% of tumors exhibited PTCH1 inactivation, which was the most prevalent gene-level alteration. Germline loss-of-function variants in ELP1 or TP53 were observed in ~30% of cases, highlighting the clinical utility of germline testing. Three patients exhibited hypermutator phenotypes, with tumor mutational burdens 100x higher than the cohort median. These individuals carried pathogenic germline variants in mismatch repair genes MSH6 or PMS2, consistent with Lynch/Turcot type I syndromes. Notably, two of the three patients also harbored somatic POLE mutations, likely contributing to replication error–driven mutagenesis. In the nonWNT/nonSHH stratum (n=452), beyond known drivers, we identified a novel mechanism of GFI1B enhancer hijacking resulting from an interchromosomal translocation, as opposed to proximal structural variants. The molecular results presented here will be further integrated with survival outcomes and used to inform the design of future clinical trials.

The NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines) for Pediatric Central Nervous System Cancers provide multidisciplinary diagnostic workup, staging, and treatment recommendations for diffuse high-grade gliomas and medulloblastomas in children and adolescents. This article summarizes the studies and panel discussion that serve as the rationale for comprehensive care recommendations included in the NCCN Guidelines for Pediatric Central Nervous System Cancers.

The NCCN Clinical Practice Guidelines in Oncology (NCCN Guidelines) for Pediatric Central Nervous System Cancers provide multidisciplinary diagnostic workup, staging, and treatment recommendations for diffuse high-grade gliomas and medulloblastomas in children and adolescents. This article summarizes the studies and panel discussion that serve as the rationale for comprehensive care recommendations included in the NCCN Guidelines for Pediatric Central Nervous System Cancers.