bluuebunny/crossref_metadata_2025 huggingface dataset:
DIPG-16. THE BIG AND SMALL OF PROTEIN POST-TRANSLATIONAL MODIFICATIONS: TARGETING SUMOYLATION AND CITRULLINATION IN DIFFUSE MIDLINE GLIOMA
Authors: Rayner Benjamin S , Samadder Arjayeeta , Holliday Holly , Chen Yongjuan , Nguyen Hieu , Lee Alvin , Liu Jie , Hayden Elisha , Ogunmodede Oluwaseyi S , Rye Kerry-Anne , Tsoli Maria , Bertoldo Jean B , Ziegler David S
Published: June 2024

BACKGROUND

Protein post-translational modifications (PTMs) alter the properties of protein through proteolytic cleavage, or the addition of modifying groups to one or more amino acids. Histones are major targets of PTMs, including through SUMOylation of lysine residues and citrullination of arginine residues, with both processes able to regulate gene transcription. We aimed to delineate the potential role of SUMOylation and citrullination in the epigenetic dysregulation that drives Diffuse Midline Glioma (DMG) tumorigenicity.

METHODS AND RESULTS

DMG tumours demonstrated an upregulation of SUMOylation machinery pathway components and increased expression of peptidylarginine deiminases (PADs), the enzymes responsible for citrullination. Nanomolar doses of the SUMOylation inhibitor TAK-981 blocks DMG cell cycle G2/M transition and results in potent decreases in cell viability and clonogenic potential, triggering ‘viral mimicry’ in DMG cells, resulting in increased expression of interferon stimulated genes and inflammatory mediators including IL-1β, IL-6 and TNFα, whilst also causing a pro-inflammatory M1 macrophage response. These data suggest that SUMOylation inhibition may lead to a double-punch effect against DMG by inhibiting tumorgenicity and stimulating an immune response. In further studies, we have shown that pharmacological inhibition of PAD activity in DMG decreases histone citrullination and restores histone methylation – considered the Achilles heel of DMG. This was accompanied by induction of reactive oxygen species and an unfolded protein response, resulting in DMG cell death in a dose dependent manner. In addition to fully characterising the epigenetic and global changes within DMG initiated through SUMOylation or PAD inhibition, we are currently performing preliminary drug efficacy studies in orthotopic DMG models, providing the necessary pre-clinical research pipeline towards an effective treatment for DMG.

CONCLUSIONS

Protein SUMOylation and citrullination are previously unrecognised therapeutic targets in DMG and potentially play key roles in tumour initiation and pathogenesis. Our data suggests a novel therapeutic approach for children with DMG.
DIPG-17. DYSREGULATED PURINE METABOLISM IN A LETHAL CHILDHOOD BRAINSTEM TUMOR REVEALS NEW TREATMENT STRATEGIES WITH OLD DRUGS
Authors: Mersich Ian , Congrove Sunny , Horchar Matthew , Caceres Roman , Desai Janki , Desai Pankaj , Sallans Larry , Haines Julie , Menon Ranjith , Stevenson Charles , de Blank Peter , Pillay-Smiley Natasha , Hummel Trent , Tavassoli Ali , D’Alessandro Angelo , Phoenix Timothy , Dasgupta Biplab
Published: June 2024

The median survival of children with Diffuse Intrinsic Pontine Glioma (DIPG), an incurable brainstem cancer is less than 1 year. Radiation therapy is the only treatment that offers a transient benefit. The preclusion of surgery and biopsy due to the anatomical location of the tumor impeded our understanding of tumor biology for a long time. Genomic analysis during the past decade revealed that over 80% of DIPGs harbor an oncogenic histone H3K27M point mutation. However, the pathways altered by the mutation are still poorly understood. Metabolites serve as proxies to the state of biochemical pathways in a tissue. Given that unbiased a priori knowledge about metabolic alterations in DIPG is limited, we set out for the first time to identify metabolic dependencies intrinsic to DIPGs by leveraging untargeted and targeted metabolomics, transcriptomics, ChiP-Seq and ATAC-seq and integrating multi-omics datasets. We discovered that the mutant histone reprograms chromatin landscape causing aberrant misregulation of genes including a family of transcription factors that in turn causes misregulation of purine biosynthesis and folate metabolism. Specifically, the mutation alone is sufficient to upregulate a rate limiting bifunctional enzyme of de novo purine biosynthesis called ATIC, and the reduced folate carrier SLC19A1 to enhance purine synthesis and folate import, respectively, causing antifolate resistance. Genetic or pharmacological inhibition of ATIC greatly reduced tumor burden and extended survival in mice. Finally, folate removal from the microenvironment sensitized DIPGs to the antifolate methotrexate. Our studies identify ATIC as a novel target in DIPG and reveal a potential treatment strategy with antifolates in children with this lethal brain tumor.
DIPG-19. SURGICAL STATUS IN THE ERA OF NEW MOLECULAR DIAGNOSTICS: INSIGHTS FROM A SINGLE-CENTER CASE SERIES REPORT IN CHINA
Authors: Li Juan , Ai Ruyu , Lai Mingyao , Cai Linbo
Published: June 2024

The therapeutic value of surgery for DIPG has been a subject of controversy. Surgical biopsy provides valuable information on molecular markers of brain gliomas, aiding in further personalized treatment. This retrospective study aimed to elucidate the real-world application of surgery in Diffuse Intrinsic Pontine Glioma (DIPG) in the era of new molecular diagnostics. This study from Guangdong Sanjiu Brain Hospital investigates the surgical landscape in the treatment of Diffuse Intrinsic Pontine Glioma (DIPG) amid the new molecular diagnostics era (Jan 2017 to Feb 2023). Among 184 cases, 58.6% underwent surgery. Notably, 97.2% were introduced to surgery, with 65.8% opting for open surgeries and 34.2% utilizing the Rosa robot. Patients aged 18 and above showed a higher surgery rate (78.3%) than those below 18 (49.1%), revealing a significant difference (P<0.05). The median time from surgery to radiotherapy varied between open (19 days) and stereotactic biopsy (7 days). Complications included increased cranial nerve dysfunction in 20 open biopsy cases. All 108 patients received pathological diagnoses, with 79 having diffuse midline gliomas. Targeted therapy based on second-generation sequencing was administered to seven patients, involving drugs like ribociclib and rigosertib. The study concludes that compared to a decade ago, there is an increased willingness among oncologists to perform brainstem surgery, especially in older patients. Stereotactic biopsy, with fewer side effects, allows swift initiation of radiotherapy. Obtaining lesion tissue during surgery holds promise for new treatments and future research.
DIPG-20. SURGICAL INTERVENTION’S CORRELATION WITH SUBSEQUENT METASTASIS DEVELOPMENT IN CHILDREN WITH DIFFUSE INTRINSIC PONTINE GLIOMAS (DIPG)
Authors: BASHIR SHOAIB , Zhou Jiangfen , Lai Mingyao , Cai Linbo
Published: June 2024

BACKGROUND

Diffuse intrinsic pontine glioma (DIPG) is a highly aggressive pediatric brain tumor with a poor prognosis. Currently, there is no curative treatment option available for these patients. These patients often undergo surgery to confirm the diagnosis pathologically. However, the relationship between surgery and dissemination is to be confirmed. This retrospective cohort determines if surgical intervention increases the risk of dissemination and what other factors may also be associated.

METHODS

The final analysis included 142 pediatric (age ≤ 12) DIPG patients treated at Sanjiu Brain Hospital between January 2017 and June 2023. The data were retrospectively collected following the initial diagnosis. The diagnosis was established based on imaging studies. Three radiologists analyzed the imaging data. The endpoint was the occurrence of subsequent metastasis. Univariable and multivariable logistic regressions were performed to identify predictors of subsequent dissemination. All statistical assessments were two-tailed; only P < 0.05 was considered statistically significant.

RESULTS

Of 142 patients, 28 (19.7%) developed metastasis, and 114 (80.3%) had no evidence of metastasis. There were 77 (54.2%) males and 65 (45.8%) females. The patients’ median age was 7 years (1-12 yrs.). Patients were followed up for at least 6 months or until they developed subsequent metastasis or died without developing metastasis. Univariate logistic regression analysis demonstrated that surgery (OR 4.277, 95%CI 1.753 - 11.607, P=0.002), adjuvant temozolomide (OR 2.891, 95%CI 1.097 - 9.106, P=0.045), and adjuvant bevacizumab (OR 10.278, 95%CI 3.759 - 9.444, P=0.003) were the potential predictor of metastasis development. Multivariate logistic regression analysis showed that surgery (OR 5.061, 95%CI 1.966 - 14.508, P=0.001) and administration of adjuvant bevacizumab (OR 3.020, 95%CI 1.196 - 8.065, P=0.022) were the independent predictors of dissemination.

CONCLUSION

Surgery and adjuvant bevacizumab may increase the risk of subsequent metastasis in the pediatric DIPG population.
DIPG-21. PRECLINICAL ASSESSMENT OF A MULTIMODAL TREATMENT APPROACH WITH GIVINOSTAT, PAXALISIB, AND RADIOTHERAPY FOR DIFFUSE MIDLINE GLIOMA (DMG)
Authors: du Chatinier Aimée , Meel Michaël H , Waranecki Piotr , Metselaar Dennis S , Hulleman Esther
Published: June 2024

BACKGROUND

Despite an extensive body of clinical research conducted, the prognosis for patients with Diffuse Midline Glioma (DMG) has remained stagnant for the past five decades. Years of investigations underscore the robust resistance of DMG tumors to treatment, rendering monotherapy an improbable route to a cure. Recognizing the need for a multimodal approach, this study evaluated a treatment strategy incorporating radiotherapy and two brain-penetrable compounds with potential immunomodulatory effects, Givinostat and Paxalisib.

METHODS

We selected Givinostat, a pan-HDAC inhibitor undergoing clinical studies in children with Duchenne muscular dystrophy, by comprehensive screening of a panel of HDAC inhibitors for their efficacy in DMG neurosphere cultures, ability to penetrate the blood-brain barrier, and clinical utility. in addition, we selected the PI3K/mTOR inhibitor Paxalisib based on a phase II clinical trial currently undergoing in DMG patients (PNOC DMG Adaptive Combination Trial, PNOC022). The anti-tumor, radio-sensitizing, and immunomodulatory effects of both compounds were assessed in vitro and in immunocompetent mice, using various cytotoxicity and proteomic assays and (single-cell) sequencing techniques.

RESULTS

Our investigations revealed that the combination of Givinostat and Paxalisib exerts a profound cytostatic and cytotoxic effect on DMG cells, by reducing cell viability, enhancing DNA damage, and inducing apoptosis. Moreover, this combination therapy sensitized DMG cells to ionizing radiation, potentially by diminishing DNA repair mechanisms and elevating oxidative stress. Additionally, the combination of Givinostat and Paxalisib considerably reduced tumor growth in vivo and transformed the immune-cold microenvironment of DMG tumors into a pro-inflammatory phenotype.

CONCLUSIONS

This study positions Givinostat as a potential addition to the backbone of Paxalisib in forthcoming clinical trials, either as a radio/chemotherapy combination or as a foundation for immunotherapy.
DIPG-25. RELATIONSHIP BETWEEN MOLECULAR CLASSIFICATION, ORIGIN AND PROGNOSIS IN DIFFUSE MALIGNANT GLIOMA IN CHILDREN
Authors: Kagawa Naoki , Nakagawa Tomoyoshi , Kuroda Hideki , Tachi Tetsuro , Hirayama Ryuichi , Kijima Noriyuki , Okita Yoshiko , Kishima Haruhiko
Published: June 2024

INTRODUCTION

Diffuse malignant gliomas that occurs in childhood is recognized as a biologically different disease from that in adults. Furthermore, with the recent development of molecular diagnostics, diffuse midline glioma (DMG), which occurs mainly in the thalamus and brainstem, frequently occurs in childhood and have poor prognosis. We report the relationship between imaging findings, origin, and prognosis of diffuse malignant glioma in children or young adult based on our case series.

PATIENTS AND METHODS

43 cases of malignant glioma that occurred from childhood to young adult (below 30 years old) and were treated at our hospital since 2004 were registered in the analysis. Of the 42 cases, 24 cases were in brain stem, 11 cases in cerebral hemisphere, 6 cases in thalamus, and 1 case in cerebellar hemisphere. Histological diagnosis was possible in 31 cases, but without histological verification in 11 brain stem lesions. Histological diagnosis included 13 high-grade glioma (GBM, including NOS and G34R mutations), 11 DMG, and 7 anaplastic astrocytoma.

RESULTS

Of 13 cases of hemispheric high-grade gliomas, 2 cases had long-term survival (both cases received WT1 peptide vaccination), and 1 case survived for more than 5 years (total resection, MGMT methylation). Among the DMGs, 7 cases were found in brain stem (5 pons, 2 pontomedullary junction, 1 cerebellar peduncle, and 1 midbrain), 3 cases in the thalamus. Two cases of onset at the pontomedullary junction were found incidentally, and the symptoms gradually increased during follow-up, and multidisciplinary treatment was performed. Bilateral thalamus DMG with HIST1H3B mutation showed slowly growing and invasion.

There were cases of pediatric hemisphere glioblastoma with long-term survival. In addition, the origin and imaging findings of DMG were various, and tumors that originate at the pontomedullary junction may be relatively favorable.
DIPG-27. MATCHED PRIMARY-RELAPSE PEDHGG PATIENT-DERIVED XENOGRAFT MODELS TO IDENTIFY THERAPY RESISTANCE PROFILES
Authors: Barnerjee Srijita , Zhao Miao , Rosen Gabriela , Olsen Thale , Michno Wojciech , Swartling Fredrik J , Holmberg Karl O , Giraud Géraldine
Published: June 2024

BACKGROUND

We established patient-derived xenografts (PDXs) from matched primary-relapse pairs of pediatric high-grade gliomas (pedHGGs) in collaboration with the Swedish Childhood Tumor Biobank. Further, we used human iPSC derived cortical organoids to evaluate interactions between glial and tumor cells under treatment pressure potentially lost in xenograft models. Using this setup, we hope to follow tumor relapse evolution and to identify novel mechanisms behind treatment evasion and relapse formation.

METHODS

Tumoral singel cell suspensions from matched primary-relapse pairs are injected orthotopically in immunodeficient NOG mice. We establish in vitro 2D models in stem cell conditions from the first generation of animals transplanted. Lentivirus-based labelling with GFP/Luciferase was done on PDX-derived cell lines to allow cell tracking and to quantify efficacy of drug screening efforts. Co-culture of these cells with normal cells in 3D assembloids were performed using 150-250 days old cortical organoids. Single-cell RNA sequencing and bulk RNA sequencing (RNA-seq) on patient biopsies, their corresponding PDXs from first generation of mice, and PDX-derived cell lines, was performed to track potential changes in vivo and in vitro and to further validate primary-relapse specific evolution.

RESULTS

2D and 3D cell cultures offered efficient labeling and we quantified differences in sensitivity to standard therapy between primary and relapse samples. PDX models and assembloids highlighted pedHGG growth patterns, modes of invasion and cell interactions, especially astrocyte-tumor cell interactions. These 3D models also allow for further validation of targeted drug testing based on multiomics data (WGS, methylation and RNA-seq) from the patient biopsies.

CONCLUSION

PedHGG primary-relapse in vitro 2D cell line/ in vivo 3D assembloid models together with PDXs is a robust approach to study different aspects of relapse-specific tumor biology. Our results offer insights aiding the development of novel therapeutics inhibiting relapse mechanisms already in the first line of treatment.
DIPG-33. COMBINING CLONAL TRACKING WITH SINGLE CELL RNA SEQUENCING REVEALS TEMPORAL SHIFTS IN CELL STATES ASSOCIATED WITH TRAMETINIB RESISTANCE IN MAPK ALTERED DIFFUSE MIDLINE GLIOMA
Authors: Das Molina , Grabovska Yura , Collins Jared , Rogers Rebecca , Burford Anna , Mackay Alan , Beroukhim Rameen , Bandopadhayay Pratiti , Jones Chris
Published: June 2024

BACKGROUND

H3K27-altered diffuse midline glioma (DMG) is an aggressive and incurable paediatric high-grade glioma with median overall survival of 9-12 months. A distinct subset of DMG harbour alterations in the MAPK pathway, and we have previously shown that DMG-MAPK may exhibit sensitivity to MEK inhibitors but develop resistance under long-term exposure to drug. Elucidating these resistance mechanisms could spearhead better treatment strategies for these patients.

METHODS

To study clonal evolution under trametinib, we used CloneSifter lineage tracking technology to uniquely barcode 300,000 DMG-MAPK cells which were then treated with a GI80 dose of trametinib for 11 weeks. We extracted DNA to quantify barcode enrichment at endpoint across six technical replicate experiments and leveraged CloneSifter’s compatibility with single cell RNA sequencing (scRNA-seq) to couple clonal identity with transcriptional states in ~90,000 trametinib treated cells at varying timepoints.

RESULTS

DNA sequencing revealed 0.5% (1500/300,000) of the initial barcodes were enriched across trametinib replicates and were also detectable by scRNA-seq. Conducting cNMF analysis on the single cell dataset showed populations of trametinib treated cells associated with each other in a temporal manner, whereby early and late-stage resistant populations were defined by neuronal and stem cell like markers, respectively. A time-dependent enrichment of CloneSifter barcodes was also observed, suggesting dynamic expansion of clones over the treatment window. Notably, week 11 trametinib treated cells showed higher expression of resistance driver genes related to EMT, stemness and p38/MAPK upregulation.

CONCLUSIONS

Utilizing the sequence-specific reporter construct built in to the CloneSifter system allows for retrieval of resistant clones marked by enriched barcodes under treatment prior to drug exposure, enabling further characterization of monoclonal populations of trametinib-resistant cells. In summary, we have employed novel tools to study resistance development in DMG-MAPK that could be exploited to devise rational treatment strategies for these patients in an evolutionary-guided manner.
DIPG-35. OVERCOMING THE BLOOD-BRAIN BARRIER CHALLENGE IN DIFFUSE MIDLINE GLIOMA
Authors: Tsoli Maria , Ung Caitlin , Upton Dannielle H , Venkat Pooja , Salerno Antonietta , Pandher Ruby , Mayoh Chelsea , Vittorio Orazio , Ziegler David S
Published: June 2024

Diffuse midline gliomas (DMGs) are aggressive paediatric brainstem tumours with no known cure. The development of effective treatments is greatly impeded by the failure of most anti-tumour agents to penetrate the blood-brain barrier (BBB). Our research, along with that of others, has revealed that the BBB remains intact in DMG. We employed DMG orthotopic models to explore potential region-specific variations in response to the therapeutic agent temsirolimus as well as confocal microscopy and single-cell RNA sequencing (scRNAseq) to elucidate the impact of DMGs on brain endothelial morphology and pathways governing BBB integrity. We first assessed the effectiveness of the mTOR inhibitor temsirolimus when administered in cortex-injected DMG models compared to brainstem-injected DMG. We found significantly increased survival rates, elevated temsirolimus levels, and reduced tumour cell proliferation in the cortex versus the brainstem. Confocal imaging analysis indicated no structural differences between DMG and Matrigel-injected animals. However, scRNAseq unveiled distinct transcriptomic changes: brainstem-injected DMG exhibited downregulation of genes related to inflammatory and apoptotic pathways, whereas cortex-injected DMG showed downregulation of interferon pathways. To overcome the tightened barrier in the brainstem, we explored three MCL1 inhibitors and SNGR-TNFa (targeting CD13) as potential blood-brain barrier modulators. These inhibitors significantly reduced transendothelial electrical resistance, increased tracer dye leakage in an in vitro DMG BBB model, and decreased the protein expression of claudin-5. Moreover, in vivo, we found that administration of a single dose of either SNGR-TNFa or the MCL1 inhibitor S63845 improved penetration of Texas-Red (3KDa) in DMG-engrafted animals, indicating the effective opening of the BBB. In summary, our study revealed that DMG directly affects BBB pathways in endothelial cells, tightening the barrier and reducing treatment efficacy. MCL1 inhibitors and SNGR-TNFa show promise in modulating the BBB and have the potential to enhance therapeutic activity.
DIPG-37. AUGMENTING RADIATION SENSITIVITY IN DMG/DIPG BY TARGETING TP53 DEPENDENCY AND INHIBITING DNA PROTEIN KINASE PATHWAY
Authors: Ciani Ludovica , Fernandez Solène , Grotzer Micheal , Müller Sabine , Nazarian Javad
Published: June 2024

BACKGROUND

Diffuse midlines gliomas (DMGs) are lethal pediatric brain tumors, with an overall survival rate of less than 12 months. Radiotherapy treatment (RT) only provides a transient clinical response, with an average survival increase of about 3 months. At the cell level it induces DNA damages, leading to tumor cell death. Although, DMGs are tumors known to be resistant to any therapy, as the tumor recurs most often within a few months post radiation. Peposertib, a DNA Protein Kinase (DNA-PK) inhibitor, is a potent radiosentitizer that crosses the blood brain barrier and has shown RT-combination efficacy in other cancers by enhancing DNA double-strand breaks (DSB) inducing agents including ionising radiation. Methods and

RESULTS

DNA-PK plays a key role in one of the main pathways involved in the repair of DNA double-strand breaks (DSB), allowing tumor cells to repair their DNA and continue proliferation. To further investigate Peposertib and its potential for clinical intervention, we explored Peposertib’s radiosensitizing activity on DMG preclinical in vitro models. In vitro viability assays with RT alone and in combination with Peposertib, were carried out using DMG cell lines carrying either the wild-type or mutated version of TP53. To further validate our results, immunofluorescence assays post-combination treatment were performed to assess DDR pathway disruption by studying the number of foci per nucleus of DNA damage markers yH2AX and P53BP1.

RESULTS

Remarkably, combinational treatment of Peposertib with RT resulted in a significant decrease in human primary DMG cells viability. Interestingly, Peposertib was effective in increasing RT sensitivity in TP53WT DMGs. However, the same good response was not observed in the DMG cell lines carrying TP53MUT, indicating a possible correlation between TP53 mutational status and response to therapy. Additionally, we demonstrated that Peposertib induces cell death. This was observed and determined through quantification analysis of DNA damage markers yH2AX and P53BP1 therefore to DDR pathway disruption.

CONCLUSIONS

Our current findings underscore Peposertib’s potential as a targeted therapeutic agent against DMGs, offering a better understanding of what drives resistance to radiotherapy. By targeting DDR pathway key factors we will increase sensitivity to radiation in vitro and in vivo in DMGs.
DIPG-39. ONCOHISTONE H3.3K27M-DRIVEN CREB5/ID1 AXIS DICTATES THE MALIGNANT CELL STATES OF DIFFUSE INTRINSIC PONTINE GLIOMA (DIPG)
Authors: Zhou Wei , Xu Cheng , Tang Yujie , Pan Changcun , Zhang Liwei , Xi Qiaoran
Published: June 2024

BACKGROUND

Diffuse intrinsic pontine glioma (DIPG) is a highly aggressive pediatric cancer characterized by the presence of H3K27M-oncohistones, which drive relentless proliferation and differentiation arrest. This study focuses on the H3.3K27M DIPG subtype and investigates the role of the oncohistone-driven CREB5/ID1 axis in promoting malignancy.

METHODS

We employed a variety of biological in vitro assays and xenograft mouse models utilizing several H3.3K27M and H3.1K27M DIPG cell lines to evaluate the impact of the studied factors on the growth of DIPGs. Additionally, we employed methodologies in molecular biology, bioinformatics, drug screening, and biochemistry to elucidate the underlying mechanisms of tumorigenesis.

RESULTS

Through inhibition of ID1, this study explores the induction of cellular differentiation and apoptosis in H3.3K27M DIPG, shedding light on the oncogenic mechanisms at play. The intricate regulation of ID1, linked to CREB5—an oncogenic contributor independent of BMP signaling—is elucidated. The involvement of the SWI/SNF complex as a co-regulator of CREB5 is also examined. Moreover, our findings reveal that H3.3K27M-oncohistone triggers CREB5 overexpression, facilitating the formation of a super-enhancer promoter loop at the CREB5 locus. Disruption of this loop demonstrates potent anti-tumor efficacy in treating DIPG. Additionally, the combination of a compound targeting the CREB5/ID1 axis and a BRG1 inhibitor shows promise for DIPG treatment.

CONCLUSIONS

This research unveils key molecular pathways specific to the H3.3K27M DIPG subtype, offering valuable insights for potential therapeutic interventions against this lethal pediatric cancer. Inhibiting the CREB5/ID1 axis represents a promising strategy for inducing differentiation and apoptosis in DIPG cells, highlighting the potential for targeted therapies in combating this disease.
DIPG-44. DIFFUSE MIDLINE GLIOMA, H3K27-ALTERED: A COMPREHENSIVE CLINICAL AND BIOMOLECULAR LANDSCAPE OF THE NON-PONTINE TUMORS
Authors: Mariet Cassandra , Tauziède-Espariat Arnault , Auffret Lucie , Ajlil Yassine , Roux Alexandre , Pallud Johan , Palfi Stéphane , Métais Alice , Beccaria Kevin , Blauwblomme Thomas , Puget Stéphanie , Sanson Marc , Peloux Marie , Richard Juliette , Thalhammer Julian , Carvalho Alice , Puiseux Chloé , Pereira Victor , de Carli Emilie , Lesieur Thomas , Rouger Jeremie , Page Philipe , Doz François , Bourdeaut Franck , Dufour Christelle , Abou Samuel , Guerrini-Rousseau Léa , Dhermain Frédéric , Bolle Stéphanie , Varlet Pascale , Grill Jacques , Castel David
Published: June 2024

BACKGROUND

Since 2016, the WHO classification of CNS tumors groups all midline gliomas with H3K27me3 loss under the term diffuse midline glioma (DMG), H3K27-altered. Molecular characteristics enable their sub-classification based on driver mutations, co-driver mutations and epigenetic alterations. Predominantly studied in the pons (Diffuse Intrinsic Pontine Glioma, DIPG), where these are mainly pediatric tumors with a median overall survival (OS) of one year, fewer studies have been conducted on non-pontine DMG (npDMG), prompting this study to clinically and biologically characterize these gliomas and compare them with DIPG.

METHODS

Retrospective monocentric study from 1994 to 2022, including pediatric and adult patients diagnosed with npDMG, H327-altered with neuropathological confirmation at Sainte Anne Hospital in Paris, France. Comparative literature review of cases of npDMG and series of DIPG.

RESULTS

108 patients were included (54.7% male, median age 13.05-years [2.4-59.3]). The most frequent location was monothalamic (62.9%), followed by bithalamic, spinal, cerebellar, bulbo-medullar and other. H3K27M mutation was found in 85.2% of cases, EZHIP overexpression in 14.8% and EGFR alteration (mutation and/or amplification) in 16.7%. EZHIP-overexpressing patients were younger than H3K27M-mutated patients, as were EGFR-altered patients compared to those without EGFR alteration. Along with data from the literature, npDMG are older at diagnosis and have a slightly longer median OS (15 months [0.1-247.2]) than DIPG. Positive prognosis factors upon univariate analysis included longer time from symptom onset until diagnosis, treatment with ONC201, re-irradiation upon relapse, microcalcifications, proliferative index >10%, MAP kinase alteration and absence of TP53 mutation and EGFR amplification. However, surgery quality did not impact OS.

CONCLUSIONS

npDMG is a heterogeneous group of gliomas, affecting all ages, which can develop in diverse locations, with an overall poor prognosis. Despite shared clinical, immuno-histochemical and molecular properties with DIPG, recent molecular biology advancements reveal distinct subtypes within DMG, with varying prognoses, challenging the unified classification.
DIPG-52. CEREBROSPINAL FLUID DIVERSION DOES NOT PROLONG OVERALL SURVIVAL IN CHILDREN WITH DIFFUSE INTRINSIC PONTINE GLIOMA (DIPG): A REPORT FROM THE INTERNATIONAL DIPG REGISTRY
Authors: Manoharan Neevika , Cooney Tab , DeWire-Schottmiller Mariko , Lane Adam , Nellan Anandani , Leary Sarah , Partap Sonia , Kilburn Lindsay , Wright Karen D , Hummel Trent , Ziegler David S , Fouladi Maryam , Hoffman Lindsey M , Warren Katherine E
Published: June 2024

BACKGROUND

The role of permanent cerebrospinal fluid (CSF) diversion in the management of patients with diffuse intrinsic pontine glioma (DIPG) is poorly elucidated. There are no standard practice guidelines regarding the role of these neurosurgical interventions in patients with DIPG or large studies regarding the impact of CSF diversion on overall survival (OS) or palliation in this disease.

METHODS

Data were extracted from subjects registered in the International DIPG registry (IDIPGR). Univariable analyses were performed using the Fisher’s exact test or Wilcoxon rank sum test. Survival was estimated using the Kaplan-Meier method.

RESULTS

Of n=1104 evaluable patients with DIPG registered in the IDIPGR, n=303 (27.4%) had permanent CSF diversion. Median time from diagnosis to CSF diversion was 1 month (range 0-4 months). There were no significant differences in age, gender or race in subjects with CSF diversion versus no CSF diversion. Patients with CSF diversion had a higher incidence of hydrocephalus at diagnosis (64.1% vs 11.3%, p<0.001). There was no significant difference in OS or post-progression survival (PPS) at 1,2 and 3 years between subjects with permanent CSF diversion compared to those without (p=0.5 and p=0.8, respectively). Median OS was 11 months in both. Median PPS was 7 months (no CSF diversion) and 6 months (CSF diversion). Patients reported less headache and vomiting after permanent CSF diversion compared to pre-diversion (p<0.0001), however steroid use was also significantly higher after CSF diversion (p<0.001).

CONCLUSIONS

In this, the largest reported international cohort of patients with DIPG with permanent CSF diversion, we show that permanent CSF diversion was not associated with an improvement in OS or PPS. CSF diversion improved headache and vomiting but was also associated with increased rates of steroid use. This highlights the need for consensus guidelines regarding the use of these diversion strategies in patients with DIPG.
DIPG-54. PREDICTING EARLY DEATH IN DIFFUSE MIDLINE GLIOMA
Authors: Moloney Sam , Carruthers Vickyanne , Davis Melissa J , Bhuva Dharmesh D , Mignone Cristina , Singh Susan , Gardiner Rebecca E , Valvi Santosh , Eisenstat David D , Hansford Jordan R
Published: June 2024

BACKGROUND

Diffuse midline glioma (DMG) is an invariably fatal diagnosis with a median survival of 9-11 months. While early death is uncommon, such cases do exist. Identification of children at risk of short-term survival can support clinicians to provide informed counselling to families regarding important treatment decisions.

METHODS

We identified patients with a radiological diagnosis of DMG at the Royal Children’s Hospital (Melbourne) and the Women’s and Children’s Hospital (Adelaide) between 2001-2023. Clinical, imaging, and histopathological data were abstracted and compared between short-term survivors (STSs) and longer survivors (LSs).

RESULTS

Among 89 patients included, 21 (24%) had an overall survival (OS) of < 6 months of which 7 (8%) had an OS < 3 months. 5 of these cases had dramatic early death with < 1 month since diagnosis. While neither the pattern nor presence of contrast enhancement were predictive of poor survival, STSs were found to have a larger volume of contrast enhancement on T1-weighted imaging compared to LSs (1.3 cm3 vs 0.2 cm3, p < 0.01). Evaluation of the presence and extent of intratumoural susceptibility signals (ITSS) on susceptibility weighted imaging (SWI) demonstrated higher grades of ITSS (> 5 foci) were strongly associated with poor survival (p = 0.01, median OS = 5.5 months). Furthermore, 50% (6/12) of STSs had ≥ 11 dot-like or fine linear ITSSs in a maximum tumour cross section. From a treatment standpoint, as expected, patients who did not receive upfront radiotherapy experienced significantly worse OS (3.4 months vs 10.4 months, p < 0.01). Notably, a number of STSs did not receive palliative therapy in the context of their rapid and unforeseen disease progression.

CONCLUSIONS

The preliminary findings from this multi-centre study provide the foundation for future exploratory analysis alongside international collaborators at the DMG/DIPG registry to improve prognostication for patients with high-risk features of early death.
DIPG-57. INITIAL AND EARLY METASTATIC SPREAD IN DIPG AND DIFFUSE MIDLINE GLIOMA, H3 K27-ALTERED - REPORT FROM THE EUROPEAN SOCIETY FOR PEDIATRIC ONCOLOGY (SIOPE) AND INTERNATIONAL DIPG/DMG REGISTRIES
Authors: Perwein Thomas , Gielen Gerrit H , Karremann Michael , Bison Brigitte , Damen-Korbijn Carin M , Baugh Joshua N , Hummel Trent R , Blank Courtney , Biassoni Veronica , Bellón Marina Caballero , Szychot Elwira , Papusha Lyudmila , Jones David T W , Sturm Dominik , Hoffmann Marion , Kullmann Silke , Steffen Laura S , Simon Thorsten , Friker Lea L , Pietsch Torsten , Seidel Clemens , Eyrich Matthias , El-Ayadi Moatasem , Nussbaumer Gunther , Massimino Maura , La Madrid Andres Morales , Bailey Simon , von Bueren André O , Benesch Martin , Veldhuijzen van Zanten Sophie E M , Hoffman Lindsey , Kramm Christof M
Published: June 2024

BACKGROUND

We aimed to assess frequency, patterns and potential prognostic impact of initial and early dissemination in pediatric patients with diffuse intrinsic pontine glioma (DIPG) and diffuse midline glioma, H3K27-altered (DMG).

METHODS

199 patients (age, 0-18 years) with DIPG/DMG and positive CSF cytology and/or metastases on MRI at diagnosis (Types 1-3) were identified within the SIOPE and International DIPG/DMG Registries. Data on metastatic first progression (PD) in DIPG/DMG (Type 4) were analyzed from the HIT-HGG-2007/2013 trials.

RESULTS

Dissemination at diagnosis was present in 5% of patients with DIPG/DMG (16.9% in non-pontine DMG). For all 199 patients with initially disseminated pontine (n=166) and non-pontine DMG (n=33), median overall survival (OS) was 10±0.5 months and progression-free survival (PFS) was 6.6±0.2 months. Tumor cells in the CSF without metastases on initial MRI (Type 1) were detected in 12 patients. Four patients had no primary tumor but primary extensive leptomeningeal dissemination resembling diffuse leptomeningeal glioneuronal tumor, molecularly confirmed as DMG, with aggressive course (Type 2). 183 DIPG/DMG patients were identified with primary tumor and metastases on MRI at diagnosis (Type 3). OS and PFS was superior in patients receiving additional treatment to radiotherapy (n=124), without significant differences between focal and craniospinal irradiation. Metastatic spread at first progression (Type 4) occurred in 32.1% (65.3% in non-pontine DMG) of 224 HIT-HGG patients with DMG and recorded PD. OS of patients with metastatic PD was inferior to local PD (median, 10.8±0.9 vs. 13.3±0.9 months, p=0.005), as was post-progression survival (median, 3.1±0.4 vs. 5.2±0.5 months, p=0.003).

CONCLUSIONS

We observed a propensity for early dissemination in (especially non-pontine) DMG. While additional treatment beyond radiotherapy seems beneficial, preventing dissemination at progression (e.g., by upfront CSI) may help to prolong survival of patients with DMG and possibly provide options for further therapy.
DIPG-60. OPERATIVE PERFORMANCE OF RAPNO DIPG CRITERIA FOR IMAGING REVIEW- A STUDY FROM THE INTERNATIONAL DIPG/DMG REGISTRY (IDIPGR)
Authors: Gorsi Hamza S , Guryildirim Melike , Kuwabara Michael , Dhatt Jovan , Hoffman Lindsey M , Cohen Kenneth J
Published: June 2024

BACKGROUND

The Response Assessment in Pediatric Neuro-Oncology (RAPNO) working group for diffuse intrinsic pontine glioma (DIPG) recently published its recommendations. Response criteria incorporate radiologic findings, neurologic status, and use of concurrent steroids and antiangiogenic drugs to define response. To test the operative performance of the imaging component, we retrospectively applied RAPNO DIPG recommendations to a convenience sample of 46 patients enrolled in the International DIPG/DMG Registry(IDIPGR).

METHODS

MRIs were independently reviewed by 2 pediatric neuro-radiologists. These measurements were analyzed utilizing RAPNO DIPG imaging criteria to categorize the response as complete response (CR), partial response (PR), stable disease (SD), or progressive disease (PD). For PR evaluation, each MRI was compared to the baseline MRI, and for PD evaluation, each MRI was compared to the MRI with the smallest cross-sectional measurement. The response category for each MRI was compared between 2 neuro-radiologists and was classified as concordant if they agreed, minor discordant if one reported SD and the other reported PR or PD, and major discordant if one reported PR and the other reported PD.

RESULTS

A total of 276 paired MRIs were analyzed. Sixty-seven percent of the time, the analyzed response assessment was concordant. In 33% discordant reads, 26% were minor discordant, and 7% were major discordant. There were no CRs reported. Further analysis of minor discordant cases showed that 62% of these cases were within 10% of the boundary for PR or PD. The median difference between 2 neuro-radiologists’ long axis measurements was 2 mm (range 0-27mm) and short axis measurements was 2mm (range 0-29mm). These small differences in measurements account for all discordant response categories between 2 neuro-oncologists.

CONCLUSION

This study demonstrated that RAPNO DIPG imaging criteria can be successfully applied with high concordance between readers. When discordant, measurements were largely at the boundaries of response type.
DIPG-71. XPEDITE: A STUDY TO EXPEDITE DIPG AND DMG RESEARCH
Authors: Stuhlmiller Timothy J , Ward Lisa , Lewis Sarah , Cole Zachariah D , Chuyka Donald , Ginn Sarah , Federowicz Bryan , Shapiro Mark A
Published: June 2024

BACKGROUND

Diffuse midline glioma (DMG), particularly the subtype known as diffuse intrinsic pontine glioma (DIPG), poses a significant challenge due to its aggressive nature and low survival rates, especially in children. Despite ongoing research and clinical trials, the survival rate remains alarmingly low. The scarcity of cases and siloed nature of current registry efforts have hindered the establishment of a reliable natural disease history for benchmarking studies and led to the need for randomization in clinical trials.

METHODS

xPedite (NCT06140719) is a fully virtual, patient-driven, nationwide, real-time observational study to collect, annotate, standardize, and report the critical data elements of DMG and DIPG in a regulatory-compliant framework. Patients of any age can join via electronic consent, with recruitment supported by provider referrals and patient advocacy organizations DDRFA and My DIPG Navigator. Medical records are accessed from institutions, sequencing vendors, and health information networks. Notably, data elements are annotated comprehensively and longitudinally in a 21 CFR Part 11-compliant clinical database by xCures remote study staff, eliminating the need for data entry by treating site staff or physicians. Data collected include anti-cancer interventions, non-cancer medications, biomarker results, radiological endpoints, steroid use, vitals, demographics, raw DICOM images, and genomics. In addition to clinical data, patient- and caregiver-reported outcome questionnaires (PROs) and video assessments of mobility and neurological status are collected for clinician review. The objective of the study is to collect these clinical, imaging, biomarker, and assessment data in a manner suitable to provide to health authorities to reduce randomization in future trials. In a commitment to scientific understanding and collaborative research, the data collected through the xPedite study is openly available to academic, government, and non-profit researchers. As of January 2024, 182 patients were enrolled in xPedite. Real-time enrollment and data statistics are available at https://xcures.com/dmgdashboard/.
DIPG-83. RBPJ-SIX1 AS A TUMOR SUPPRESSOR AXIS IN DIFFUSE INTRINSIC PONTINE GLIOMA
Authors: Racanelli Anna , Shen Chen , Gadd Samantha L , Chizhik Mili , Setara Ruby , Ge Yi , Schüpper Alexander J , Puigdelloses Montserrat , Hambardzumyan Dolores , Tsankova Nadejda M , Becher Oren J
Published: June 2024

BACKGROUND

Diffuse intrinsic pontine glioma (DIPG) is a rare, incurable pediatric brain tumor that arises in the brainstem. Despite decades of clinical trials, no treatments have proven effective at prolonging survival for children with this disease. A deeper insight into the epigenetic and genetic changes driving DIPG are imperative for identifying therapeutic targets. A defining feature of DIPG is the histone H3 lysine to methionine mutation at residue 27 (H3K27M), which prevents H3K27 trimethylation. The H3.1K27M mutation, specifically, co-segregates with mutations in the Activin A receptor ACVR1. Tumor cells with both mutations have significantly higher levels of the transcripts and proteins that inhibit proneural gene expression in the NOTCH signaling pathway.

METHODS

We used the replication-competent avian sarcoma-leukosis virus long terminal repeat with splice acceptor/tumor virus A (RCAS/Tv-a) system to evaluate the role of various NOTCH signaling proteins in a genetically engineered mouse model of DIPG. We also performed RNA sequencing of specific NOTCH signaling protein knockout DIPG models.

RESULTS

Knocking out Hairy of Enhancer Split-1 (Hes-1) in combination with H3K27M and ACVR1 mutations did not significantly reduce survival compared to control. However, knocking out Recombination Signal Binding Protein for Immunoglobulin kappa J region (RBPjk)—a transcriptional regulator upstream of Hes-1—did significantly reduce survival compared to wild-type RBPjk expression. RNA sequencing of RBPjk knockout tumors revealed that Sine Oculis-related Homeobox 1 (Six1) was the most top most differentially expressed gene. To assess whether Six1 downregulation drives DIPG tumorigenesis in RBPjk knockout tumors, we overexpressed Six1 in a murine model of DIPG with H3.3K27M and p53 loss. Six1 overexpression did not significantly prolong survival or decrease tumor grade but trended toward significance. However, Six1 overexpression significantly increased tumor cell proliferation.

CONCLUSIONS

These results indicate that Six1 could function as a potential tumor suppressor in DIPG, necessitating further exploration.
DIPG-86. INVESTIGATING THE TUMOR ENVIRONMENT AND SIGNALING DEPENDENCIES DRIVING DIFFUSE MIDLINE GLIOMA TUMOR INITIATION, PROGRESSION, AND RESISTANCE: A COMPREHENSIVE APPROACH LEVERAGING BULK MULTIOMICS DECONVOLUTION AND SINGLE-CELL OMICS VALIDATION
Authors: Patel Palak , Ryall Scott , Kukreja Bharti , Turinski Andrei , Siddaway Robert , Brudno Michael , Ramani Arun , Pajovic Sanja , Kalish Brian , Hawkins Cynthia
Published: June 2024

BACKGROUND

Diffuse Midline Glioma (DMG) is a incurable tumor affecting children. Recent genomic investigations have identified a recurrent H3K27M mutation which induces global alterations in histone methylation patterns and DNA methylation. These epigenetic modifications hint at pivotal roles in DMG pathogenesis, yet effective therapeutic strategies remain elusive, with median survival rates stagnant at approximately one year. This shortfall stems from two main factors: 1) Inadequate multiomics studies hindering our understanding of DMG evolution and tumor progression, and 2) Limited comprehension of the tumor microenvironment in DMG.

METHODS

To elucidate DMG’s clonal evolution, we adopted a comprehensive multi-region sampling approach, acquiring 33 specimens from seven patients. Whole-exome and transcriptome sequencing, alongside DNA methylation profiling, were performed. Additionally, we generated one of the most extensive single-cell multiomics datasets (>300,000 cells from 21 additional samples), with a focus on pre- and post-treatment, and dissemination effects in DMG.

RESULTS

Our analysis delineated a tumor-promoting microenvironment characterized by hypoxia and pro-inflammatory conditions, nurturing genomic alterations and specialized biological processes such as proliferation and epithelial-mesenchymal transition. Notably, a majority of immune cells exhibited M1-like polarization, bolstering pro-inflammatory programs within the tumor milieu. Subclones within the DMG cohort demonstrated dissemination potential, often exhibiting up-regulation of NOTCH, P53, and WNT beta-catenin signaling pathways. These findings suggest that DMG clones harboring dissemination capabilities outside the pons acquire additional phenotypic features, possibly mediated by epigenetic or transcriptional alterations, contributing to enhanced migratory and aggressive behaviors.

CONCLUSIONS

Our study unveils the parallel evolution of DMG at genetic, epigenetic, and transcriptional levels, unveiling novel subclonal phenotypes governing tumor behavior. Crucially, we identify critical environmental shifts such as hypoxia and inflammatory changes, coupled with specialized signaling programs, driving dissemination and resistance phenotypes. These insights may pave the way for generating accurate genetically mouse models and targeted therapeutic interventions to combat this challenging malignancy.
DIPG-88. TARGETING THE ANTIOXIDANT RESPONSE ELEMENT AXIS SENSITIZES DIFFUSE MIDLINE GLIOMA CELLS TO ONC201
Authors: Jackson Evangeline R , Duchatel Ryan J , Fish Cameron J , Persson Mika L , Staudt Dilana E , Findlay Izac J , Dun Matthew D
Published: June 2024

BACKGROUND

Diffuse midline glioma (DMG) is a universally fatal CNS tumor, with a median overall survival of <12-months. Recent studies, including our own, have identified overexpression of the mitochondrial protease ClpP in DMG patient samples. ClpP plays a pivotal role in regulating energy production. Brain-penetrant ClpP agonists, including ONC201, ONC206, and TR107, induce non-specific degradation of the mitochondrial electron transport chain. However, there is considerable variability in the response of DMG cell lines to these therapies, with only about half reaching IC50. Notably, TP53-mutant cell lines exhibit reduced sensitivity.

METHODS/RESULTS

Further analysis using quantitative proteomics of TP53-mutant DMG cell lines exhibited hallmark activation of ClpP, increased mitochondrial stress and dysfunction. Importantly, ClpP agonism increased the activity of the transcription factor NRF2, driving expression of NQO1 (p=0.0017). NQO1 is a cytoprotective enzyme, promoting redox balance and cell survival. This highlights a potential adaptive response to ClpP agonist therapy in DMG cells. Analysis of DMG patient samples through publicly available RNA sequencing data identified ubiquitously high NRF2 expression compared to other pediatric CNS tumors. This led us to hypothesize that a clinically relevant, selective NQO1 inhibitor could elicit a strong response to ClpP agonism. Therefore, we examined ARQ761, a CNS-active therapy that disrupts NQO1 leading to the formation of an unstable hydroquinone, causing oxidative DNA damage and apoptosis. When ARQ761 was used concurrently with a ClpP agonist, the combination showed additive effects, average Bliss Score of 9.91 (n=4). Interestingly, administering ARQ761 24 h after ClpP agonist exposure combined synergistically, average Bliss Score of 15.87 (n=4). This synergy was further potentiated using ARQ761 48 h after ClpP agonism, when NQO1 expression peaked, resulting in an average Bliss Score of 20.30 (n=4).

CONCLUSIONS

This study has unveiled an intrinsic antioxidant defense mechanism in DMG, which facilitates evasion from treatments inducing oxidative stress. This defense mechanism appears particularly robust in cell lines harboring TP53-mutations, which are also known to exhibit resistance to radiotherapy.