bluuebunny/crossref_metadata_2025_split huggingface dataset:
P02.12.B UNDERSTANDING THE ROLES OF EPHRIN A5 IN GLIOBLASTOMA AND ITS POTENTIAL AS A THERAPEUTIC TARGET
Authors: Skarne N , D'Souza R , Harris L , Baumgartner U , Offenhauser C , Akgül S , Beckett K , Boyd A , Day B 
Published: September 2023

Abstract

BACKGROUND
Glioblastoma (GBM) is the most common and aggressive malignant primary brain cancer and is associated with very poor patient outcomes. GBM has been classified into three cell subtypes (proneural (PN), classical (CL), and mesenchymal (MES)), which have been further defined into four plastic cell states (neural-progenitor-like (NPC-like), oligodendrocyte-progenitor-like (OPC-like), astrocyte-like (AC-like), and mesenchymal-like (MES-like)). Existing treatment options have had limited success in improving overall survival, with the extreme heterogeneity present in GBM presenting a major barrier to effective therapy. Thus, there is an urgent need to develop more effective therapeutic strategies which better capture heterogeneity and delay tumour relapse. Several erythropoietin-producing human hepatocellular (Eph) receptors and ephrin ligands have been shown to be functionally overexpressed in cancer and have become attractive therapeutic targets. EphA3 has previously been shown to be a functional, tumour-specific target, frequently elevated in the MES subtype. We identified ephrin A5, the high-affinity ligand of EphA3, to be frequently co-expressed with EphA3 in GBM tumours. However, unlike EphA3, the mechanisms of ephrin A5 in GBM remain understudied. We therefore believe that ephrin A5 serves as an attractive synergistic target for the treatment of GBM.

MATERIAL AND METHODS
We examined the expression of ephrin A5 in a panel of GBM patient specimens. A panel of GBM tumours were also selected for immunohistochemical and spatial transcriptomics analysis. For in vitro functional studies, ephrin A5 was overexpressed in several GBM primary cell lines. Assays examining proliferation, stemness and differentiation were performed.

RESULTS
Expression of ephrin A5 was comparable to EphA3 in GBM. 64% of GBM tumours analysed showed higher levels of ephrin A5, relative to the highest normal brain specimen. Initial immunohistochemistry analysis suggests EphA3 and ephrin A5 to be expressed discretely on over 75% of the total tumour mass. Spatial transcriptomics analysis suggests ephrin A5 to be enriched in cells of the AC-like cell state. In vitro overexpression of ephrin A5 was shown to reduce neurosphere formation and induce differentiation of GBM cells.

CONCLUSION
EphA3 and ephrin A5 are overexpressed in a portion of GBM tumours. Critically, there was a strong mutually exclusive expression to different compartments of the GBM tumour, covering the vast majority of the total tumour mass. Our data suggests that ephrin A5 overexpression drives a less aggressive, more differentiated AC-like tumour phenotype. Thus, dual targeting of EphA3 and ephrin A5 in GBM could provide an opportunity to better capture tumour heterogeneity and achieve more durable therapeutic responses.
P02.22.B CELLULAR COMPOSITION DETERMINED BY STEMNESS MARKERS IN MONOLAYER GLIOBLASTOMA CULTURES IS TUMOR SPECIFIC, HIGHLY DYNAMIC AND RELATED TO TMZ SENSITIVITY
Authors: Verploegh I S C , de Kroon A , Verhoeven F A M , Loriaux S , e. Hassnaoui H , Lamfers M , Huylebroeck D , Leenstra S 
Published: September 2023

Abstract

BACKGROUND
Different models are used to study pathophysiology and potential new therapeutic strategies of glioblastoma (GBM) in vitro. Generally, monolayer cultures are chosen because of their high reproducibility, while spheroid cultures are preferred to better recapitulate tumor heterogeneity and effects of the micro-environment. A common assumption is that monolayer cultures display less plasticity than spheroid cultures. In this study we examine the plasticity of stem cell-like cells in monolayer cultures and their relationship to temozolomide (TMZ) sensitivity.

MATERIAL AND METHODS
Three patient-derived GBM cultures: one TMZ sensitive, one induced TMZ-resistant and one naturally TMZ-resistant were used. All cultures were sorted in four subpopulations (CD133+/CD44+, CD133+/CD44-, CD133-/CD44+ and CD133-/CD44-) and subsequently cultured. Cellular composition was determined at multiple timepoints by immunofluorescent imaging and FACS-analysis. The effect of seeding density on cellular composition was assessed. Furthermore the effect of original culture composition on sphere forming capacity and TMZ sensitivity was assessed.

RESULTS
Culture composition was tumor-specific and the pre-sorting composition was acquired within two passages after sorting, irrespective of original subpopulation or sorting density. Even the CD133-/CD44- subpopulation was able to rapidly re-acquire the initial tumor composition. Furthermore, sphere-forming capacity was restored four passages after sorting. Interestingly, cells that were inherently resistant to TMZ expressed higher levels of CD44 than TMZ-sensitive cells. There was no relationship between expression of CD133 and sensitivity to TMZ. Sensitivity to TMZ was similar in all subpopulations four passages after sorting.

CONCLUSION
Monolayer cultures display rapid plasticity with regard to the expression of the stemness markers CD133 and CD44. Culture composition is tumor specific and inherently programmed in each tumor cell. TMZ sensitivity seems to be mostly related to CD44 expression, and not to CD133 expression. Sphere forming capacity and TMZ sensitivity are restored soon after sorting, even in initially differentiated GBM cells.
P02.24.B A DRUG DISCOVERY PLATFORM FOR COMBINATORIAL TARGETING OF CELL STATES AND ENTITIES
Authors: Jiang B , Schmitt M , Gargiulo G 
Published: September 2023

Abstract
Glioblastoma (GBM) is a lethal brain tumor with limited therapeutic options. Therapeutic resistance arises from the collaboration among heterogeneous and highly plastic cellular entities and states conserved across GBM patients and the tumor microenvironment.Tumor cells adapt in response to the current standard of care and infiltration of innate immune cells, recurrently acquiring a mesenchymal state that drives therapeutic resistance.Moreover, the blood-brain barrier (BBB) poses challenges to the effectiveness and bioavailability of approved therapeutics, as well as the discovery of new ones.Our lab has developed synthetic locus control regions (sLCRs) as transcriptional reporters for glioblastoma subtypes, enabling the study of cell identity and state transitions in vitro and in vivo. We have combined this tool with validated cellular models for glioblastoma mesenchymal transition and therapeutic resistance to establish an in vitro phenotypic screening platform. Currently, we are utilizing this platform to identify treatment combinations and dosing schemes that aim to overcome innate immunity-driven resistance and synergize with the standard of care. As a proof-of-concept for a new generation of brain tumor target discovery platforms, we will present the setup of our platform and preliminary results for a class of BBB-penetrant drugs.
P03.02.B VIABLE TUMOR RECURRENCE IS A MAJOR CAUSE OF LOCAL FAILURE AFTER BEVACIZUMAB THERAPY FOR RADIATION NECROSIS IN BRAIN METASTASES TREATED WITH STEREOTACTIC RADIOSURGERY
Authors: Cho Y , Yoon K , Lee D , Song S , Kim Y , Hong C , Kim J 
Published: September 2023

Abstract

BACKGROUND
Bevacizumab (BVZ) is known to be effective to control radiation necrosis (RN) following stereotactic radiosurgery (SRS) for brain metastases (BMs), although treatment failure may occur. Here, we investigated the incidence and pattern of local failure after BVZ therapy for RN and its underlying biological mechanism.

MATERIAL AND METHODS
We conducted a retrospective analysis on 17 patients who had been treated with BVZ for RN following SRS for BMs between 2016 and 2021. In each patient, the diagnosis of RN was made based on the conventional and advanced MR with or without positron emission tomography. Median 5 cycles (range, 2-10 cycles) of BVZ (5 mg/kg) were administered at 2-week intervals. Treatment response was assessed by volumetric changes of the lesions on MR and patients’ neurological status.

RESULTS
Treatment response was typically brisk and substantial. Best MR response was seen at median 13 weeks (range, 3-56 weeks) after the start of BVZ with a median volume decrease of 84.5% (range, 38.7-100%) of perilesional brain edema on T2WI and of 54% (range, 2.9-100%) of contrast enhancing lesions on T1WI. Patients’ neurological status improved in 16 patients (94.1%) and was stationary in 1 (5.9%). During the median follow-up of 12 months (range, 2-60 months), delayed local failure was observed in 6 patients (35.3%) at median 10 months (range, 6-14 months) after starting BVZ treatment, where viable tumor recurrence was demonstrated in all of them. No reconstitution of RN without viable tumor was observed during the follow-up.

CONCLUSION
Although BVZ was highly effective to control RN following SRS for BMs, delayed local failure frequently occurs owing to viable tumor recurrence. This may imply that much predominant vascular stabilizing effect of BVZ over anti-tumor effect transiently obscures the presence of potential viable tumor cells but does not prevent them from eventual recurrence.
P03.04.B EFFICACY OF NEOADJUVANT STEREOTACTIC RADIOSURGERY IN SOLID CANCER BRAIN METASTASES: A SYSTEMATIC REVIEW AND METANALYSIS
Authors: Gagliardi F , De Domenico P , Snider S , Nizzola M , Mortini P 
Published: September 2023

Abstract

BACKGROUND
Neoadjuvant stereotactic radiosurgery (NaSRS) is a novel strategy for brain metastasis (BMs) management, promising to achieve lower rates of toxicity, leptomeningeal progression, and high percentage of local control.

MATERIAL AND METHODS
An extensive systematic literature review and meta-analysis of proportions were performed. A total of 8 papers were eligible for final analysis after searching MEDLINE via PubMed, Web-of-science, Cochrane Wiley, and Embase databases.

RESULTS
A total of 551 patients undergoing NaSRS to treat 583 lesions were included. Median age was 57.7 [56.5-61.1] years, with a median tumor volume of 9.1 [8.3-14.4] cc. The most frequent histology was non-small cell lung cancer (40.7%), followed by breast (20.5%), and melanoma (15.6%). Most of the lesions had a supratentorial location (77.6%). Treatment parameters were homogenous across the studies and showed a median dose of 16 [14.9-20.5] Gy at 80% isodose.Surgery was performed after a median of 1 [1-2] day and achieved gross-total extent in 92.4% of cases. Median follow-up was 12.9 [9.9-18.9] months.Pooled proportion of patients showed an overall mortality rate of 50% (CI 37-63; I2=87%), local failure of 15% (CI 12-20%; I2=12.1%) and a rate of distant brain failure of 42% (CI 35-49%; I2=55.3%). The median local progression-free survival (PFS) was 10.4 months, while median survival without distant failure was 8 months.Rate of radionecrosis (RN) and leptomeningeal dissemination (LMD) were 4% respectively. Median overall survival (OS) was 14.4 months.

CONCLUSION
Existing data suggest that NaSRS achieves good local control on BMs with relatively lower dose than adjuvant treatment and low incidence of RN and other complications. These results need to be confirmed by larger studies with higher level of evidence to address formal comparisons with other regimens.
P03.05.B PRIMARY GAMMA KNIFE RADIOSURGERY FOR PINEAL REGION TUMORS, A SYSTEMATIC REVIEW AND POOLED ANALYSIS OF AVAILABLE LITERATURE WITH HISTOLOGICAL STRATIFICATION
Authors: Gagliardi F , De Domenico P , Garbin E , Bailo M , Snider S , Mortini P 
Published: September 2023

Abstract

BACKGROUND
Pineal region tumors (PTs) represent extremely rare pathologies, characterized by highly heterogeneous histological patterns. Most of the available evidence for Gamma Knife radiosurgical (GKSR) treatment of PTs arises from multimodal regimens, including GKSR as an adjuvant modality following surgery or as a salvage treatment at recurrence. We aimed to gather the existing evidence on the topic and analyze single-patient level data to address the efficacy and safety of primary GKSR when only a biopsy was attempted or biochemical or neuroimaging diagnosis was available.

MATERIAL AND METHODS
This is a systematic review of literature (Pubmed, Embase, Cochrane, Science Direct) and pooled analysis of single-patient-level data. A total of 1054 original works were retrieved. After excluding duplicates and irrelevant works, we included 13 papers in our final analysis comprising 64 patients. Additional 12 patients were included from the authors’ original series.

RESULTS
A total of 76 patients reached the final analysis. Presumptive diagnoses included intrinsic pineal low-grade (46%), glial low-grade (LGG, 5%), pineoblastoma (4%), germ cell tumors (GCTs, 4%), pineal tumors of intermediate differentiation (3%), other pineal tumors (3%), glial high-grade (HGG, 2%) and meningioma (1%). In 32% of cases, the diagnosis was unclear. Only 57% of cases received histological confirmation. The mean age at treatment was 38.7y and the mean lesional volume was 4.2 ± 4 cc. All patients received GKSR with a mean marginal dose (50% isodose) of 14.7 ± 2.1 Gy. At a median 36mo follow-up, local control was achieved in 84% of cases. Ten patients showed progression within a mean time of 24.2mo. Progression was more common in HGG and GCTs (p=0.03) compared to intrinsic pineal tumors. Overall mortality was 13.2%. The median OS was not reached for all included lesions, except high-grade gliomas (8mo). The 3y-OS was 100% for LGG, 90% for the intrinsic pineal, 60% for GCTs, 50% for HHG, and 82% for undetermined tumors. The 3y-PFS was 100% for LGG, 90% for the intrinsic pineal, 32% for GCTs, and 0% for HHG. Median PFS was 5mo for HHG and 34mo for GCTs. The radionecrosis rate was 6%, and cystic degeneration was observed in 2%. Ataxia as presenting symptom strongly predicted mortality (OR 104, p=0.02), while GCTs and HHG histology well predicted PD (OR: 13, p=0.04).

CONCLUSION
These results support the efficacy and safety of primary GKSR treatment of pineal region tumors. Further studies are needed to validate these results, which highlight the importance of the initial presumptive diagnosis for choosing the best therapeutic strategy.
P03.07.B DOSIMETRIC ACCURACY OF CYBERKNIFE STEREOTACTIC RADIOSURGERY FOR BENIGN PERIOPTIC TUMOR
Authors: Yoon K , Jeong C , Park M , Goh Y , Kim S , Cho B , Kwak J , Cho Y 
Published: September 2023

Abstract

BACKGROUND
The Cyberknife (CK) SRS delivers a highly focused ablative radiation dose to tumors to effectively control tumors with minimized radiation toxicity to surrounding normal tissues. This study aimed to evaluate the dosimetric accuracy of Cyberknife (CK) for benign perioptic tumor using patient specific head phantom.

METHODS
Patient specific head phantoms was fabricated using a 3D-printer to be dosimetrically equivalent to actual target regions of benign perioptic tumor case treated via Cyberknife radiosurgery. Head phantom QA plan was produced using the original CK contour set (target and optic nerve). The head phantom containing the Gafchromic EBT3 film was irradiated using Cyberknife 6D skull tracking method. The dose distributions calculated by MultiPlan Treatment Planning System (version 5.6) was compared with those measured by film dose using gamma analysis method. After moved rotating (1 degree) and translating (1-5mm) the couch table prior to beam delivery, we checked the 6D skull tracking accuracy according to the beam irradiation.

RESULTS
Gamma passing rates for the 2%/1 mm and 2%/2 mm criteria was found to be 84.11 ± 2.12% and 93.61±1.39% greater than the acceptance criteria 80% and 90%. The difference between measured and calculated dose to the optic nerve in contact with the tumor was less than 3%.

CONCLUSION
Dosimetric verification with patient-specific head phantoms could be successfully implemented as the evaluation method for CK perioptic tumor radiosurgery delivery with 6D skull tracking system.
P04.08.A PATIENT-DERIVED ORGANOIDS FOR MODELING AND CHARACTERIZING PEDIATRIC BRAIN TUMORS
Authors: Lago C , Federico A , Leva G , Ballabio C , Gianesello M , Abballe L , Kool M , Miele E , Tiberi L 
Published: September 2023

Abstract

BACKGROUND
Central nervous system (CNS) tumors are the most common solid tumors in the pediatric population and the leading cause of death among childhood cancers. They present common features related to the development and produce symptoms based on their location, rate of tumor growth and age of the child. Tumors of embryonal origin, such as medulloblastoma or primitive neuroectodermal tumor, have a higher incidence in younger patients, whereas older ones tend to present tumors of glial origin. Most of CNS-embryonal tumors are highly malignant and consequently leading to poor prognoses. Despite the understanding of their underlying molecular landscape that has helped in clinical advances to improve life expectancies of the patients, there is still an emerging need to minimize morbidity and manage the long-term therapy effects.

MATERIAL AND METHODS
Human samples were collected according to protocols approved by the institutional review board with written consent. Patient-Derived Organoids (PDOs) and Patient-Derived Xenograft Organoids (PDXOs) were obtained using scalpels by chopping in small pieces the tumor tissue and were grown in PDOs medium. PDOs and PDXOs were analyzed through DNA methylation, immunostainings, sc-RNA sequencing and orthotopic nude mice engraftment. Therapeutic regimens were adapted according to patients’ treatment protocols depending on the tumor of origin.

RESULTS
Developing new models to better understand the complexity and heterogeneity of pediatric tumors and to test personalized treatment strategies remains a clinical challenge. We tried to answer this medical need with PDOs and PDXOs established through the direct in vitro culture of different kinds of primary tumors (ependymoma EPN, medulloblastoma MB and low-grade glial tumors) and MB patient derived xenograft (PDX)-derived tumors. We have developed a method that recapitulates mutational profiles, histological features, and cellular heterogeneity of the corresponding parental tumors. We have obtained promising results from the DNA methylation profile analysis, nowadays used for an accurate routine CNS tumors diagnosis. Through this new model, we confirmed the action of clinically used therapeutic regimens, i.e., for EPN with ZFTA-RELA fusion and G3 MB-derived PDOs. PDOs were subjected to the same treatment the patients underwent and responded similarly to the patients.

CONCLUSION
We found a suitable in vitro culture condition to generate and maintain PDOs from different kinds of pediatric brain tumors. Their reliability has been proved by different analyses, confirming they still resemble the parental tumors. Through this new model, we confirmed the action of clinically used therapeutic regimens, suggesting its potential at the translation level as a reliable tool for testing personalized medicine, drug screening and development of newer and more specific therapeutic strategies.
P06.02.B GSDME-MEDIATED PYROPTOSIS IN GLIOBLASTOMA
Authors: Solel E , Brudvik E , Afroz S , Choudhury R , Bjerkvig R , Hossain J , Miletic H 
Published: September 2023

Abstract

BACKGROUND
Immunotherapeutic strategies have largely failed for glioblastoma , the most frequent primary brain tumor. Beyond apoptosis and autophagy, little is known about cell death mechanisms in glioblastoma, despite their potential impact on the immune microenvironment. Pyroptosis, unlike apoptosis, is an immunogenic and lytic type of programmed cell death, where gasdermin proteins form pores into the plasma membrane, releasing cell content to the tumor microenvironment. Gasdermin E (GSDME), often inactivated in cancers by mutations/low expression, is proteolytically cleaved by caspase-3 upon apoptotic stimuli leading to a shift from apoptosis to pyroptosis. Here, we investigated the relevance of GSDME-mediated pyroptosis for glioblastoma. According to The Cancer Genome Atlas, GSDME is highly expressed in glioblastomas compared to many other cancers and no mutations are found.

MATERIAL AND METHODS
Using the drug raptinal to induce apoptosis, we assessed expression of full length and cleaved GSDME as well as cleaved caspase-3 in syngeneic mouse and human patient-derived glioblastoma stem-like cell lines by western blotting. Lytic cell death and viability were measured by lactate dehydrogenase (LDH) and WST-1 assays respectively. PI uptake assay was performed to capture pyroptotic morphology by Incucyte®️ S3. BAPTA-AM, a fast Ca2+ chelator, was used to block plasma membrane repair in human glioblastoma cell lines and analyze its effect on pyroptosis. GSDME was knocked out by CRISPR/Cas9.

RESULTS
GSDME and caspase-3 cleavage were confirmed after raptinal treatment in mouse and human glioblastoma cell lines, indicating that the molecular pyroptotic machinery is intact. A significant drop in viability with raptinal was shown indicating successful execution of cell death. A significant LDH release was detected in mouse glioblastoma cell lines, which could be blocked by GSDME knockout. In contrast, human glioblastoma cells, although executing GSDME cleavage, showed less or no LDH release. Pyroptotic morphology was detected after raptinal treatment in mouse glioblastoma cell lines, while it was delayed/inhibited in human glioblastoma cells. Application of BAPTA-AM in human glioblastoma cell lines enhanced pyroptotic morphology, indicating a role for plasma membrane repair in inhibiting pyroptosis.

CONCLUSION
The molecular machinery to execute pyroptosis is functioning in mouse and human glioblastoma cells. However, while mouse glioblastoma cells show clear execution of pyroptosis upon apoptosis induction, human glioblastoma cells indicate less efficient pyroptotic ability which partly can be reverted by blocking plasma membrane repair. While pyroptosis can induce anti-tumor immunity in experimental cancer models, future in vivo experiments must delineate if pyroptosis is a viable strategy for immunotherapy of glioblastoma.