bluuebunny/crossref_metadata_2025 huggingface dataset:
MDB-01. A POSITIVE FEEDBACK LOOP BETWEENLNC-HLX-2-7, HLX, AND MYC STRONGLY PROMOTES GROUP 3 MEDULLOBLASTOMA
Authors: Katsushima Keisuke , Joshi Kandarp , Yuan Menglang , Batish Mona , Stapleton Stacie , Jallo George , Seal Sudipta , Taylor Michael D , Wechsler-Reya Rober J , Eberhart Charles G , Perera Ranjan J
Published: June 2024

Recent studies suggest that long non-coding RNAs (lncRNAs) contribute to medulloblastoma formation and progression. We have identified a lncRNA, lnc-HLX-2-7, as a potential therapeutic target in group 3 (G3) medulloblastomas. lnc-HLX-2-7 RNA specifically accumulates in the promoter region of HLX, its host gene, which activates HLX expression by recruiting multiple factors, including enhancer elements. RNA sequencing and chromatin immunoprecipitation reveal that HLX binds to and activates the promoters of several oncogenes, including TBX2, LIN9, HOXM1, and MYC. Intravenous treatment with cerium oxide nanoparticle–coated antisense oligonucleotides targeting lnc-HLX-2-7 (CNP-lnc-HLX-2-7) inhibits tumor growth by 40-50% in an intracranial medulloblastoma xenograft mouse model (p<0.01). Combining CNP-lnc-HLX-2-7 with standard-of-care cisplatin further inhibits tumor growth and significantly prolongs mouse survival compared with CNP-lnc-HLX-2-7 monotherapy (p<0.01). Thus, the lnc-HLX-2-7–HLX–MYC axis is important for regulating G3 medulloblastoma progression, providing a strong rationale for using lnc-HLX-2-7 as a therapeutic target for G3 MBs in children.
MDB-12. NANOPORE SEQUENCING AS A CUTTING-EDGE TECHNOLOGY FOR MEDULLOBLASTOMA CLASSIFICATION
Authors: Filser Mathilde , Torrejon Jacob , Merchadou Kevin , Girard Elodie , Bourneix Christine , Lemaître Elisa , Brillet Riwan , Wong Jennifer , Gentien David , Rapinat Audrey , Servant Nicolas , Vasiljevic Alexandre , Bertozzi Anne-Isabelle , Raimbault Sandra , Tauziède-Espariat Arnault , Lhermitte Benoit , Faure-Conter Cécile , Icher Céline , Berger Claire , Maurage Claude-Alain , Bodet Damien , Meyronet David , Uro-Coste Emmanuelle , De Carli Emilie , Forest Fabien , Palenzuela Gilles , Chotard Guillaume , Gauchotte Guillaume , Sudour Hélène , Mansuy Ludovic , Deparis Mariana , Machet Marie-Christine , Faisant Maxime , Entz-Werle Natacha , Varlet Pascale , Leblond Pierre , Michalak-Provost Sophie , Houdemont Stéphanie Proust , Rigau Valérie , Doz François , Bourdeaut Franck , Delattre Olivier , Ayrault Olivier , Masliah-Planchon Julien
Published: June 2024

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-19. THE 3D CHROMATIN LANDSCAPE OF MEDULLOBLASTOMA
Authors: Wu Stephanie C , Diaz Eduardo Ferreira Martins , Mileti Enrichetta , Bihannic Laure , Smith Kyle S , Rodríguez-González Francisco G , Sidiropoulos Nicos , Weischenfeldt Joachim , Northcott Paul A
Published: June 2024

BACKGROUND

Medulloblastoma (MB) is a highly malignant childhood cerebellar tumor comprised of four molecularly and clinically distinct subgroups. Sporadic yet subgroup-specific genetic and epigenetic landscapes suggest that higher-order chromatin topology may contribute to distinct subgroup identities and disease biology. Herein, we investigated the 3D chromatin landscape of MB by performing genome-wide chromosome conformation capture (Hi-C) across molecular subgroups using a cohort of n=24 patient-derived xenografts and primary tumor specimens, aiming to elucidate regulatory mechanisms facilitated by genome organization.

METHODS

Integrative analyses were adapted to incorporate a wide variety of sample-paired datasets. Chromatin contact matrices generated from Hi-C allowed for the mapping of active/inactive compartments (A/B compartments), topologically associated domains (TADs), and distal pairwise chromatin contacts (loops). Topological insights of chromatin architecture were complemented with whole-genome sequencing, RNA-seq, ATAC-seq, DNA methylation, CTCF and histone modification (H3K27ac/H3K27me3) ChIP-seq datasets to dissect mechanisms of gene regulation.

RESULTS

Unsupervised analysis revealed that chromatin compartmentalization is subgroup stratified. Gene loci contained in subgroup-unique active compartments correspondingly exhibited subgroup-specific chromatin contact patterns, adopted open chromatin conformations, and remained transcriptionally active. As an example, LIN28B resides within a Group 3-specific ‘A’ compartment where its local topology displays higher complexity compared to that of other subgroups. This consequentially facilitates a Group 3-unique loop connecting the LIN28B promoter with a distal enhancer, resulting in subgroup-specific transcriptional upregulation. Targeted displacement of CTCF that anchors and stabilizes chromatin looping at the LIN28B locus was sufficient to alter its transcriptional activity. Using similar lines of investigation, mechanistic insights into the topological determinants of known enhancer-hijacking events, including GFI1, GFI1B and PRDM6, were resolved.

CONCLUSIONS

Pioneering this integrative approach centered on comprehensive Hi-C maps, our study provides a high-resolution blueprint of the 3D chromatin landscape defining MB subgroups, informing the mechanistic basis of recurrent driver gene activation, and disclosing putative subgroup-specific vulnerabilities.
MDB-31. THE CLINICAL SIGNIFICANCE OF SUB-TOTAL SURGICAL RESECTION IN CHILDHOOD MEDULLOBLASTOMA: A MULTI-COHORT ANALYSIS OF 1100 PATIENTS
Authors: Keeling Claire , Davies Simon , Goddard Jack , Ramaswamy Vijay , Schwalbe Edward C , Bailey Simon , Hicks Debbie , Clifford Steven C
Published: June 2024

BACKGROUND

Medulloblastoma patients with sub-total surgical resection (STR; >1.5 cm2 primary tumour residuum) typically receive intensified treatment. However, the association of STR with poor outcomes has not been observed consistently, questioning the validity of STR as a high-risk disease feature.

METHODS

We collected extent of resection (EOR) data from 1110 patients (UK CCLG centres (n=416), published cohorts (n=694)), the largest cohort with clinico-molecular annotation assembled to specifically assess the significance of EOR to date. We performed association and survival analyses, assessing overall survival (OS) cohort-wide with reference to the consensus medulloblastoma molecular groups and clinico-molecular features.

RESULTS

STR was reported in 20% (226/1110) of the cohort and was enriched in patients (i) <5 years at diagnosis (p=0.021), (ii) with metastatic disease (p<0.0001) or (iii) with non-WNT tumours (p=0.047). In cohort-wide analysis, STR was associated with worse survival in univariable analysis (p<0.0001), though, outcomes for patients with STR as their only risk-feature were as per standard-risk disease. Examination of specific disease contexts showed STR was prognostic in univariate analysis for patients (i) receiving cranio-spinal irradiation (CSI) and chemotherapy (p=0.016) and (ii) with Group 3 tumours receiving CSI (p=0.039). In non-metastatic CSI treated patients, STR was not prognostic, independent of CSI dose. Crucially, STR was not independently prognostic in multivariable analyses when considered alongside established clinico-molecular high-risk features.

CONCLUSIONS

In a cohort of 1100 molecularly characterised medulloblastoma patients, STR (n=226) predicted significantly lower OS in univariable analysis, but was not an independent prognostic factor. Our data suggest that maximal safe resection can continue to be carried out for patients with medulloblastoma and suggest STR should not inform patient management when observed as a sole, isolated risk feature. These findings provide necessary evidence to inform the selection criteria for forthcoming risk-adapted medulloblastoma clinical trials.
MDB-32. PRDM6 PROMOTES MEDULLOBLASTOMA FORMATION IN NEUROEPITHELIAL STEM CELLS AND IS A CHROMATIN REPRESSOR BY BINDING TO HISTONE H3 LYSINE 27 TRIMETHYLATION MARKS
Authors: Schmidt Christin , Cohen Sarah , Gudenas Brian L , Husain Sarah , Carlson Annika , Westelman Samantha , Wang Linyu , Phillips Joanna J , Northcott Paul A , Weiss William A , Schwer Bjoern
Published: June 2024

A substantial fraction of Group 4 MBs are characterized by enhancer hijacking through tandem duplication of SNCAIP, resulting in high expression of PRDM6, a putative transcriptional repressor and histone methyltransferase. PRDM6 is a poorly characterized member of the PRDF1 and RIZ1 homology domain-containing (PRDM) family of transcription factors. Here, we investigated the function of PRDM6 in human hindbrain neuroepithelial stem cells and its potential role as a driver of Group 4 medulloblastoma. We report that human PRDM6 localizes predominantly to the nucleus, where it causes widespread repression of chromatin accessibility and complex alterations of gene expression patterns. Genome-wide mapping of PRDM6 binding reveals that PRDM6 binds to chromatin regions marked by histone H3 lysine 27 trimethylation that are located within, or proximal to, genes. Moreover, we show that PRDM6 expression in neuroepithelial stem cells promotes medulloblastoma. Surprisingly, medulloblastomas derived from PRDM6-expressing neuroepithelial stem cells match human Group 3, but not Group 4, medulloblastoma. We conclude that PRDM6 expression has oncogenic potential but is insufficient to drive Group 4 medulloblastoma from neuroepithelial stem cells. We propose that both PRDM6 and additional factors, such as specific cell-of-origin features, are required for Group 4 medulloblastoma. Given the lack of PRDM6 expression in normal tissues and its oncogenic potential shown here, we suggest that PRDM6 inhibition may have therapeutic value in PRDM6-expressing medulloblastomas.
MDB-38. AN INTEGRATED MODEL FOR RESISTANCE TO BROMODOMAIN INHIBITORS IN CHILDHOOD MEDULLOBLASTOMA
Authors: Choi Seung H , Lupien Leslie , Boynton Adam N , Bandopadhayay Pratiti , Prensner John R
Published: June 2024

BACKGROUND

Bromodomain inhibitors (BETi) have emerged as a potential treatment option for children with medulloblastoma, where hyperactivation of gene transcription by the MYC and MYCN transcription factors may lead to a transcriptional dependency. Yet, drug resistance limits the utility of these agents when employed as a monotherapy. We therefore sought to identify mechanisms of BETi resistance that inform rational development of multimodal therapy options.

METHODS

We used CRISPR screening of D458 medulloblastoma cells that have acquired resistance to the JQ1 inhibitor. We subsequently used small molecule inhibition and knockdown experiments to validate Menin as a target. Finally, we used RNAseq, ribosome profiling, and mass spectrometry to characterize transcriptional and post-transcriptional changes in D458 cells compared to D458 with JQ1 resistance.

RESULTS

In the setting of BETi resistance, we observed dependency on key transcriptional actors, such as MLL (KMT2A) and the Mediator complex, which we are now validating in mini-pool experiments. We further characterized aberrant MLL complex function through knockdown, small molecule inhibition, and RNA-sequencing of Menin, a critical MLL cofactor. Next, we were surprised to observe ribosome and proteosome genes as CRISPR dependencies in our initial screening of BETi-resistant cells, suggesting post-transcriptional regulation. Indeed, MYC, whose transcription is potently blocked by BETi, remained robust at the protein level in BETi resistance. We therefore performed integrated transcriptional, translational, and proteomic profiling to characterize gene regulation in BETi-resistance. We found that the BETi-resistant state leads to an “uncoupling” of transcriptional and post-transcriptional regulation for a subset of genes, leading us to test small molecule inhibitors of post-transcriptional processes in BETi-resistant medulloblastoma.

CONCLUSIONS

Taken together, our data inform an integrated model of BETi resistance based on both transcriptional and post-transcriptional mechanisms, suggesting new therapeutic angles that may be effective in combination with BETi therapies.
MDB-46. ARE RAB40 SMALL GTPASES DRIVERS OF MEDULLOBLASTOMA PATHOGENESIS?
Authors: Allen Joseph , Entwistle Susannah , Coyle Beth , Hume Alistair N
Published: June 2024

BACKGROUND

Medulloblastoma is the most common solid paediatric neoplasm and is among the leading causes of cancer-related deaths in children. Patients associated with high-risk molecular subgroups convey poor outcomes due to frequent relapse and metastasis, with two thirds of survivors experiencing life-changing developmental deficits. Poor patient outcomes and significant co-morbidities highlight an urgent need for specific, molecularly targeted therapies. The RAB40 subfamily of atypical small GTPases regulate cytoskeletal structure and cell adhesion via their ubiquitin ligase activity. Although previous studies have linked the RAB40 subfamily with cell invasiveness, their role in paediatric brain cancer remains undefined.

METHODS

Patient methylation and survival data from Cavalli et. al (2017) was analysed via the R2 Genomics Analysis and Visualisation Platform. RAB40 gene and protein expression were determined in eight in vitro models of medulloblastoma via qRT-PCR and western blotting. Cells were transfected with EGFP-Cas9 mRNA and RAB40-targeted CRISPR sgRNA and seeded singly using fluorescence-activated cell sorting. RAB40 knockout clones were identified via western blot and Sanger sequencing.

RESULTS

Patient data demonstrated that higher RAB40 gene expression correlates with worse outcomes in group 3 and 4 medulloblastoma. RAB40 genes were also significantly more highly expressed in high-risk groups 3 and 4 compared to low-risk WNT group medulloblastoma. RAB40 mRNA was enriched in cell lines derived from high-risk group 3 and 4 medulloblastoma relative to SHH controls. RAB40 knockout models were subsequently generated in HD-MB03, an in vitro model of metastatic Group 3 medulloblastoma using CRISPR/Cas9.

CONCLUSIONS

Our data supports the hypothesis that RAB40 small GTPases are enriched in high-risk medulloblastoma subgroups, and that their expression correlates with worse patient outcomes. We have generated knockout models which will be used to characterise the impact of RAB40 GTPases on cell growth, adhesion, invasiveness, and chemotherapeutic tolerance.
MDB-48. A SINGLE INSTITUTION EXPERIENCE OF CIPN MEASURED BY THE PED-MTNS IN CHILDREN AND ADOLESCENTS WITH MEDULLOBLASTOMA TREATED ON A TRIAL USING SIMILAR DOSING OF VINCRISTINE AND CISPLATIN FOR ALL PATIENTS
Authors: Richards Jill M , Olsen Monica R , Bendel Anne E , Gilchrist Laura S , Tanner Lynn R
Published: June 2024

BACKGROUND

Standard treatment for medulloblastoma in patients >3 years of age consists of tumor resection, radiation and chemotherapy using vincristine, cisplatin, cyclophosphamide +/- CCNU. Vincristine and Cisplatin are known to cause chemotherapy induced peripheral neuropathy (CIPN). The prevalence of CIPN in children with non-CNS cancers can be measured using the pediatric-modified Total Neuropathy Score (ped-mTNS). The purpose of this observational, retrospective pilot design is twofold: to investigate the prevalence of CIPN in children with medulloblastoma receiving neurotoxic chemotherapy, and to examine the feasibility of using the ped-mTNS as a measurement tool in this population.

METHODS

Data was manually extracted from medical records and entered into a REDCap database for analysis. The percent of patients successfully tested and clinical inferences made from ped-mTNS testing results were used to determine feasibility and barriers to use. A total of 33 medulloblastoma patients >3 years of age were treated from 6/2013 - 6/2021 at Children’s Minnesota. Of those, 18 received testing using the ped-mTNS during chemotherapy treatment.

RESULTS

During cancer treatment, 54.5% of the cohort of patients received ped-mTNS testing. The average age of children tested was 10.26 (4.39) years with a majority being male (79%), and non-Hispanic white race/ethnicity (89%). Of the tested participants, 61.1% had more than one measurement. For those patients tested at multiple timepoints, 72.7% demonstrated worsening ped-mTNS scores while on chemotherapy treatment. Therapists recorded inferences relating to neuropathy in 88.1% of testing occurrences with questions of accuracy recorded in 31.3% of testing occurrences. Barriers to use of results included confounding impairments that may occur as a result of the primary tumor, post-surgical changes, and radiation.

CONCLUSIONS

Using the ped-mTNS is feasible in children and adolescents with medulloblastoma. The ped-mTNS scores indicated that children with medulloblastoma, treated with Vincristine and Cisplatin, experience worsening neuropathy while undergoing chemotherapy treatment.
MDB-52. MYC-DRIVEN PEDIATRIC BRAIN TUMORS SHARE A COMMON PHOTORECEPTOR IDENTITY
Authors: Zhao Miao , Rosén Gabriela , Sundstrom Anders , Olsen Thale Kristin , Furukawa Takahisa , Gudenas Brian L , Northcott Paul A , Swartling Fredrik J
Published: June 2024

Mouse models serve as invaluable tools for improving the understanding and treatment of pediatric brain tumors. We recently published a highly aggressive MYC-driven brain tumor model (GMYC), which exhibits approximately 70% penetrance. Our transcription profiling indicates that GMYC tumors accurately resemble human Group 3 medulloblastoma (MB-G3). Interestingly, a strong photoreceptor program was activated in the tumor model. This is commonly upregulated in MB-G3 but also in pineoblastoma (PB), another malignant pediatric brain tumor. CRX, a transcription factor critical for pineal cell development is also a master regulator for MB-G3 maintenance. By studying early tumor initiation using Crx-lineage tracing and scRNA-seq, we confirmed that GMYC tumors originate from early pinealocytes. Exome sequencing of GMYC tumors further confirmed mutations of epigenetic regulators commonly reported in either PB or MB. To investigate tumor development from photoreceptor-positive progenitors in the developing brain, we performed Crx-lineage tracing with tamoxifen injections, confirming that Crx-positive cells exist in pineal gland progenitors. However, Crx-traced cells also marked granule neurons in the flocculonodular lobe of the cerebellum, which is the suggested site of MB-G3 origin arising from the developing rhombic lip. To investigate if the Myc oncogene can generate different types of pediatric brain tumors in Crx-positive cells, an XMYCT58A-Tomato mouse model was established by crossing Crx-CreERT2; LSL-MycT58A and Rosa26LSL-tdTomato strains. Here, MycT58A was turned on with tamoxifen injection after birth and tumor development followed using the tdTomato reporter. XMYCT58A-Tomato mice developed brain tumors from both the pineal gland and cerebellum after 2-6 months. Histologically, tumors were non-glial (GFAP-, Olig2-), showed neuronal activity (Neurod1+), and stained positive for photoreceptor identity markers (CRX+, OTX2+) similar to both MB-G3 and PB-MYC. Collectively, our data suggest that both MYC-driven MB-G3 and PB-MYC originate from photoreceptor-positive cells, which has implications for developing novel treatments targeting these devastating childhood malignancies.
MDB-54. GENOMIC VARIANCE OF MATCHED PRIMARY AND RECURRENT HUMAN MEDULLOBLASTOMA
Authors: Su Yu N/A , Liu Hailong N/A , Li Jiankang N/A , Qiu Xiaoguang N/A , Jiang Tao N/A , Tian Yu N/A
Published: June 2024

BACKGROUND

To investigate the difference between matched primary and recurrent human medulloblastoma across four molecular subgroups and to further expand the existing mutational spectrum in the recurrence or dissemination lesions.

METHODS

We analyzed the whole-exome sequencing (WES) data of 56 matched samples from 28 pairs with primary and recurrent/disseminated medulloblastoma. WES was performed using the Illumina HiSeq 2500 system, and the sequencing data were then subjected to variant detection, genome annotation, GO functional enrichment analysis, and GWAS.

RESULTS

All tumor samples were classified as four subgroups, including 4 cases of WNT-MB, 8 cases of SHH-MB, 20 cases of G3-MB and 24 cases of G4-MB. Conical and non-conical chromosomal variations were observed in each subgroup. Recurrent G3-MB cohort exhibited the loss of chr1p and chr22q and gain of the chr16p. The gain of chr2p, 2q, and 9p, as well as the loss of chr9q were found in the recurrent lesions of G4-MB. Recurrence of SHH-MB exhibited the loss of chr19q and chr4p, and loss of chr5p was found in the WNT-MB recurrences. CTNNB1, IDH1, and ARID1A were only observed in recurrent tumors of G3-MB; MYC was only observed in the recurrent tumors of G4-MB; IDH1 and TERT were only observed in recurrent tumors of the SHH-MB. Compared to primary medulloblastoma, recurrent tumor samples obtained a higher level of tumor mutation burden. The mutant gene profiles were mainly enriched in cell adhesion function.

CONCLUSIONS

When relapsing, medulloblastoma subgroup generally remains consistent, but exhibits the specific CNV, SNV and mutated gene profiles.
MDB-56. CLINICAL OUTCOME AND SURVIVAL OF INFANT MEDULLOBLASTOMA PATIENTS: THE BAMBINO GESÙ CHILDREN’S HOSPITAL EXPERIENCE
Authors: Baldo Giada Del , Ruscio Valentina Di , Sasso Annalisa Lo , Megaro Giacomina , Cacchione Antonella , Alessi Iside , De Salvo Andrea , Albino Giulia , Cipri Selene , Agolini Emanuele , Miele Evelina , Rossi Sabrina , Giovannoni Isabella , Colafati Giovanna Stefania , Vennarini Sabina , Vinci Maria , Carai Andrea , Mastronuzzi Angela
Published: June 2024

Young children diagnosed with medulloblastoma (MB) pose a significant challenge due to the high susceptibility of the developing brain to neurotoxic agents such as craniospinal irradiation (CSI). Different innovative strategies were developed with the aim to enhance outcomes while either delaying or avoiding CSI. We describe a case series of patients affected by infant MB treated according to the Italian Infants High Risk recommendation. Retrospective data from 42 children with MB younger than 5 years of age at diagnosis treated between 2007 and 2023 at Bambino Gesù Children Hospital were recorded. Demographic, clinical, surgical, radiological, molecular, and histopathological data were collected. Overall survival (OS) and event-free survival (EFS) were calculated using the Kaplan-Meier method. Seventeen patients were excluded for first-line radiotherapy. Conclusively, 25 patients (13 male, 12 female) with a median age of 25.3 months at diagnosis were eligible for the analysis. Eleven patients had metastatic disease at diagnosis. Histotype: 13/25 classic, 6/25 desmoplastic nodular (DNMB), 4/25 extensive nodularity (MBEN) and 2/25 large cell/anaplastic. DNA methylation profile: SHH-subgroup: 12/25, group 3: 8/25 and group 4: 5/25. MYC amplification was detected in 3 cases. In five cases a cancer predisposition syndrome was found. The median duration of follow-up was 48 months. EFS at 12, 24 and 60 months was 80% (95%CI: 58.4-91.1), 72% (95%CI: 50.1-85.5), and 68% (95%CI: 46.1-82.5), respectively. OS at 12, 24 and 60 months was 80% (95%CI: 67.3-96), 80% (95%CI: 58.4-91.1) and 68% (95%CI: 46.1-82.5) respectively. Significant better survival was found for MBEN/DNMB histotype, with no significant difference between molecular subgroups. We reported high EFS and OS in a population of infant MB treated according Italian Infants High Risk recommendation. Our patients presented an average intelligence quotient in the normal range at a median follow-up of 4 years from diagnosis and no long-term toxicity after treatment.
MDB-57. PROTEOMIC ANALYSIS REVEALS AMBRA1 AS A NEW CRL3REN/KCTD11 SUBSTRATE PROMOTING SONIC HEDGEHOG-DEPENDENT MEDULLOBLASTOMA
Authors: Basili Irene , Infante Paola , Conenna Marilisa , Bernardi Flavia , Marsaud Veronique , Cancila Gabriele , Bufalieri Francesca , Severini Ludovica Lospinoso , Yu Hua , Diaz Jacob Torrejon , Re Valentina Lo , Adabbo Gennaro , Guardavaccaro Daniele , Ayrault Olivier , Marcotullio Lucia Di
Published: June 2024

BACKGROUND

Sonic Hedgehog (SHH) signaling pathway is fundamental for a proper embryonic development and its uncontrolled activation is responsible for various cancers. One the most relevant SHH-dependent tumors is medulloblastoma (MB), a highly heterogeneous brain malignancy that predominantly occurs in childhood. Understanding the complex networks that govern SHH aberrant activation represents a dramatic challenge for the development of efficient personalized therapies. Here, we identify a new role of the pro-autophagic protein AMBRA1 as interactor of REN/KCTD11 (here REN), the substrate-receptor subunit of the Cullin3-RING ubiquitin ligase complex (CRL3REN) and tumor suppressor lost in ~30% of human SHH-MBs. We show a novel mechanism of action of AMBRA1 in SHH-MB pathogenesis whereby its highly expression, due to loss of REN, drives the activation of GLI1, the final effector of SHH signaling.

METHODS

Proteomic analysis, co-immunoprecipitation, ubiquitylation and cycloheximide assays have been performed to demonstrate that AMBRA1 is a substrate of ubiquitylation of CRL3REN and to define AMBRA1 mechanism of action in SHH-MB tumorigenesis. Transcriptomic analysis, in vitro and in vivo proliferation assays have been carried out on murine and patient-derived xenograft (PDX) models to examine the effect of AMBRA1 modulation on SHH-MB growth.

RESULTS

We demonstrate that CRL3REN binds and induces polyubiquitylation and proteasome-mediated degradation of AMBRA1. Interestingly, we found that genetic depletion of AMBRA1 strongly reduces SHH-dependent MB growth both in vitro and in vivo, an effect associated with the decrease in the expression of GLI1 and other SHH target genes. We hypothesize that AMBRA1 could impact on the stability of GLI1 by counteracting the activity of E3 ubiquitin ligases regulating GLI proteins. Remarkably, transcriptomic analysis of SHH PDX upon AMBRA1 knockdown revealed an impairment of SHH activity and an upregulation of cerebellum differentiation.

CONCLUSION

Our findings identify AMBRA1 as a novel CRL3REN substrate and promising therapeutic target for SHH-dependent cancer.
MDB-58. REDUCED CRANIOSPINAL DOSE FOR NEWLY DIAGNOSED HIGH-RISK MEDULLOBLASTOMA IN CHILDREN YOUNGER THAN 5 YEARS OLD: RETROSPECTIVE EXPERIENCE FROM SFCE
Authors: Oozeerally Jihann , Dufour Christelle , Faure Cécile , Cardine Aude , De Carli Emilie , Merlin Marie-Sophie , Andry Leslie , André Nicolas , Icher Céline , Klein Sébastien , Pagnier Anne , Puiseux Chloé , Briandet Claire , De Benaze Gwenaelle Duhil , Bolle Stéphanie , Padovani Laetitia , Flumian Clara , Bertozzi Anne-Isabelle
Published: June 2024

BACKGROUND

High Risk medulloblastoma (MB) treatment strategy is based on an intensive multimodal treatment including surgery, chemotherapy and radiotherapy. French adapted PNET HR +5 is based on high-dose chemotherapy and age-stratified (18 Gy < 3 yo, 23 for 3-5 yo, 36 Gy for > 5 yo) craniospinal dose irradiation to reduce long-term side-effects of radiotherapy.

METHODS

To estimate progression-free (PFS) in a national multi-institutional retrospective study, we enrolled children younger than 5 yo with newly diagnosed high-risk MB treated according to adapted PNET HR +5 trial.

RESULTS

Thirty-seven children from 15 French hospitals (median age, 3.4 yo; range, 0.7-4.8) with confirmed high-risk MB were included. 15 patients (39%) were less than 3 years old. 29 patients (81%) had metastatic disease. Group 3 was the most represented molecular subgroup. 5-year PFS and OS was 59 % and 77 %, respectively. According to dose of craniospinal irradiation, PFS for 18 Gy, 23.4 Gy and 36 Gy patient groups were 100 %, 75 % and 45%, respectively. Treatment was well tolerated. One toxic death was declared. Analysis of longitudinal neurocognitive performance is ongoing.

CONCLUSIONS

the French strategy comprising high-dose chemotherapy and age-adapted craniospinal irradiation is promising with a high rate of survival in young children and must be confirmed by a prospective trial.
MDB-75. MOLECULAR PATHOLOGY, TREATMENT AND PROGNOSIS OF INFANT SONIC HEDGEHOG MEDULLOBLASTOMA: A GLOBAL MULTI-COHORT STUDY
Authors: Richardson Stacey , Hicks Debbie , Gough Melissa , Castle Jemma , Crosier Stephen , Garcia-Ariza Miguel , Martín-Guerrero Idoia , LA Plasschaert Sabine , Bourdeaut Franck , Dufour Christelle , Masliah-Planchon Julien , Buttarelli Francesca R , Garre Maria-Luisa , Biassoni Veronica , Massimino Maura , Kanemura Yonehiro , Ramaswamy Vijay , Gajjar Amar , Northcott Paul , Robinson Giles , Korshunov Andrey , Pfister Stefan M , Mynarek Martin , Rutkowski Stefan , Schwalbe Edward C , Bailey Simon , Clifford Steven C
Published: June 2024

BACKGROUND

Clinical studies in infant medulloblastoma (iMB; <5 years) have, to date, focussed on modestly-sized or national trials cohorts and have not directly compared therapeutic approaches or placed these in the context of dedicated biomarker studies, to inform future treatment strategies.

METHODS

We assembled a multi-national cohort of molecularly and clinically-annotated iMBs (<5 years at diagnosis; n=646), the largest to date. We investigated molecular pathology, treatments received, and relationships to outcome within this population.

RESULTS

The SHH group (iMBSHH; n=267, 40%) predominated, encompassing SHH-1 (37.7%, median age 2.0 years), SHH-2 (47.0%, 1.4 years) and SHH-3 (14.4%, 3.0 years) WHO subgroups. MBEN histology was significantly enriched in SHH-2, and SUFU mutation and MYCN amplification in SHH-1 and SHH-3, respectively. Notably, TP53 mutations were identified in all subgroups; in SHH-1 and SHH-2 (each n=3) tumours lacked features typically associated with TP53-mutated SHH-3 (LCA histology and MYCN amplification) and did not have a worse survival. Upfront radiation-sparing treatments were used in 132/267 children and comprised regimens founded on intraventricular methotrexate (IVT-MTX; 54.5%), high-dose (HDCTx; 22.0%) or standard-dose (23.5%) chemotherapy. Across all radio-naive iMBSHH, non-DN/MBEN histology (HR 2.76, CI 1.26-6.01, p=0.011) and SHH-1 (vs SHH-2, HR 2.51, CI 1.25-5.01, p=0.009) conferred worse PFS in univariable analysis; subgroup was the only independently prognostic risk-factor (multivariable analysis; SHH-1 HR 2.65, CI 1.27-5.51, p=0.009). 5-year OS for HDCTx recipients was very favourable in SHH-1 (100%) and SHH-2 (93.3%), and subgroup-dependent for IVT-MTX (SHH-1; 76.4%, SHH-2; 100%, p=0.009). In CSI-treated iMBSHH (n=49), non-DN/MBEN histology (HR 10.23, CI 2.22-47.04, p=0.003) and MYCN amplification (HR 7.35, CI 2.01-26.87, p=0.003) conferred worse PFS.

CONCLUSION

iMBSHH outcomes in this cohort are dependent upon WHO subgroup, histology and therapy received. These findings provide an evidence-based foundation for selection of cohorts and therapies for prospective assessment in forthcoming iMBSHH clinical trials (e.g. SIOP-CONNECT-YC-MB-LR).
MDB-79. THE DEUBIQUITINATING ENZYME USP7 COOPERATES WITH SONIC HEDGEHOG SIGNALING TO PROMOTE MEDULLOBLASTOMA
Authors: Cigna Sara Maria , Basili Irene , Bernardi Flavia , Severini Ludovica Lospinoso , Forget Antoine , Chang Chia-Hsiang , Yu Hua , Cancila Gabriele , Marsaud Veronique , Talbot Julie , Mercier Audrey , Lasgi Charlene , Puget Stéphanie , Beccaria Kevin , Blauwblomme Thomas , Kawauchi Daisuke , Kumar Suresh , Tsai Jin-Wu , Marcotullio Lucia Di , Ayrault Olivier
Published: June 2024

BACKGROUND

Constitutive activation of the Sonic Hedgehog (SHH) signaling pathway during cerebellar development may lead to medulloblastoma (MB), one of the most common malignant pediatric brain tumor. Clinically, SHH-MB with mutations downstream of SMO or TP53-mutated are a real challenge and alternative targeted therapies are sorely needed for this group of patients. In SHH-MB, the transcription factor Atoh1 is constitutively overexpressed and essential for tumor initiation and progression. Previous work demonstrated that proteasomal-mediated degradation of Atoh1 induces tumor regression in vitro and in vivo, therefore deciphering mechanisms controlling Atoh1 stabilization should provide new avenues for drug development in SHH-MB.

METHODS

Using a proteomic approach, we identified the deubiquitinating enzyme Usp7 as a positive regulator of Atoh1 in MB cells. We then applied a SILAC-based quantitative ubiquitinomics to investigate the Usp7 target landscape in SHH-MB. We also employed several biochemical approaches to demonstrate the functional interaction between Atoh1 and Usp7 in SHH-MB. In parallel, we validated the impact of Usp7 depletion in vitro and in vivo using relevant SHH-MB models. Last, we investigated large transcriptomic and proteomic datasets to unveil the prognostic value of Usp7 in SHH-MB human tumors.

RESULTS

We demonstrate that Usp7 binds and accumulates Atoh1 protein by counteracting its ubiquitylation. Post-natal electroporation in vivo directly into the developing cerebella also indicates that overexpression of Usp7 promotes proliferation, whereas in the absence of Atoh1 Usp7 was no longer able to spread cell growth, demonstrating a functional link between the Usp7-Atoh1 axis. In vivo, disruption of Usp7 expression in SHH-MB blocks tumor proliferation. Pharmacological inhibition of Usp7 triggers tumor growth in a syngeneic SHH MB mouse model. Lastly, SHH-MB cells with either TP53 or SMO downstream mutations are sensitive to Usp7 inhibition.

CONCLUSIONS

Overall, our data pinpoint Usp7 as a promising target against Atoh1, a master regulator of SHH-MB.
MDB-82. TRANSFUSION-RELATED IRON OVERLOAD MIMICKING METASTATIC DISEASE IN A PEDIATRIC PATIENT WITH MEDULLOBLASTOMA ON A RADIATION-DELAYING CHEMOTHERAPY REGIMEN
Authors: Pytel Nicholas , Glanz Hannah
Published: June 2024

BACKGROUND

Transfusion-related iron overload (TRIO) is a known complication of administering numerous packed red blood cell (pRBC) transfusions in pediatric oncology patients undergoing myelosuppressive therapy. Infant medulloblastoma regimens attempt to avoid or delay radiation and its adverse effects in this population, with intensive myelosuppressive chemotherapy. We report a case of TRIO first suspected as hepatic metastatic disease at time of CNS recurrence in a young patient with medulloblastoma after completion of a high-dose chemotherapy protocol.

METHODS

Case Report

RESULTS

A five-year-old male presented with four months of headaches, dizziness, imbalance, and emesis. MRI revealed a posterior fossa mass with near total resection of the mass confirming classic medulloblastoma, non-WNT, non-SHH. The patient received five cycles of induction chemotherapy. This was followed by a single cycle of high-dose chemotherapy (HDCSCR) with autologous stem cell rescue. During therapy, he received 26 pRBC transfusions to correct severe anemia. CNS recurrence occurred 19 months after therapy. Abdominal imaging revealed enlarging hepatic lesions concerning for metastases. Ferritin was 1,268 ng/mL (normal: 7-142 ng/mL) and MRI of liver revealed severe iron load with LIC of 11 mg Fe/g dry weight (dw) (normal: 0.4-2.2 mg Fe/g dw). Ultimately, hepatic lesions were thought to be non-cancerous, and he proceeded with craniospinal irradiation. He was diagnosed with TRIO. He underwent phlebotomy for 33 months with ferritin and MRI liver LIC within normal limits.

CONCLUSION

This case illustrates an occurrence of TRIO presenting as hepatic lesions concerning for metastatic disease during CNS recurrence in a patient who underwent an infant medulloblastoma regimen with HDCSCR. Our patient required phlebotomy for nearly 3 years to return ferritin and LIC to normal limits. Further investigation on the prevalence of TRIO in medulloblastoma survivors treated with radiation delaying/sparing protocols is necessary to improve outcomes and avoid end organ damage due to iron overload.
MDB-85. THE MIGRATORY ROLE OF BAIAP2 IN MEDULLOBLASTOMA
Authors: Ruiz Luz C , Jangde Nitish , Suter Robert , Jermacowikz Anna , Ayad Nagi
Published: June 2024

BACKGROUND

Medulloblastoma is the most common malignant brain tumor in adolescents. There is still a dire need for a more effective treatment despite advances in technology. One challenge in treating medulloblastoma is its migratory and invasive properties. The underlying mechanisms that lead to these characteristics are still not understood.

METHODS

We sought to identify targets related to invasion and migration in medulloblastoma, first by mining single-cell RNA sequencing and microarray data.

RESULTS

We determined that BAIAP2 is differentially expressed in medulloblastoma compared to non-tumor brain cells and other brain cancers. In addition, there is a significant increase in BAIAP2 in metastatic medulloblastoma compared to non-metastatic. We found that CDC42 and RAC1, small GTPases, are protein-protein interactors with BAIAP2. In three medulloblastoma cell lines, we have demonstrated that these genes modulate each other’s expression. Furthermore, we show that the knockdown of BAIAP2, CDC42, and RAC1 decreases medulloblastoma cell migration and filopodia length. Using surface plasmon resonance, we identified compounds that target BAIAP2 protein. After characterizing these compounds, we have identified potential compound candidates that target these genes and impede their roles in migration and invasion in medulloblastoma.

CONCLUSIONS

These findings demonstrate a migratory role for BAIAP2 in medulloblastoma through the modulation of CDC42 and RAC1. We also demonstrate that BAIAP2 is a therapeutic target and have identified novel compounds that target it.
MDB-86. ISOTRETINOIN HAS NO EFFECT ON EVENT FREE SURVIVAL ACROSS HIGH-RISK MEDULLOBLASTOMA MOLECULAR SUBGROUPS WHEN ADDED TO MAINTENANCE CHEMOTHERAPY: A SECONDARY ANALYSIS OF THE CHILDREN’S ONCOLOGY GROUP ACNS0332 DATA
Authors: Raskin Scott I , de Blank Peter M K , Billups Catherine A , Li Yimei , Olson James M , Leary Sarah E S
Published: June 2024

BACKGROUND

Medulloblastoma has been redefined into four molecular subgroups (WNT, SHH, Group 3, and Group 4), with distinct prognoses and responses to treatment. The ACNS0332 trial investigated the impact of carboplatin and isotretinoin in high-risk medulloblastoma finding a differential effect of carboplatin by molecular subgroup. An interim futility analysis showed no benefit from isotretinoin, but did not evaluate results by molecular subgroup in the primary analysis. We performed a secondary analysis to examine whether the effect of isotretinoin on event-free survival (EFS) differed by molecular subgroup.

METHODS

Evaluable patients with known molecular subgroups were analyzed. Molecular subgroup analysis was conducted retrospectively in 231 patients at a central laboratory by methylation array. Kaplan-Meier survival analysis compared EFS (time from enrollment to disease progression or recurrence, second malignancy, death from any cause, or to date of last follow-up for patients without events).

RESULTS

Based on data from the 2014 interim analysis, 164 patients were eligible for secondary analyses. Group 3 (n=56, 34.1%) and Group 4 (n=75, 45.7%) were the most common groups. SHH (n=22, 13.4%) and WNT (n=11, 6.7%) groups were less common. There was no difference in isotretinoin (p=0.87) or carboplatin (p=0.49) randomization by subgroup. Race was well-balanced with expected differences in age (p=0.017) and sex (p=0.016) among molecular subgroups. Based on the interim analysis, there was no evidence of a difference in EFS by isotretinoin randomization (p=0.34; one-sided log-rank test stratified by carboplatin randomization). Based on secondary analysis data, there was no difference in EFS between patients receiving isotretinoin or no additional therapy in any molecular subgroup: Group 3 (p=0.68), Group 4 (p=0.55), SHH (p=0.68), WNT (p=1) based on two-sided log-rank tests.

CONCLUSIONS

Despite differential efficacy of carboplatin among molecular subgroups, no difference was found in EFS between patients receiving isotretinoin and those who did not, regardless of molecular subgroup.
MDB-90. PRIMARY LEPTOMENINGEAL MEDULLOBLASTOMA: A SINGLE-SITE RETROSPECTIVE STUDY OF 5 CASES
Authors: Wells Olivia , Bhatia Kanika , Campbell Marty , Hansford Jordan R , Sullivan Michael , Williams Molly , Mignone Cristina , D’Arcy Colleen , Eisenstat David D , Khuong-Quang Dong-Anh
Published: June 2024

BACKGROUND

Primary leptomeningeal medulloblastoma without mass is a rare entity with fewer than 10 reported paediatric cases. Establishing an accurate diagnosis is challenging and reported outcomes are very poor.

METHODS

We conducted a retrospective analysis of cases presenting to the Royal Children’s Hospital Melbourne between 2013 and 2023.

RESULTS

Between 2013 and 2023 five patients, all male, were identified, with an average age of 6.6 years [1.9 - 12.2 years]. The presenting history was short in all cases (2-3 weeks) and all had symptoms of raised intracranial pressure. Diagnostic imaging (MRI brain and spine) showed non-specific leptomeningeal thickening and enhancement with no primary posterior fossa mass seen; interestingly only two imaging reports considered malignancy in the differential diagnosis. Surgical biopsy confirmed the diagnosis of medulloblastoma on histology; 3 anaplastic and 2 with classic histology. Subgrouping by IHC, available for 4 patients, showed non-WNT/non-SHH, with one reclassified as SHH by DNA methylation (performed in 2 patients). MYC was amplified in one patient (assessed by FISH in all tumours). Four patients were treated with craniospinal irradiation (36-39.6Gy with 18Gy boost), followed by chemotherapy as per SJMB12 stratum N3 (n=3) or HeadStart III (n=1). One case, an infant, was treated according to COG ACNS0334 with concurrent intraventricular topotecan, followed by maintenance with irinotecan/temozolomide/IVent topotecan. At follow up (average 54 months; 11 – 126 months), four children with residual disease on end of treatment MRI are alive without disease progression. One patient with classical SHH medulloblastoma and MYCN amplification progressed on treatment and died of disease 11 months after diagnosis.

CONCLUSIONS

Contrary to the historical literature, our case series suggests a more favourable outcome if this disease is recognised early. Further studies are warranted to better understand the molecular profile of these tumours.
MDB-93. A TRANSCRIPTOME-BASED APPROACH TO DRUG REPURPOSING FOR AGGRESSIVE GROUP 3 MEDULLOBLASTOMAS
Authors: Doss David , Kanchan Ranjana K , Perumal Naveenkumar , Batra Surinder K , Mahapatra Sidharth
Published: June 2024

BACKGROUND

Medulloblastoma (MB) is the most common malignant brain tumor in children, representing 20% of pediatric brain tumors. International consensus has identified molecular subgroups: WNT, SHH, group 3 (G3), and group 4 (G4). Current treatments, guided by histologic tumor grade and stage, do not fully harness these subgroups, resulting in varying prognoses. G3MB particularly faces challenges with rapid recurrence and aggressive disease, necessitating focused intervention.

METHODS

We conducted differential expression analysis on MB patient datasets (GSE148389, GSE164677) to generate a G3MB gene expression signature. This gene expression signature was analyzed against the LINCS database, yielding candidate antineoplastic compounds. Candidates were filtered for blood-brain barrier permeability and FDA approval. Functional assays evaluated drug effects on cytotoxicity, clonogenicity, wound healing, cell cycle, and apoptosis in G3MB cell lines (HDMB03 and D425). RNA sequencing of treated cells identified differentially regulated pathways to infer mechanism, followed by in vitro validation. Mitochondrial function and membrane potential were measured by Seahorse and TMRM assays, respectively.

RESULTS

Eighty-one candidates passed filtering. The leading drug class identified was antidepressants. Based on MTT assays, the top four candidates were nortriptyline, simvastatin, fluoxetine, and sertraline. Nortriptyline (NT) emerged as the lead compound, with IC50 values of ~7uM (HDMB03) and ~11uM (D425). Amongst the four drugs, NT induced consistent anti-neoplastic effects on wound healing, colony formation, medullosphere generation, and apoptosis. Mechanistically, NT induced mitochondrial stress, escalated mitochondrial superoxide, and decreased mitochondrial membrane potential. CHRM3, a receptor antagonized by NT and upregulated in G3MB samples was downregulated by NT treatment. Western blotting confirmed the inhibition of CHRM3 downstream effectors.

CONCLUSIONS

Our pipeline identified cytotoxic compounds against G3MB, including NT, which we hypothesize induces mitochondrial stress via CHRM3 modulation. This approach offers repurposable FDA-approved candidates for high-risk G3MB, potentially reducing current treatment toxicity and improving outcomes.