bluuebunny/crossref_metadata_2025_split huggingface dataset:
TMET-01. DISRUPTION OF THE SERINE-PRODUCING PATHWAYS SLOWS PROGRESSION OF THE SONIC HEDGEHOG-DRIVEN MEDULLOBLASTOMA
Authors: Tikunov Andrey , Bates Dale , Macdonald Jeff , Gershon Timothy 
Published: November 2022

Abstract
We have found that medulloblastoma metabolism is specialized to promote growth in the CNS and can be targeted for anti-tumor therapy. Prior studies show that serine is scarce in the CNS and that metastatic tumors require serine-producing enzymes to grow in the brain. We analyzed whether medulloblastoma, a primary brain tumor, showed similar serine dependency. We found that Sonic Hedgehog signaling, which induces proliferation in cerebellar granule neuron progenitors in normal development and medulloblastoma formation when hyperactivated, also induces PHGDH and SHMT1, which each catalyze key steps in different serine-production pathways. To define the functional role of PHGDH and SHMT1 in medulloblastomas, we bred mice genetically engineered to develop SHH medulloblastomas with mice carrying deletions of each of these genes. We found that mice with Phgdh-deleted medulloblastomas showed slower progression and longer survival times, compared to mice with Phgdh-intact medulloblastomas. Medulloblastomas with combined deletion of Phgdh and Shmt1 show even slower tumor progression. Stable-isotope flux analysis suggests that different serine-producing pathways compensate for the loss of individual serine-producing enzymes, providing a rationale for the greater anti-tumor effect of disrupting multiple serine-producing enzymes simultaneously. Our data show that serine-producing enzymes can be targeted as a novel approach to medulloblastoma therapy, and underscore the importance of targeting multiple serine-production pathways in complex metabolic interventions.
CNSC-20. PROTEOMIC ANALYSIS OF GLIOMA BIOPSIES SHOWS CONSISTENCY WITH GLIOMA GRADING
Authors: Leu Severina , Boulay Jean-Louis , le Amer Michè , Ritz Marie-Françoise , Halbeisen Florian Samuel , Schmidt Alexander , Buczak Katarzyna , Hutter Gregor 
Published: November 2022

Abstract

BACKGROUND
Glioma are currently classified according to integrated histopathologic and molecular genetic (WHO 2021 classification) as well as methylomic criteria (Capper 2018). However, little is known about proteomic profiling of these subgroups , which may reveal specific biomarkers and pathways potentially activated or inactivated. Material and

METHODS
Eighty-six glioma biopsy samples were prospectively collected from patients undergoing surgery at the University Hospital of Basel between 2017 and 2019. Samples were classified into seven distinct glioma subgroups based on integrated WHO 2016 and methylome profiling criteria. Proteins were extracted from fresh frozen samples devoid of necrosis/haemorrhage and processed for mass spectrometry analysis.

RESULTS
For each tumour sample, a total of 8’855 proteins were identified and quantified. 950 proteins were differentially expressed between at least two glioma subgroups. Strikingly, most significant differences in expression are related to tumor grading rather than to glioma subgroups. Proteins SSDH and GABT, involved in Gamma-aminobutyric acid (GABA) catabolism, show the strongest increase between low grade glioma (LGG) and high grade glioma (HGG), whereas S10AA and ANXA2, part of pathways involved in coagulation, cell adhesion and immune response, appear increased in HGG.

CONCLUSION
Tumor grading turns out to be dominant over genetic and epigenetic criteria for proteomic classification. Most significant protein level variations may represent potential glioma grade-specific therapeutic targets.
TMET-12. HYPERPOLARIZED Δ- [1-13C] GLUCONOLACTONE IMAGING VISUALIZES TERT-ASSOCIATED CHANGES IN DYNAMIC PENTOSE PHOSPHATE PATHWAY METABOLISM IN GLIOBLASTOMA
Authors: Minami Noriaki , Hong Donghyun , Taglang Celine , Batsios Georgios , Gillepspie Anne Marie , Stevers Nicholas , Viswanath Pavithra , Costello Joseph F , Ronen Sabrina 
Published: November 2022

Abstract

BACKGROUND
TERT promoter mutations are a hallmark of glioblastoma (GBM). We recently reported that the expression of TERT and its upstream transcriptional factor, GABPB1, are strongly correlated with redox in GBM with TERT promoter mutations. However, further imaging biomarkers which visualize redox-associated metabolism and TERT expression are needed. Here we demonstrate that 13C magnetic resonance spectroscopy (MRS) of hyperpolarized δ-[1-13C] gluconolactone metabolism is a useful imaging tool to visualize changes in dynamic pentose phosphate pathway (PPP) metabolism that reflect TERT-associated changes in redox in GBM.

METHODS
U251 human GBM cells stably expressing shRNA targeting TERT or GABPB1 were compared to controls. Doxycyclin-inducible shTERT or shGABPB1 U251 cells were also examined. For in vivo studies, cells were injected into immunodeficient rat brains and tumors confirmed by T2-weighted MRI. δ-[1-13C] gluconolactone was polarized using a Hypersense DNP polarizer and injected into live cells or tumor-bearing rats. For cells 13C-MRS was acquired using a 500MHz Agilent spectrometer and analyzed using Mnova and Matlab software. For in vivo13C-MRS studies, spectra were acquired using a 3T Bruker scanner and a spectral spatial echo-planar spectroscopic imaging sequence. Spectral signal to noise was improved using Tensor denoising. Spectra were processed using a custom-written Matlab script.

RESULTS
Hyperpolarized 6-phosphogluconolactone (6PG), the metabolic product from δ-[1-13C] gluconolactone via the PPP, was significantly reduced in TERT or GABPB1 silenced cells compared to control cells in both U251 models. The positive correlation between TERT expression and 6PG level was confirmed by linear regression analysis. Hyperpolarized 6PG production in both tumor models was also significantly reduced in TERT or GABPB1-silenced tumors compared to controls.

CONCLUSION
We successfully visualized TERT-associated changes in dynamic PPP metabolism in a GBM model in cells and in vivo. Hyperpolarized δ-[1-13C] gluconolactone is a potential tool for monitoring TERT expression in GBM with TERT promoter mutations.
TMET-19. DEFINING A NOVEL ROLE FOR CD97 IN REGULATING GBM GLYCOLYTIC METABOLISM
Authors: Ravn-Boess Niklas , Bhowmick Nainita , Hattori Takamitsu , Prakash Varsha , Stephan Gabriele , Frenster Joshua , Bready Devin , Lawson Christopher , Wilcox Jordan , Placantonakis Dimitris 
Published: November 2022

Abstract
Glioblastoma (GBM) is the most common and aggressive primary brain malignancy. Despite multimodal therapy, disease recurrence is inevitable. To identify novel vulnerabilities of GBM, we performed an arrayed CRISPR/Cas9 screen against select adhesion G protein-coupled receptors (aGPCRs), many of which we found to be de novo expressed in GBM. Knockout of CD97, previously implicated in GBM cell migration, produced the most striking proliferative disadvantage in patient-derived GBM cultures (PDGC) among aGPCRs tested. We found high CD97 surface expression in all our PDGCs, while levels remained nearly undetectable in non-neoplastic brain cells, confirming that CD97 is de novo expressed in GBM. Upon shRNA-mediated knockdown of CD97 in PDGCs from all three TCGA transcriptional subtypes, we observed reduced proliferation, as measured by cell cycle analysis. Notably, CD97 knockdown also significantly reduced tumorsphere formation capacity as measured by limiting dilution assays; an effect that was partially rescued upon CD97 overexpression. To elucidate mechanisms of action of CD97, we performed RNA-sequencing and GO pathway enrichment analysis from PDGCs following CD97 knockdown. The top downregulated pathways involved glycolytic metabolism, specifically involving many genes relevant for glucose-6-phosphate (G6P) and fructose-6-phosphate (F6P) processing. Indeed, when we measured metabolite levels under both steady-state and flux conditions using mass spectrometry, we observed an accumulation of G6P and a depletion of most downstream glycolytic and Krebs cycle metabolites upon CD97 knockdown. Furthermore, Seahorse metabolic assays revealed deficits in both glycolytic metabolism and oxygen consumption. We aim to interrogate the activity of specific glycolytic enzymes involved in processing G6P and F6P, pinpointing how these are influenced by CD97 signaling pathways (MAPK or Akt). Overall; our studies suggest a novel role of CD97 in regulating GBM metabolism (Warburg effect), and provide a strong scientific rationale for developing biologics to target CD97 which appears to be universally and de novo expressed in GBM.
CNSC-22. IDENTIFICATION OF MECHANISMS INVOLVED IN THE RECURRENCE OF GLIOBLASTOMA BY TRANSCRIPTOMIC, PROTEOMIC AND SPATIAL PROFILING ANALYSIS
Authors: Shekarian Tala , Ritz Marie-Françoise , Martins Tomas A , Hogan Sabrina , Schmassmann Philip , Burger Bettina , Hutter Gregor 
Published: November 2022

Abstract

BACKGROUND
Glioblastoma (GBM) is the most aggressive and common brain tumor. Despite multimodal therapy which consists of complete tumor removal, chemo- and radiation therapy in most patients, the tumor recurs, and the overall survival of only 14 months remains very poor. To identify tumor-induced immune evasion and treatment resistance mechanisms induced during GBM recurrence, we aimed at characterizing the immunological/molecular changes in patient-matched treatment-naïve and recurrent GBM samples.

MATERIAL AND METHODS
Proteins and total RNA were extracted from 16 patient-matched treatment-naïve (primary) and recurrent GBM fresh frozen tumor samples. The expression of genes involved in brain immune responses were compared using Nanostring panels encompassing Cancer and Neuroinflammation genes. Proteomic was conducted using 6 patient-matched primary and recurrent tumor samples. Immunostaining was performed on a tissue microarray containing matched primary and recurrent tumor samples to identify tumor cells and microglia in order to assess spatial transcriptome of these specific cell types in situ using GeoMX technology. Moreover, correlation analysis of differentially genes and proteins with time to relapse was performed.

RESULTS
Genes significantly differentially expressed between primary and recurrent tumors revealed genes overexpressed in recurrent tumors belonging to “FCGR-mediated Phagocytosis” and “synapse pruning” pathways. Proteomic confirmed the synaptic signalling considering overexpressed proteins. Correlation studies with the time to relapse indicated that higher expression of FCG receptors, complement molecules and synaptic components correlate with shorter time to relapse, sharing similarities with neurodegenerative/cognitive disorders such as schizophrenia and Alzheimer disease. The source for these molecules is under evaluation with the spatial transcriptomic analysis.

CONCLUSION
Upregulation of specific proteins such as FCG receptors, complement and synaptic components may result in more favourable conditions for tumor regrowth and may therefore be identified as negative predictors patient outcome. Interfering with these molecules may represent novel therapeutic options after initial GBM surgery.
TMET-25. MEK/ERK AND AMPK SIGNALING DICTATE VULNERABILITY TO FATTY ACID DESATURATION INHIBITION IN GLIOBLASTOMA
Authors: Sammarco Alessandro , Eyme Katharina , Jha Roshani , Mnatsakanyan Hayk , Neustadt Rudolph , Moses Charlotte , Alnasser Ahmad , Tardiff Daniel , Su Baolong , Williams Kevin , Bensinger Steven J , Chung Chee Yeun , Badr Christian E 
Published: November 2022

Abstract
Lipid metabolism is altered in many cancers. Glioma stem cells (GSCs), a subpopulation that contributes to molecular heterogeneity and therapeutic resistance of glioblastoma (GBM), are highly dependent on de novo lipid synthesis (DNLS) for energy production and proliferation. Although DNLS is a promising therapeutic target in GBM and other tumors, the molecular mechanisms that determine the dependence on DNLS remain largely unknown. In this study, we aimed to define the genetic drivers of vulnerability to DNLS inhibitors in GSCs and to explore the molecular mechanisms dictating resistance to a brain-penetrant inhibitor of stearoyl-CoA desaturase (SCD), a key enzyme involved in DNLS. Using an astrocyte model of gliomagenesis, we discovered that activation of RAS/MEK/ERK promotes vulnerability to DNLS inhibitors, whereas defective MEK/ERK activity drives resistance to SCD inhibition. Also, we identified the energy sensor AMPK as a downstream target of MEK/ERK and discovered an adaptive metabolic reprograming of GSCs dictated by AMPK activation. Shotgun lipidomics revealed that constitutive AMPK activation protects from SCD inhibition-induced lipotoxicity and endoplasmic reticulum stress and preserves the ability of transformed cells to form brain tumors in mice. Mechanistically, we found that AMPKs ability to circumvent lipotoxicity was achieved by altering the lipid composition and saturation state of cell membranes, and channeling potentially lipotoxic fatty acids into lipid droplets (i.e., lipid storage). These results strongly suggest that the efficacy of DNLS-targeted therapy can be dictated by the MEK/ERK and AMPK signaling status of GSCs. These data also suggest that MEK/ERK and AMPK signaling could be used as predictive biomarkers to understand which GBM patients are likely to respond to DNLS-targeted therapy. In conclusion, our data uncover an unexpected mechanism by which GSCs avoid a metabolic vulnerability and provides a rationale for integrating DNLS-targeted therapies for GBM treatment.
TMET-28. A METHOD FOR EX VIVO STABLE ISOTOPE TRACING IN SURGICALLY EXPLANTED GLIOMA ORGANOIDS
Authors: El Shami Mohamad , Savani Milan , Gattie Lauren , Hicks William , Richardson Timothy , McBrayer Samuel , Abdullah Kalil 
Published: November 2022

Abstract
Stable isotope tracing is a powerful method for elucidating tumor metabolism. While in vivo stable isotope tracing is one of the most faithful approaches for studying metabolic activity under pathophysiologically relevant conditions, this method is technically challenging and costly to perform. We therefore sought to optimize an ex vivo stable isotope tracing approach under conditions that recapitulate the in vivo tumor microenvironment (TME), focusing on three key features: cellular heterogeneity, nutrient availability, and oxygenation. Surgically eXplanted Organoids (SXOs) are 3-dimensional glioma models that retain the diverse cell populations in human brain tumors. We tested whether glioma SXOs can be cultured in Human Plasma-Like Medium (HPLM) under brain-relevant oxygen levels (5%) for the purpose of stable isotope tracing. We cultured SXOs in conventional media, HPLM for 24 hours, or HPLM for 120 hours under 5% oxygen. We observed no significant differences in markers of tumor architecture, cellular proliferation rates, or stemness profiles. We then performed stable isotope tracing in SXOs cultured in HLPM at 5% oxygen. We acclimated SXOs to HPLM for 24 hours or 120 hours before replacing media with either HPLM (unlabeled) or HPLM with 15N2 glutamine (labeled). 15N isotopic labeling patterns in various metabolites were compared between the 24- and 120-hour conditions by linear regression. These patterns were strongly concordant over time (r2 = 0.9544), indicating that steady-state metabolic fluxes are stable in these cultures. We then analyzed intermediates that occupy key nodes in amino acid metabolism, redox homeostasis, and nucleotide synthesis pathways and found similarly strong concordance between labeling patterns at the individual metabolite level. Our findings outline an effective approach for conducting ex vivo stable isotope tracing in heterocellular glioma models under conditions that mimic the TME. This approach may complement and enhance analogous in vivo methods to provide new insights into glioma metabolism.
TMET-31. CHOLESTEROL ESTERIFICATION IN A PATIENT WITH DIFFUSE ASTROCYTOMA PROGRESSING TO GLIOBLASTOMA
Authors: Ijare Omkar , Baskin David , Pichumani Kumar 
Published: November 2022

Abstract
Metabolic profiles of diffuse astrocytoma and recurrent tumor tissues were compared in a patient with diffuse astrocytoma who showed tumor recurrence and progressed to glioblastoma (GBM). The patient presented with a diffuse infiltrating tumor and was operated in April 2018. The pathology report showed that the tumor was a diffusely infiltrating astrocytoma with Ki-67 of 5% (WHO Grade II). In addition, the molecular testing results indicated that the tumor was IDH1wt with TERTp mutation and showed an indeterminate MGMT promoter status. Given diffuse brain involvement and IDH1wt status, an aggressive behavior was suggested for this tumor. After ~3.5 years (January 2022), the MRI examination showed tumor recurrence and the patient underwent craniotomy for the resection of the recurrent tumor. The pathology report showed a high-grade glial neoplasm composed of ovoid tumor cells with nuclear pleomorphism (GBM, WHO Grade IV). We collected the tumor specimens from both surgeries and compared the metabolic profiles using 1H NMR spectroscopy. The tumor specimens were extracted using methanol-chloroform (2:1 ratio) extraction-method. The methanol-layer of the recurrent tumor showed decreased levels of N-acetylaspartate (NAA) and myo-inositol, and the elevated levels of phosphocholine (PC) and glycine, compared to the diffuse astrocytoma. Decreased NAA and elevated PC and glycine are metabolic features of high-grade gliomas, suggesting progression of the low-grade astrocytoma to GBM. The chloroform-layers of both tumors showed the presence of cholesterol; however, cholesteryl ester (CE) was detected only in the recurrent tumor (GBM). The ratio of CE to cholesterol was 0.44, suggesting that 44% of cholesterol pool was esterified in GBM. Cholesterol esterification is dysregulated in GBM and targeting this metabolic pathway using AcylCoA:cholesterol acyltransferase (ACAT) inhibitors would be promising in the treatment of GBM and astrocytomas with TERTp mutation which transform to GBM.
TMET-33. ELONGATION CONTROL OF MRNA TRANSLATION SUPPORTS GROUP 3 MEDULLOBLASTOMA ADAPTATION TO NUTRIENT DEPRIVATION
Authors: Delaidelli Alberto , Yao Betty , Negri Gian Luca , Huang Yue Zhou , Huang Albert , Leprivier Gabriel , Sorensen Poul 
Published: November 2022

Abstract
Group 3 affiliation and MYC genetic amplification are associated with poor life expectancy and substantial morbidity in children suffering from medulloblastoma (MB). However, the high metabolic demand induced by MYC-driven transformation sensitizes MYC-overexpressing MB to cell death under conditions of nutrient deprivation (ND). Additionally, MYC-driven transformation is known to promote mitochondrial oxidative phosphorylation (OXPHOS). We previously reported that eukaryotic Elongation Factor Kinase 2 (eEF2K), the master regulator of mRNA translation elongation, promotes survival of MYC-overexpressing tumors under ND. Interestingly, eEF2K is overexpressed in MYC-driven MB and our preliminary proteomics data highlight large-scale alterations in OXPHOS components affecting eEF2K deficient MB cells. We therefore hypothesized that eEF2K activity is required for the selective translation of mRNAs needed for efficient OXPHOS, and for the progression of MYC-driven MB. We performed Multiplexed enhanced Protein Dynamic Mass Spectrometry in eEF2K knockdown MYC-overexpressing D425 MB cells to identify mRNAs selectively translated upon eEF2K activation. Messenger RNAs encoding multiple (9 out of 10 detected) components of the mitochondrial OXPHOS pathway are selectively translated upon eEF2K activation. Inactivation of eEF2K by genetic KO leads to the disassembly of electron transport chain (ETC) complexes I-IV without affecting mRNA levels of their respective components. Consistently, eEF2K KO MB cells display decreased mitochondrial membrane potential and 20% increased proton leak thorough the mitochondrial membrane. In addition, eEF2K inactivation results in increased Group 3 MB cell death under ND and doubles survival of MB bearing mice fed with calorie restricted diets (p&lt; 0.05). Control of mRNA translation elongation by eEF2K is critical for mitochondrial ETC complex assembly and efficient OXPHOS in MYC-overexpressing MB, likely representing an adaptive response by which MYC-driven MB cells cope with acute metabolic stress. Future therapeutic studies will aim to combine eEF2K inhibition with caloric restriction mimetic drugs as eEF2K activity appears critical under metabolic stress conditions.
TMET-34. RADIATION METABOLOMICS IN PRIMARY HUMAN MENINGIOMA AND SCHWANNOMA: EARLY EXPERIENCE AND INITIAL RESULTS
Authors: Dougherty Mark , Taylor Eric , Hansen Marlan 
Published: November 2022

Abstract

Introduction
Meningiomas and schwannomas account for 45% of primary CNS tumors. Yet when surgery and radiation fail, no further treatments exist. Metabolomics has been used to discover new cancer therapies; however, to date few have used metabolomics to study meningiomas and schwannomas. Here we present initial results and lessons learned from this novel endeavor.

METHODS
Primary tumors were obtained from patients during surgery and immediately taken for culturing or xenograft implantation. Upon reaching &gt;90% confluence, cultures were treated with 0gy, 3gy, 10gy, or 20gy gamma radiation, then flash frozen 6 or 72 hours post-treatment. Xenograft tumors were implanted in nude mice. MRI 4 weeks post-implantation confirmed tumor viability. Mice were then given 10gy, 20gy, or sham radiation treatment. Xenografts were harvested 72 hours post-treatment. Metabolites were measured with a ThermoISQ gas chromatography-mass spectrometer.

RESULTS
Eleven meningiomas and nine schwannomas were successfully cultured. Unsupervised hierarchical clustering of cultures demonstrated greater influence from tumor of origin than from radiation. Univariate analysis of schwannoma xenografts demonstrated elevated ornithine following radiation (fold change 1.62; P = 0.008). However, principal component analysis did not show significant between-group differentiation. Orthotopic meningioma xenografts did not produce sufficient tissue for metabolomics; however, subsequent subcutaneous implants have been successful (data forthcoming).

CONCLUSION
Standard cell cultures did not reveal significant metabolic changes following radiation; it is unclear whether this was due to culture technique or inter-tumor heterogeneity. In radiated schwannoma xenografts, elevated ornithine may implicate related pathways such as ornithine decarboxylase-mediated polyamide synthesis for DNA double-strand break repair. Compared to other ‘-omics’ studies, metabolomics requires more tissue per sample ( &gt;10mg) and is more sensitive to environmental conditions. Thus, large sample sizes are needed to detect significant changes, and xenografts are likely superior to cell culture. Future plans include increased xenograft sample size and stable isotope tracing for pathway analysis.