bluuebunny/crossref_metadata_2025_split huggingface dataset:
P01.02.B CLONAL ARCHITECTURE, 3D TUMOR GROWTH, TRANSCRIPTIONAL STATES AND IN-VITRO DRUG RESPONSE IN GLIOBLASTOMA
Authors: Song Y , Miller-Michlits Y , Greutter L , Nenning K , Kiesel B , Loetsch-Gojo D , Redl V , Gesperger J , Jana Breu J , Nam D , Widhalm G , Sa J K , Woehrer A 
Published: October 2024

Abstract

BACKGROUND
Glioblastoma (GBM) is one of the deadliest types of cancer with median overall survival of little more than a year. It is the prototypic example of a molecularly diverse tumor associated with multiple subclones and high transcriptional plasticity. We here set out to map the clonal architecture and link it with growth patterns, transcriptional states and drug response in-vitro.

MATERIAL AND METHODS
Seventy-eight multiregional samples from 24 GBM patients (F/M ratio = 0.60, median age= 65.5ys) were profiled using deep whole-exome sequencing with a nominal coverage of &gt;300x. Clonal complexity was assessed by the Euclidean distance based on somatic mutations between regions. Molecular distance was integrated with 3D neuronavigation-based physical distances and patient survival. Fresh tumor slices of 8 patients were cultured in minimal essential media and treated with 4.2Gy plus 200µM temozolomide for 48h. Spatial Transcriptomics was performed on matched tumor slices with /without treatment.

RESULTS
Integrative characterization revealed two evolutionary trajectories termed expansive and stochastic diversification based on the correlation between physical and molecular distance (RE=0.6, RS=-0.2). High molecular distance was associated with reduced PFS (R=-0.5538, p=0.026). Tumors with expansive evolution demonstrated volume-based growth that directly correlated with increased molecular diversification, while stochastic models showed irregular and discordant patterns. Patients with stochastic evolution were characterized by unfavorable prognosis (p= 0.35) and tumors appeared more frequently in the frontal lobe. In-vitro drug response varied considerably between patients based on cell proliferation (minus 0.5-13.8%), apoptosis (30.1-44.4%), and viability (minus 7.8-34.1%). Under treatment an increase of mesenchymal states with peculiar redistribution of transcriptional patterns and increased clonal complexity was observed.

CONCLUSION
In sum, we report on an ongoing study in glioblastoma that seeks to link the clonal architecture with tumor growth, transcriptional patterns and in-vitro drug response. Our study will provide new insights into the clinical relevance of the clonal composition of this deadly cancer.
P01.03.A DECIPHERING THE TUMOUR ECOSYSTEM IN GLIOMAS USING SPATIAL TRANSCRIPTOMICS
Authors: Seferbekova Z , Calafato D , Rukhovich G , Patel A , Ritter M , Ippen F , Suwala A , Sahm F , Gerstung M 
Published: October 2024

Abstract

BACKGROUND
Tumours are complex ecosystems comprised of tumour cells and various normal cell types. The interactions among these cells with diverse molecular characteristics lead to intratumoral heterogeneity and cancer evolution. However, little is known about the spatial organisation of the glioma tumour ecosystem, the cellular interactions therein, and their influence on patient outcomes. Spatially resolved transcriptomics, which measures the activity of hundreds of genes in situ, offers new avenues to address these long-standing questions.

MATERIAL AND METHODS
We assembled a clinically annotated cohort of glioma patients spanning 278 specimens from low- and high-grade IDH-mutant astrocytoma, IDH-mutant and 1p/19q co-deleted oligodendroglioma, ependymoma, and glioblastoma. Using tissue microarrays from archival FFPE, we performed spatial transcriptomics profiling with the 10x Xenium platform. A custom panel of 344 genes, selected based on publicly available single-cell RNA sequencing datasets of gliomas, was interrogated.

RESULTS
The spatial transcriptomics data enabled annotation and spatial mapping of more than 20 tumour microenvironment (TME) cell types and various tumour cell types present across the 5 glioma entities. We observed broadly conserved transcriptional profiles of TME cell types across gliomas, but TME cell type composition and spatial organisation varied depending on tumour entity. In particular, lower amounts of macrophages and microglia as well as oligodendrocytes and oligodendrocyte progenitor cells were observed in oligodendroglioma tumours compared to other entities. High-grade astrocytoma samples contained more myeloid dendritic cells and monocytes in comparison to low-grade astrocytomas. A lower proportion of vascular cells correlated with better overall survival for glioblastomas and low- and high-grade astrocytomas. Tumour cells exhibited mostly entity-specific gene expression, but also distinct tumour cell states including AC-, OPC-, NPC-, and MES-like cell states.

CONCLUSION
Our work demonstrates the utility of the 10x Xenium platform for high-resolution spatial profiling of expression in archival FFPE samples. Preliminary mapping of the TME and tumour cell types unravelled a complex spatial architecture of the glioma ecosystems. Distinct compositions of the TME were found in each glioma subtype, which may help understand different mechanisms of malignancy. Associations between cellular spatial patterns and clinical data available for FFPE samples have the potential to provide better insights for improved patient stratification and treatment.
P01.04.B DISSECT THE ROLE OF DRIVER GENE MUTATIONS IN THE INITIATION AND PROGRESSION OF GLIOMA
Authors: Tian A 
Published: October 2024

Abstract

BACKGROUND
Cancer initiation and progression are always associated with the acquisition and accumulation of genetic mutations which drive the malignant transformation of the cancer cells of origin. It has been shown that different driver mutations can generate tumors with distinct pathological features. How driver mutations interact with tumor cells of origin and how such interactions dictate the fate of the cell of origin and the tumor pathological features are the fundamental questions in cancer biology.

MATERIAL AND METHODS
We developed a set of novel “all-in-one” glioma lineage-tracing model system based the intra-brain ventricle electroporation and transposon-mediated conditional gene editing techniques. By using this system in genetically engineered mouse models, we generated glioma models with the top driver mutation combinations found in IDH-WT GBM patients and trace their initiation and progression with single-cell resolution, therefore we combined lineage tracing with multi-omics analysis almost recapitulating the full driver-mutation spectrum found in glioblastoma patients.

RESULTS
We found that driver gene mutations combinatorially dictate the early cell fate of tumor development. Driver gene mutations and cell fate combinatorially determine the prognosis and malignancy of glioma. Driver gene mutations combinatorially affect the molecular phenotypes and the treatment of end-stage tumor via deterministic CNVs. Synergistic or exclusive patterns of driver gene mutation combinations regulate the initiation and progression of gliomagenesis.

CONCLUSION
The results will help to address those fundamental questions pertinent to the relationships between driver mutations, tumor evolution pathways and the final tumor phenotypes. The results will also help to understand the nature of gliomagenesis and to develop novel approaches for glioma intervention and treatment.
P01.07.A HOW IS A RECURRENT GLIOBLASTOMA SELECTED?
Authors: Göhring K , Kässer A , Konig S , Strobel H , Karpel-Massler G , Mulaw M A , Debatin K M , Westhoff M A 
Published: October 2024

Abstract

BACKGROUND
Glioblastoma is the most common primary brain tumour in adults and one of the most lethal cancers per se. Despite aggressive therapeutic approaches, patient survival remains dismal, typically not exceeding 15 months. Standard treatment involves maximal safe resection followed by irradiation of the surgical cavity and chemotherapy with Temozolomide. However, recurrence is inevitable, and additional interventions provide only limited survival benefits.

MATERIAL AND METHODS
We conducted a study using two pairs of stem cell-like cell populations and their differentiated progeny obtained from the same patient, either before standard therapy or upon recurrence. Our objective was to investigate the selective forces driving the recurrent phenotype and identify potential therapeutic targets.

RESULTS
Our findings revealed an increased resistance to apoptosis in cells harvested after recurrence, albeit less pronounced than anticipated. The population doubling time was also reduced in post-recurrence cell populations, suggesting that they proliferate faster. However, the most striking changes we observed was a phenotypic shift towards a more mesenchymal phenotype. This was corroborated by increased expression of epithelial-mesenchymal transition (EMT) markers, the presence of oncostreams in differentiated progeny, and enhanced migratory velocity, suggesting heightened invasiveness post-standard intervention. Interestingly, detailed expression pattern analysis suggested that the recurrent phenotype might be expressed by cells more closely related to the cells of origin than those found in the primary tumour, implying recurrence may occur due to therapeutic clearance of the ecological niche.

CONCLUSION
Analysis of signalling networks identified potential druggable targets upregulated in cells from recurrent tumours, including phosphorylated STAT1 and various extracellular matrix components. Pharmacological inhibition of these targets exhibited a greater effect on recurrent tumour cells, indicating promise for inclusion in complex combination therapies. Our findings underscore the need to consider recurrent glioblastoma as an independent entity from the primary tumour and suggest further analysis may unveil additional therapeutic targets for novel combination therapies.
P01.13.B SPATIALLY RESOLVED FUNCTIONAL PROFILING FOR GLIOBLASTOMA
Authors: Buck A , Lee S , Wegmann R , Mena J , Vasella F , Friesen J , Regli L , Le Rhun E , Zeitlberger A , Neidert M , Tatari N , Hutter G , Weller M , Snijder B , Weiss T 
Published: October 2024

Abstract

BACKGROUND
Glioblastoma is the most common malignant primary brain tumor in adults with a poor survival and an unmet need for more effective therapies. Macroscopically, as observed through MR imaging and microscopy, these tumors are characterized by distinct tumor regions: a contrast-enhancing tumor core with a high cancer cell density and an infiltration zone that appears T2/FLAIR hyperintense without the uptake of contrast agent and consists of infiltrating tumor cells as well as non-malignant cells of the microenvironment. Previous genomic and transcriptomic studies revealed molecular differences between the tumor core and the infiltration zone. However, functional differences, such as drug sensitivity, between tumor regions are still largely unknown.

MATERIAL AND METHODS
To address this gap, we collected region-specific glioblastoma patient tissue samples from both the contrast-enhancing tumor core and the non-contrast-enhanced but 5-aminolevulinic acid (5-ALA) fluorescent infiltration zone directly from surgery. We utilized pharmacoscopy, a microscopy-based single-cell drug screening approach, to identify drug responses across regions and patients.

RESULTS
By profiling a library of 60 drugs across regions in a cohort of 23 patients, we identified region-specific drug responses and developed a complementarity score to pinpoint drug combinations capable of targeting both the tumor core and the infiltration zone. These combination treatments exhibited highly promising anti-tumor activity across regions and patients.

CONCLUSION
Given the high levels of functional spatial intra- and inter-tumor heterogeneity, relying on a single drug across patients may not be feasible for effective patient treatment. Thus, there is a strong rationale to administer drug combinations that not only consider regional heterogeneity but also account for patient-specific variations.
P02.01.B 3-PHOTON IN VIVO IMAGING REVEALS BREAKDOWN OF MICROGLIA SURVEILLANCE UPON GLIOMA INVASION IN THE CORPUS CALLOSUM
Authors: Nebeling F C , Fuhrmann F , Mittag M , Deli A , Stork M , Clements M , Garcia Diaz C , Parrinello S , Hölzel M , Herrlinger U , Salomoni P , Fuhrmann M 
Published: October 2024

Abstract

BACKGROUND
Glioblastomas are the most malignant primary brain tumors with median survival of about 17 month after diagnosis despite extensive treatment. Molecular heterogeneity and the tumor microenvironment contribute to the challenges in treating GBs. Glioma-associated microglia and macrophages, comprising 30-50% of non-neoplastic cells in the microenvironment, play a crucial role in facilitating tumor growth and inducing tissue necrosis. However, the intricate interplay between these immune and glioma cells remains poorly understood. This study focuses on understanding the dynamic role of microglia during glioblastoma invasion at the tumor’s invading front in corpus callosum.

MATERIAL AND METHODS
In order to investigate glioma and microglial dynamics, we use an autochthonous model of mesenchymal GBs employing a CRISPR/Cas mediated approach in recipient Cx3Cr1 knock-in mice. Previous studies using two-photon in vivo imaging were limited to superficial cortical regions, hindering investigations into deeper brain areas like the corpus callosum. Recently, three-photon microscopy has emerged as a powerful tool to image between 1100-1600µm deep into the intact brain.

RESULTS
By implanting chronic cortical windows and taking advantage of three photon in vivo imaging, we reveal differential regulation of microglial motility based on glioma infiltration levels in the corpus callosum. Moreover, we find distinct clusters of microglial movement in dependence to the distance of tumor cells. Conclusion: This approach provides insights into microglia-glioma interactions at the invading front in the corpus callosum, offering a valuable tool for studying deep-seated cellular dynamics and identifying potential therapeutic targets. Funding Source: CANcer TARgeting (CANTAR)
P02.03.A BLOOD-BRAIN BARRIER AFFECTING THE METABOLOMICS PROFILES IN THE TUMOR MICROENVIRONMENT, CSF AND BLOOD OF IN VIVO HUMAN GLIOBLASTOMA XENOGRAFT
Authors: Altendorfer-Kroath T , Schimek D , Monedeiro F , Zügner E , Prugger E , Magnes C , Birngruber T 
Published: October 2024

Abstract

BACKGROUND
Detection of glioblastoma (GBM) before clinical signs appear can significantly improve treatment outcome and patients’ quality of life. To date, there are various biomarker candidates for early detection of GBM but most of them can only be detected in cerebrospinal fluid (CSF) and in brain tissue biopsies. Brain tissue biopsies are very rarely taken in routine hospital practice while CSF or blood samples are part of the standard clinical routine in hospitals. However, no reliable biomarker has yet been identified for the early detection of most central nervous system (CNS) tumors in liquid biopsies, probably due to the limited ability of the biomarkers to cross the blood-brain barrier (BBB). In contrast to blood, CSF provides good access to the CNS. However, previous studies on pharmacokinetics of CSF and the interstitial fluid in the brain tissue (ISF) have shown that the equilibrium between the CSF and the ISF is slow and substance concentrations in the CSF do not reflect the concentrations that are locally present in brain tissue. We aimed to assess the metabolome from ISF collected with Open Flow Microperfusion (cOFM) from the tumor microenvironment (TME), CSF and blood from human GBM-bearing animals and compare it to respective samples collected from non-GBM-bearing animals (non-GBM controls) in order to identify potential metabolic biomarker or biomarker profiles that reflect the local situation of the TME.

MATERIAL AND METHODS
We applied a recently established atraumatic access to human GBM xenograft with cOFM that allows sampling of metabolites inside the TME with functional BBB. Tumor growth was monitored with magnetic-resonance-imaging. On day 18 post-inoculation, ISF from TME was collected with cOFM and blood, CSF and brain tissue were collected from the same animals at terminal time point. The collected samples were analyzed using a UHPLC-HRMS/MS metabolomics platform. Metabolomics data from GBM-bearing animals were analyzed and compared with that from non-GBM controls.

RESULTS
The metabolome from the GBM-bearing animals showed distinct metabolic patterns when compared to that from the non-GBM controls. GBM-relevant metabolites such as Kynurenine, Carnosine or Guanidinoacetate exhibited different levels in ISF from GBM-bearing animals than in CSF or blood samples from these mice.

CONCLUSION
This suggests that the BBB plays an important role when obtaining biomarkers from CSF or blood as it might affect the assessment of the local condition in the TME.
P02.12.B IMMUNOMODULATORY EFFECT OF MESENCHYMAL CELLS ON T-LYMPHOCYTES IN A 3D SPHEROID MODEL OF GLIOBLASTOMA
Authors: Šváblová T , Ternerová N , Houdová Megová M , Vymolová B , Balážiová E , Bušek P , Šedo A 
Published: October 2024

Abstract

BACKGROUND
Glioblastoma (GBM) represents the most aggressive form of primary brain tumors, typically resulting in a median survival of less than two years. Immunosuppression characterized by macrophage polarization into a protumorigenic phenotype and T cell dysfunction belong to key features of glioblastoma microenvironment. Recently, mesenchymal cells were identified in GBM, but their immunomodulatory effect has not been elucidated yet. This study aimed to establish a 3D GBM spheroid model to investigate the influence of mesenchymal cells derived from GBM patients on the phenotype of human T cells.

MATERIAL AND METHODS
Glioma stem-like cells (GSCs) and paired mesenchymal cell cultures were derived from 3 human GBMs. CD4+ T-lymphocytes were isolated from buffy coats of healthy donors using Ficoll Paque gradient separation, followed by negative magnetic isolation. T-lymphocytes were activated using anti-CD2, anti-CD3, and anti-CD28 antibodies. 3D spheroids were generated by encapsulating the GSC and T-lymphocytes in extracellular matrix (Geltrex) with or without mesenchymal cells. After co-culture for 5-days, the spheroids were dissociated, and the expression of T-cell activation markers CD69 and CD25, along with immune checkpoint molecules LAG3, TIM-3, and PD-1 was analyzed by flow cytometry. Immunohistochemistry was performed on frozen sections of the spheroids to investigate the spatial distribution of the cell populations.

RESULTS
In spheroids with mesenchymal cells, CD4+ T-lymphocytes exhibited elevated expression of the activation marker CD69, whereas no significant changes were observed in CD25 expression. PD-1 expression was higher in spheroids containing mesenchymal cells, whereas no difference was noted in TIM3 and LAG3 expression levels. Uniform distribution of glioblastoma-derived cells was revealed in spheroids containing glioma stem-like cells with and without mesenchymal cells and T-lymphocytes did not exhibit any specific spatial distribution pattern.

CONCLUSION
We have successfully established a 3D spheroid model to investigate the impact of mesenchymal cells on the GBM microenvironment. Our findings suggest that mesenchymal cells can influence the phenotype of CD4+ T-lymphocytes in glioblastoma. This research was funded by project National Institute for Cancer Research (Programme EXCELES, ID Project No. LX22NPO5102) by the European Union - Next Generation EU, and Charles University Cooperatio Program,,Oncology and Haematology“.
P02.13.B IMPLEMENTATION OF A MULTIPLEXED TISSUE IMAGING PROTOCOL FOR CHARACTERIZATION OF THE TUMOR MICROENVIRONMENT IN MELANOMA BRAIN METASTASES
Authors: Sagerer A , Westphal D , Meinhardt M , Schulz A , Meier F , Eyüpoglu I Y , Schackert G , Juratli T A 
Published: October 2024

Abstract

BACKGROUND
Melanoma-specific genetic changes are being identified for targeted therapy. Unfortunately, patients receiving such therapies often develop resistance and experience recurrence. Current data suggest that the tumor-specific microenvironment has a decisive influence on therapy resistance. We therefore established a multiplexed immunofluorescence staining procedure to precisely characterize the tumor microenvironment in melanoma brain metastases (BM).

MATERIAL AND METHODS
Co-detection by indexing (CODEX) technology relies on DNA-conjugated antibodies and the cyclic addition and removal of complementary fluorescently labeled DNA probes and has been used so far to simultaneously visualize up to 100 biomarkers in situ. It enables a deep view into the single-cell spatial relationships in tissues and is intended to spur discovery in disease and therapeutic design. Hence, CODEX was provided by Akoya Biosciences. To quickly handle a larger scale of samples, a tissue microarray (TMA) was created both of tumor bulk as well as positive and negative surgical margins.

RESULTS
At first, a panel of yet 30 antibodies were conjugated with its definite DNA oligonucleotide sequence (reporter). On our part, these antibodies were previously validated on conventional IHC using normal and gliotic brain along with tonsil tissue, including CD molecules (e.g. CD4, CD8, CD20, CD31, CD68), signal transducing antibodies (e.g. AKT, ERK, EGFR, IGFR and their phosphorylated isoforms) as well as tumor (e.g. HMB-45, Ki67, Melan-A) and neuronal (e.g. GFAP, NeuN, Olig2, PSD-95) markers. Subsequently, TMAs of different BM composition were generated (25 cores (1.5 mm), per slide 6x4 samples + 1 positive control). Within only a single working day TMAs were prepared and CODEX staining protocol fully assembled (single step staining to preserve sample integrity). CODEX run (3 reporter dyes imaged with complimentary barcodes) was performed overnight truly smooth. Lastly, analysis of spatial information has been received via QuPath (open source).

CONCLUSION
Once individually designed, multiplex fluorescence imaging platform is an easy and especially in case of TMAs fast-performing tool to gain insight into the tumor microenvironment of small sample volumes in particular. For the prediction of possible therapy resistance distinct further studies are warranted to define the potential of high throughput multiplexed imaging in a clinical routine.