bluuebunny/crossref_metadata_2025_split huggingface dataset:
IMMU-13. PRECLINICAL EFFICACY OF MULTI-ANTIGEN (HER2/EGFR806/B7-H3/IL13RA2) TARGETING QUAD-CAR T CELLS AGAINST DIFFUSE MIDLINE GLIOMA
Authors: Myers Carrie , Timpanaro Andrea , Song Edward , Meechan Michael , Chen Alex , Biery Matthew , Reid Aquene , Gwiazda Kamila , Baldwin Michael , Gustafson Joshua , Gardner Rebecca , Jensen Michael , Vitanza Nicholas 
Published: June 2023

Abstract

BACKGROUND
Diffuse midline glioma (DMG), including diffuse intrinsic pontine glioma (DIPG), remains a fatal disease. We have published the preclinical efficacy of HER2, EGFR806, and B7-H3 CAR-T cells and opened 3 trials (BrainChild-01, -02, and -03, respectively) that have delivered &gt;350 intracranial CAR T doses to &gt;70 enrolled children with recurrent/refractory CNS tumors and DIPG/DMG. While these trials have demonstrated the feasibility and tolerability of repeated, locoregional CAR-T cell dosing, responses have not been uniform in part due to tumor heterogeneity.

METHODS
Seattle Children’s Therapeutics generated multi-antigen (HER2, EGFR806, B7-H3, IL13ra2) quad-CAR T cells and we performed IND-enabling preclinical studies using CNS tumor models including DMG models (PBT-22, PBT-27, and PBT-29).

RESULTS
Quad-CAR T cells were generated by transducing T cells with a mixture of four vectors. The specificity of these CARs is conferred by scFv binding domains derived from the monoclonal antibodies Herceptin, mAb806, and MGA271 for HER2, EGFR806, and B7-H3, respectively. The IL13-zetakine, aligned with NCT00730613, carries an E13Y mutation in place of an scFv for specificity. In vitro, the quad-CAR T cells demonstrated significant in vitro cytolysis and cytokine (IL-2, IFNy, TNFa) production compared to mock (untransduced) CAR-Ts. In vivo, pilot studies in a GFP:ffLuc+Her2 U87 orthoptic xenograft mouse model showed significant survival (p&lt;0.001). To validate against DMG, we are conducting a larger study of single dose intracranial quad-CAR T cells in an orthotopic xenograft PBT-29 DMG mouse model. While ongoing, at day 19 post-treatment there is already a significant bioluminescence decrease (p&lt;0.001). In vivo studies will be completed and complimented by tumor spatial transcriptomics.

CONCLUSION
Quad-CAR T cell targeting of DMG is highly effective preclinically. A phase I trial of repeated, locoregional quad-CAR T cells for children with recurrent/refractory CNS tumors and DIPG/DMG is FDA-approved and will open in 2023.
IMMU-14. WAKING THE SLEEPING GIANT: TLR7/8 AGONIST TREATMENT STIMULATES TUMOR-ASSOCIATED MYELOID CELLS FOR MEDULLOBLASTOMA IMMUNOTHERAPY
Authors: Pauneto Coral Alicea , Hwang Duhyeong , Malawsky Daniel , Kim Kyoungtea , Park Christopher , Sokolsky Marina , Gershon Timothy 
Published: June 2023

Abstract
Medulloblastomas contain large myeloid populations and we have identified a novel therapy that effectively recruits these non-tumor cells for anti-cancer effect. Medulloblastoma myeloid cells have bivalent functions, supporting tumor growth through IGF1 secretion and immunosuppression, or inhibiting tumor growth through diverse mechanisms including phagocytosis. We investigated a nanoparticle formulation of the TLR7/8 agonist Resiquimod (POx-Resiquimod), administered to Gfap-Cre/SmoM2 (G-Smo) mice that are engineered to develop SHH medulloblastomas, using animal survival time studies, flow cytometry and single-cell trancriptomics (scRNA-seq). POx-Resiquimod treatment increased the survival of G-Smo mice as a single agent and sensitized the typically radiation-resistant G-Smo tumors to radiation therapy. Mechanistically, POx-resiquimod increased the proportions of myeloid cells in medulloblastoma, while blocking IGF1 secretion and re-polarizing the myeloid population from anti-inflammatory to pro-inflammatory. POx-resiquimod induced phagocytosis-related genes, including CD72 and the scavenger receptor MARCO. Functional studies confirmed that POx-resiquidmod increased tumor cell phagocytosis, linking molecular changes to changes in myeloid cell behavior. Our data show that the innate immune function of myeloid cells can be harnessed for medulloblastoma treatment and identify the nanoparticle-delivered TLR7/8 agonist POx-Resiquimod as a specific and effective new agent for medulloblastoma immunotherapy that can be combined with existing modalities.
IMMU-16. IDENTIFICATION OF TUMOR REJECTION ANTIGENS FOR PRECISION ADOPTIVE CELLULAR THERAPY FOR PEDIATRIC GLIOMAS
Authors: Trivedi Vrunda , Yang Changlin , Ogando-Rivas Elizabeth , Dyson Kyle , Mitchell Duane 
Published: June 2023

Abstract

BACKGROUND
Identifying tumor rejection antigens remains a major barrier to developing effective antigen-directed immunotherapeutics and their application to pediatric brain tumors. Recent progress in the field of cancer immunogenomics has facilitated the search for tumor-specific antigens by applying comprehensive cancer genomics to tumor antigen discovery. Using a custom antigen prediction pipeline, we performed in silico antigen prediction across a broad array of antigen classes including neoantigens, tumor-associated antigens (TAAs), and fusion proteins for human and murine medulloblastoma tumors. We generated antigen-specific T-cells and validated the antigen-mediated immune responses for the development of precision adoptive cellular therapy.

METHODS
We evaluated WES and RNA-seq data from 170 medulloblastoma patients available through ICGC and performed a comprehensive immunogenomic analysis of subgroup-specific transcription profiles to identify candidate antigen targets. Antigen-specific T-cells were generated from healthy donor PBMCs stimulated against commonly recurring neoantigens and TAAs and immune response was evaluated using IFNγ secretion. Furthermore, we performed tumor antigen discovery for preclinical medulloblastoma models Ptch1 (sonic hedgehog driven) and NSC (Group 3 MYC-driven) and validated the antigen-mediated immune responses of ex vivo expanded antigen-specific T-cells.

RESULTS
Antigen prediction analysis revealed most tumors expressed a higher proportion of immunogenic TAAs than neoantigens and fusion proteins. The neoantigens and TAAs evaluated in this study showed significantly higher IFN-γ secretion in an antigen-dependent manner for both the human and murine platform validation. We thus confirmed the capability to produce functional tumor antigen-specific T-cells that can be leveraged into adoptive cellular therapy for the treatment of medulloblastoma.

CONCLUSION
Medulloblastomas are immunologically heterogeneous, and identifying potential tumor rejection antigens is desirable to develop personalized immunotherapies for the patients. Using a custom antigen prediction and ex vivo antigen-specific T-cell expansion pipeline, we identified potential tumor rejection antigens with important implications for the development of antigen-directed cellular therapies.
LGG-10. TREATMENT-RELATED MYELODYSPLASTIC SYNDROME FOLLOWING CHEMOTHERAPY FOR PEDIATRIC LOW-GRADE GLIOMA
Authors: Power Phoebe , Payne Susannah , Walsh Rebecca , Manoharan Neevika , Nelson Adam 
Published: June 2023

Abstract
Treatment-related myeloid neoplasms are rare though well-defined secondary malignancies associated with prior cytotoxic treatment, (particularly alkylating agents and topoisomerase inhibitors). They are rarely associated with primary central nervous system tumours and have historically carried a very poor prognosis. Pediatric low-grade gliomas (pLGG) are the most common CNS tumours of childhood, and up to 50% of patients will require adjuvant therapy. This has traditionally consisted of low-dose metronomic chemotherapy, although the advent of genomic profiling and identification of molecular drivers of pLGG means novel targeted therapies are changing this paradigm. There has only been a single reported case of treatment-related myeloid leukaemia secondary to pLGG treatment. We present a novel case of a 17-year-old girl with treatment-related myelodysplastic syndrome following chemotherapeutic treatment for pLGG. The patient was initially diagnosed at age 4 with a metastatic pilomyxoid astrocytoma (primary suprasellar lesion with spinal metastases) and received first-line treatment with a vincristine and carboplatin regimen. She had disease progression at age 7 and received second-line treatment with thioguanine, procarbazine, lomustine and vincristine. Following further progression at age 16, a biopsy of the suprasellar mass showed a pilocytic astrocytoma with a KIAA1549-BRAF fusion. Prior to commencing a targeted treatment as third-line therapy, she was noted to have a macrocytic anaemia, which subsequently evolved to a bicytopenia. Bone marrow analysis showed a hypocellular marrow with an abnormal myeloid clone with a chromosomal 1q gain, consistent with a diagnosis of treatment-related myelodysplastic syndrome. Germline analysis was negative for bone marrow failure predisposition gene abnormalities. She is currently undergoing workup for bone marrow transplant. Given the poor prognosis of treatment-related myeloid neoplasms, this case represents an important note of caution when choosing appropriate therapy for pLGG, especially given the evolving role for targeted treatments in this disease with otherwise very favourable long-term survival.
LGG-16. NOVEL COMBINATION THERAPIES FOR PEDIATRIC LOW-GRADE GLIOMA AND CHARACTERIZATION OF THE UNDERLYING CELL DEATH MECHANISMS
Authors: Zeiser Constantia , Rösch Lisa , Herter Sonja , Peterziel Heike , Selt Florian , Milde Till , Witt Olaf , Oehme Ina 
Published: June 2023

Abstract

BACKGROUND
The majority of pediatric low-grade gliomas (pLGGs) harbor a MAPK/ERK pathway overactivating alteration, making them suitable for MAPK-targeted therapies such as MEK inhibition. However, MEK inhibitor treatment alone is in most cases not sufficient to induce complete response, and a rapid rebound of the tumor is the consequence after withdrawal of the therapy.

METHODS
To identify novel cytotoxic drug combinations, we screened 89 clinically relevant drugs in combination with the MEK inhibitors trametinib and binimetinib. We assessed the regression of cultured 3D microtumors of patient-derived BT40 and BT314 cells, both harboring a BRAFV600E mutation. Depending on the inducible expression of SV40 large T antigen, the BT314 model can reflect two states, proliferation and oncogene-induced senescence. The area of spheroids was measured both in brightfield and with the fluorescent dye TMRE (mitochondrial polarization) at the start and the end of the treatment (6 days). Moreover, metabolic activity was determined through bulk ATP measurement. Drug hit combinations were further analyzed for the successful induction of cell death using high-content microscopy (HCM) imaging and enzymatic caspase activity assays. The microscopic readouts included Hoechst 33342 for the detection of nuclear morphology, BODIPY for sensitizing lipid peroxidation and CM-H2DCFDA as a general oxidative stress indicator. The images were analyzed with the automated image analysis program CellProfiler and the machine learning application CellProfiler Analyst.

RESULTS
The screening identified the following drug hits: navitoclax (BCL-2 family inhibitor), BRAF inhibitor dabrafenib, mTOR inhibitor everolimus and selinexor, a selective inhibitor of nuclear export. All drugs increased apoptotic cell death and ROS levels at clinically achievable drug concentrations, except the mTOR inhibitors. These results are currently validated in vivo with BT40 zebrafish embryo xenograft models.

CONCLUSION
Combinations of MEK inhibitors with navitoclax, dabrafenib or selinexor resulted in sufficient apoptosis to induce regression of pLGG microtumors in vitro.
MDB-01. ELUCIDATING THE ROLE OF THE BAIAP2-CDC42 INTERACTION MEDULLOBLASTOMA
Authors: Ruiz Luz , Suter Rober , Jangde Nitish , Ayad Nagi 
Published: June 2023

Abstract
Medulloblastoma is the most common malignant pediatric brain tumor. Despite advancements in treatment, there is still no significant improvement in survival rates. This indicates novel, more effective treatment options are necessary. In addition, when medulloblastoma is metastasized outside the central nervous system, there is worst mortality rate. The regulatory properties of metastasis and invasion are still not well understood. We sought to identify potential key regulators for these processes in medulloblastoma. Single-cell RNA sequencing data was collected for medulloblastoma patients and an experiment with a Tg (Neurod-Smoothened*A1) mouse. We identified BAIAP2 and CDC42 to be differentially expressed compared to non-tumor brain cells. Previous literature shows that these genes are implicated in oncogenic properties including invasion, migration, and proliferation in different cancers and neurological diseases. We hypothesize that distinct cell populations within medulloblastoma should show different expressions of BAIAP2 and CDC42, specifically those involved in invasion and migration. We determined that BAIAP2 and CDC42 have higher expression in medulloblastoma tumors compared to the control. In our data, the cell population enriched for microtubule depolymerization had a high expression of BAIAP2, unlike the other clusters. This study is important because BAIAP2 and CDC42 have yet to be characterized in medulloblastoma. Our data suggest a potential role for BAIAP2 and CDC42 in the pathophysiology of medulloblastoma which can possibly be a target for future studies.
MDB-02. NUCLEAR ENVELOPE PHOSPHATASE CTDNEP1 MUTATIONS POTENTIATE AGGRESSIVE MEDULLOBLASTOMA BY TRIGGERING MYC ACTIVATION AND GENOMIC INSTABILITY
Authors: Lu Richard , Luo Zaili 
Published: June 2023

Abstract
MYC-driven medulloblastomas are highly aggressive childhood brain tumors, however, the molecular and genetic events triggering MYC amplification and malignant transformation remain elusive. Here we report that mutations in CTDNEP1, a CTD nuclear-envelope-phosphatase, are the most significantly enriched recurrent alterations in MYC-driven medulloblastomas, and define high-risk subsets with poorer prognosis. Ctdnep1 ablation promotes the transformation of murine cerebellar progenitors into Myc-amplified medulloblastomas, resembling their human counterparts. CTDNEP1 deficiency stabilizes and activates MYC activity by elevating MYC serine-62 phosphorylation, and triggers chromosomal instability to induce p53 loss and Myc amplifications. Further, phosphoproteomics reveals that CTDNEP1 post-translationally modulates the activities of key regulators for chromosome segregation and mitotic checkpoint regulators including topoisomerase TOP2A and checkpoint kinase CHEK1. Co-targeting MYC and CHEK1 activities synergistically inhibits CTDNEP1-deficient MYC-amplified tumor growth and prolongs animal survival. Together, our studies demonstrate that CTDNEP1 is a potent tumor suppressor in highly aggressive MYC-driven medulloblastomas by controlling MYC activity and mitotic fidelity, pointing to a CTDNEP1-dependent targetable therapeutic vulnerability.
MDB-17. THE PTBP2 SPLICING FACTOR IS ESSENTIAL IN GROUP 3/4 MEDULLOBLASTOMA
Authors: Caisova Veronika , Rivero-Hinojosa Samuel , Chintala Navya , Lu I-An , Huizhen Zhang , Mochizuki Aaron , Yang Jianhua , Rood Brian 
Published: June 2023

Abstract
Medulloblastoma (MB) is a common malignant pediatric brain tumor with significant heterogeneity among its four molecular subgroups WNT, SHH, Group 3 and 4. Group 3/4 MB are aggressive with a relatively poor prognosis indicating a critical need for novel therapeutic targets. Since the transcriptomic profiling of MB subgroups has not resulted in the expected identification of therapeutic potential, we focused on a proteomic approach. Building upon our previously published quantitative proteomic study, we selected 157 proteins significantly over-expressed in Group 3/4 MB. By using targeted loss-of-function CRISPR-Cas9 screen in four MB cell lines, we identified 17 essential genes shared by at least 2 of the cell lines. The PTBP2 splicing factor was essential to the survival of all cell lines. PTBP2 is an RNA binding protein involved in splicing regulation playing an important role during neuronal maturation. We hypothesize that persistent expression of PTBP2 results in a differentiation arrest and preservation of proliferative potential. To evaluate the role of PTBP2 in group 3/4 MB biology, we used single target CRISPR-Cas9 to stably knock-out PTBP2 expression in D556 and MB002 MB cells. We performed bulk RNA-seq and quantitative proteomics of these KO clones along with CLIP-seq to map PTBP2 targeted transcripts. We also performed phenotype assays in KO clones. RNA-seq in PTBP2 KO MB cells revealed 2,213 differentially expressed genes predominantly involved in neuronal differentiation and antigen presentation. These genes were cross-referenced with quantitative proteomics and eCLIP data. Phenotype assays of PTBP2 KO cells revealed reduced proliferation and migration compared to controls. To study the effect of PTBP2 on MB differentiation, we stimulated MB002 PTBP2 KO cells with FBS and observed increased surface attachment and differentiated morphology compared to controls. This suggests that persistent PTBP2 expression inhibits differentiation in MB cells with an attendant increase of proliferation activity.
MDB-28. DEEP LEARNING-BASED PERSONALIZED SURVIVAL PREDICTION FOR MEDULLOBLASTOMA
Authors: Stefan Sabina , Northcott Paul , Hovestadt Volker 
Published: June 2023

Abstract
Medulloblastoma is one of the most common malignant brain tumors in children and a leading cause of cancer-related death in this age group (5-year survival rate of ~70%). However, survival outcomes are heterogeneous, and currently associated with only a small number of clinical and molecular parameters. Instead, in this work we aim at exploiting the wealth of hidden information in genome-wide DNA methylation array data to predict accurate personalized survival probability curves for Group 3 and 4 medulloblastoma patients, the subgroups associated with the most heterogeneous outcomes. To accomplish this, we implemented a sparse neural network trained on DNA methylation and copy-number variation (CNV) profiles of over 900 medulloblastoma patients. Our network is designed specifically to leverage censored data for training, and makes no assumptions about the underlying distribution of the output survival probability curves. Furthermore, the architecture of our network is biologically interpretable, enabling us to probe the underlying biology driving survival. Our results demonstrate excellent discrimination and calibration, with a 5-year AUROC of 0.8. We compare our results to currently used stratification schemes on validation cohorts, including SJMB03 clinical trial data, and demonstrate significantly improved risk stratification (c-index of 0.66 vs 0.74, p &lt; 0.0001). We find several cases in which patients assumed to be low-risk were predicted to progress rapidly, and vice versa, motivating possible refinement of risk categories in clinical practice. Our approach, which is based entirely on DNA methylation array data, may facilitate the de-escalation of therapy for low-risk patients to minimize treatment-related side effects and the intensification of treatment for high-risk individuals. The approach can be extended to other cancer types, and highlights the value of AI in healthcare towards realizing the goal of personalized medicine.
MDB-31. PRELIMINARY STUDY ON BIOLOGICAL SIGNIFICANCE OF WTX G435D MUTATION IN MEDULLOBLASTOMA
Authors: Hu Yuanjun , Wang Jing , Chen Zhongping 
Published: June 2023

Abstract
BACKGROUND:
Previously, we reported a case of familial medulloblastoma without hereditary tumor syndrome. Through analysis of the whole genome sequencing of the family members, we found a common germline WTX (c.1863C&gt;T, p.Gly435Asp) mutation in this family: cytosine (C) at position 63411863 on X chromosome was replaced by thymine (T), and the corresponding amino acid was glycine Gly (G) at position 435, which was converted to aspartic acid Asp (D). We conduct this study to explore the biological significance of this mutation in medulloblastoma cell lines.
METHODS:
Medulloblastoma cells (cell lines and primary cells) with stable WTX (c.1863C &gt; T) hybrid mutation were contstrcuted by lentivirus transfection. The effects of WTXG435D on proliferation, invasion and chemosensitivity of medulloblastoma cells were investigated in vitro and in vivo.
RESULTS:
Proliferation and invasion ability of WTXG435D medulloblastoma cell were significantly weaker than WTX WT cells, and they were more sensitive to radiation and chemotherapy(cisplatin, vincristine) treatment in vitro. In vivo, animal experiments also showed that intracranial in situ tumors formed by WTXG435D medulloblastoma cells were more sensitive to radiotherapy and chemotherapy than in situ tumors formed by WTXWT ones, and the survival was longer.
CONCLUSION:
WTX (c.1863C&gt;T, p. Gly435Asp) mutation could reduce the proliferation and invasion ability of medulloblastoma cells, and enhance the sensitivity of radiotherapy and chemotherapy. Which could be a potential therapeutic target for further investigation.
SURG-03. LAYER-BY-LAYER CORE-SHELL NANOPARTICLES FOR THE DELIVERY OF HDAC INHIBITORS TO THE CENTRAL NERVOUS SYSTEM
Authors: Marand Michael H , Han Emily , Kim Seung Hyun , Hammond Paula T , Straehla Joelle P 
Published: June 2023

Abstract
Diffuse midline gliomas (DMGs) present unique challenges in pediatric cancer care. The location of DMGs often renders the tumor inoperable and radiation produces only a temporary effect. One promising approach in DMG treatment is histone deacetylase (HDAC) inhibitors. However, HDAC inhibitors are difficult to deliver to primary central nervous system (CNS) tumors because of low blood-brain barrier (BBB) permeability and poor accumulation in tumor tissues. The BBB permeability is particularly restricting for DMGs, which maintain a relatively intact BBB. To overcome these delivery challenges, we developed a new polymer-lipid core-shell nanoformulation for the delivery of HDAC inhibitors to the CNS that does not rely on polyethylene glycol (PEG) for colloidal stability. Removing PEG allows for surface charge manipulation to generate charged nanoparticles amenable to layer-by-layer assembly with alternately charged polyelectrolytes. This modular platform allows the addition of targeting moieties to enhance the delivery of HDAC inhibitor core-shell nanoparticles across the BBB and into DMG cells. HDAC inhibitors panobinostat and quisinostat were loaded into core-shell nanoparticles comprised of a poly(lactide-co-glycolide) (PLGA) core and a phospholipid shell. Nanoparticles were characterized using dynamic light scattering and transmission electron microscopy exhibiting favorable properties for in vivo delivery (panobinostat nanoparticle diameter 91.6 ± 1.0 nm, polydispersity index 0.115 ± 0.013). The release of HDAC inhibitors from layered and unlayered core-shell nanoparticles was investigated using high-performance liquid chromatography. The deacetylating activity of nanoencapsulated agents was confirmed in vitro in patient-derived DMG neurosphere models. CNS pharmacokinetic parameters are currently being investigated after intravenous delivery of drug-loaded nanoparticles in non-tumor-bearing mice, including cerebrospinal fluid and brain tissue concentrations of the encapsulated agents. Future studies will investigate brain biodistribution of core-shell HDAC inhibitor nanoparticles in orthotopic DMG tumor-bearing mice, comparing intravenous and local delivery strategies.