bluuebunny/crossref_metadata_2025_split huggingface dataset:
EXTH-55. DELIVERY OF CAS9/SGRNA RIBONUCLEOPROTEIN CONJUGATE VIA VIRUS-LIKE PARTICLES FOR EFFICIENT GENE EDITING OF TRANSCRIPTION FACTOR BRACHYURY IN GLIOBLASTOMA
Authors: Hsu Wesley , Lu Baisong , Hu Yunping 
Published: November 2022

Abstract
Brachyury, a transcription factor that is a critical driver during normal embryonic development, is highly expressed in glioblastoma (GBM) but rarely expressed in normal adult tissues. Recent evidence suggests that brachyury plays a crucial role in regulating GBM cell proliferation and stemness, implying the potential of brachyury as a therapeutic target. However, like other transcription factors, brachyury is not readily inhibited pharmacologically due to the absence of ligand-accessible small-molecule binding pockets, and a direct inhibitor of brachyury has not been identified. Our team has recently developed a novel virus-like particle (VLP)-based system by fusing aptamer-binding protein to the lentiviral nucleocapsid protein within the group-specific antigen, which allows for efﬁcient packaging of Cas9/guide RNA (gRNA) ribonucleoprotein (RNP) for gene editing. Our data suggests that our VLP-mediated Cas9/gRNA RNP allows for transient expression of Cas9 in patient-derived GBM cells while maintaining effective gene editing efficiency (∼40%). Our strategy results in 90% knockdown of brachyury in vitro with subsequent inhibition of GBM cell proliferation and stemness as well as enhanced susceptibility of tumor cells to radiation via increased cleavage of caspase-3. In addition, our in vivo results using a xenograft mouse model further demonstrates that transduction with VLP-packaged Cas9/brachyury gRNA inhibits GBM progression. Our findings demonstrate that our novel VLP-based system allows for efficient delivery of Cas9/gRNA RNP-based brachyury gene editing technology and represents a novel treatment strategy for GBM.
EXTH-57. IDENTIFICATION OF NOVEL IMMUNE CHECKPOINT MOLECULE IN GLIOMA, LAIR1
Authors: Tao Haipeng , Chen Dongjiang , Yang Changlin , Jin Linchun , Von Roemeling Christina , Mendez-Gomez Hector , Liu Ruixuan , Nguyen Duy , Hou Alicia Y , Pepe Alfonso , Weidert Frances , Ghiaseddin Ashley , Mitchell Duane , Deleyrolle Loic , Sayour Elias , Sawyer W , Tran David , Huang Jianping 
Published: November 2022

Abstract

BACKGROUND
In recent years, emerging evidence indicates that tumor-associated myeloid cells (TAMCs) affect cancer progression. But the molecular mechanisms and signaling pathways involved in the TAMCs’ interaction with extracellular matrix (ECM) and tumor stroma are incompletely understood. Checkpoint inhibition therapy targets immune inhibitory receptors, such as CTLA-4 and PD-1, which have become a major weapon in fighting cancer. These antibodies demonstrate apparent advantages such as being easy to use, broad applicability, and durable clinical response. Leukocyte-Associated Immunoglobulin-like Receptor 1 (LAIR1), also called CD305, a collagen-binding immunoreceptor tyrosine-based inhibition motifs (ITIM)-bearing inhibitory receptor, was identified to present on almost all immune cell populations and overexpressed in cancers of human patients. However, LAIR1’s role and regulation in solid cancers are poorly defined.

OBJECTIVE
To identify LAIR1 as a new class of immune checkpoints in cancers that impacts TAMCs-associated tumor immunity. Further functional investigation warrants understanding the crosstalk between LAIR1-related immune checkpoint blocking agent(s), immune micro-environment, and its underlying mechanisms for targeted therapy development.

METHODS
Murine GBM cell line KR158-Luciferase cell line was used to set up the mouse model. Tumor-bearing mice were administered the Anti-LAIR1 blockade and IgG, followed by IVIS imaging for tumor growth and survival were recorded. The presentation and phenotype of immune cell populations in tumors and spleens were measured through scRNA-Seq.

RESULTS
GBM is one of these tumors that overexpress LAIR1 based on the TCGA analysis. LAIR1 molecules express highly on macrophages, monocytes, DCs, and activated T cells, not on the naive T cells. We found that LAIR1 blockade enhanced survival in preclinical GBM models. LAIR1 blockade reduced the presentation and function of TAMCs in tumors. What's more, LAIR1 blockade provided a synergetic antitumor effect with CD70 CAR T cells.

CONCLUSION
This study suggests that we identified a novel immune checkpoint molecule, LAIR1, which can limit tumor progression.
EXTH-74. ATR INHIBITION IN EXPERIMENTAL GLIOMA
Authors: Walter Bianca , Schnabel Leonard , Hirsch Sophie , Stahl Aaron , Haeusser Lara , Tsiami Foteini , Kuhlburger Laurence , Nahnsen Sven , Templin Markus , Schenke-Layland Katja , Merk Daniel , Tabatabai Ghazaleh 
Published: November 2022

Abstract
Glioblastoma are incurable primary tumors of the central nervous system. Novel treatment options are urgently needed. Recently, we investigated molecular effects of disrupting the (b)HLH network, a frequently altered and tumor-promoting transcriptional network in glioma, by introducing a dominant-negative variant of the E47 protein. The identified downstream anti-tumor effects included the DNA damage response pathway. In the present study, we aimed at investigating the anti-glioma effects of ATR inhibition in vitro and in vivo with a particular focus on treatment-induced vulnerabilities and potential combination therapies. We performed acute cytotoxicity, clonogenic survival assays, flow cytometry-based determination of cell cycle status and apoptosis induction in human and murine glioma cell lines in vitro, and investigated tumor growth in the SMA560/VMDk syngeneic mouse model in vivo. Furthermore, we applied RNA sequencing, DigiWest protein profiling analyses and genome-wide CRISPR/Cas9 loss-of function and activation screens to determine potential combination therapies. Combination treatments were further analyzed for synergism by the Bliss and ZIP synergy models. ATR inhibition in glioma cells resulted in reduced viability and clonogenic survival. However, differential patterns of gene expression deregulation (e.g. p53 signaling) and differing modes of cell cycle arrest in distinct glioma model systems argue for a substantial heterogeneity with regard to response to ATR inhibition. Based on our data we designed combinatorial treatments. We will present novel insights into promising combinatorial treatments using ATR inhibition and their underlying molecular mechanisms.
EXTH-79. HIJACKING A NEURODEVELOPMENTAL EPIGENOMIC PROGRAM IN METASTATIC DISSEMINATION OF MEDULLOBLASTOMA
Authors: Zou Han , Poore Bradley , Brown Emily , Yash Patel , Hu Zhongliang , Li Xuejun , Pollack Ian F , Friedlander Robert M , Hainer Sarah , Taylor Michael , Hu Baoli 
Published: November 2022

Abstract
Normal brain development relies on precise genetic and epigenetic spatiotemporal regulation of gene expression. How dysregulation of neurodevelopment relates to medulloblastoma, the most common pediatric brain tumor, remains elusive. Here, we uncovered a novel neurodevelopmental epigenomic program that regulates Purkinje cell migration in developing cerebellum is hijacked to induce tumor metastatic dissemination in medulloblastoma. Integrating publicly available datasets with our in-house data, unsupervised analyses revealed that BAF60C/SMARCD3, a subunit of SWI/SNF chromatin remodeling complex, promotes tumor cell migration in vitro and metastasis in vivo. Based on analyzing the single-cell RNAseq data of cerebellum developmental trajectory in mice and humans, aligning with the medulloblastoma patients’ datasets, we found that BAF60C/SMARCD3 regulated DAB1-mediated Reelin signaling is involved in Purkinje cell positioning during cerebellum development and medulloblastoma metastasis by orchestrating the cis-regulatory elements (CREs) at the DAB1 gene locus. Moreover, analysis of spatiotemporal gene expression and chromatin architecture in the human and mouse cerebellum demonstrated that transcription activity of the BAF60C/SMARCD3-DAB1 circuit is downregulated in a mature state of cerebellar development, however, is upregulated in metastatic medulloblastoma. We further identified that a core set of transcription factors, enhancer of zeste homolog 2 (EZH2) and nuclear factor I X (NFIX), bi-directionally control BAF60C/SMARCD3 transcriptional regulation by coordinating with the CREs at the BAF60C/SMARCD3 gene locus to form a chromatin hub during developing cerebellar development and medulloblastoma metastatic dissemination. Highly expressed BAF60C/SMARCD3 activates the Reelin/DAB1 signaling pathway downstream Src kinase, which was validated in the pair-wised primary and metastatic tumors from medulloblastoma patients. Preclinical medulloblastoma mouse models revealed that inhibiting Src activity reduces tumor cell migration and metastatic dissemination at a lower and safe dose. Together, these data deepen our understanding of how the developmental program influences disease progression and provide an opportunity for the development of therapeutics for this devastating brain cancer in children.
EXTH-80. THE MOLECULAR FUNCTION AND THE THERAPEUTIC POTENTIAL OF MICRORNA-211 IN GROUP 3 MEDULLOBLASTOMAS IN CHILDREN
Authors: Yuan Menglang , Katsushima Keisuke , Pokhrel Rudramani , Lee Bongyong , Stapleton Stacie , Jallo George , Raabe Eric , Eberhart Charles , Perera Ranjan 
Published: November 2022

Abstract
Medulloblastoma is a central nervous system tumor that primarily affects children and requires aggressive therapy. Patients often suffer from treatment-related side effects, and treatment-resistant recurrences are common, with high mortality rates. There are four major molecular MB subgroups (Wnt- and Shh-activated MBs G3 and G4 MBs). G3 MB is the most aggressive subtype, and the diagnosis and management remain challenging. Medulloblastomas develop through various genetic, epigenetic, and noncoding (nc) RNA-related mechanisms. However, the roles played by ncRNAs, (microRNAs, long noncoding RNAs, circular RNAs, etc.) in MB development remain poorly defined. Here we address this knowledge gap with an exemplar microRNA, microRNA 211 (miR-211) implicated in G3 MB development and progression. Our preliminary results support that miR-211 is an attractive therapeutic agent to treat this aggressive MB subtype. miR-211 is significantly downregulated in medulloblastoma cell lines compared to normal cerebellum, underscoring its important role as a therapeutic agent and a biomarker. miR-211 ectopic expression in G3 MB cells significantly reduced cell proliferation and 3D colony formation and induced apoptosis. In vivo, miR-211force-expression in G3 MB cells injected into mouse cerebellum produce smaller tumors than those derived from parental cells. We identified that Long-chain-fatty-acid—CoA ligase 4 (ACSL4), and the oncogene Ras-related protein Rab22A are miR-211 targets genes for G3 medulloblastomas. The preliminary results of this study are encouraging and will provide a pre-clinical foundation for further therapeutic testing.
EXTH-84. A NOVEL DUAL-SPECIFIC CHIMERIC ANTIGEN RECEPTOR T CELL WITH HIGH SPECIFICITY FOR EGFR AND EGFRVIII IMPROVES SURVIVAL IN EGFR EXPRESSING MEDULLOBLASTOMA
Authors: Wilson Joseph , Wilkinson Daniel , Ryan Katherine , Bigner Darell , Chambramohan Vidya , Fecci Peter 
Published: November 2022

Abstract

BACKGROUND
Medulloblastoma is the most common malignant brain tumor in children 0-19 years of age and current treatment requires surgical resection, followed by high dose chemotherapy and radiotherapy. Therapy carries high morbidity, with late effects including neurocognitive decline, endocrine dysfunction, and subsequent malignancies. Chimeric antigen receptor (CAR) T cell therapy presents the potential for less toxic and more effective treatment for medulloblastoma. Epidermal growth factor (EGFR) is expressed in medulloblastoma derived cell lines and appears to serve an important role in the metastatic potential of this tumor type. D2C7 is a recombinant monoclonal antibody short chain variable fragment (scFv) with dual specificity, binding to wild type EGFR (EGFRwt) and its mutant EGFR variant III (EGFRvIII). We previously developed a novel, third-generation chimeric antigen receptor T-cell construct utilizing the D2C7 scFv and performed analysis on both glioblastoma and medulloblastoma tumor models.

METHODS
U87 and U87vIII glioblastoma and DAOY medulloblastoma cell lines were characterized by flow cytometry to evaluate for EGFRwt and EGFRvIII expression. Cytotoxicity assays were performed utilizing flow cytometry on serially diluted effector: target ratios of U87, U87vII, and DAOY mixed with D2C7 CAR. DAOY was orthotopically implanted into the frontal lobe of NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ (NSG) mice. After tumor implantation, D2C7 CAR was introduced within the tumor bed.

RESULTS
EGFR was detected in U87, U87vIII, and DAOY on flow analysis. EGFRvIII was expressed only on U87vIII. D2C7 CAR demonstrated effective in vitro cytotoxicity against all three cell lines with &gt;90% cytotoxicity achieved at an effector: target ratio of 2.5:1. In vivo, D2C7 CAR treatment significantly prolonged survival compared to treatment with mock CAR.

CONCLUSION
D2C7 CAR demonstrates efficacy in vitro and in mouse models of medulloblastoma and glioblastoma. Future steps in our work will set out to assess the D2C7 CAR in NSG mice with DAOY in the posterior fossa.
EXTH-89. EXPLORING ANTI-TUMOR EFFECTS OF SMALL MOLECULE INHIBITORS OF CDK4/6, FAK, AND RAF/MEK IN PRECLINICAL MODELS OF MENINGIOMA
Authors: Ijad Nazanin , Waller Katie L , Nayyar Naema , Larson Juliana M , Spear Joshua , Cahill Daniel , Barker Frederick G , Galanis Evanthia , Sarkaria Jann , Wakimoto Hiroaki , Vaubel Rachael , Brastianos Priscilla 
Published: November 2022

Abstract

BACKGROUND
Adequate treatment options are limited for patients with meningiomas once they have exhausted surgery and radiation. As our molecular understanding of meningiomas and their progression has improved, CDK, FAK, and RAF/MEK pathways have emerged as potential avenues for therapeutic intervention. However, patient-derived cell and xenograft models of meningiomas are difficult to establish, presenting a challenge in preclinical testing and validation of new therapeutic targets. The objective of this study is to develop informative preclinical models of meningiomas and use those to evaluate the activity of potent and selective inhibitors of CDK4/6, FAK, and RAF/MEK.

METHODS
We used CellTiter-Glo assay to assess the anti-tumor effects of CDK4/6 inhibitor abemaciclib, FAK inhibitor defactinib (VS-6063) and dual RAF/MEK inhibitor RO5126766 (VS-6766) in BEN-MEN-1 (WHO Grade I, NF2-mutant), CH157-MN (WHO Unknown, NF2-mutant), and IOMM-Lee (WHO Grade III, CDKN2A loss) cell lines. Additionally, we used CyQUANT assays to quantify anti-proliferation activity of these inhibitors in a newly established patient-derived cell line MN14 (WHO Grade II, NF2 mutant). In parallel, we continue our attempts to establish patient-derived cell and xenograft models using freshly isolated surgical specimens of progressive meningioma.

RESULTS
Defactinib monotherapy significantly decreased cell viability across a 10nM-10μM range for all three established cell lines. RO5126766 showed improved efficacy (by 10-fold) in IOMM-Lee, compared to CH157-MN and Ben-Men-1. Abemaciclib showed potent effects at 10nM in IOMM-Lee and BEN-MEN-1, compared to 100nM for CH157-MN. In the MN14 patient-derived cell line, established from an NF2-mutant atypical meningioma, we observed a significant decrease in cell proliferation with defactinib. A decrease in cell proliferation was also observed with RO5126766 and abemaciclib.

CONCLUSION
Selective inhibitors of CDK4/6, FAK, and RAF/MEK showed anti-meningioma activity against traditional and patient-derived cell models in vitro. Molecular characterization and establishment of patient-derived models that encompass the diversity of clinical meningiomas are ongoing.
EXTH-99. SYSTEMIC AND BRAIN PHARMACOKINETICS OF A GABAA RECEPTOR AGONIST, AMLAL-101, AS AN INVESTIGATIONAL THERAPEUTIC FOR THE TREATMENT OF PRIMARY AND METASTATIC BRAIN CANCERS
Authors: Karve Aniruddha , Ahmed Taukir , Desai Janki , Bhattacharya Debanjan , Toukam Donatien Kamdem , Gadgil Sidharth , Pemberton Merissa , Plas David , Gudelsky Gary , Sallans Larry , Cook James , Desai Pankaj , Krummel Daniel Pomeranz , Sengupta Soma 
Published: November 2022

Abstract

PURPOSE
AMLAL-101 is a novel agent which preferentially targets α3, α5 subtypes of ɣ-amino butyric acid receptors and shows anti-tumor activity against disparate cancer types. AMLAL-101 is being advanced as an ‘add-on’ to potentiate treatment of primary and metastatic brain cancers. However, AMLAL-101 must penetrate the blood-brain barrier (BBB) and show sufficient brain retention. The primary purpose of this study was to determine the plasma pharmacokinetics (PK) and quantitative estimate of the BBB permeability of AMLAL-101.

METHODS
We performed intracranial microdialysis, employing jugular vein cannulated Sprague-Dawley rats which facilitated simultaneous serial blood and brain extracellular fluid (ECF) sampling. AMLAL-101 was injected i.p. at 5 mg/kg and serial blood and brain ECF samples collected up to 10 h post-dosing. Plasma and ECF samples were analyzed by LC/MS-MS and plasma and ECF concentration vs time PK profiles determined. In vivo recovery analysis was performed using retrodialysis and rapid equilibrium dialysis employed to determine the extent of protein binding.

RESULTS
AMLAL-101 plasma protein binding was 85% and in vivo recovery from ECF was 25%. AMLAL-101 peak concentration (Cmax) in plasma and brain ECF were 15 µM and 13.8 µM, respectively. The plasma and brain ECF area under the concentration (AUC0-10) were 27.5 h.µg/mL and 24.10 h.µg/mL, respectively. The brain partitioning of unbound AMLAL-101 (Kp,uu; determined either as a ratio of brain ECF Cmax:unbound plasma Cmax or brain ECF AUC: unbound plasma AUC), were 6.13 and 4.13, respectively. The elimination half-life of AMLAL-101 was 3 h for both brain ECF and plasma.

CONCLUSIONS
These results suggest that AMLAL-101 has the requisite BBB permeability required for brain cancer therapeutics. AMLAL-101 shows significant brain retention when compared to a chemically similar agent that does not show anti-cancer activity, which may contribute to efficacy of AMLAL-101 as an anti-tumor agent for treatment of brain cancers.
EXTH-101. MECHANISTIC INSIGHTS INTO CYTOTOXIC EFFECTS OF LETROZOLE AGAINST GLIOBLASTOMA CELLS
Authors: Karve Aniruddha , Desai Janki , Dave Nimita , Phoenix Timothy , Gudelsky Gary , Plas David , Dasgupta Biplab , Wise-Draper Trisha , Desai Pankaj 
Published: November 2022

Abstract

PURPOSE
Despite significant advances in multimodal approaches for the treatment of glioblastoma (GBM), the outcome for the patients remains dismal. Previous research from our lab has shown that aromatase (CYP19A1), an enzyme that catalyzes in situ production of estrogens, is markedly elevated in GBM tissues. Additionally, letrozole (LTZ), an aromatase inhibitor with extensive record of efficacious use in breast cancer, exhibited significant activity against GBM in preclinical studies. The purpose of this study was to investigate the mechanism(s) of LTZ activity in GBM.

METHODS
Patient-derived GBM cells with varying aromatase expression and other genotypic/phenotypic characteristics were employed including aromatase expressing G43, G75, G76, BT-142-gfp-luc and aromatase-null G80 cells. The effects of LTZ on cell viability, neurosphere growth, DNA damage (assessed by measuring phosphorylated histone product ỴH2A.X using flow cytometry) and induction of apoptosis (caspase3/7 activity) were assessed. The effects of exogenous addition of estradiol (E2) was also examined.

RESULTS
LTZ treatment (72 hours) caused a marked reduction in viability and growth of all GBM cells (IC50 ranging from 0.08 to 0.7 µM) except in G80 cells where LTZ had no effects. LTZ treatment also resulted in inhibition of E2 synthesis in a time and dose-dependent fashion. Furthermore, in aromatase expressing cells LTZ treatment induced apoptosis as measured by increase in Caspase-3/7 activity and triggered significant DNA double strand break as determined by ỴH2A.X formation. The addition of exogenous E2 (250 pg/ml) abrogated the effects of LTZ on cell viability, DNA damage and induction of apoptosis.

CONCLUSIONS
Our study suggests that the cytotoxic effects of LTZ against GBM are mediated, at least partially, by the inhibition of aromatase activity which results in estrogen deprivation. These mechanistic insights are important to facilitate repurposing LTZ as a novel anti-GBM drug.
EXTH-103. DISCOVERY AND PHARMACOLOGICAL CHARACTERIZATION OF SKL27969, A NOVEL BRAIN-PENETRATING PRMT5 INHIBITOR WITH STRONG ANTITUMOR ACTIVITY, IN GLIOBLASTOMA AND BRAIN METASTASIS PRECLINICAL MODELS
Authors: Moon Mijin , Shin Yongje , Lee Ho Yeon , Hong Beomjin , Kim Jinhee , Lee Jun , Laramy Janice K , Vashi Vijaykumar , Park Sook-Kyung 
Published: November 2022

Abstract
Protein arginine methyltransferase 5 (PRMT5), a type II methyltransferase, regulates cellular processes such as survival, proliferation, and apoptosis by inducing symmetric arginine dimethylation of multiple cellular proteins involved in the regulation of cellular transcription and RNA splicing. Elevated tumor PRMT5 protein level has recently been correlated with poor prognosis and poor patient survival in various human cancers, including glioblastoma multiforme (GBM), lung cancer, breast cancer, and ovarian cancer. SKL27969 is a highly selective and potent PRMT5 inhibitor that is designed for brain penetration. It strongly binds to S-adenosylmethionine (SAM)-bound PRMT5/MEP50 (methylosome protein 50), leading to potent cellular symmetric dimethylarginine (SDMA) inhibition with an IC50 of 3.4 nM. Cell line panel profiling of SKL27969 revealed a broad spectrum of anti-proliferative activity in various cancer cell types, including CNS tumors, solid cancers prone to spread to brain, other solid cancers, and hematologic cancers. SKL27969 showed excellent brain exposure, with total and unbound partition coefficient (Kp and Kpuu) values of 6.57 and 1.08, respectively, and high tumor-to-plasma ratio, which resulted in significant survival benefit in orthotopic GBM xenograft models and metastatic brain cancer models such as intracranial xenograft models of non-small cell lung cancer (NSCLC) or triple-negative breast cancer (TNBC). Administration of SKL27969 showed strong tumor growth suppression in subcutaneous xenograft models of GBM, NSCLC, or TNBC. High correlation between tumor and plasma SDMA levels after SKL27969 treatment were observed in the preclinical models. Furthermore, SKL27969 showed potentiation of DNA damage when used in combination with DNA-damaging agents, which may suggest possible benefit as a combination therapy. Given the therapeutic target potential of PRMT5 in glioblastoma and other brain metastatic cancers, and the preclinical efficacy of SKL27969, these results have enabled the initiation of Phase 1/2 open-label, multicenter dose-finding study in patients with advanced solid tumors (NCT05388435).