bluuebunny/crossref_metadata_2025_split huggingface dataset:
CBMT-34. MODULATING DIETARY TRYPTOPHAN OR GUT MICROBIOTA LEVELS DOES NOT IMPROVE THE EFFICACY OF COMBINED TREATMENT WITH RADIATION, ANTI-PD-1 mAb, AND AN IDO1 ENZYME INHIBITOR IN A MODEL OF GLIOBLASTOMA
Authors: Ladomersky Erik , Zhai Lijie , Lauing Kristen , Qian Jun , Bell April , Otto-Meyer Sebastian , Savoor Rohan , Wainwright Derek 
Published: November 2019

Abstract

INTRODUCTION
Immunotherapy has failed to improve glioblastoma (GBM) patient survival in all phase III clinical trials to-date. We recently discovered that a combination immunotherapeutic approach that simultaneously administers radiation (RT), anti-PD-1 mAb, and an IDO1 enzyme inhibitor resulted in long-term survival of ~40% of mice intracranially-engrafted (i.c.) with syngeneic GBM (Ladomersky et al., 2018; CCR). In the same study we discovered that the survival benefit depended on the inhibitor of IDO1 in non-GBM cells. Our current study further interrogates the role of non-GBM cell IDO1 and its role in the brain tumor microenvironment.

METHODS
Mice were engrafted with 2×105 GL261 cells and treated at 14 days post-intracranial injection (i.c.) with RT and/or anti-PD-1 mAb, followed by sampling tissue at 1- and 7-days post treatment. Questioning whether dietary supplementation of Trp would overcome the immunosuppressive activity of the Trp metabolic enzyme, IDO1, mice were placed on ad libitum dietary formulations containing 0–500% normal daily intake. Since Trp is only taken in through the diet, and since bacteria also metabolize Trp, we questioned the role of the gut microbiome on immunotherapy of GBM by injecting mice with an antibiotic cocktail beginning 4 weeks prior to i.c.-injection.

RESULTS
Both RT and anti-PD-1 treatment increase IDO1 expression in the GBM-draining cervical lymph nodes at day 1 post-ic. (P< 0.001). Mice receiving normal (100%) dietary Trp, but no other supplementation levels, achieve a long-term survival benefit (P< 0.001). Unexpectedly, the depletion of gut microbiota has no effect on therapeutic efficacy (P=0.4808).

CONCLUSION
These data suggest that although both systemic and tissue-selective alterations in Trp metabolism occur after RT and anti-PD-1 therapy, modulating dietary Trp levels or gut microbiota composition does not alter treatment efficacy against GBM.
CBMT-35. METABOLIC REWIRING BY ONC201/TIC10 AND 2-DEOXYGLUCOSE HAS SYNERGISTIC ANTI-GLIOBLASTOMA ACTIVITY
Authors: Pruss Maximilian , Dwucet Annika , Hlavac Michal , Kast Richard , Westhoff Mike-Andrew , Wirtz Rainer , Halatsch Marc-Eric , Siegelin Markus , Karpel-Massler Georg 
Published: November 2019

Abstract

BACKGROUND
Metabolic dysregulation is a common feature of cancers such as primary brain malignancies. In this work we examined whether a rewiring of the metabolome by a multi-targeting approach would induce enhanced anti-neoplastic activity in glioblastoma.

METHODS
Preclinical testing of a combined treatment with ONC201/TIC10 and 2-Deoxyglucose was performed in established and primary cultured glioblastoma cells. Extracellular flux analysis was used to determine real-time effects on OXPHOS (OCR) and glycolysis (ECAR). Expression of respiratory chain complexes was analysed by Western blotting. Biological effects on tumor formation were tested in patient-derived model systems on the chorion allantoic membrane (CAM). Protein array analyses were performed to determine effects on phospho protein kinase expression.

RESULTS
Treatment with ONC201/TIC10 leads to impaired mitochondrial respiration and a dose-dependent increase of glycolysis. ONC201/TIC10 combined with 2-Deoxyglucose, induces a state of energy deprivation characterized by a significant decrease in ATP levels. On the molecular level, pAMPK α1 was significantly up-regulated while a hypo-phosphorylation signature was noted including mTOR signaling, src family kinases and receptor tyrosine kinases such as EGFR and PDGFR-β. As a result, synergistic anti-proliferative and anti-migratory effects were observed among established and primary cultured glioblastoma cells. In addition, tumor formation on the CAM was significantly impaired following the combination treatment.

CONCLUSIONS
Simultaneous treatment with ONC201/TIC10 and 2-Deoxyglucose causes a reprogramming of the metabolic circuitry and results in a synergistic anti-glioblastoma activity. Since both agents were tested in clinical trials with good tolerability, and they both penetrate the blood-brain barrier, further clinical evaluation of this therapeutic strategy seems promising.
CBMT-42. A GENOME-WIDE CRISPR-Cas9 SCREEN FOR GENES REGULATING QUIESCENT-LIKE STATES IN GLIOBLASTOMA
Authors: Feldman Heather , Mihalas Anca , Plaisier Christopher , Patel Anoop , Paddison Patrick 
Published: November 2019

Abstract
Current standard of care therapy for glioblastoma (GBM) includes cytoreduction followed by ablative therapies that target rapidly dividing cell types. However, non-cycling, quiescent-like states (G0 phase cells) are present in both normal tissue and tumors and play important roles in maintaining heterogeneity and cellular hierarchies. The presence of quiescent-like/G0 states therefore represents a natural reservoir of tumor cells that are resistant to current treatments. Quiescence or G0 phase is a reversible state of “stasis” cells enter in response to developmental or environmental cues. However, it remains largely unclear to what degree or by what mechanisms tumor cells enter into or exit from quiescent-like states. To gain insight into how glioblastoma cells might regulate G0-like states, we performed a genome-wide CRISPR-Cas9 screen in patient-derived GBM stem-like cells (GSCs) harboring a p27-mVenus reporter construct, which is stabilized when cells enter a G0-like state. By assaying p27 reporteractivity, we were able to identify sgRNAs enriched in p27hipopulations and, which upon retest, trigger a G0-like arrest in GSCs. Among the top screen hits were members of the Tip60/KAT5 histone acetyltransferase complex, including KAT5 itself. Remarkably, we show that downregulation of KAT5 in vitro and in vivo dramatically increases the pool of cells in G0-like states in GSC cultures and GSC-induced tumors. Using single cell RNA-sequencing, we show that this cell state is characterized by gene expression signatures similar to those found in non-dividing subpopulations of GBM tumors and quiescent neural stem cells. In addition, we perform in-depth molecular and phenotypic characterization of these induced G0-like states, including epigenetic and metabolic profiles. These suggest a key role for KAT5 in regulating genes related to protein synthesis. In summary, our results suggest that Tip60/KAT5 activity plays key roles in G0 ingress/egress for GBM tumors and may provide novel therapeutic opportunities.
CBMT-43. INTEGRATED LIPIDOMIC AND MOLECULAR ANALYSIS REVEALS A SUBSET OF GLIOBLASTOMA VULNERABLE TO FERROPTOSIS
Authors: Morrow Danielle , Bayley Nicholas , Williams Kevin , Muranaka Hayato , Prins Robert , Liau Linda , Cloughesy Timothy , Bensinger Steven , Nathanson David 
Published: November 2019

Abstract
Cancers, including the universally lethal glioblastoma (GBM), have reprogrammed lipid metabolism to fuel tumor growth. However, the molecular alterations responsible for aberrant lipid metabolism, and the potential for identifying new therapeutic opportunities are not fully understood. To systematically investigate the GBM lipidome, we performed integrated transcriptomic, genomic and shotgun lipidomic analysis of a library of molecularly diverse patient-derived GBM cells (n=30). Using this comprehensive approach, we discovered two GBM sub-groups defined by their combined molecular and lipidomic profile. Polyunsaturated fatty acids (PUFAs) were among the most significant lipids that distinguished these two groups of GBM tumors. Intriguingly, this lipid metabolic phenotype was associated with heightened sensitivity to ferroptosis – a newly discovered form of regulated cell death. As PUFA oxidation is a critical feature of ferroptosis, our findings suggest a novel association between specific molecular signatures of GBM, lipid metabolism and ferroptosis. This relationship may present a new therapeutic opportunity to target ferroptosis in a molecularly-defined subset of GBMs.
CBMT-44. COMPREHENSIVE CHARACTERIZATION OF THE INTRINSIC APOPTOTIC MACHINERY REVEALS THE MOLECULAR BLOCKS RESPONSIBLE FOR RESISTANCE TO CELL DEATH IN GLIOBLASTOMA
Authors: Fernandez Elizabeth , Mai Wilson , Bayley Nicholas , Liau Linda , Cloughesy Timothy , Letai Anthony , Nathanson David 
Published: November 2019

Abstract
Conventional therapies (e.g., temozolomide (TMZ), Irradiation (IR)) transiently halt tumor growth of glioblastoma (GBM) but fail to induce cell death through apoptosis. Consequently, the inability to kill GBM tumor cells ultimately leads to disease progression and a poor patient survival. The precise molecular mechanisms by which GBM are refractory to apoptosis remain enigmatic. We preformed BH3 profiling to functionally characterize the intrinsic apoptotic machinery and define the molecular ‘blocks’ that obstruct GBM apoptosis under both basal and treatment states. Using a molecularly diverse panel of freshly purified patient tumors, patient-derived neurospheres and patient-derived orthotopic xenografts, we identified that nearly all GBMs have two anti-apoptotic blocks, BCL-xL and MCL-1, which are essential for GBM survival in an untreated state. TMZ or IR (TMZ/IR) disabled the MCL-1 block in a subset of GBMs, leaving tumors exclusively dependent on BCL-xL for survival. Mechanistic studies revealed that TMZ/IR treatment induced p53-dependent expression of the pro-apoptotic protein, PUMA, which subsequently bound to and neutralized MCL-1. Consequently, pharmacological inhibition of BCL-xL in combination with TMZ/IR initiated intrinsic apoptosis and was synergistically lethal in p53 wild-type GBM. These studies identify the existence of two anti-apoptotic proteins that are critical for GBM survival, which can be therapeutically exploited in a molecularly defined subset of GBMs for tumor eradication.
CBMT-49. OXALOACETATE ALTERS GLUCOSE METABOLISM IN GLIOBLASTOMA: 13C ISOTOPOMER STUDY
Authors: Ijare Omkar , Conway David , Cash Alan , Baskin David , Pichumani Kumar 
Published: November 2019

Abstract
Anhydrous enol-oxaloacetate (AEO) has demonstrated the ability to enhance neuronal cell bioenergetics and activate brain mitochondrial biogenesis. Since oxaloacetate has demonstrated positive effects on brain bioenergetics in neurodegenerative diseases we have begun to investigate whether AEO may also have a positive effect on the altered cellular metabolism found in cancer cells, particularly Glioblastoma multiforme. The “Warburg effect” describes an abnormal metabolic state in cancer, distinct from normal tissue, in which energy is generated through enhanced conversion of pyruvate to lactate even in the presence of oxygen during glycolysis. Oxaloacetate (OAA) is a key anaplerotic substrate that is required to maintain TCA cycle flux. The role of oxaloacetate supplementation on the energy metabolism is not known in cancer cells. Goal of this study is to investigate the changes in metabolic fluxes in glucose metabolism with and without the presence of OAA in patient-derived GBM cells. We use GC-MS based 13C isotopomer analysis for this study. GBM cells are grown in 15mM glucose containing DMEM medium supplemented with 2mM oxaloacetate for 10 days. 6 hours prior to harvesting, [U-13C]glucose is introduced to the medium. 13C isotopomer analysis of GC-MS data showed that OAA supplementation for 10 days drastically decreased Warburg glycolysis by reducing 13C labeling (M+3) by 19.7% and 48.8% in pyruvate and lactate pools respectively in comparison with cells not treated with OAA. M+3 13C labeled pyruvate entered TCA cycle via acetyl-CoA, where we also observed reduced levels of M+2 13C labeled citrate (20.5%) and glutamate (23.9%) isotopomers. Pyruvate can also enter TCA cycle via pyruvate carboxylation pathway and this activity was also found to be slightly decreased in the OAA treated cells. All the differences were statistically significant. These results indicate that OAA can be used to alter bioenergetics of GBM cells, specifically glucose oxidation.
CSIG-01. NA-K-CL COTRANSPORTER PROTEIN IN THE PATHOGENESIS OF LOW-GRADE GLIOMAS
Authors: Luo Lanxin , Guan Xiudong , Begum Gulnaz , Ding Dawei , Kohanbash Gary , Sun Baoshan , Sun Dandan 
Published: November 2019

Abstract
Low-grade gliomas (LGGs) present a high incidence of epilepsy, infiltrative growth, resistance to therapy, and high risk of transforming into high-grade gliomas (HGGs). Temozolomide (TMZ) is a conventional chemotherapy drug for adjuvant treatment of patients with high-risk LGGs. Na+-K+-2Cl- co-transporter 1 (NKCC1) regulates cell volume and intracellular Cl- concentration which promotes glioma cell migration, resistance to TMZ, and tumor-associated epilepsy. Our new bioinformatic analysis of TCGA and CGGA datasets shows that LGGs expressed higher SLC12A2 gene mRNA (encoding NKCC1 protein) than HGGs, which was confirmed at NKCC1 protein levels. In syngeneic mouse glioma models with intracranial transplantation of two different mouse glioma cell lines (GL26 and SB28), we detected upregulated NKCC1 protein expression in glioma tumor cells as well as in peri-tumor reactive astrocytes in response to TMZ monotherapy. This provided a rationale for us to test the efficacy of combining NKCC1 pharmacological blockade with TMZ on improving therapeutic outcomes. 5-day combination therapy of TMZ with potent NKCC1 inhibitor bumetanide reduced tumor proliferation, potentiated TMZ-mediated apoptosis, and decreased peri-tumor reactive astrocyte formation, which collectively led to suppressed tumor growth and prolonged survival of tumor-bearing mice. Taken together, these results demonstrate that NKCC1 protein plays multifaceted roles in regulating the pathogenesis of LGGs and its blockade presents therapeutic potentials for reducing TMZ-mediated resistance.
CSIG-02. TARGETING CONSTITUTIVE ACTIVATION OF THE EGFR-STAT1 AXIS DECREASES PROLIFERATION OF MENINGIOMA TUMOUR CELLS
Authors: Ferluga Sara , Baiz Daniele , Ercolano Emanuela , Sharma Vikram , Dunn Jemma , Kurian Kathreena M , Hilton David , Oliver Hanemann Clemens 
Published: November 2019

Abstract
Meningiomas are the most frequent primary brain tumours of the central nervous system. The standard of treatment is surgery and radiotherapy, but the lack of knowledge of their tumorigenesis contributes to delay the development of additional therapeutic options. We found STAT1 widely overexpressed in meningioma tumours and in patient-derived meningioma cells but not in the corresponding healthy controls. The protein showed a constitutive phosphorylation on both phosphosites (Y701 and S727), which was not dependent on the JAK/STAT pathway. STAT1 knocked down resulted in a significant reduction of cellular proliferation, showed as a decrease in Ki67-positive cells and Cyclin D1, and deactivation of AKT and ERK 1/2. By studying STAT1 binding partners we isolated a complex composed by STAT1, STAT2, PRMT5 and MEP50. As PRMT5 is known to interact with EGFR, we tested the tyrosine kinase and found that EGFR was constitutively active in meningioma and was responsible for the aberrant phosphorylation of STAT1 on both phosphosites. We tested different drugs inhibiting of EGFR phosphorylation, Canertinib was most effective. It caused a significant reduction in meningioma cells proliferation and a reduction of overall levels of Cyclin D1, phospho- AKT and phospho-ERK 1/2. Hence, STAT1 constitutive phosphorylation, initiated by EGFR activation, is responsible for inducing a positive feedback loop causing its own overexpression and consequently an increased proliferation of the tumour cells. These findings underline a pivotal role of the EGFR and STAT1 axis in meningioma and provides the rationale for further studies aiming to identify novel and effective therapeutic options.
CSIG-06. DEPATUXIZUMAB MAFODOTIN (ABT-414)-INDUCED GLIOBLASTOMA CELL DEATH REQUIRES EGFR OVEREXPRESSION, BUT NOT PHOSPHORYLATION
Authors: von Achenbach Caroline , Blot Vincent , Weiss William , Weller Michael 
Published: November 2019

Abstract
Glioblastomas commonly (40%) exhibit epidermal growth factor receptor (EGFR) amplification, half of these tumors carry the EGFRvIII deletion variant characterized by an in-frame deletion of exons 2–7, resulting in constitutive EGFR activation. Although EGFR tyrosine kinase inhibitors had only modest effects in glioblastoma, novel therapeutic agents targeting amplified EGFR or EGFRvIII continue to be developed. Depatuxizumab mafodotin (ABT-414) is an antibody drug conjugate consisting of the monoclonal antibody 806 and, as its toxic payload, monomethyl auristatin F, designed to target EGFR-overexpressing tumor cells. Since long-term glioma cell lines and patient-derived glioma-initiating cell lines appeared to express too little EGFR in vitro to be sensitive to ABT-414, we generated glioma sublines overexpressing EGFR or EGFRvIII to explore the determinants of ABT-414-induced glioma cell death. Overexpression of EGFRvIII induces sensitization to ABT-414 more readily than overexpression of EGFR. There is no bystander killing of cells devoid of EGFR or EGFRvIII expression. Surprisingly, either exposure to EGF or to EGFR tyrosin kinase inhibitors reduce EGFR protein levels and are thus no strategies to promote ABT-414-induced cell killing. Furthermore, glioma cells overexpressing kinase-dead EGFR or EGFRvIII retain binding of mAb 806 and sensitivity to ABT-414, allowing to dissociate EGFR phosphorylation from the emergence of the “active” EGFR conformation required for ABT-414 binding. The combination of EGFR-targeting antibody drug conjugates with EGFR tyrosine kinase inhibitors carries a high risk of failure. Promoting mere EGFR expression rather than phosphorylation should result in glioblastoma cell sensitization to ABT-414 or related agents.
CSIG-09. PROTEOMIC ANALYSIS OF MENINGIOMA CELL-DERIVED EXTRACELLULAR VESICLES: FIRST OF A KIND
Authors: Ricklefs Franz , Fuh Manka , Maire Cecile , Holz Mareike , Kolbe Katharina , Westphal Manfred , Schlüter Hartmut , Lamszus Katrin 
Published: November 2019

Abstract

BACKGROUND
Extracellular vesicles (EVs) play an important role in cell-cell communication in different types of tumors, carrying multiple layers of biological functional molecules, including proteins, RNA, DNA and lipids. Their implication as biomarkers in tumor disease is under current investigation. We previously showed that EVs in glioblastoma reflect the tumor subtype and that glioblastoma patients have elevated circulating particle counts. Regarding to meningioma, it is not known to what extent these usually benign tumors secrete EVs and how these EVs reflect the tumor. Here we report the first study that analyzed meningioma cell-derived EVs.

METHODS
Meningioma tissue, short-term cell cultures and cell culture-derived EVs (menEVs), (n=4) were analyzed by global mass-spectromety, immunoblotting and imaging flow cytometry and compared to EVs from glioblastoma short-term cell cultures (gEVs), (n=4). Plasma EVs from meningioma patients (n = 12) were analyzed for their tetraspanin marker expression (CD9, CD81 and CD63). EVs were further analyzed by nanoparticle analysis (NTA) and electron microscopy.

RESULTS
menEVs were 110-140nm in size and exhibited vesicular structures by electron microscopy. We identified 269 proteins in menEVs through mass spectometry. 45 proteins were upregulated in menEVs compared to short-term cell culture and original tumor tissue. 99 proteins were exclusively found in menEVs compared to gEVs, with osteopontin being the top highly expressed protein within the mEV fraction. Both meningioma and glioblastoma patients have elevated circulating plasma EV counts (p< 0.01), as measured by NTA.

CONCLUSION
The increase in circulating plasma EVs in meningioma patients suggests that tumor cell-derived EVs augment the pool of circulating EVs and could be utilized to obtain information on the tumor by liquid biopsy. Osteopontin is known to be expressed at high levels in meningiomas and its association with menEVs may facilitate isolation of circulating meningioma-specific EVs for analysis.
CSIG-20. EFFECT OF TUMOR-TREATING FIELDS (TTFIELDS) ON EGFR PHOSPHORYLATION IN GBM CELL LINES
Authors: Luisa D’Angelo Maria , Piffaretti Deborah , Burgio Floriana , Chiappini Alessio , Marchi Francesco , Reinert Michael 
Published: November 2019

Abstract
Glioblastoma (GBM) is the most common high grade and most devastating brain tumor among adults. About 50% of GBM express EGFR (epidermal growth factor receptor) and from these another 50% the mutated form EGFRvIII which is associated with a more aggressive disease and poorer prognosis. We have previously shown that expression of different status of EGFR in GBM cell lines drives the 5-ALA induced fluorescence downstream by influencing the rate-limiting enzyme heme oxygenase-1 (HO-1) probably via PI3K/Akt signalling. We demonstrated that 5-ALA-induced protoporphyrin IX (PpIX) fluorescence is pharmacologically influenced by adding different drugs such as deferoxamine (DFO) and tin-protoporphyrin (SnPP), which are an iron chelator of Fe2+ and an inhibitor of HO-1 respectively, or genistein that promotes PpIX accumulation via functional repression of ABCG2 (ABC transporter G2). Our aim is to increase these pharmacological effects on PpIX fluorescence using tumor-treating fields (TTFields). TTFields, a new therapeutic technology for treating newly diagnosed or recurrent GBM, is able to suppress the growth of cancer cells destabilising microtubule elongation and increasing membrane permeability. Interestingly, TTFields, like as other destabilising drugs and compounds, is able to inhibit the phosphorylation of EGFR and subsequent downregulation of EGFR-induced signalling acting for example on the mechanism that regulate the HO-1 activity. Here, we investigate the effects of TTFields on glioma cells, with different EGFR status and consequently different PpIX fluorescence. Exposure to TTFields during or after pharmacological treatments may represent a novel strategy to block or diminish the phosphorylation of EGFR to ameliorate the visualization of PpIX fluorescence in patients where it is not enough to ensure a safe and precise removal of the tumor bulk. In fact, if a combination of TTFields and drug treatment should give the desired results, this strategy could be applied on patients before being subjected to surgical resection.