tbuckley/cabot-search huggingface dataset:
MDB-12. NANOPORE SEQUENCING AS A CUTTING-EDGE TECHNOLOGY FOR MEDULLOBLASTOMA CLASSIFICATION
Authors: Mathilde Filser, Jacob Torrejon, Kevin Merchadou, Elodie Girard, Christine Bourneix, Elisa Lemaître, Riwan Brillet, Jennifer Wong, David Gentien, Audrey Rapinat, Nicolas Servant, Alexandre Vasiljevic, Anne-Isabelle Bertozzi, Sandra Raimbault, Arnault Tauziède-Espariat, Benoit Lhermitte, Cécile Faure-Conter, Céline Icher, Claire Berger, Claude-Alain Maurage, Damien Bodet, David Meyronet, Emmanuelle Uro-Coste, Emilie De Carli, Fabien Forest, Gilles Palenzuela, Guillaume Chotard, Guillaume Gauchotte, Hélène Sudour, Ludovic Mansuy, Mariana Deparis, Marie-Christine Machet, Maxime Faisant, Natacha Entz-Werle, Pascale Varlet, Pierre Leblond, Sophie Michalak-Provost, Stéphanie Proust Houdemont, Valérie Rigau, François Doz, Franck Bourdeaut, Olivier Delattre, Olivier Ayrault, Julien Masliah-Planchon
Journal: Neuro-Oncology (2024)

Abstract INTRODUCTION Medulloblastoma (MB) is one of the most prevalent embryonal malignant brain tumors, divided into four distinct molecular subgroups (WNT, SHH, group 3, and group 4) with its individual prognosis. A more extensive classification of MB has recently been provided, identifying numerous subtypes, some with poor prognosis, leading to a significant number of patients succumbing to the disease while many survivors experience long-term side effects, highlighting the need for efficient subgrouping methods. Distinct genome-wide DNA methylation profiles characterize each MB subgroup, and the advent of Nanopore DNA sequencing has positioned itself as a reliable technique for comprehensive copy number and methylation profiling. MATERIAL AND METHODS We evaluated the ability of Nanopore sequencing to provide clinically relevant methylation and copy number profiles of MB, on a discovery cohort of 45 frozen MB samples, benchmarked against the gold standard EPIC array, and subsequently validated on a cohort of 116 MB samples. RESULTS The majority of MB in both the discovery cohort (43/45; 95.6%) and the validation cohort (110/116; 94.8%) were accurately subgrouped by Nanopore sequencing. Flongle flow cells for 18 MB allowed for a more rapid and cost-effective analysis, with 94.4% (17/18) being correctly classified. Additionally, Nanopore sequencing allowed us to correctly subtype 24/30 (80.0%) within MB groups. DISCUSSION We provided proof of concept that Nanopore sequencing can subgroup MB using DNA extracted from formalin-fixed paraffin-embedded (FFPE) samples and cell-free DNA obtained from cerebrospinal fluid. This first large-scale study establishes the proof of concept that this modern and innovative technology is well-suited for MB classification. We confirm the use of Nanopore sequencing on circulating DNA and present an initial exploration of its application on DNA extracted from FFPE samples. Its ability to deliver quick and cost-effective results firmly establishes Nanopore sequencing as a game-changer in the field of MB classification.
MDB-108. MRI-GUIDED FOCUSED ULTRASOUND BLOOD BRAIN/TUMOR BARRIER DISRUPTION FOR AUGMENTATION OF ANTIBODY DELIVERY TO HIGH-RISK MEDULLOBLASTOMAS
Authors: Stefanyda Maslova, Zehra Demir, Thomas Sherlock, Beyzanur Ak, Serge Yaacoub, Dalia Haydar, Natasha Sheybani
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Medulloblastoma is the most common pediatric brain cancer, and among its four molecular subgroups, group 3 (G3MB) harbors the worst prognosis (5-year survival rate &amp;lt;50%). Effective treatment of G3MB remains difficult due to disease complexity, its challenging niche within the immunologically- and pharmacologically-privileged central nervous system, and significant risk of long-term developmental deficits due to treatment toxicities. MRI-guided focused ultrasound (MRgFUS) holds the potential to revolutionize care as a non-invasive, non-ionizing approach for spatially precise intervention in G3MBs. Given that the utility of FUS in MBs remains unknown, we implemented FUS-mediated blood brain/tumor barrier disruption (BBB/BTB-D) in a high-fidelity model of G3MB and evaluated its impact on localized antibody delivery – with an eye toward future applications in immuno-oncology. METHODS An orthotopic syngeneic model was established via stereotactic implantation of G3MB cells (Gfi1/c-MYC driven) into the cerebellum. G3MB growth kinetics were validated via serial MRI. Multiple protocols for naive cerebellum or G3MB BBB/BTB-D were established with a 1.15 MHz FUS system under 9.4T MRI guidance. Across both settings, mice were sonicated with concomitant intravenous administration of microbubbles and fluorescently-labeled IgG. Safety was confirmed by acoustic emissions monitoring, susceptibility-weighted imaging, histology, and acute activity monitoring. Antibody delivery was evaluated via ex vivo epifluorescence imaging. RESULTS Post-contrast T1w-MRI depicted clear evidence of localized BBB/BTB-D in naive and G3MB mice. No damage was noted by imaging or histopathologic analysis, and mice displayed normal cardiorespiratory and motor functions following FUS. FUS conferred significantly increased epifluorescence signal in targeted cerebellum (1.3x) and G3MBs (1.5x) relative to controls, indicating improved antibody access. CONCLUSIONS Our findings support the safety and feasibility of FUS BBB/BTB-D in naive and G3MB-bearing cerebellum. We demonstrate that FUS markedly improves antibody penetrance within G3MBs. Ongoing studies are evaluating FUS in combination with novel immunotherapeutic antibodies for G3MB therapy.
CP-15. LONG-TERM OUTCOMES OF PAEDIATRIC CRANIOPHARYNGIOMAS: MULTI-CENTRE COMPARATIVE STUDY
Authors: Vitor Nagai Yamaki, Jai Sidpra, Bruno Santanna Peres, Flavia Massey, Catuto Domingos Alexandre Quianga, Guilherme Jose Costa da Borsatto, Joao Paulo Mota Telles, Inês Nobrega Silva, Juan Pedro Martinez-Barbera, Asthik Biswas, Kshitij Mankad, Hoong-Wei Gan, Hani J Marcus, Noor ul Owase Jeelani, Darren R Hargrave, Hamilton Matushita, Suely Kazue Nagahashi Marie, Kristian Aquilina
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Craniopharyngioma (CP) account for 80% of childhood hypothalamopituitary tumours and, though benign, their proximity to the visual pathways, hypothalamus, and pituitary incurs significant long-term neurologic and endocrine morbidity. Informed surgical decision-making has the potential to reduce morbidity and improve outcomes. METHODS This multicentre retrospective observational study aimed to compare long-term outcomes of CP at two paediatric neurosurgery units (GOSH, UK, and USP, Brazil), delineating management paradigms associated with better prognosis. CP treated over a 20-year period were identified and standardized clinical outcomes recorded for comparative analysis. RESULTS 123 patients were identified (USP=52; GOSH=71). No demographic differences were identified between series (mean age: USP=9.2±4.6; GOSH=8.2±4.1 years). Headaches (USP=75%; GOSH=62%) and visual deficits (USP=46%; GOSH=38%) were the most common presenting symptoms. Growth hormone deficiency at presentation was more common in the USP cohort (USP=62.5%; GOSH=35%; p=0.009). There was significantly more hypothalamic invasion in the GOSH cohort (Paris grade 2: GOSH=67.0%; USP=37.5%; p=0.026), but significantly larger cystic volumes in the USP cohort (USP=27.5±37cm3; GOSH=10.5±8.9cm3;p=0.004). These differences led to different surgical approaches, with more subtotal resections at USP (78%) and more cyst aspirations at GOSH (62%). 80% of patients at GOSH received RT compared to 32% at USP. Mean follow-up at GOSH was 7.5±4.8 years compared to 10.1±5.6 years at USP. Cognitive outcome did not vary. GOSH cohort presented higher rates of levothyroxine replacement (USP=67%; GOSH=86%;p=0.017). GOSH exhibited better hypothalamic outcomes (p=0.08) and better visual outcomes (field deficits: GOSH=44%; USP=66%;p=0.01). In multivariate analysis (n=123), subtotal/gross-total resection (OR0.17; 95%CI:0.05-0.55;p&amp;lt;0.01) and older age at diagnosis (OR0.91, 95%CI:0.82-0.99;p=0.04) were associated with longer progression-free survival. CONCLUSIONS Comparative analysis of two large craniopharyngioma series identified different paradigms of surgical management driven by clinical-radiological presentation with similar long-term outcomes. Subtotal/gross-total resection was not related to worse clinical outcome or better progression-free survival.
TRLS-02. SAFETY AND EFFICACY OF BRAF INHIBITOR PLIXORAFENIB (FORE8394; PLX8394) IN CHILDREN AND ADULTS WITH RECURRENT, BRAF-ALTERED PRIMARY CENTRAL NERVOUS SYSTEM TUMORS (PCNST)
Authors: Macarena de la Fuente, Nicholas A Butowski, Jennie Taylor, Rona Yaeger, Frank Y-C Tsai, Filip Janku, Carl E Allen, Natraj Ammakkanavar, Glenn Michelson, Ping Jiang, Emil Samara, Michael Paz, Alexia Tussay-Lindenberg, Kongming Wang, Stacie P Shepherd, Irina Kline, Eric Sherman, Jordi Rodon
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Plixorafenib is a selective, potent BRAFi that inhibits V600 and non-V600 alterations by disrupting BRAF monomers/dimers. Plixorafenib evades paradoxical MAPK pathway activation, eliminating the need for MEKi combination, and is potentially less prone to toxicities and resistance typical of approved BRAFis. METHODS Phase 1/2a study (NCT02428712) in children (n=5, aged 4-17 years) and adults (n=108) with BRAF-altered solid tumors assessing safety, PK, and efficacy of oral plixorafenib 900-3600mg/day ± cobicistat (PK enhancer). RESULTS Five children, 3 LGG (1 V600E, 1 fusion, 1 amplification), 1 neuroblastoma (G469A) and 1 ND-LCH (V600E), 4-8 prior therapies, received plixorafenib 300-1200 mg/day without cobicistat. Four received BSA-adjusted dosing, which delivered lower exposures than RP2D. All had SD, with duration 66.9+ months for ND-LCH. The fifth (neuroblastoma, 1200 mg/day) had PD. No DLTs, dose reductions, or LFT elevations. One had possibly related Grade 3 headache. Of 9 evaluable MAPKi-naïve adults with V600E PCNST, 4/5 with HGG and 2/4 with LGG/GNT responded; ORR 67% (95% CI: 29.9-92.5), mDOR 13.9 months. Responses in patients with only non-enhancing lesions and/or pilocytic astrocytoma indicate clinically relevant CNS penetration. Responses occurred across V600 solid tumors, ORR 42% (10/24); 1-year PFS 54.4%, 2-year PFS 48.4% in the MAPKi-naïve (non-CRC) subgroup. Of 14 evaluable adults with fusions, 2 responses (melanoma: CR, DOR 62.6+ months; PTC: PR, DOR 4.0+ months), 8 SD (5 with PCNST), 5 PD (3/5 with co-occurring driver mutations). Safety, efficacy, and PK in adults and children and insight into factors affecting plixorafenib exposure (not age or weight dependent) determined the optimal RP2D of plixorafenib as 900 mg QD with cobicistat for ages ≥10 years. CONCLUSIONS Plixorafenib had low rates of symptomatic TEAEs with a manageable safety profile, and high ORR in MAPKi-naïve BRAFV600 PCNST. A phase 2 study in children and adults is ongoing (NCT05503797).
MDB-96. CEREBROSPINAL FLUID LEVELS OF THE MIR216B-5P AS A MINIMALLY INVASIVE MONITORING TOOL FOR MEDULLOBLASTOMA
Authors: Mateusz P Koptyra, Anne Coyle, Madison Hollawell, Komal Rathi, Shehbeel Arif, Maura Diamond, Noni Diarra, Ammar Naqvi, Peter Madsen, Phillip B Storm, Jessica B Foster, Adam C Resnick
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Current management of pediatric brain tumor patients relies on imaging as the only non-invasive tool available for monitoring therapeutic response and tumor progression. Results are often inconclusive and unable to capture biologically relevant changes that presage progression on imaging. Circulating miRNA provides an attractive non-invasive biomarker because miRNA has an essential role in regulating gene expression, stability in bio-fluids, selective over-expression in tumors, and is actively released from tumors into the extracellular environment. We aimed to establish the diagnostic and monitoring value of miRNA biomarkers for pediatric brain tumors in cerebrospinal fluid (CSF). METHODS We performed HTG EdgeSeq miRNA panel profiling (2100 targets) of CSF (n=33) samples across pediatric brain tumors from 5 histologies (low grade glioma, ependymoma, germ cell tumor, medulloblastoma, high-grade glioma). We utilized unsupervised consensus clustering, differential expression analysis, and machine learning approaches to investigate miRNA biomarkers. Results were validated using qPCR and ddPCR methods. RESULTS CSF miRNA signatures revealed a correlation with leptomeningeal disease and higher disease burden (p&amp;lt;0.05). Differentially expressed miRNA revealed histology-specific signatures for low grade glioma, medulloblastoma, and germinoma (p&amp;lt;0.05). miR216 expression was significantly elevated in medulloblastoma patients with active disease, indicating a potential novel biomarker for tumor monitoring. The miR216b-5p expression in CSF was confirmed using qPCR and ddPCR. The analysis of independent tumor tissue-based dataset confirmed miR216b-5p and its coding gene MIR217HG expression upregulated in medulloblastoma. MiR-216b-5p expression was also confirmed in patient-derived medulloblastoma cell lines and detected in cell free-media composition indicating extracellular release of miR216b-5p. CONCLUSIONS These results support the use of circulating miRNA in CSF as a potential biomarker for diagnosis and clinical monitoring of medulloblastoma. Further utilization of this approach provides a unique platform to inform liquid biopsy-based management in the clinical setting.
PATH-07. SPINAL OCCURRENCE OF INTRACRANIAL MESENCHYMAL TUMOR WITH FUS::CREM FUSION
Authors: Sarah Edminster, Hung N Tran, Jianling Ji, Zied Abdullaev, Kenneth Aldape, Jennifer Cotter
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND Intracranial Mesenchymal Tumor (IMT), with FET::CREB fusion is a rare and recently characterized tumor. Previously, they were classified as intracranial myxoid mesenchymal tumor and angiomatoid fibrous histiocytoma-like neoplasms. Presence of FET::CREB gene fusion defines the entity, however the incidence and prognosis are not well understood. CASE PRESENTATION Our 6-year-old female patient presented with 6 weeks of neck pain. MRI revealed a posterior intradural extramedullary cervical spine mass (2.4 x 1.4 x 1 cm). She subsequently underwent complete tumor resection. H&amp;E showed a round cell neoplasm, in a lobulated architecture with thick interstitial collagen. Mitotic activity was rare. Immunohistochemistry for CD99 and desmin were positive, with focal synaptophysin expression. The tumor lacked specific immunohistochemical features to enable further classification. OncoKids next generation sequencing revealed an in-frame FUS::CREM fusion. DNA methylation profiling matched to an IMT with FET::CREB fusion. Post-operatively, she had pain resolution and is without recurrence 3 months later. DISCUSSION IMTs have gene fusions of the FET RNA-binding protein family (EWSR1 or FUS) with a member of the CREB family of transcription factors (CREB1, ATF1, or CREM). This fusion creates a chimeric protein with altered function. EWSR1 is the most common fusion partner. IMTs have a broad morphological spectrum that overlaps with other entities making molecular studies essential for the diagnosis. While most IMT are supratentorial, few spinal cord examples have been reported. Optimal treatment strategy remains undefined, but adjuvant radiation therapy may be considered, especially for incompletely resected tumors. CONCLUSION FET::CREB fusion defines IMT, a tumor type that is typically intracranial. Spinal cord occurrence is possible, although less well-recognized. They have a broad morphological spectrum, immunohistochemical overlap with other entities, and molecular studies are required to make the diagnosis. Further studies to characterize their clinical behavior to guide future treatment are needed.
PATH-10. ACCELERATING COMPREHENSIVE CNS TUMOR MOLECULAR DIAGNOSTICS WITH RAPID-CNS2 AND MNP-FLEX: A PROSPECTIVE MULTI-CENTER VALIDATION
Authors: Areeba J Patel, Kirsten Göbel, Felix Hinz, Helin Dogan, Daniel Schrimpf, Pauline Göller, Henning Leske, Skarphéðinn Halldórsson, Graeme Fox, Simon Deacon, Simon Paine, Stuart Smith, Natalie Jäger, Christel Herold-Mende, Wolfgang Wick, Stefan M Pfister, Einar Vik-Mo, Andreas von Deimling, Matthias Schlesner, David T W Jones, Matthew Loose, Martin Sill, Felix Sahm
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND The 2021 WHO classification update highlights the necessity of integrating molecular alterations for precise central nervous system (CNS) tumor diagnoses. However, current molecular reporting methods are hindered by significant initial investment, labor-intensive protocols, and lengthy turnaround times. Methylation-based classification has emerged as a pivotal diagnostic tool but is currently limited to array-based techniques. This necessitates exploration of novel technologies to streamline molecular analysis. METHODS We implemented Rapid-CNS2 - our adaptive sampling-based nanopore sequencing workflow- on 190 adult and pediatric samples at University Hospital Heidelberg and University of Nottingham. Intraoperative potential was assessed through real-time analysis followed by 24-hour sequencing for comprehensive genomic insights. Additionally, we developed MNP-Flex, a platform-agnostic version of the Heidelberg methylation classifier covering 184 CNS tumor classes. We evaluated MNP-flex on a global cohort of over 78,000 samples from methylation arrays, whole genome bisulfite sequencing, nanopore whole genome sequencing, methylation panels and Rapid-CNS2. RESULTS Rapid-CNS2 validation yielded accurate integrated diagnoses in all 190 samples. Within a crucial 30-minute timeframe, we reported accurate methylation families and arm-level copy number profiles followed by next-day reporting of fine-grained methylation classification, SNVs, focal CNVs, MGMT status, fusions and novel structural variants. Moreover, MNP-Flex achieved 92% accuracy over the validation dataset spanning over 78,000 samples from five different technologies. CONCLUSIONS The adoption of Rapid-CNS2 and MNP-Flex enables rapid intraoperative broad methylation classification and copy number alteration reporting within 30 minutes, with additional clinically relevant, fine-grained molecular insights available the following day. It offers clinicians rapid access to comprehensive molecular information critical for treatment decisions. Furthermore, MNP-Flex extends the utility of the Heidelberg methylation classifier to diverse sequencing-based data. By overcoming the limitations of currently available methods, our workflow represents a paradigm shift in the field, promising improved management of CNS tumor patients. Rapid-CNS2 can be executed with a single command, while MNP-Flex is publicly available as a web service, enhancing accessibility and usability for clinical applications.
DIPG-34. IDENTIFICATION OF CAR-T CELL ENHANCING COMPOUNDS USING HIGH THROUGHPUT SCREENING
Authors: Amber B Jones, Michaela Meehl, Rebekah DeVries, Anang Shelat, Giedre Krenciute
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND The infiltrative growth patterns and restrictiveness of the blood brain barrier greatly limit therapeutic options for children diagnosed with diffuse intrinsic pontine gliomas (DIPGs). However, clinical trials using chimeric antigen receptor (CAR)-T cells have shown promise in DIPG patients. Despite some success, the biological limitations plaguing CAR-T cells hinder their overall efficacy. We propose that leveraging chemical compounds in tandem with CAR-T cells may provide an increased therapeutic benefit against this lethal brain tumor. Our study is aimed at using high throughput screening of &amp;gt;2,000 FDA approved and investigational compounds to identify candidates that enhance CAR-T cell mediated cytotoxicity of DIPGs. METHODS In an initial limited screen (32 compounds) used for optimization, compounds were serially diluted using the Echo 650 acoustic liquid handler. Subsequent co-culture conditions of renilla luciferase transduced DIPG007 tumor cells and clinically relevant firefly luciferase transduced CAR-T cells were plated at a 2:1 effector: target (E: T) ratio. In comparison to DMSO control, drug induced changes on cell viability were measured using cell-specific luciferase activity at 24 and 72 hours. RESULTS The limited screen revealed the BET bromodomain inhibitor, GSK-046, as an initial target based on its ability to increase CAR-T cell viability after 72 hours. Preliminary validation of these findings supports this CAR-T enhancing role of GSK-046 via MTS assays. We also identified nonlethal concentrations of GSK-046 for DIPG007 cells that could be further tested in combination with CAR-T cells to determine therapeutic synergy. CONCLUSIONS Our initial findings warrant further investigation into the combinatorial benefit of GSK-046 and CAR-T cells against DIPGs. Currently, we are optimizing and expanding our drug screen to identify additional compounds that may complement CAR-T cell therapy. Furthermore, we intend to validate the cytotoxic benefit of our prioritized combinations in additional in vitro and in vivo settings.
PATH-08. PRE-OPERATIVE TUMOUR CLASSIFICATION USING NANOPORE SEQUENCING OF CEREBROSPINAL FLUID
Authors: Natalia Stepien, Daniel Senfter, Julia Freund, Sarah Machreich, Katharina Bruckner, Leah Mager, Anna Laemmerer, Carola Jaunecker, Christine Pirker, Daniela Lötsch-Gojo, Lisa Mayr, Cora Hedrich, Maria Aliotti Lippolis, Hannah Schned, Clara Köller, Barbara Kiesel, Christian Dorfer, Amedeo A Azizi, Andreas Peyrl, Maria T Schmook, Philipp Euskirchen, Marc Pagès-Gallego, Carlo Vermeulen, Bastiaan Tops, Christine Haberler, Leonhard Müllauer, Sibylle Madlener, Johannes Gojo
Journal: Neuro-Oncology (2024)

Abstract BACKGROUND The introduction of tumour classification by analysis of DNA methylation has changed the landscape of paediatric neuro-oncology. With the launch of Oxford Nanopore sequencing, generation of sequencing data is possible in real-time, including information on base-modifications. Diverse classification tools for tumour tissue have been developed recently. While this is an important step towards optimization of surgical treatment, pre-operative knowledge on tumour entity would further facilitate surgical planning. Therefore, we investigated the possibility of liquid biopsy for rapid pre-operative tumour classification. METHODS CSF samples from patients with pre-operative extra-ventricular drain (EVD) (n = 5) as well as samples obtained via Ommaya reservoirs in cases of suspected recurrence (n = 3) were analysed. Additionally, samples from non-tumour patients (epilepsy, inflammation) who had lumbar puncture were available as controls. Samples were sequenced on MinIon devices (Oxford Nanopore) and subsequently classified with previously published tumour tissue classification pipelines (NanoDx, crossNN, Sturgeon). RESULTS Even though CSF derived from EVD has been stored on room temperature for up to 72 hours before processing, cfDNA isolation was possible with the advantage of higher CSF input amounts. DNA fragments showed a distribution characteristic for cell-free DNA, with a maximum peak at 170bp. The amount of total input DNA ranged from 0.35ng to 12ng, with higher input amounts generating more sequencing data but not resulting in better classification. Classification by sturgeon and crossNN/nanoDx using CSF derived cfDNA was possible in exemplary cases, such as a medulloblastoma patient with a new inoperable lesion, where CSF analysis resulted in medulloblastoma Group 4 within the first hour of sequencing. In total, 38% of samples were correctly classified. CONCLUSION Liquid biopsy as a method to pre-operatively classify tumours and thereby enable risk-adjusted surgery is feasible. Ongoing efforts focus on optimisation of ctDNA based tumour classification by extension of the cohort.