bluuebunny/crossref_metadata_2025 huggingface dataset:
P11.22.A Prognostic and predictive relevance of immunohistochemically determined p53 mutation in glioblastoma
Authors: Kempter J , Gempt J , Wiestler B , Combs S E , Schlegel J , Liesche-Starnecker F , Schmidt-Graf F
Published: September 2022

Background

It can be expected that molecular biomarkers will increasingly affect clinical decisions and lead to the development of more personalized therapies in glioblastoma (GBM) in the future. In several other tumor entities TP53 gene mutation or p53 immunoreactivity (IR) serve as a prognostic marker, significantly affecting overall survival (OS) and progression-free survival (PFS). Such an association has not yet sufficiently been demonstrated in GBM. However, there are known prognostic markers in GBM, notably MGMT promotor methylation (mMGMT) which also serves as an important predictive marker leading to a better response to temozolomide chemotherapy. Our aim was to evaluate retrospectively if p53 mutation determined via immunohistochemistry (IHC) could act as a prognostic or predictive marker in GBM.

Material and Methods

Tumor samples of 195 treatment-naïve patients with IDHwt GBM that had been stained with the p53 antibody DO-7 were subdivided into 2 different groups by p53 IHC. Samples were considered as p53mut when strong p53 IR was detected in ≥10% of all tumor cells and as p53wt when in <10%. Treatment, further molecular and survival data were gathered retrospectively for all patients. Statistical analyses were performed with SPSS.

Results

The frequency of p53mut was 36.4% (71/195). p53mut tumors showed a significantly higher IR with Ki-67 proliferation marker (p=0.005) and p53wt seemed to be associated with multifocal primary tumor localization, though not statistically significant (p=0.107). There was no significant difference between p53wt and p53mut regarding gender, age, extent of resection, adjuvant therapy, occurrence of seizures, mMGMT or ATRX loss. The p53 status was not associated with OS or PFS. Factors that univariately led to significantly longer OS and PFS were younger age, unilateral or unifocal primary tumor localization, gross-total resection, higher Karnofsky Performance Status (KPS), mMGMT and adjuvant treatment via Stupp regimen instead of radiotherapy alone, the latter being significantly better than best supportive care. In multivariate survival analyses only age <65 years, the Stupp regimen more than radiotherapy alone and KPS ≥80% significantly prolonged both OS and PFS. Unifocal primary tumor localization led to longer OS and mMGMT led to longer PFS independently. The p53 status did not significantly affect the response to different adjuvant therapy regimens neither concerning OS nor PFS.

Conclusion

Based on our study, p53 IR has no prognostic or predictive significance in IDHwt GBM. There have been previous studies with similar and others with contradicting results. Remarkable is the discordance of the used IR thresholds between different studies. Further studies should aim to revalidate the staining threshold and improve the concordance between TP53 gene sequencing and p53 IHC in IDHwt GBM.
P11.26.A STAT3 expression in brain metastases from breast cancer: correlations with different molecular subtypes and clinical outcome
Authors: Pellerino A , Bruno F , Mo F , Bertero L , Bellini E , Beano A , Montemurro F , Valiente M , Rudà R , Soffietti R
Published: September 2022

Background

STAT3 expression in peritumoral reactive astrocytes (RA) of brain metastases (BM) may favor a pro-metastatic environment. The aim of the study was to evaluate in a retrospective cohort of surgically resected BM from breast cancer (BC) the expression of pSTAT3 in RA of peritumoral tissue of BM, identify different patterns of expression according to molecular subtypes, and correlate with intracranial progression-free survival (i-PFS).

Material and Methods

Patients with histologically proven BM diagnosis from BC were identified from the biobank of Pathology Unit of University of Turin and Spanish national BrM network (RENACER). pSTAT3 expression was evaluated and scored in RA of peritumoral tissue using GFAP and STAT3 immunohistochemistry, according to Priego et al. (Nat Med 2018). Data on histological diagnosis, molecular subtypes, and i-PFS were retrieved by chart review. Intracranial progression was defined based on MRI reports.

Results

Eighty-five BM specimens from BC of 85 female patients with a median age of 54 years (range 30-81 years) were available for analysis. Immunohistochemistry for GFAP and pSTAT3 was feasible in 68/85 (80%). Fifteen out of 68 patients (21.1%) had BM from luminal BC, 27/68 (39.7%) from HER2-positive BC, and 26/68 (39.2%) from TNBC. Fifty-six out of 68 (82.4%) showed positive staining of pSTAT3 in peritumoral RA, of which 9/68 (13.3%) scored with 3, 26/68 (38.2%) with 2, and 21/68 (30.9%%) with 1, while pSTAT3 expression was negative (score 0) in 12/68 (17.6%). High pSTAT3 expression (score 2-3) was observed in 17/27 (62.9%) BM from HER2-positive BC and in 15/26 (57.7%) BM from TNBC, while most of BM from luminal BC (12/15 - 80%) had low or absent pSTAT3 (score 0-1) (p=0.021). Overall i-PFS was 16 months (range 7-41): low pSTAT3 BM (score 0-1) had a median i-PFS of 21 months versus 12 months for high pSTAT3 BM (score 2-3). A shorter median i-PFS was observed in high pSTAT3 BM from TNBC (4 months) as compared with low pSTAT3 BM (11 months). Conversely, i-PFS of high pSTAT3 BM (7 months) was similar to low pSTAT3 BM (6 months) in HER2-positive BC.

Conclusion

pSTAT3 expression in RA of peritumoral tissue of BM from TNBC and HER2-positive BC is higher than in BM from luminal BC. Of note, patients with high pSTAT3 BM from TNBC progressed earlier in comparison with those with low pSTAT3, suggesting that pSTAT3 expression has an influence on the outcome.
P11.43.B The impact of neutrophils-to-lymphocytes ratio on survival in patients affected by brain metastases and treated with Gamma Knife Radiosurgery
Authors: Gagliardi F , Snider S , Roncelli F , Pompeo E , De Domenico P , Klungtvedt V , Barzaghi L , Bulotta A , Lazzari C , Del Vecchio A , Mortini P
Published: September 2022

Background

The Neutrophil-to-Lymphocyte ratio (NLR) is a biomarker for host systemic inflammation. Elevated NLR may result from neutrophilia, lymphopenia or both and is associated with poor prognosis and treatment responses in various malignant tumors, such as breast, gastric, colorectal and lung cancer. It was found that in patients treated with GK-SRS for brain metastases (BM) from non-small cell lung cancer (NSCLC), median survival was significantly longer in patients with NLR < 5 at the time of radiosurgery and after treatment (within 1 month from SRS). High percentage of neutrophils, low percentage of lymphocytes and elevated platelet-to-lymphocyte ratio were predictive of poor OS in patients with brain metastases. Further evidences are needed to support and better define these findings.

Material and Methods

We retrospectively examined 111 consecutive patients with BMs from NSCLC (95 patients) and melanoma (16 patients) treated with GK-SRS. A total of 695 lesions were treated (median lesion number per patient 6; range 1-18), the median volume was 0.108 cc (range 0.004-33.44 cc).

NLR was calculated using N/L, where N and L, respectively, refer to peripheral blood neutrophils (N) and lymphocyte (L) counts at the timepoint nearest to the SRS treatment. Kaplan-Meier analysis described the survival time according to NLR. Univariable and multivariable Cox regression analyses were used to confirm the impact of NLR on overall survival.

Results

Median (IQR) age at diagnosis of brain metastases was 64 yrs (55-70), median (IQR) NLR was 6.7 (1.3-32), and median (IQR) overall survival was 13.5 months (1-42). At univariable Cox-regression analyses, lower NLR was associated with improved overall survival (HR: 1.05; p=0.004). Total number of lymphocytes, neutrophils and monocytes were not associated with improved overall survival (all p>0.1). At multivariable Cox regression analyses, after adjusting for patient age, sex and the use of DEX therapy, NLR represented an independent predictor of overall survival (HR: 1.06; p=0.003).

Conclusion

NLR represents an independent prognostic factor in patients affected by BMs from NSCLC and melanoma. Inflammation and immunity may play a critical role in these patients. Further analysis examining more specific neutrophils or lymphocytes subsets may increase our understanding of cancer etiology and progression.
P11.47.B A description of inpatient admissions of adult glioblastoma patients in England 2103 - 2018 from the GlioCova project
Authors: Le Calvez K , Mauricaite R , Williams M
Published: September 2022

Background

The Gliocova dataset uses linked English national cancer data on all 51,775 adult primary brain tumour patients diagnosed between 2013-2018. Here we present early analysis of inpatient admissions of adult glioblastoma (GBM) patients.

Material and Methods

We identified all adults patients diagnosed with a GBM and extracted all the inpatient admissions for 1 night or more after the date of diagnosis. We focused on number and length of admissions, variation in those numbers, and place of discharge.

Results

Between 2013 and 2018, 15,294 patients were diagnosed with a glioblastoma in England (60% male) with a median age of 66, of whom, 12,441 (61% male) were admitted overnight with a total of 49,384 admissions post-diagnosis. Half of these patients were less than 64 at the time of their admission. The mean number of post-diagnosis admissions was 4, with a mean length of stay of 9.5 days. However, for half of the admissions, patients stayed 5 days or less in hospital (IQR = 10). Most of the procedures done were treatment-related, such as surgery for which patients stayed an average of 6.3 days (median = 4; IQR = 5). Patients who were admitted for non-treatment reasons stayed on average almost 10 days (median = 5; IQR = 11). Fewer than 3,000 admissions resulted in a patient death (5.5% of all admissions), whereas over 23,000 admissions for 10,426 patients ended with patients being discharged at home. About 2,000 patients were discharged to another hospital, hospice or a nursing home.

Conclusion

Most of the patients diagnosed with a glioblastoma will be admitted at some point after their diagnosis. Although the average length of stay is not that long, there is a considerable tail of longer-staying patients, for whom improved services and support might enable quicker discharge. To our knowledge, this is the first time inpatient admissions in adult brain patients are being looked at.GlioCova is supported by the Imperial/NIHR BRC, and the members of the GlioCova EAG. This work uses data provided by patients and collected by the NHS as part of their care and support.More information on the Gliocova project can be found on https://blogs.imperial.ac.uk/gliocova.
P11.51.B Initial temozolomide monotherapy without radiotherapy might be of limited benefit in the treatment of astrocytoma, IDH-mutant, CNS WHO grade 2 and 3
Authors: Weller J , Katzendobler S , Blobner J , Quach S , Egensperger R , Suchorska B , Thon N , Weller M , Tonn J
Published: September 2022

Background

The role of temozolomide chemotherapy alone in isocitrate dehydrogenase (IDH-) mutant astrocytomas has not been conclusively determined. Radiotherapy might be superior to temozolomide alone. Recent studies have linked temozolomide therapy with poor clinical course and induction of hypermutation in IDH-mutant gliomas.

Material and Methods

In this retrospective, single-center study, 183 patients with astrocytoma, IDH-mutant, CNS WHO grade 2 or 3 according to WHO 2021 and diagnosed between 2000 and 2019 were included. Patients initially monitored by means of a wait-and-scan strategy, or treated with radiotherapy alone, or receiving temozolomide alone after histological sampling through biopsy or tumor resection were studied. Patient-related, clinical and imaging data were correlated with progression-free and overall survival. A matched-pair analysis accounting for post-surgical tumor volume was conducted.

Results

No significant differences in median age and clinical status at diagnosis was seen. WHO grades were balanced between patients treated with radiotherapy and temozolomide, but the proportion of WHO grade 2 gliomas was higher in the wait-and-scan cohort. Radiotherapy was associated with significantly longer overall survival than temozolomide (in years, 14.4 vs 10.7; p=0.02) and longer progression-free survival than temozolomide (in years, 6.2 vs 3.4, p=0.02) and wait-and-scan strategies (in years, 6.2 vs 4; p=0.03). Comparing wait-and-scan with temozolomide, survival was significantly longer in the wait-and-scan cohort (in years, not reached vs 10.7, p<0.0001). An analysis of WHO grade 2 astrocytomas only also showed superior survival in the wait-and-scan cohort as compared to temozolomide (p=0.02). Of note PFS was similar overall and in WHO grade 2 astrocytomas.

Conclusion

The results suggest superiority of radiotherapy alone over temozolomide alone or wait-and-scan strategies in IDH-mutant WHO grade 2 and 3. Recent study results indicating that temozolomide might compromise prognosis in some IDH-mutant gliomas might be supported by our data, although caution in interpretation of retrospective data is required. The potential negative effects may only be apparent in the long-term.
P11.62.A The link between gliomas infiltrationandwhite matter architecture investigated with electron microscopy and diffusion tensor imaging
Authors: Latini F , Fahlström M , Beháňová A , Sintorn I , Hodik M , Staxäng K , Ryttlefors M
Published: September 2022

Background

Diffuse low-grade gliomas (DLGG) display different preferential locations in eloquent and secondary associative brain areas. The reason for this tendency is still unknown. We hypothesized that the intrinsic architecture and water diffusion properties of the white matter bundles in these regions may facilitate gliomas infiltration.

Material and Methods

Magnetic resonance imaging of sixty-seven diffuse low-grade gliomas patients were normalized to/and segmented in MNI space to create three probabilistic infiltration weighted gradient maps according to the molecular status of each tumor group (IDH mutated, IDH wild-type and IDH mutated/1p19q co-deleted). Diffusion tensor imaging (DTI)- based parameters were derived for five major white matter bundles, displaying regional differences in the grade of infiltration, averaged over 20 healthy individuals acquired from the Human connectome project (HCP) database. Transmission electron microscopy (TEM) was used to analyze fiber density, fiber diameter and g-ratio in 100 human white matter regions, sampled from cadaver specimens, reflecting areas with different gliomas infiltration in each white matter bundle. Histological results and DTI-based parameters were compared in anatomical regions of high- and low grade of infiltration (HIF and LIF) respectively.

Results

We detected differences in the white matter infiltration of five major white matter bundles in three groups. IDHm infiltrated left fronto-temporal subcortical areas. IDHwt were detected in the posterior-temporal and temporo-parietal regions bilaterally. IDHm/1p19q infiltrated anterior subcortical regions of the frontal lobes bilaterally. Regional differences within the same white matter bundles were detected by both TEM- and DTI analysis linked to different topographical variables. Our multimodal analysis showed that HIF regions, common to all the groups, displayed a smaller fiber diameter, lower FA and higher RD compared with LIF regions.

Conclusion

Our results suggest that the both morphological features and diffusion parameters of the white matter may be different in regions linked to the preferential location of DLGG.
P11.67.B Tumor centrality is associated with a declining overall survival in irradiated diffuse glioma
Authors: van der Boog A T J , David S , Nagtegaal S H J , Bruil D E , Cialdella F , Snijders T , Robe P A J , Verhoeff J J C
Published: September 2022

Background

For diffuse glioma, tumor location is an important factor that influences symptomatology, treatment strategy and, ultimately, survival. Our previous research showed an association between overall survival (OS) and local involvement in eloquent gray and white matter areas in the left hemisphere. Also, involvement of central anatomical structures, such as midline corpus callosum, seem to relate to poorer survival rates, when predefined categories of survival groups (>6 months, 6-24, 24< months) are established. In this study, we investigated OS in relation to centrality of the tumor and its microstructural environment defined as the Clinical Target Volume (CTV) in diffuse glioma treated with radiotherapy in a data-driven fashion.

Material and Methods

We retrospectively included 273 adult patients with histologically proven diffuse glioma who received first radiotherapy between November 2014 and July 2020 at University Medical Center Utrecht. CTV was spatially normalized to stereotaxic MNI152 space and its center of gravity (CoG) was subsequently calculated. Then, the distance between CoG of CTV to the center of the MNI brain was measured in millimeters (mm) for every tumor. A multivariable Cox-regression model included the distance between CoG of CTV and center of MNI brain, age, sex, total intracranial volume (TIV), Karnofsky Performance Status (KPS), WHO grade and extent of resection.

Results

During follow-up, 183 patients (67%) deceased and median OS of the population was 13.7 months (range 0.03 - 65.25) from start of radiotherapy. CoG of CTV to center of brain distance had a significantly inverse association with OS in a multivariable Cox-regression model (P < 0.001) with a hazard ratio 0.97 [95% Confidence Interval: 0.95-0.98] per mm.

Conclusion

In line with literature, centrality of diffuse glioma was associated with a poor survival in irradiated diffuse glioma. In fact, this association was linear, showing a better OS the greater the distance between center of CTV and center of MNI brain after correcting for known prognostic factors such as WHO grade, KPS and extent of resection. Tumor centrality is not a biomarker of involvement in specific eloquent brain regions in and of itself, but a poorer OS in these areas might indicate a proximity to large bulk of commissural white matter tracts and tumor progression along these tracts. Structures of the memory circuit like the fornix and cingulum are also centrally located. Damage to these areas are highly associated with cognitive decline in the memory domain, which is independently related to decreased survival in diffuse glioma patients. Another factor could be tumor involvement of the subventricular zone along the walls of caudate and the lateral ventricles, in which we previously showed that is also associated with worsened survival.
P11.69.B IDH1/2wildtype gliomas grade 2 and 3 with molecular glioblastoma-like profile have a distinct course of epilepsy compared toIDH1/2wildtype glioblastomas
Authors: van Opijnen M P , Tesileanu C M S , Dirven L , van der Meer P B , Wijnenga M M J , Vincent A J P E , Broekman M L D , Dubbink H J , Kros J M , van Duinen S G , Smits M , French P J , van den Bent M J , Taphoorn M J B , Koekkoek J A F
Published: September 2022

Background

IDH1/2 wildtype (IDHwt) glioma WHO grade 2 and 3 patients with pTERT mutation and/or EGFR amplification and/or +7/−10 chromosome gain/loss have a similar overall survival time as IDHwt glioblastoma patients, and are both considered glioblastoma IDHwt according to the WHO 2021 classification. However, differences in seizure onset have been observed. This study aimed to compare the course of epilepsy in the two glioblastoma subtypes.

Material and Methods

We analyzed epilepsy data of an existing cohort including IDHwt histologically lower-grade glioma WHO grade 2 and 3 with molecular glioblastoma-like profile (IDHwt hLGG) and IDHwt glioblastoma patients. Primary outcome was the incidence proportion of epilepsy during the disease course. Major secondary outcomes included onset of epilepsy, number of seizure days and antiepileptic drug (AED) polytherapy.

Results

Out of 254 patients, 78% (50/64) IDHwt hLGG and 68% (129/190) IDHwt glioblastoma patients developed epilepsy during the disease course (p=0.121). Epilepsy onset before histopathological diagnosis occurred more frequently in IDHwt hLGG compared to IDHwt glioblastoma patients (90% versus 60%, p<0.001), with a significantly longer median time to diagnosis (3.5 versus 1.3 months, p<0.001). Median total seizure days was also longer for IDHwt hLGG patients (7.0 versus 3.0, p=0.005), and they received more often AED polytherapy (32% versus 17%, p=0.028).

Conclusion

Although the incidence proportion of epilepsy during the entire disease course is similar, IDHwt hLGG patients show a significantly higher incidence of epilepsy before diagnosis and a significantly longer median time between first seizure and diagnosis compared to IDHwt glioblastoma patients, indicating a distinct clinical course.
P11.75.B Survival benefit of radiotherapy and surgery in patients with lung cancer brain metastases with poor prognosis factors
Authors: Liang L , Wang Z , Duan H , Lu J , Jiang X , Hu H , Li C , Yu C , Zhong S , Cui R , Guo X , He Z , Chen L , Mou Y
Published: September 2022

Background

Radiotherapy and surgery are the standard treatments for lung cancer brain metastases (BMs). However, limitted studies focused on the treatments for patients with lung cancer BMs with poor prognosis factors. The purpose of this study was to investigate the effects of radiotherapy and surgery in patients with lung cancer BMs with poor prognosis factors, providing reference for clinical strategies.

Material and Methods

We analyzed retrospectively 714 patients with lung cancer BMs. A 1:1 propensity score matching (PSM) was performed to balance potential confounders. Analyses of overall survival (OS) and risk factors for OS were assessed by log-rank test and Cox proportional hazard model.

Results

Age ≥65 years, Karnofsky Performance Scale (KPS) score ≤70, anaplastic large-cell lymphoma kinase (ALK)/epidermal growth factor receptor (EGFR) wild type, extracranial metastases, non-surgery and non-radiotherapy led to poor prognosis. Patients were stratified according to these factors. Radiotherapy and surgery showed no survival benefit in patients with aged ≥65 years or pretreatment KPS score ≤70 before and after PSM. Before PSM, whole brain radiotherapy (WBRT) improved the OS and predicted good prognosis in patients with ALK/EGFR wild type or extracranial metastases. WBRT also predicted good prognosis in patients with non-surgery. Stereotactic radiosurgery (SRS) improved the OS and predicted good prognosis in patients with ALK/EGFR wild type or non-surgery. WBRT plus SRS improved the OS and predicted good prognosis in patients with extracranial metastases or non-surgery. WBRT plus SRS also predicted good prognosis in patients with ALK/EGFR wild type. Surgery improved the OS and predicted good prognosis in patients with non-radiotherapy. After PSM, SRS improved the OS and predicted good prognosis in patients with non-surgery. WBRT plus SRS improved the OS and predicted good prognosis in patients with non-surgery or extracranial metastases. WBRT plus SRS also predicted good prognosis in patients with ALK/EGFR wild type. Surgery improved the OS of patients with non-radiotherapy. We defined that the treatment would provide significant survival benefit if it both prolonged the OS and predicted good prognosis. Meanwhile, the results after PSM were more convincing than the results before PSM.

Conclusion

Radiotherapy has significant survival benefit in patients with lung cancer BMs with poor prognosis factors, including patients with ALK/EGFR wild type or extracranial metastases or non-surgery. Surgery only has significant survival benefit in patients with non-radiotherapy.
P12.02.A Landscape of surfaceome and endocytome in human glioma is divergent and depends on cellular spatial organization
Authors: Governa V , Talbot H , Gonçalves de Oliveira K , Cerezo-Magaña M , Bång-Rudenstam A , Forsberg-Nilsson K , Malmström J , Bengzon J , Welinder C , Belting M
Published: September 2022

Background

Cell-surface proteins have a key role in drug development, and tumor cell-surface proteins integrated with the plasma membrane (tumor surfaceome, TS) have attracted considerable attention as targets for immunotherapies in cancer. Checkpoint inhibitor blocking antibodies, antibody drug conjugates (ADCs), and CAR-T cells are all directed at the TS. However, a remaining challenge is the lack of strategies to comprehensively map potential TS target antigens for the design of more rational, individualized treatments. Of particular relevance, ADC and other intracellular drug delivery strategies rely on targets that functionally engage in endocytic internalization.

Material and Methods

With the aim to address these challenges and to provide insight into the complexity of the TS, we have developed a versatile technology for TS mapping (TS-MAP). As proof-of-concept, we focused on primary brain tumors that remain among the most aggressive forms of cancer, and for which attempts to conquer the most common variant, glioblastoma (GBM) have failed so far. TS-MAP is compatible with primary 3D cultures and intact patient glioma tumors with preserved tissue architecture, and specifically identifies proteins capable of endocytosis as tractable targets for ADCs and other modalities requiring toxic payload internalization. Moreover, we have curated a TS classifier (SURFME) for categorization of bona fide membrane proteins.

Results

We show how cellular spatial organization (2D vs. 3D) fundamentally transforms the surfaceome and endocytome in glioma with general implications for target screening approaches. The TS-MAP platform was further applied to profile the surfaceome and endocytome landscape in a cohort of freshly resected patient gliomas. We found a highly diverse TS repertoire between patient tumors, not directly associated with grade and histology, which highlights the need for individualized approaches.

Conclusions

The TS-MAP platform and SURFME classifier should be widely applicable to a variety of brain tumors. Our findings in GBM provide new layers of understanding fundamental to the future development of immunotherapy strategies, as well as new procedures for proteomics-based target identification aimed at a better understanding of how to harness the TS for personalized immunotherapy. Our results reflect the oncogenetic multiclonality and transcriptional diversity of gliomas, which warrants future studies that elucidate how the oncogenetic profile and spatial organization synergize to shape the full complexity of the tumor microenvironment.
P12.09.B Extracellular vesicle derived-miR-146a increases melanoma brain metastasis progression via Notch signalling pathway dysregulation
Authors: Rigg E K , Wang J , Xue Z , Lunavat T , Hoang T , Parajuli H , Han M , Liu G , Bjerkvig R , Nazarov P , Nicot N , Kreis S , Wurth C , Miletic H , Sundstrøm T , Li X , Thorsen F
Published: September 2022

Background

Melanoma has the highest tropism of any cancer to metastasize to the brain, and 40% of late-stage patients develop brain metastasis. Invasion, survival, and progression of tumors is dependent on the support of the surrounding microenvironment; therefore, modulation of neighboring cells is a key factor in metastasis. Extracellular vesicles (EVs) are important in cell-to-cell signalling, shuttling proteins, RNA and DNA to alter the surroundings into a favorable tumor microenvironment. Our aims were to investigate the role of melanoma brain metastasis (MBM) derived EVs in MBM development to find possible contributing mechanisms to cancer progression for eventual therapeutic targeting.

Material and Methods

MBM-EVs isolated via sequential ultracentrifugation were injected into mice as a pre-treatment prior to intracardial injection of MBM cells. EVs were co-cultured with normal human astrocytes (NHA) to investigate phenotypic changes. MiRNA sequencing was performed on EVs collected from MBM cells and compared to NHA and melanocytes to determine a candidate miRNA for targeting. In situ hybridization was utilized to evaluate the level of miRNA in clinical patient MBM samples. Functional in vivo validation was performed by injecting miRNA knockout MBM cells into mice. Sequencing of NHA in the presence or absence of target miRNA mimic was used to determine downstream targets.

Results

Mice primed with EVs had a significant increase in MBM tumor burden, compared to non-primed mice. Co-culture with MBM-EVs resulted in NHA activation in vitro, with increased proliferation, invasion, cytokine production, and upregulation of GFAP. MiR-146a was highly upregulated in MBM EVs, and miR-146a mimics activated NHA. Patient samples had a significant increase in miR-146a expression, compared to healthy brain controls. MiR-146a knockdown in MBM mice models reduced MBM burden and prolonged animal survival. Sequencing of NHA determined NUMB, an inhibitor of the Notch signalling pathway, as a target of miR-146a. Numb and other downstream Notch proteins expression was significantly altered in NHA in the presence of both MBM-EVs and miR-146a.

Conclusion

In conclusion, EVs are important regulators of MBM and establish tumor-supporting reactive astrocytes by delivery of miR-146a. MiR-146a alters Notch signalling in astrocytes via inhibition of the tumor suppressor gene NUMB. Elevated miR-146a levels in patients suggests a potential clinical intervention is possible via miR-146a targeting.
P12.13.B Investigating the tumor - immune cell crosstalk inex vivo glioblastoma models
Authors: Rezaeipour M , Klein E , Dording C , Niclou S P , Golebiewska A
Published: September 2022

Background

Immunotherapy is a promising therapeutic approach to fight cancer by activating the immune system. Multiple immune-based strategies are under development that aim at recruiting or re-activating cellular components of the immune system. While immunotherapies have recently revolutionized cancer therapy, they have shown so far little therapeutic success in glioblastoma patients. To enhance the efficacy of novel strategies, we need to better understand the immunogenic status of glioblastoma cells and their cross-talk with immune cells in different microenvironmental niches.

Material and Methods

We assessed expression of molecules related to antigen processing and presentation as well as immune checkpoints in patient tumor databases as well as in a series of glioblastoma patient-derived organoids, 3D stem-like cultures and adherent cell lines under varying microenvironmental conditions (varying oxygen levels, inflammation). We further established an allogenic co-culture protocol for glioblastoma organoids with immune cells isolated from HLA matched donor blood, allowing for the functional assessment of the crosstalk between tumor and immune cells.

Results

Analysis of a large cohort of patient tumors and patient-derived glioblastoma preclinical models shows inter-patient heterogeneity at the level of components of major histocompatibility complex (MHC)-MHC-II, immune checkpoints. Glioblastoma cells in general express MHC-I machinery, albeit at different levels. MHC-II and immune checkpoints are variably expressed across glioblastoma cells. Different tumor microenvironment conditions, including hypoxia and interferon-γ, impact the expression of immune-related molecules. Upon co-culture, HLA-matched donor-derived T cells integrate well into the core of glioblastoma tumor organoids and display reciprocal crosstalk with tumor cells.

Conclusion

Assessing antigen presentation and immune cell responses at the functional level are key to improve patient-specific responses to immunotherapies. Advanced glioblastoma organoids incorporating the immune compartment appear as clinically-relevant models for ex vivo efficacy studies.
P14.05.B Plasma-EV based liquid biopsy for precision medicine in the treatment of glioblastoma
Authors: Faletti S , Zaccheroni E , Osti D , Del Bene M , DiMeco F , Pelicci G
Published: September 2022

Background

Glioblastoma (GBM) is a devastating and protean brain tumor. Diagnosis and molecular characterization rely on magnetic resonance imaging and brain biopsy which are challenged by low specificity/sensitivity and surgical risks. This hampers the longitudinal monitoring of patients with GBM, whose molecular dynamism ideally requires to refine the therapeutic plan in different disease stages. Extracellular vesicles (EVs) shed by GBM act as a reservoir of circulating biomarkers and can be easily collected through a blood sample, placing the rational to exploit them as a platform for liquid biopsy in GBM.

Material and Methods

We isolated EVs from 2ml of pre-operative platelet-free plasma by Size Exclusion Chromatography (SEC). EVs are characterized by (i) immunoblot and Fluorescent-Activated Cell Sorter (FACS) for the presence of EV markers; (ii) TEM for morphology and size; (iii) Nanoparticle Tracking Analysis (NTA) for size and concentration; (iv) RNAseq and ddPCR for EV-associated nucleic acid profiling.

Results

SEC allowed to define EV-enriched fractions expressing the EV-markers CD9, CD81, Flotillin1, lacking the non-EV marker Calnexin and depleted of the majority of plasma contaminants. EV quantification by NTA confirmed their enrichment in the plasma from GBM patients. EV-RNA length (< 200nt) and total yield (1-10ng among different samples) allowed a successful RNAseq library preparation for downstream identification of splicing isoforms, translocations, fused mRNAs, and expression level of GBM-specific transcripts. The ddPCR-based analysis of mutations and copy number alterations proved the ability of cell-derived EV-DNA to recapitulate the alterations of the parental cells. Although the yield of plasma-EV-DNA is low (around 1ng), this paves the way to optimize the isolation/analysis of plasma EV-DNA to permit tumor profiling.

Conclusion

Plasma-EV based liquid biopsy could implement the personalisation of GBM care for every timepoint of the disease course. Increased plasma-EV concentration is an effective biomarker for GBM presence and its measurement well suits with the clinical routine. EV-RNA/DNA mirror the genomic profile of parental tumor and their analysis could allow the identification of actionable molecular aberrations and the monitoring of GBM mutability during follow-up. Thus, the multilayered analysis of plasma-EV concentration and cargo could enable GBM early diagnosis and to monitor therapy response and the whole tumor evolution.
P15.07.A Predicting sites of local tumour progression - what should be our imaging biomarker?
Authors: Mayrand R C , Wei Y , Li C , Perry Mayrand R , Wan Y , Simon N , Sinha R , Sravanam S , Price S J
Published: September 2022

Background

Glioblastoma is the most aggressive primary brain tumour diagnosed in adults. Despite intensive treatment of maximal safe resection and chemoradiotherapy, the prognosis remains grim due to invasive tumour cells. Current treatment and standard imaging methods are highly limited in terms of managing these invasive cells as they are often located outside the area of surgical resection and are generally resistant to chemoradiation. DTI appears to be a promising tool for imaging tumour cell invasion and predicting the site of recurrence especially when decomposed into its anisotropic (q) and isotropic (p) components. The aim of this study is to investigate the sensitivity of imaging biomarkers as predictors of recurrence.

Material and Methods

All pre-op and recurrence sequences were co-registered to the pre-op post-contrast T1-weighted images as reference. Co-registration of images was performed using FSL and ANTs. The ROIs for 49 patients with a primary diagnosis of GBM were segmented using 3DSlicer. Each voxel was assigned to one of four status: true negative, false negative, true positive and false positive. Sensitivity and specificity between the pre-op ROIs and the progression region were calculated using FSL. The significance of the differences in sensitivity and specificity between the ROIs was computed in MATLAB.

Results

The sensitivity for the contrast enhancing region was 48.77 ± 26.13 (Mean ± SD) and 62.40 ± 23.07 (Mean ± SD). The abnormal q alone has a significantly greater sensitivity than the contrast enhancing region (t = -2.7327, df = 96, p-value = 0.0075). The sensitivity for the ROI of combined contrast enhancement and abnormal q was 65.86 ± 23.29 (Mean ± SD). There is an even more significant increase in sensitivity when the contrast enhancing ROI is combined with abnormal q region (t = -3.4133, df = 96, p-value = 9.4123e-04) compared to when it is alone. There was no statistical difference in the specificities of the different ROIs.

Conclusion

Current surgical and radiation volumes focus solely on pre-op contrast enhancement. However, these results suggest that combining the abnormal q with the standard contrast enhancing region is a more sensitive predictor of tumour recurrence than contrast enhancement alone, while still retaining high specificity. The higher sensitivity is an indicator of correct identification of tumour recurrence while the high specificity correctly identifies normal brain, or non-recurrent regions. These results are currently being prospectively assessed in a multi-centre study (PRaM-GBM).
P16.02.B Leptomeningeal dissemination in gliomas in National Research Institute of Oncology in Gliwice experience
Authors: Nowicka E , Grzbiela H , Widera K , Zarudzki L , Grządziel A , Tarnawski R
Published: September 2022

Background

The leptomeningeal dissemination (LMD) in the course of gliomas is rare. According to literature it can affect 4.7% of all patients with gliomas. LMD can be diagnosed at the initial presentation or at the time of recurrence. The standard therapy has not yet been determined. We aimed to present the clinical features and prognosis in retrospective analysis of patients with LMD in gliomas in National Institute of Oncology in Gliwice, Poland experience.

Material and Methods

Twenty patients with leptomeningeal dissemination in course of treatment of brain gliomas were identified. There were 16 men and 4 women. Median age of pts was 35,5 y (range 18-51). All tumours were supratentorial at diagnosis. There were 6 Diffuse Astocytomas, 3 oligodendrioglimas and 11 glioblatomas. In two patients LMD was found at initial diagnosis of GBM. The majority of patients (15 pts) presented clinical symptoms and in 5 asymptomatic pts LMD was diagnosed in MRI. The diagnosis was confirmed in craniospinal MRI. One patient was operated due to rapid clinical progression of spinal compression symptoms. All but one patient were offered palliative therapy. Radiotherapy was prescribed to 16 patients: in 7 pts for whole brain to C2, in 3 pts for craniospinal axis, in 7 for spinal canal and in 2 pts local therapy was performed. Mean dose of RT was 28,7 Gy/g (range 18-45 Gy/g). In 8 patients radiotherapy was combined with chemotherapy and chemotherapy as sole treatment was prescribed to 2 pts (Lomustin, EP).

Results

Mean time from initial diagnosis to LMD diagnosis was 46 mo (range 0,4-199 mo) for all patients and for non-GBM and GBM patients were 81,4 mo (16-199 mo) and 14,0 mo (0,4-27 mo) respectively. In 7 patients LMD was diagnosed concurrently with local relapse. In remaining mean time from local relapse to LMD was 4,6 mo (1,2-20,0 mo). Mean OS dated from time of LMD diagnosis for whole group was 5,9 mo (0,4-27 ), for non-GBM and GBM patients were 9,2 mo (3,6-27 mo) and 2,94 (0,4-12,7 mo) respectively. The palliative effect as improvement in clinical symptoms was achieved in 15 pts.

Conclusion

LMD in gliomas is rare but should be kept in mind while clinical symptoms in glioma patients occur. The prognosis of LMD is poor. Radiotherapy have palliative intent and it can affect the clinical symptoms and survival.
P17.01.A Reradiation by Glioblastoma recurrence with VMAT technique
Authors: Theodorou M , Polycarpou I
Published: September 2022

Background

Patients who have been treated with reirradiation for recurrent glioma reported survival benefits.

This study aims to investigate whether re-irradiation of recurrent glioma with 45Gy dose can increase the overall survival of patients.

Material and Methods

A retrospective analysis of 35 patients re-irradiated for high-grade glioma recurrence between 2012 and 2020 was performed. All included patients met the following criteria: a) histopathological confirmation of primary brain cancer at initial diagnosis; b) a history of initial primary radiation; c) histological and/or imaging modality confirmation of recurrence. Outcome metrics included overall survival, prognostic factors for survival, and treatment-related toxicity.

Results

After the end of re-irradiation the median overall survival was 11 months (95% confidence interval, 7-14 months). From the patients evaluated in the current study after the end of re-irradiation the progression free survival was 6 months (3.8 - 8 months) while after the end of first radiation was 13 months (8 - 17.9 months). Our findings suggest that re-irradiation might prolong survival rates.

Conclusion

Recurrent Glioblastoma WHO IV is associated with a median overall survival of less than a year and the majority of patients have profound tumor-related symptoms .The results of this study suggest that re-irradiation may prolong the overall survival.
P17.03.B Modelling migration of glioblastoma patient-derived cells using human iPSC-derived neural spheroid and high content analysis
Authors: Tsang V S , Lieberam I , Danovi D
Published: September 2022

Background

Glioblastoma multiforme (GBM) is the most common and aggressive brain tumour in adults. Despite current advances, the existing standard of treatment is ineffective, and the survival prognosis remains just over a year from diagnosis. Migrating tumour cells have been implicated in the therapeutic resistance of GBM. They spread by interacting with structures such as white matter tracts and inevitably cause recurrence of the tumour. Valuable cell models able to capture the invasiveness of GBM are critically needed to develop innovative therapies targeting migrating GBM cells.

Material and Methods

We established an in vitro model mimicking the GBM microenvironment by co-culturing patient-derived GBM cells and human induced pluripotent stem cell-derived cortical neural spheroids with radiating axons. Using high content imaging, we developed a robust workflow to quantify the GBM cells infiltration of the neural spheroid in endpoint assays. Images were acquired on the Operetta CLS high content device (Perkin Elmer) and analysed using the built-in Harmony Imaging and Analysis Software. We also performed live imaging assays using the Livecyte quantitative phase imager (Phasefocus), in which we studied the directionality, displacement and speed of the GBM cells engaged on axons.

Results

Our data indicate that GBM cells change morphology when cultured on axons and that they migrated towards the neural spheroid once engaged on axons. We showed that cell lines from different patients vary in migratory properties as well as in levels of infiltration capability. Finally, we used this model to test several antagonists to pathways involved in migration of GBM cells.

Conclusion

The main deliverable of this project is the setup of a novel model able to mimic the GBM migration on axons and able to screen for compounds affecting cell migration. This could potentially offer innovative precision-medicine therapies.
P17.07.A Metronomic temozolomide therapy in heavily pretreated patients with recurrent glioblastoma: a large mono-institutional retrospective study
Authors: Bosio A , Padovan M , Caccese M , Cerretti G , Zagonel V , Lombardi G
Published: September 2022

Background

Glioblastoma (GBM) is the most common and aggressive primary brain cancer. Despite advances in surgical and first-line treatment, all pts relapse. The aim of this study is to evaluate the benefit of metronomic Temozolomide (mTMZ) for recurrent GBM.

Material and Methods

All pts treated at Veneto Institute of Oncology from September 2013 to March 2021 were retrospectively reviewed. Major inclusion criteria were: first-line therapy with Stupp protocol, relapse after first or subsequent line of therapy, treatment with mTMZ schedule (50mg/m2 continuously), hystologically confirmed diagnosis of GBM. RANO criteria and CTCAE v 5.0 were used for response and toxicity assessment.

Results

120pts were enrolled. Median follow-up was 15.6ms. Median age was 59ys (range 18-81), ECOG-PS was 0-2 in 107pts (89%) and 3 in 11 (9%). MGMT was methylated and IDH mutated in 66 of 105 (62%) and in 9 of 106 (8%) evaluable pts, respectively. Median number of prior lines of treatment was 2 (range 1-7) and 41% of pts received the therapy beyond the third line. Median time between the last standard maintenance TMZ (sTMZ) cycle and the mTMZ administration was 6ms (range 1-50) and 40% of pts started mTMZ after 3ms from sTMZ. All pts were evaluable for response: 3 (2%) and 48 (40%) showed PR and SD. mOS from the start of mTMZ was 5.4ms (95% CI 4.3-6.4), mPFS was 2.6ms (95% CI 2.3-2.8). At univariate analysis, MGMTmet and MGMTunmet pts had a mOS of 5.6 and 4.4ms (p=0.03); mOS for patients with ECOG-PS > or ≤2 was 2.3 and 6.0ms (p<0.001); number of prior lines of therapies, time between sTMZ and mTMZ and age were not significant. At multivariate analysis, MGMT methylated status (HR=2.3, 95% CI, p=0.004) and ECOG-PS (HR=0.5, 95% CI, p=0.017) remained statistically significant for PFS, while ECOG-PS (HR=0.4, 95% CI, p=0.001) was the only factor significantly associated with OS. The most common grade 3-4 hematologic toxicities were lymphopenia (10%) and thrombocytopenia (3%). Grade 3-4 nonhematologic toxicities were uncommon.

Conclusion

Rechallenge with mTMZ can be a well tolerated treatment option for recurrent GBM, even in heavily pretreated pts. Pts with MGMTmet and good ECOG-PS might report the major benefit.