bluuebunny/crossref_metadata_2025_split huggingface dataset:
TAMI-14. NANOPARTICLE DELIVERY OF PD-L1 CRISPR/CAS9 PLASMID DNA FOR ANTI-GLIOBLASTOMA IMMUNOTHERAPY
Authors: Fierro Javier , Tran An , Factoriza Chris , Chin Brandon , Dou Huanyu 
Published: December 2020

Abstract
Glioblastoma multiforme (GBM) is a devastating cancer that develops from astrocytes in the brain. GBM is fast acting and kills 90% of patients within 5 years. Several immunotherapies have been developed to treat GBM, however, major challenges still persist. For example, checkpoint proteins such as programmed cell death protein 1 (PD-1) and its ligand, programmed death ligand 1 (PD-L1), are upregulated in GBM cells to evade the immune system. Targeting PD-L1 for genetic knockdown is thus a promising avenue for the treatment of GBM. However, PD-L1 protein inhibitors have been shown to cause immune overreaction and toxicity, therefore requiring new technologies. CRISPR/Cas9 gene editing has been widely used for the study and treatment of many diseases, but has not been extensively studied for the treatment of GBM. The main challenge is developing a gene delivery platform for the delivery of CRISPR/Cas9 plasmid DNA (pDNA). Many viral vectors have been used for the delivery of pDNA, but unfortunately are associated with high toxicity. Nanotechnology is emerging as a new platform for the delivery of pDNA as it shows high transfection efficiency with low cytotoxicity. We developed a cationic core-shell nanoparticle (NP) capable of carrying CRISPR/Cas9 pDNA. This plasmid contains multiple guide RNA (gRNA) expression cassettes for the knockdown of PD-L1. PDL1gRNA-CRISPR/Cas9pDNA-NPs were taken up by U87 cells within 30 minutes, and entered into the nucleus at 2 hours. The effective delivery of PDL1gRNA-CRISPR/Cas9pDNA-NPs led to the expression of PD-L1 gRNA and Cas9 enzyme, and the knockdown of PD-L1. Regulation of immune balance was determined after PD-L1 knockdown in vitro and in vivo. Our study shows the potential of NP-based PDL1gRNA-CRISPR/Cas9 delivery as an anti-GBM immunotherapy for clinical applications.
TAMI-30. LOCAL TARGETING OF NAD+ SALVAGE PATHWAY ALTERS THE IMMUNE TUMOR MICROENVIRONMENT AND ENHANCES CHECKPOINT IMMUNOTHERAPY IN GLIOBLASTOMA
Authors: Li Ming , Kirtane Ameya , Kiyokawa Juri , Nagashima Hiroaki , Traverso Giovanni , Cahill Daniel , Wakimoto Hiroaki 
Published: December 2020

Abstract
The aggressive primary brain tumor glioblastoma (GBM) is characterized by aberrant metabolism that fuels its malignant phenotype. Diverse genetic sub-types of malignant glioma are sensitive to selective inhibition of the NAD+ salvage pathway enzyme nicotinamide phosphoribosyltransferase (NAMPT). However, the potential impact of NAD+ depletion on the brain tumor microenvironment has not been elaborated. In addition, systemic toxicity of NAMPT inhibition remains a significant concern. Here, we show that microparticle-mediated intratumoral delivery of NAMPT inhibitor GMX1778 induces specific immunological changes in the tumor microenvironment of murine GBM, characterized by upregulation of immune checkpoint PD-L1, recruitment of CD3+, CD4+ and CD8+ T cells and reduction of M2-polarized immunosuppressive macrophages. NAD+ depletion and autophagy induced by NAMPT inhibitors mediated the upregulation of PD-L1 transcripts and cell surface protein levels in GBM cells. NAMPT inhibitor modulation of the tumor immune microenvironment was therefore combined with PD-1 checkpoint blockade in vivo, significantly increasing the survival of GBM bearing animals. Thus, the therapeutic impacts of NAMPT inhibition extended beyond neoplastic cells, shaping surrounding immune effectors. Microparticle delivery and release of NAMPT inhibitor at the tumor site offers a safe and robust means to alter an immune tumor microenvironment that could potentiate checkpoint immunotherapy for glioblastoma.
TAMI-35. AUTOPHAGY MEDIATED LIPID CATABOLISM FACILITATES GLIOMA PROGRESSION TO OVERCOME BIOENERGETIC CRISIS
Authors: Wang Chenran , Haas Michael , Yeo Syn , Paul Ritama , Yang Fuchun , Vallabhapurapu Subrahmanya , Qi Xiaoyang , Plas David R , Guan Jun-Lin 
Published: December 2020

Abstract
Activation of mTORC1 plays a significant role in cancer development and progression. However, the metabolic mechanisms to sustain mTORC1 activation in stressed cancer cells are still underappreciated. Autophagy, one downstream process of mTORC1, is proposed to be suppressed under the condition of mTORC1 hyper-activation. Interestingly, we recently revealed higher autophagy activity in various Tsc-deficient tumor cells with mTORC1 hyper-activity. Nevertheless, the functions and mechanisms of autophagy in regulating mTORC1 in cancer cells are not well understood. In this study, we revealed a strong association of altered mRNA levels in mTORC1 upstream and downstream genes with poor prognosis of glioma patients. Our metabolic and molecular studies indicated that autophagy mediated lipid catabolism was essential to sustain mTORC1 activity in glioma cells under energy stresses. We found that autophagy inhibitors or fatty acid oxidation (FAO) inhibition in combination with 2-Deoxy-D-glucose (2DG) decreased oxidative phosphorylation, ATP production, mTORC1 activity, and survival of glioma cells in vitro. Consistently, the combination of chloroquine (CQ) or FAO inhibitors with 2DG effectively suppressed the progression of xenografted glioma with mTORC1 hyperactivation in mice. This study established a novel autophagy/lipid degradation/FAO/ATP pathway that maintains high mTORC1 signaling and tumor progression in brain cancer cells under energy stresses. The requirement of lipophagy in brain cancers may provide an opportunity to develop new molecular therapeutic targets to counteract mTORC1 for tumor progression.
TAMI-40. TUMOR MICROBIOME AND GLIOBLASTOMA (GBM)
Authors: Yust-Katz Shlomit , Gigi Elinor , Rosenberg Deborah , Kanner Andrew A , Laviv Yoseph , Benouaich-Amiel Alexandra , Siegal Tali , Barda Adva Levi , Straussman Ravid 
Published: December 2020

Abstract
Bacteria have been found to play major roles in many physiologic/disease processes including cancer. The presence of bacteria within brain tumors has never been explored. The aim of this study was to examine the microbiome of Glioblastoma. A cohort of 40 glioblastoma samples (FFPE), from two medical centers served for DNA extraction using a specialized extraction protocol that includes a bead beating step to ensure complete bacterial DNA recovery. A set of negative controls was introduced at different steps of the assay to identify and monitor contaminating bacterial DNA. We measured the levels of bacterial DNA in the samples using a RT-qPCR assay, amplifying the bacterial 16S rRNA gene and detected bacterial DNA in over 40% of the samples. To characterize the bacterial taxa that are present in GBM tumors, we applied 16S DNA sequencing on the samples. After implementing a stringent set of filters on the sequencing data, eliminating contaminating signal, we detected a total of 22 bacterial taxa in GBM tumors. To visualize bacteria in GBM tissues and learn about their localization within the tissue we used immunohistochemistry staining with anti-lipopolysaccharide (LPS) and anti-lipoteichoic acid (LTA) antibodies detecting gram negative and gram positive bacteria (correspondingly). Bacteria were also visualized by staining bacterial RNA using a 16S rRNA in situ hybridization assay. Staining of a human GBM tissue microarray (TMA) containing 32 cases of GBM showed that the majority of cases stained positive for LPS and ~40% were positive for 16S rRNA staining. Bacterial LPS and 16S rRNA were localized mainly inside the tumor cells. Our study demonstrates, for the first time, that bacteria or bacterial components are present in human Glioblastoma tumors. We are currently expanding our study cohort in order to better define the bacteria found within glioblastoma samples and assess their possible effects
TAMI-47. EVALUATION OF THE EFFECT OF MICROENVIRONMENTAL STRESSES ON GLIOMA STEMNESS IN VITRO AND EX VIVO
Authors: Pantazopoulou Vasiliki , Berg Tracy , Pietras Alexander 
Published: December 2020

Abstract
Glioblastoma is the highest-grade glioma and most aggressive brain tumor, affecting both adults and children. Treatment includes surgery followed by chemotherapy, most frequently in the form of temozolomide treatment, radiation and in some cases by application of tumor-treating fields. Tumor recurrence is very common and is driven by cells exhibiting a treatment-resistant cancer stem cell phenotype, located in specific niches, namely the hypoxic and perivascular niche. While previous studies have delved into the role of the microenvironment in supporting glioma stemness and therapy resistance, little is known about how the stromal cells themselves respond to microenvironmental stresses. Here, we show that one cell type of the tumor microenvironment, the astrocyte, becomes activated after treatment with radiation, temozolomide or hypoxia in vitro, showing an increase in GFAP and Vimentin expression as well as morphological changes. Activated astrocytes also change their secretome, secreting cytokines such as IL6, IL7, CCL2 and CCL8, all associated with worse prognosis in several patient cohorts. Importantly, conditioned medium or extracellular matrix from activated astrocytes induce stemness features in glioma cells, measured by increased side population or radiation resistant fractions. To evaluate our findings in a more relevant model, we use an ex vivo organotypic culture system for brain sections from our experimental mouse model of glioma. Cultures are maintained for up to 21 days in 21%, 5% or 1% O2 with a high proportion of surviving tumor and stromal cells. We intend to use the ex vivo system to evaluate the effect of microenvironmental stresses on glioma stemness. Our study suggests that the tumor microenvironment that has been exposed to stressors such as radiation, chemotherapy or hypoxia is important in maintaining stemness and therefore treatment resistance in glioblastoma. Further research is required to address these stress-induced changes and provide better treatment options for glioblastoma patients.
TAMI-52. G-CSF SECRETED BY EPIGENETICALLY REPROGRAMMED MUTANT IDH1 GLIOMA STEM CELLS, REVERSES THE MYELOID CELLS’-MEDIATED IMMUNOSUPPRESSIVE TUMOR MICROENVIRONMENT
Authors: Alghamri Mahmoud , Avvari Ruthvik , Thalla Rohit , Zhang Li , Ventosa Maria , Taher Ayman , Núñez Felipe , Heth Jason , Orringer Daniel , Lowenstein Pedro , Castro Maria 
Published: December 2020

Abstract
Mutation in isocitrate dehydrogenase (mIDH) is the main genetic lesion that defines clinical glioma subtypes and prognosis. This gain of function mutation is associated with the production of the oncometabolite, R-2-hydroxyglutarate, that inhibits α-ketoglutarate dependent enzymes such as TET2 and the Jumonji-C domain containing demethylases. The resultant epigenetic modifications elicit profound effects on the tumor biology and on the glioma-infiltrating immune cells. Here, we report that in genetically engineered mouse glioma models(1), IDH1 mutation caused an expansion of tumor infiltration granulocytes. Upon phenotypic and functional characterization, we uncovered that granulocytes in mIDH1 glioma express low level of immunosuppressive molecules and did not inhibit T-cell function. Single-cell sequencing revealed that these granulocytes are heterogeneous and composed of three distinct populations; neutrophils, pre-neutrophils, and a small fraction of immunosuppressive PMN-MDSCs. Moreover, primary human gliomas showed a higher cellular fraction exhibiting the PMN-MDSCs gene signature in wtIDH1 tumors than the mIDH1 tumors. The mechanism by which mIDH1 mediates non-immune suppressive granulocytes expansion involves epigenetic reprogramming which leads to enhanced expression of granulocyte colony-stimulating factor (G-CSF) in stem-like cells. High G-CSF gene expression is correlated with favorable patient outcome solely in LGG-astrocytoma with mIDH1. Thus, G-CSF represents a potential therapeutic that can be harnessed to improve immunotherapeutic responses in wild type IDH1 glioma patients.
TAMI-53. THE ONCOMETABOLITE D-2HG INDUCE INFLAMMATORY ASTROGLIOSIS CAUSING NEUROTOXICITY AND SEIZURES IN IDH MUTATED GLIOMA
Authors: Behringer Simon , Ravi Vidhya , Joseph Kevin , Beck Juergen , Schnell Oliver , Heiland Dieter 
Published: December 2020

Abstract
The role of tumor-associated astrocytes in the microenvironment of glioma has long been underestimated but is moving into the focus of current research. We explored the role of reactive astrocytes in IDH-mutated glioma using RNA-sequencing of purified astrocytes and microglia and single-nucleus RNA-sequencing of infiltrating tumor regions. Mapping of the transcriptional phenotype of astrocytes along developmental and reactive trajectories revealed an inflammatory transformation of IDH-mutated associated astrocytes. The major proportion of astrocytes is marked by complement-activation similar to findings in neuroinflammatory diseases. A human neocortical slices model with injected IDH-mutated patient-derived cells or D-2HG treatment (+/- microglia depletion) was used to map shared and unique transcriptional adaptation in astrocytes promoted by either tumor cells or metabolic alteration. High-dimensional electrophysiological profiling was used to investigate alterations in neural response to tumor-induced microenvironmental transformation. We showed that 2HG alone promote the inflammatory pattern of astrocytes, which causes neurotoxicity and seizures in our neocortical slice model. Depletion of microglia rescued the neurotoxicity suggesting that microglia predominantly drive inflammatory astrogliosis as a response to metabolic alteration the tumor environment. We showed that neurotoxic astrogliosis induced by the oncometabolite D-2HG via distinct microglia activation promote the evolution of frequently observed seizures in IDH-mutated glioma patients.
TAMI-64. PHYSICAL STRESSES IN BRAIN TUMORS
Authors: Kumar Ashwin , Nia Hadi , Datta Meenal , Chatterjee Sampurna , Ferraro Gino , Nahed Brian , Jain Rakesh 
Published: December 2020

Abstract
Solid stress, distinct from fluid pressure, is a physical force contained in and transmitted by solid components of the brain tumor, including cells and the matrix they produce. Solid stress has been shown to promote tumor progression, and decrease anticancer therapy efficacy. This is especially relevant in brain tumors, as the rigid skull results in these trapped forces, increasing intracranial pressure, and potentially leading to other complications, including neuronal cell death. Here we present a novel method of quantifying these physical stresses in situ in both mice (glioblastoma [U87], brain metastasis [BT474], and ependymoma models) and patients. Briefly, following a craniotomy, mechanical forces that include solid stress are released, which causes the tissue to deform in peaks (areas under compression) and valleys (areas originally under tension). This tissue deformation is imaged via high-resolution ultrasound, and analysed via custom code to produce an accurate 3D model of the entire mouse brain, including the tumour region. For human samples, a pre-operative MRI is used to generate a detailed 3D model of the human brain. During surgery, the trapped physical stresses results in a bulge of the dura post craniotomy, which is mapped via Brainlab. These deformations are analysed in an identical fashion to the murine model. We further show that in the brain metastases model, chemotherapy reduces compression stresses by 51%. Further, our technique results in fast processing time (~ 15 minutes), and has the potential to prevent the need for intraoperative MRI based on position simulations. As such, solid stress measurements provide a new class of mechanical biomarkers that can be correlated to clinical outcomes for predictive and prognostic value.
TMOD-06. MALIGNANT TRANSFORMATION OF PLEXIFORM NEUROFIBROMAS IN AN NF1 MUTANT MOUSE MODEL OF SPINAL IRRADIATION
Authors: Nakamura Jean , Laurent Danny , Smith Abbi , Bessler Waylan , Clapp Wade , Horvai Andrew 
Published: December 2020

Abstract

PURPOSE
Understanding the relationships between ionizing radiation (IR) and carcinogenesis, as well as other variables, including genetic background and sex, may minimize risks of transformation from benign to aggressive cancers while maintaining treatment efficacy. We aim to understand the relationship between IR, tumor microenvironments, and effect of sex on malignant transformation using a conditional knockout mouse model of Neurofibromatosis type 1 (NF1).

METHODS
Conditional knockout mouse models of plexiform neurofibromas (PeriCre+; Nf1fl/fl and PeriCre+; Nf1fl/-, n = 172) were treated with 0 Gy; 3 Gy x 5; 3 Gy x 10 of focal, fractionated spinal irradiation (SI) and aged until signs of illness. Histopathological analysis was performed on images from H&E-stained FFPE peripheral nerve tissues.

RESULTS
SI affected the survival of mice in a dose-dependent manner, with a significant decrease at 30Gy (median survival 254 and 218 days for 0 Gy and 30 Gy, p &lt; 0.01, Log-rank test). Male mice had reduced overall survival at 15 and 30 Gy (median survival 368, 278 and 245 days for 0, 15 and 30Gy, p &lt; 0.01, Log-rank test), but not in female mice, indicating sex-specific radiosensitivity. Differences in tumour microenvironments did not affect mouse survival after SI. Histopathological analysis is ongoing (43 out of 76 FFPE samples analyzed). Preliminary findings showed a total of 43 malignancies ranging from plexiform neurofibromas (PNs) to high-grade malignant peripheral nerve sheath tumours (MPNSTs). Increasing histologic aggressiveness was observed in higher SI doses, with high-grade MPNST (2.3%) exclusively found in mice irradiated at 30 Gy.

CONCLUSION
We present a novel mouse model to study malignant transformation in the context of NF1. Preliminary findings suggest increased malignant transformation at high doses of SI.
TMOD-14. CREATION OF A GENETICALLY ENGINEERED MOUSE MODEL OF ANAPLASTIC ASTROCYTOMA DRIVEN BY THE IDH1-R132H ONCOGENE
Authors: Shi Diana , Wang Adam , Gao Wenhua , Khanal Januka , Levitt Michael , Jennings Rebecca , Bonal Dennis , Signoretti Sabina , Nguyen Quang-De , Cahill Daniel , Abdullah Kalil , Ligon Keith , Kaelin William , McBrayer Samuel 
Published: December 2020

Abstract
Despite the high prevalence of IDH1-R132H mutations in lower grade gliomas, the ability to study this mutation in vivo has been hampered by a lack of faithful mouse models. Therefore, we used a CRISPR/Cas9- and AAV-based strategy to create a genetically engineered mouse model (GEMM) of astrocytoma driven by IDH1-R132H that recreates the genetic landscape of human IDH1 mutant astrocytoma. IDH1 mutations in astrocytomas often co-occur with mutations in TP53, ATRX, and either PIK3R1 or PIK3CA. Using human astrocytes immortalized via expression of telomerase (which phenocopies ATRX loss) and HPV E6 and E7 oncoproteins (which phenocopy p53 and pRb loss, respectively), we found that PIK3R1 and IDH1 oncogenes cooperate to promote anchorage-independent cell growth in vitro and orthotopic brain tumor formation in vivo. These data identified a combination of clinically relevant mutations that we hypothesized could be leveraged to cause spontaneous astrocytoma formation in mice. To simultaneously engineer Idh1, Pik3ca, Tp53, and Atrx mutations in mouse brain tissue, we intracranially injected adeno-associated virus (AAV) expressing Cre recombinase and sgRNAs targeting murine Atrx and Tp53 genes into four mouse strains with the following conditional alleles: 1) LSL-Cas9; 2) LSL-Cas9; LSL-Pik3caH1047R, 3) LSL-Cas9; LSL-Idh1R132H, and 4) LSL-Cas9; LSL-Idh1R132H; LSL-Pik3caH1047R. Grade III anaplastic astrocytomas preferentially formed 9-14 months after injecting the mice carrying both the Idh1 and Pik3ca conditional alleles. These astrocytomas harbored all intended mutations, expressed astrocytoma lineage markers, and displayed elevated (R)-2-hydroxyglutarate, the oncometabolite produced by mutant Idh1. To create an additional model with shorter tumor latency, we transplanted glioma stem-like cells derived from our GEMM into recipient mice to produce Idh1 mutant astrocytoma allografts. These allografts provide a tractable platform for preclinical therapeutic studies. Taken together, our findings show that IDH1 and PI3K oncoproteins cooperate to promote gliomagenesis and unveil new genetically faithful mouse models of mutant IDH1-driven astrocytoma.
TMOD-29. ESTABLISHMENT OF PATIENT-DERIVED XENOGRAFTS FROM RARE PRIMARY BRAIN TUMORS
Authors: Kijima Noriyuki , Nakajima Yoshikazu , Kanematsu Daisuke , Shofuda Tomoko , Higuchi Yuichiro , Suemizu Hiroshi , Mori Kanji , Kodama Yoshinori , Mano Masayuki , Sasaki Atsushi , Inoue Takeshi , Hirato Junko , Kishima Haruhiko , Kanemura Yonehiro 
Published: December 2020

Abstract
Patient derived xenografts are essential tools for translational research and preclinical development of novel therapeutic strategies of primary brain tumors. Recent advances in genomics of primary brain tumors revealed molecular classification of primary brain tumors, thus establishment of patient derived xenografts from each subtype of primary brain tumors is urgently needed. However, currently available patient derived xenografts are limited and are from specific subtype of primary brain tumors such as glioblastoma IDH wild type. In this study, we aim to establish patient derived xenografts from primary brain tumors with various molecular characteristics, especially rare primary brain tumors. We got primary brain tumor tissues from patients, dissociated those tissue into single cells, and orthotopically injected those cells into NOD/Shi-scid IL2Rγ KO mouse. We successfully established rare patient-derived xenografts from atypical teratoid rhabdoid tumor and CNS Ewing sarcoma family tumor with CIC alteration, which is recently described as new entity of primitive neuroectodermal tumors of the CNS. We also analyzed histopathological characteristics of these xenografts and found that each xenograft well recapitulated histopathological features of original patients’ resected tumors. These xenografts have advantages for translational research and preclinical development of novel therapeutic strategies for rare primary brain tumors. In addition, further efforts are needed to establish other types of rare primary brain tumors.