Pediatric Gliomas Research Articles

LGG-48. CLINICAL CHARACTERIZATION OF FGFR-ALTERED, PEDIATRIC LOW-GRADE GLIOMAS

Abstract BACKGROUND Pediatric low-grade glioma (pLGG) is the most common central nervous system tumor of childhood. Most pLGGs are driven by BRAF alterations, but a subset harbor activating mutations or fusions of FGFR1/2/3. Significantly less is known about the biologic activity, clinical course, and response to targeted inhibition of these FGFR-altered gliomas (FGFR pLGG). We sought to define the epidemiologic characteristics, molecular signature, prognostic markers, and clinical course of patients with FGFR pLGG. METHODS We performed a multi-institution, retrospective chart review of pediatric neuro-oncology patients treated at two large, academic cancer centers from 2012 – 2023 with tumor molecular profiling demonstrating any FGFR alteration. RESULTS Sixteen eligible patients were identified; 9 (56%) were female and median age was 12 years of age (range 6 - 18 years). Reported histologies included pilocytic astrocytoma (4), rosette-forming glioneuronal tumor (3), low grade glial/glioneuronal tumor (3), low grade glioma NOS (2), intermediate grade astrocytoma (1), dysembryoblastic neuroepithelial tumor (2), and pilomyxoid astrocytoma (1). FGFR point mutations were seen in 10/16 (62.5%) patients followed by FGFR-TACC1 fusions in 4/16 (25%) patients and two patients with FGFR1 tandem duplication. Metastatic disease was noted at diagnosis in two patients and upfront gross total resection was achieved in four patients. The most common systemic therapy for the remaining patients was Vincristine and/or Carboplatin. All sixteen patients are alive at median follow up of 52 months (range 6 - 198 months). CONCLUSIONS FGFR pLGG represent a distinct subset of tumors with unique biologic properties and clinical activity. Additional studies are needed to investigate this unique patient population to appropriately adapt our therapeutic approach and improve patient outcomes.

MODL-15. CHARACTERIZATION OF SPONTANEOUS CANINE HIGH-GRADE GLIOMA MODELS AS CLINICALLY-RELEVANT SURROGATES OF PEDIATRIC HIGH-GRADE GLIOMA

Abstract BACKGROUND High-grade glioma (HGG) occurring spontaneously in canines shows histological and genetic resemblance to pediatric high-grade glioma (PHGG). Both canine HGG and PHGG are treated with combinations of surgical resection, radiation therapy, and/or chemotherapy, but outcomes for both groups are poor. Studying canine HGG both serves companion dogs and provides an orthogonal preclinical model of PHGG. METHODS Canine HGG lines J3T, SDT3G, and G06A were characterized and compared with H3K27M-mutant PHGG lines DIPG7, DIPG13, and BT245 using neurosphere dilution assay and drug screening with all FDA-approved anti-cancer agents. J3T and SDT3G were orthotopically implanted into mice and monitored using MRI and bioluminescence. RESULTS Like PHGG, the canine lines J3T and SDT3G were highly sensitive to proteasome and XPO1 inhibition; compared to PHGG, the canine lines were most sensitive to protein translation inhibitors and DNA intercalators. Canine HGG lines were not as sensitive to other targeted inhibitors effective in PHGG. Compared to PHGG cell lines, canine cell lines had greater variance in self-renewal capability. J3T and SDT3G were also injected into the cortex of immunodeficient mice. Bioluminescent signal confirmed J3T engraftment one week post-surgery with rapid tumor growth, while SDT3G presented initial engraftment by MRI two weeks post-surgery but did not exhibit sustained growth. DISCUSSION Canine HGG and PHGG showed similar sensitivity to proteasome and XPO1 inhibition; they differed in self-renewal capability and susceptibility to other anti-cancer agents. This could be because canine target proteins often differ from their human counterparts and thus may respond differently to targeted medications designed for humans. Overall, canine HGG provides a novel, primary, immunocompetent surrogate that resembles PHGG and may provide a step for clinical investigation between xenograft models and PHGG trials and/or opportunities for co-clinical trials.

HGG-20. CANCER PREDISPOSITION SYNDROMES IN PEDIATRIC PATIENTS WITH HIGH-GRADE GLIOMAS

Abstract The association between pediatric high-grade gliomas (HGG) and cancer predisposition syndromes (CPS) remains inadequately elucidated in the existing literature, while next-generation sequencing (NGS) is increasingly utilized for diagnostic support. In this retrospective analysis, we evaluated a cohort of 95 consecutive pediatric patients with HGG reclassified according to the latest 2021 WHO classification, diagnosed and treated at the Bambino Gesù Children’s Hospital between 2011 and 2023. All patients underwent genetic germline testing with NGS technology to detect the presence of germline variants predisposing to cancer and were included in this study. Diagnosis of pHGG H3-IDH wild-type was performed in 23 patients (24.2%), diffuse midline glioma in 46 patients (48.4%), astrocytoma IDH-mutant in 7 patients (7.4%), anaplastic pleomorphic xanthoastrocytoma in 4 patients (4.2%), adult type HGG in 4 patients (4.2%), neuroepithelial tumor PATZ1 fusion-positive in 3 patients (3.2%), diffuse hemispheric glioma H3 G34-mutant in 2 patients (2.1%), infant-type hemisheric glioma in 2 patients (2.1%), other rare histologies in 4 patients (4.2%). Oncological family history was investigated up to the third generation for all patients and was positive in 43/95 (45.3%) cases. We identified a total of 65 variants in 95 patients, distributed as follows: 48 variants of uncertain significance (73.8%), 10 likely pathogenic (15.4%), and 7 pathogenic variants (10.8%). Notably, our findings reveal that pathogenic/likely pathogenic variants were detected in about 17% of the entire study population, higher than the 10% incidence reported in the existing literature. The high rate of mutations in CPS associated genes described in our work, leads to consider performing NGS even in the absence of clinical or anamnestic criteria suggestive of it. However, ethical and social issues are to be considered when we propose and investigate patients for CPS. Moreover, further research are needed to determine the role of variants of uncertain significance.

NURS-20. A PATIENT AND FAMILY EDUCATION PROJECT: COMPREHENSIVE RESOURCE FOR MEK INHIBITOR-INDUCED PARONYCHIA

Abstract BACKGROUND Targeted therapies for pediatric tumors of the central nervous system have recently brought promising results as well as new challenges in the clinical management of these patients. FDA approval of MEK inhibitors has prompted a significant increase in use of these agents in pediatric neuro-oncology patients. MEK inhibitors are commonly used for treatment of pediatric low and high-grade gliomas, as well being a proven treatment for plexiform neurofibromas. Cutaneous reactions to targeted therapy occur frequently, with paronychia arising in more than half the patients. METHODS Despite providing a robust MEK inhibitor skin care in-service at the start of treatment, the multidisciplinary neuro-oncology team identified a need for specific paronychia education for patients undergoing therapy with MEK inhibitors. The education content was developed following a thorough literature review of MEK inhibitors and associated side effects. Evidence-based information was selected to create a clear, concise, one-page educational resource. RESULTS The educational flyer addressed five key educational objectives including defining and reviewing paronychia as an expected side effect of MEK inhibitor therapy, discussing strategies for prevention and management of paronychia, and identifying when to contact a healthcare provider for paronychia support. Content included written and pictorial definitions to aid in recognition of paronychia, bulleted list and visual guide for preventive nail maintenance, recommendations and instructions for over-the-counter management strategies, review of therapies that may be prescribed, and specific guidelines on when to seek prompt medical attention. This flyer served as a valuable educational tool, equipping individuals undergoing MEK inhibitor therapy with knowledge about paronychia, prevention, and effective management. CONCLUSION MEK inhibitor-induced paronychia is an undesirable side effect that can impact adherence to treatment. By empowering patients and families with this educational resource, healthcare providers may enhance patient engagement, promote adherence to treatment, and optimize quality of life.

IMG-28. AUTOMATIC BRAIN TUMOR VOLUMETRIC ANALYSIS IN MAGNETIC RESONANCE IMAGING GENERALIZABLE TO PEDIATRIC NEURO-ONCOLOGY

Abstract BACKGROUND The prognosis of brain tumors is variable in clinical practice if it only relies on human interpretation of magnetic resonance imaging (MRI). The automatic segmentation of brain tumors in MRI enables quantitative analysis in support of clinical trials and personalized patient care. We developed benchmarked deep learning-based tools that are generalizable to the volumetric quantification of various tumor types across diverse populations. METHODS We participated in the well-established international brain tumor segmentation challenge (BraTS 2023) and benchmarking competition. The challenge made available 4,500 multi-national brain tumor cases with multi-sequence MRIs, including pediatric high-grade gliomas (PED), i.e., high-grade astrocytoma and diffuse midline glioma, and adult gliomas, brain metastases (MET) and intracranial meningiomas (MEN). Each case comprises four MRI volumes: T1, contrast-enhanced T1, T2, and T2-FLAIR. Manual segmentations were provided to establish ground truth for enhancing tumor (ET), tumor core (TC), and whole tumor (WT). Our framework used a model ensemble strategy based on two state-of-the-art deep learning models: a convolutional neural network (nnU-Net) and a vision transformer (Swin UNETR) and was tested for broader applicability across multiple tumor types. The framework was trained on 99, 1,000, and 165 cases and validated on 45, 141, and 31 unseen cases for PED, MEN, and MET, respectively. Automatic segmentations were evaluated by lesion-wise volume overlap (Dice similarity score, DSC) and Hausdorff distance (HD). RESULTS In the evaluation on independent unseen test datasets, our automatic tool was ranked first for PED, third for MEN, and fourth for MET volumetric analysis. Our method resulted in PED lesion-wise DSC of 0.733, 0.782, 0.817 and HD (mm) of 75.93, 25.54, 24.18 for ET, TC, and WT, respectively. CONCLUSIONS These brain tumor volumetric analysis tools are readily available to be efficiently tested on diverse datasets. Automatic MRI analysis provides consistent quantitative data for multi-institutional protocols and clinical trials.

HGG-30. TARGETED THERAPY OF PEDIATRIC HIGH GRADE GLIOMAS

Abstract BACKGROUND The prognosis for patients with pediatric high-grade glioma (pHGG) is poor despite aggressive multimodal therapy. Recent advances in understanding tumor biology facilitate utilization of targeted therapies (TT) in pediatric neuro-oncology. METHODS We analyzed the efficacy of targeted therapy in patients with progressive pHGG treated with TT according to molecularly-identified actionable events. Molecular characterization of the tumor included DNA panel and transcriptomic RNA sequencing. RESULTS Eighteen patients were included (median age 3.7 years, range 0.1-17.7) with anaplastic pleomorphic xanthoastrocytoma (n=10), infant-type hemispheric glioma (n=4), and diffuse high-grade glioma (n=4). Metastatic disease was present in 3/18 patients. Molecular genetic alterations included BRAF V600E mutations (n=8), NTRK1/2/3 fusions (n=4), ROS1 fusions (n=4), MET fusion (n=1), KIT P627L mutation (n=1). Eight patients received an NTRK/ALK/ROS1 inhibitor (entrectinib), seven patients received combination therapy with BRAF and MEK inhibitors (dabrafenib and trametinib), and one patient anti-BRAF monotherapy (dabrafenib). Two patients were treated with a multi-tyrosine kinase inhibitor (cabozantinib). The median duration of targeted therapy was 10.6 months (1-42). Most patients tolerated treatment well with no grade 4-5 toxicities. The overall response rate was 66% (12/18), including 4 complete (22%) and 8 partial responses (44%). At 12 months, PFS was 86% with dabrafenib/trametinib and 42% with entrectinib; at 18 months PFS was 44 % with dabrafenib plus trametinib and 42% with entrectinib. Three out of four patients with ROS1 rearranged glioma experienced disease progression on entrectinib after an initial good response in all cases; two of them subsequently received lorlatinib with clinical benefit. All four patients with NTRK1/2/3 rearranged gliomas remain on entrectinib therapy without signs of progression. Cabozantinib therapy resulted in prolonged complete and partial responses in both cases. CONCLUSIONS TT for patients with progressive pHGG is feasible and provides unexpectedly good clinical efficacy.

LGG-10. RECOMBINANT HUMAN ENDOSTATIN IN COMBINATION WITH CHEMOTHERAPY FOR PEDIATRIC LOW-GRADE GLIOMAS

Abstract BACKGROUND Pediatric low-grade gliomas (pLGGs) entail highly effective treatment regimens. In this study, we aimed to examine the efficacy and safety of recombinant human endostatin (rh-ES) combined with chemotherapy for pLGGs. METHODS We collected the clinical data of pLGG patients treated with carboplatin and vincristine (CV) plus rh-ES in the Department of Neuro-oncology, Sanbo Brain Hospital Capital Medical University, Beijing, China, from July 2015 to October 2019 and retrospectively analyzed the effectiveness and safety of this regimen. RESULTS A total of 41 eligible patients were enrolled. Median age at diagnosis was 4 years. Median follow-up lasted for 22.7 months (range 8-73.4 months). Six patients achieved complete response, 7 attained partial response, 4 had minor response, the disease was stable in 3 cases, and in 1 the condition progressed. The objective response rate was 90.2% and complete response rate was 14.6%. The median time to response was 4.1 months. The duration of response was 28.1 months. The progression rate within 6 months of treatment completion was 6.9%. 5-year rates were 74% for progression-free survival and 92% for overall survival. The main toxicities included myelosuppression and carboplatin allergy. No serious rh-ES-related toxicities were observed. CONCLUSIONS Rh-ES in combination with CV in pLGG patients was effective and safe and could achieve rapid and lasting responses.

LGG-29. BEYOND MAPK SIGNALING - GENOME-SCALE CRISPR/CAS9 SCREENS REVEAL NEW TARGETS FOR TREATMENT OF KIAA1549::BRAF-DRIVEN PEDIATRIC LOW-GRADE GLIOMA

Abstract Since the KIAA1549::BRAF fusion was discovered as the most common driver of pediatric low-grade glioma (pLGG), it has been hypothesized that the fusion induces oncogenicity through BRAF activation as a result of KIAA1549 replacing the BRAF negative regulatory N-terminus. This led to the rapid translation of MAPK pathway inhibitors into the clinical setting. Despite most tumors exhibiting promising initial responses, some tumors are not sensitive and about half of responsive tumors grow back after treatment cessation. Therefore, strategies that result in sustained tumor responses are desperately needed. We have recently performed genome-scale CRISPR/Cas9 screens across isogenic neural stem cell models transduced to express pLGG-associated oncogenes (KIAA1549::BRAF, BRAFV600E, and multiple FGFR1 and MYB family alterations) to generate a dependency map of genetic vulnerabilities associated with expression of these oncogenes. We also included normal neural stem cells to allow identification of genetic dependencies specifically induced by the expression of each oncogene. Surprisingly, through these efforts, we have discovered KIAA1549::BRAF expressing cells to harbor striking and specific dependency on multiple members of an enzymatic complex that exerts its activity outside of the MAPK signaling axis*. Interestingly, this enzymatic complex has been described to modify only a few substrates, including KIAA1549, suggesting specificity. We have now validated this dependency across other isogenic models of KIAA1549::BRAF-expressing cells. Finally, our KIAA1549::BRAF-expressing models exhibit preferential sensitivity to a tool compound that targets our novel enzyme, suggesting novel therapeutic potential for KIAA1549::BRAF. These findings highlight a MAPK pathway-independent avenue for therapeutically targeting the most frequent genomic alterations in pediatric brain tumors. Consequently, we also propose that the fusion partner KIAA1549 is instrumental for the aberrant BRAF signaling driving pLGGs. (* we are currently in the process of working with our IP offices to ensure that we can disclose the names of the genes and proteins at the ISPNO meeting)

LGG-46. MEKI WITH MIRDAMETINIB: EXPLORING ADAPTIVE SIGNALING PATHWAY DYNAMICS AND IMMUNE MODULATION IN PEDIATRIC GLIOMAS

Abstract INTRODUCTION Glial neoplasms are among the most common types of solid tumors in children. The mitogen-activated protein kinase (MAPK) pathway is implicated in glioma tumorigenesis. Drugs targeting this pathway hold promise as novel therapy for this disease. Vertical inhibition of the MAPK pathway has been linked to increased inflammation in several other cancers but similar exploration in pediatric brain tumors is limited. Within the MAPK pathway, MEK plays a significant role as a modulator of ERK phosphorylation. This study investigates the therapeutic potential of the MEK inhibitor, Mirdametinib, and evaluates the drugs’ capacity to induce tumor-intrinsic inflammation by characterizing changes in PD-L1 expression and proliferation in a panel of pediatric glial neoplasms. METHODS Three cell lines were included in the panel (Res186, Res259, SF188) and were treated with Mirdametinib for 3, 5, or 7 days. P-ERK expression, analyzed by western blot, verified successful inhibition of MEK by Mirdametinib. The CCK8 cytotoxic assay was utilized to determine the drug’s 50% inhibitory concentration as well as its direct effects on cell viability. Changes in Ki67 expression assessed by flow cytometry further characterized Mirdametinib’s effects on cell proliferation. Flow cytometry was also utilized to measure changes in PD-L1 expression following treatment. RESULTS Despite minimal effects on proliferation as measured by Ki67 and CCK8, expression of P-ERK verifies successful inhibition of MEK. Western blot data also reveals increased Pi3K/AKT pathway activity in SF188 groups treated with Mirdametinib. FACS data for SF188 also demonstrates increased PD-L1 expression in response to treatment. CONCLUSIONS Data suggests that, in response to MEKi, gliomas utilize compensatory growth mechanisms through Pi3K/AKT signaling. Findings also point to immune-modulatory effects of MEKi. Supporting results in ongoing experiments are expected to further our understanding of these mechanisms and may help inform effective combination treatment strategies for pediatric gliomas.

NFS-22. INVESTIGATING MALIGNANT TRANSFORMATION IN NF1 PEDIATRIC GLIOMAS USING AN IPSCS-DERIVED CEREBRAL ORGANOID MODEL

Abstract BACKGROUND Neurofibromatosis Type 1 (NF1) is an autosomal dominant tumor predisposition syndrome affecting 1 in 3000 children around the world. Although NF1 inactivation is recognized as the cause of low-grade glioma (LGG) development, little is known about subsequent genetic alterations that lead to malignant transformation. One of the limitations in understanding the molecular events underlying the malignant transformation of NF1 gliomas is the lack of appropriate NF1 preclinical models for its comprehensive profiling. OBJECTIVE Brain organoids have emerged as powerful in vitro models for studying the interaction between tumor cells and the tumor microenvironment. In this study, we present a novel cerebral organoid model, developed to investigate the molecular events leading to malignant transformation in NF1 pediatric gliomas. METHODS Induced pluripotent stem cells (iPSCs)-derived cerebral organoids were generated using NF1wt and NF1mut iPSCs, the latter reprogrammed from patients with plexiform neurofibromas (Carlos III Health Institute, Spain). Immunofluorescence analysis was performed at four different developmental stages to confirm their correct cytoarchitectural development and differentiation into heterogeneous cell populations. Furthermore, four mCherry-labelled isogenic glioma cells lines, RES 186 wild type, NF1-/-, CDKN2a-/-,NF1-/-/CDKN2a-/- were co-cultured with the developed cerebral organoids and their migration potential and invasiveness were compared using two-photon microscopy. RESULTS/DISCUSSION NF1mut cerebral organoids display ventricular-like zone at early stages of development (d15-d30), characterized by luminal Sox2+ neural stem cells surrounded by Tuj1+ immature neuron. These organized structures disappear at later stages, while progenitor cells start to differentiate into fully mature neurons and glial cells (d45-d60). Additionally, after 10 days of co-culture, all four RES186 isogenic cell lines infiltrated the organoid structure with different invasion patterns. The established organoid model will provide a platform for evaluating the impact of sequential mutational events on LGG transformation, taking into consideration the role exerted by the tumor microenvironment in supporting tumor cell invasion and migration.

BIOM-01. CARBONIC ANHYDRASE IX INTERPLAY WITH LOCAL AND SYSTEMIC IMMUNE RESPONSE IN PEDIATRIC BRAIN TUMORS

Abstract BACKGROUND Brain tumors (BT) remain the leading cause of death among childhood cancers. There is an urgent need to identify novel targets in these patients. BT progression depends on the adaptive responses to hypoxia. Carbonic anhydrase IX (CAIX) helps cancer cells survive in a hypoxic environment and can be potentially as a therapeutic target. The role of CAIX in the interplay between the tumor and immune cells requires in-depth investigation to design effective therapies. METHODS Fresh BT, blood, and intraoperative cerebrospinal fluid (iCSF) samples were collected prospectively from pediatric patients, who underwent brain tumor surgery at Vilnius University Hospital Santaros Klinikos from March 2022 to December 2023. All tumor types were included. Immunohistochemistry (IHC) and flow cytometry (FC) were performed to analyze the expression of CAIX and selected immunological markers in the fresh tumor, iCSF and peripheral blood mononuclear cells (PBMCs). Soluble CAIX content in serum and iCSF was assessed by ELISA. In addition, a retrospective analysis of previously diagnosed pediatric glioma specimens was performed by IHC. RESULTS Twelve patients with heterogeneous BT types have been enrolled prospectively and 17 patients with glioma were reviewed retrospectively. Glioma accounted for 72% (21/29) of all samples. IHC showed that 57% of gliomas were positive for CAIX as well as a moderate correlation between the CAIX score and the expression of CD44, CD206, and CCR2 markers. PD-1/PD-L1 expression was negligible. In the prospective cohort, 83% (10/12) of BT tissue was positive for CAIX by IHC and FC. Soluble CAIX was detectable in 40% of iCSF samples, but not in serum. We found a correlation trend of CAIX expression in the fresh tumor cells and iCSF with increased CCR2 expression on CD45+ PBMCs. CONCLUSIONS This interim analysis showed a correlation between CAIX tumor expression and immune biomarkers, prompting further investigation by enrolling more patients.

METB-11. HOW METHYLATION PROFILING HELPS IN DIAGNOSING AND TREATING PEDIATRIC BRAIN TUMORS IN INDONESIA

Abstract BACKGROUNDS Pediatric brain tumors (PBT) exhibit extensive heterogeneity, impacting treatment outcomes. While traditionally classified based on histopathological features, recent WHO classifications have integrated DNA methylation array profiling amongst other molecular characteristics, which improves diagnostic accuracy and thus treatment decisions. However, access to this technology is limited in low- and middle-income countries (LMICs) like Indonesia. As part of the MNP Outreach Consortium, we aimed to introduce methylation profiling in Indonesia to aid in diagnosing and stratifying PBT patients. METHODS In 2023, we conducted a case series involving seven challenging PBT cases from Cipto Mangunkusumo National General Hospital, re-assessing them with methylation profiling. RESULTS Through collaboration with the Institute of Neuropathology Heidelberg and the DKFZ, our centre adopted a discerning approach to re-evaluate histopathological samples from complex cases. This endeavour yielded notable results, reaffirming four initial diagnoses and altering three others Amongst the findings, one case initially diagnosed as anaplastic ependymoma, CNS WHO grade 3, was reclassified as a supratentorial ependymoma with YAP1 fusion gene, based on methylation class. Noteworthy within this latter group, one case initially categorized as low-grade pilocytic astrocytoma was reclassified as high-grade diffuse midline glioma, H3 K27 altered. Moreover identifying the copy number plot could indicate a KIAA1549::BRAF fusion to reclassify a case from diffuse pediatric glioma WHO grade 2 to pilocytic astrocytoma WHO grade 1. The recurrent pigmented neuroectodermal tumor of infancy (MNTI) was reclassified as Medulloblastoma, non-WNT/non-SHH, group 3 subtype II on methylation profiling, highlighting molecular similarities and potential treatment insights. These findings offer insights into genetic mechanisms and guide treatment strategies. CONCLUSION Accurate diagnosis, facilitated by advanced molecular profiling, enhances prognostication and enables personalized treatment plans, even in LMICs. This underscores the importance of expanding access to advanced diagnostic technologies in resource-limited settings.

DIPG-55. DISSECTING THE ECOSYSTEM OF PEDIATRIC DIFFUSE MIDLINE GLIOMA

Abstract BACKGROUND Pediatric gliomas are heterogeneous tumors encompassing high grade (pHGG) and low grade (pLGG) subtypes. For tumors not amenable to surgery, in particular high grade H3K27 mutant diffuse midline glioma (DMG), no curative options exist. DMG is characterized by 1) extensive heterogeneity of tumor cells that can exist in different interchangeable cellular states mimicking normal development and 2) extensive heterogeneity of the tumor microenvironment (TME). A better understanding of the DMG ecosystem is needed for therapeutic target identification. METHODS Publicly available single cell RNA seq data of DMG (n=17, Jessa et al. Nat. Genet. 2022), high grade pediatric glioblastoma (GBM, n=16, scPCA) and low grade tumors (ganglioglioma, n=5; pilocytic astrocytoma, n=14, scPCA) were analyzed to define key molecular characteristics and differences between these pediatric brain tumors. Doublet cells were removed from each dataset and tumor and TME cells were discriminated based on marker gene expression and inferred CNVs. RESULTS In DMG, glioma cellular states and TME heterogeneity differed depending on the tumor location. DMG arising in the pons harbored a bigger fraction of astrocyte-like glioma cells whereas oligodendrocyte-like cells were more abundant in thalamic tumors. In line with this observation, Liana cellular communication analysis inferred more intratumoral (autocrine) signaling involving oligodendrocyte-like glioma cells in thalamic tumors (e.g. NCAM1-PTPRZ1, NLGN4X-NRXN1, NTN1-DSCAM) than in pons tumors. Notably, the TME of thalamic tumors contained a bigger fraction of immunotolerant macrophages expressing CD163 or VSIG4. In contrast, pons tumors harbored a unique cluster of VEGFA expressing astrocytes, not found in GBM or pLGG. Liana analysis inferred COPA/APP-CD74 interaction between VEGFA+ astrocytes and CD163/VSIG4+ macrophages and microglia, potentially contributing to an immunotolerant TME in pons DMG. CONCLUSIONS In DMG, patterns of tumor and TME composition associated with tumor location were uncovered. Furthermore, unique TME celltypes could contribute to the particularly aggressive nature of these tumors.

LGG-33. NETWORK ANALYSIS INTEGRATING MICRORNAS AND CLINICAL DATA TO DETERMINE PROGNOSTIC OUTCOME IN PEDIATRIC LOW-GRADE GLIOMA PATIENTS

Abstract BACKGROUND Low-grade pediatric gliomas (pLGGs) are the most common pediatric brain tumors. Surgical resection, when feasible, represents the standard of care. If complete resection cannot be achieved, disease progression occurs in about 30% of the cases, and these patients receive adjuvant therapy. The identification of predictive biomarkers to predict tumors that will progress after partial resection, requiring systemic treatment (Chemotherapy or target therapy), could support clinicians in the decision-making strategy. MicroRNAs have been related to tumorigenesis and tumor maintenance and can be used as important biomarkers. This study aims to develop a predictive model able to determine prognostic outcomes and to better stratify pLGG patients according to disease progression. METHODS MicroRNAs levels have been assessed in 76 human tumor tissue samples. Clinical data have been collected for statistical and network-based analyses. Using a network-based approach, we integrated clinical data and tissue microRNA expression to identify a microRNA signature able to predict disease progression. RESULTS Our preliminary microRNA expression analysis displayed new promising signatures correlating with prognostic features, such as disease progression and embryological origin of the tumor. CONCLUSION The predictive model we developed integrating biomarkers and clinical data could be crucial to support clinicians in the decision-making strategy and in advancing personalized medicine.

LGG-51. SYSTEMIC IMMUNE TOXICITY OF BRAF AND MEK-INHIBITORS IN CHILDREN: A PAN-CANADIAN SERIES

Abstract BACKGROUND BRAF/MEK-inhibitors have been transformative for pediatric gliomas with RAS/MAPK pathway aberrations. The RAS/MAPK pathway has a complex role in immune regulation. Though well-known in adults, the systemic immune-related adverse effects (irAE) of BRAF/MEKi in children are not widely reported. METHODS We performed a retrospective chart review for irAE following BRAF/MEKi in children treated at SickKids between 2016-2023. Additional patients were included through a pan-Canadian survey. RESULTS 136 patients on BRAF/MEKi were included (MEKi: 74%, BRAFi: 13%, BRAF+MEKi: 13%); irAE was reported in 6/136 (4.5%). One additional patient was included through the national survey. Median age was 14 years (range: 7-16). Male:female was 3:4. Three patients were on BRAFi, and two each on MEKi, and BRAF+MEKi. Early events developing within weeks of drug initiation included hyper-immune systemic inflammatory responses in three children on BRAFi and one child on MEKi. There was no evidence of any concomitant infection. Two children needed intensive care and showed evidence of multi-organ dysfunction. Delayed irAE developing after months to years on therapy included multi-systemic sarcoid-like reaction, parasympathetic dysautonomia (both on BRAF+MEKi) and brachial neuritis/Parsonage-Turner syndrome (MEKi). Management included drug holidays, dose reductions, and immunosuppressive treatment with steroids. Long-term steroids were needed in two of these seven patients. CONCLUSIONS Pediatric oncologists need to be aware of rare yet possibly under-reported systemic immune toxicity in children treated with BRAF/MEKi. They can present acutely, or while on chronic therapy. Management may be challenging in view of issues with rebound tumor growth on stopping some of these drugs, with select patients needing concomitant, long-term immunosuppressive therapy. Genetic susceptibility and biomarkers may need to be systematically studied in future clinical trials of novel RAS/MAPK inhibitors. Continued surveillance and reporting are needed for the marketed compounds, especially for children on long-term use of these inhibitors.

Immunotherapy for pediatric low-grade gliomas.

Pediatric low-grade gliomas (pLGGs) are the most common brain tumor types affecting children. Although gross-total resection remains the treatment of choice, many tumors are not amenable to complete removal, because they either involve midline structures, such as the optic chiasm or hypothalamus, and are not conducive to aggressive resection, or have diffuse biological features and blend with the surrounding brain. Historically, radiation therapy was used as the second-line option for disease control, but with the recognition that this often led to adverse long-term sequelae, particularly in young children, conventional chemotherapy assumed a greater role in initial therapy for unresectable tumors. A variety of agents demonstrated activity, but long-term disease control was suboptimal, with more than 50% of tumors exhibiting disease progression within 5 years. More recently, it has been recognized that a high percentage of these tumors in children exhibit constitutive activation of the mitogen-activated protein kinase (MAPK) pathway because of BRAF translocations or mutations, NFI mutations, or a host of other anomalies that converged on MAPK. This led to phase 1, 2, and 3 trials that explored the activity of blocking this signaling pathway, and the efficacy of this approach compared to conventional chemotherapy. Despite initial promise of these strategies, not all children tolerate this therapy, and many tumors resume growth once MAPK inhibition is stopped, raising concern that long-term and potentially life-long treatment will be required to maintain tumor control, even among responders. This observation has led to interest in other treatments, such as immunotherapy, that may delay or avoid the need for additional treatments. This chapter will summarize the place of immunotherapy in the current armamentarium for these tumors and discuss prior results and future options to improve disease control, with a focus on our prior efforts and experience in this field.

Ex Vivo Modeling of the Tumor Microenvironment to Develop Therapeutic Strategies for Gliomas

AbstractImproving the prognosis for patients diagnosed with aggressive tumors remains a critical challenge in clinical oncology. One crucial aspect of addressing this challenge involves the development of preclinical models that can accurately predict the effectiveness of potential therapies for clinical application. A pivotal element in enhancing these models lies in their ability to accurately recapitulate the tumor microenvironment. In this review, the authors focus on investigating the tumor microenvironment in the context of ex vivo glioma modelling, with a specific emphasis on the extracellular matrix. The review discusses the changes that occur within the extracellular matrix of pediatric and adult gliomas. Furthermore, it explores the potential of advanced cellular models, particularly those employing bioprinting technologies, to replicate the extracellular matrix ex vivo.

Predicting histological grade in pediatric glioma using multiparametric radiomics and conventional MRI features

Prediction of glioma is crucial to provide a precise treatment plan to optimize the prognosis of children with glioma. However, studies on the grading of pediatric gliomas using radiomics are limited. Meanwhile, existing methods are mainly based on only radiomics features, ignoring intuitive information about tumor morphology on traditional imaging features. This study aims to utilize multiparametric magnetic resonance imaging (MRI) to identify high-grade and low-grade gliomas in children and establish a classification model based on radiomics features and clinical features. A total of 85 children with gliomas underwent tumor resection, and part of the tumor tissue was examined pathologically. Patients were categorized into high-grade and low-grade groups according to World Health Organization guidelines. Preoperative multiparametric MRI data, including contrast-enhanced T1-weighted imaging, T2-weighted imaging, T2-weighted fluid-attenuated inversion recovery, diffusion-weighted images, and apparent diffusion coefficient sequences, were obtained and labeled by two radiologists. The images were preprocessed, and radiomics features were extracted for each MRI sequence. Feature selection methods were used to select radiomics features, and statistically significant clinical features were identified using t-tests. The selected radiomics features and conventional MRI features were used to train the AutoGluon models. The improved model, based on radiomics features and conventional MRI features, achieved a balanced classification accuracy of 66.59%. The cross-validated areas under the receiver operating characteristic curve for the classifier of AutoGluon frame were 0.8071 on the test dataset. The results indicate that the performance of AutoGluon models can be improved by incorporating conventional MRI features, highlighting the importance of the experience of radiologists in accurately grading pediatric gliomas. This method can help predict the grade of pediatric glioma before pathological examination and assist in determining the appropriate treatment plan, including radiotherapy, chemotherapy, drugs, and gene surgery.

Tovorafenib (Ojemda) for pediatric low-grade glioma.

Tovorafenib (Ojemda – Day One), a type II RAF kinase inhibitor, has received accelerated approval from the FDA for treatment of patients ≥6 months old with relapsed or refractory pediatric low-grade glioma harboring a BRAF fusion or rearrangement or a BRAF V600 mutation. Tovorafenib is the first systemic treatment to be approved in the US for pediatric low-grade gliomas with BRAF fusions. Accelerated approval of tovorafenib was based on response rates and duration of response.

European standard clinical practice recommendations for primary pediatric low-grade gliomas

Pediatric low-grade gliomas are the most common brain tumours in childhood and adolescence. Despite the excellent prognosis, pediatric low-grade glioma survivors may suffer from variable long-term complications and may require repeated therapies, implying that this is a chronic disease. The current review describes the European Standard Clinical Practice recommendations for low-grade gliomas at primary diagnosis, that were developed on behalf of SIOPe BTG LGG Working Group within the framework of European Reference Network PaedCan. The manuscript describes the diverse spectrum of pediatric low-grade gliomas in terms of location, age, underlying cancer predisposition syndromes, and special circumstances, such as infantile chiasmatic hypothalamic glioma and diencephalic syndrome, as well as current diagnostic criteria and indications for treatment. Furthermore, it provides current knowledge in histopathology and molecular pathology. Finally, the review focuses on the need for a multidisciplinary approach and treatment indications providing a guide on current treatment modalities, used as first-line therapy in Europe along with information on adverse effects, and follow-up.