Classification of Brain Tumors by Nanopore Sequencing of Cell-Free DNA from Cerebrospinal Fluid.

BACKGROUND: Molecular brain tumor diagnosis is usually dependent on tissue biopsies or resections. This can pose several risks associated with anesthesia or neurosurgery, especially for lesions in the brain stem or other difficult-to-reach anatomical sites. Apart from initial diagnosis, tumor progression, recurrence, or the acquisition of novel genetic alterations can only be proven by re-biopsies. METHODS: We employed Nanopore sequencing on cell-free DNA (cfDNA) from cerebrospinal fluid (CSF) and analyzed copy number variations (CNV) and global DNA methylation using a random forest classifier. We sequenced 129 samples with sufficient DNA. These samples came from 99 patients and encompassed 22 entities. Results were compared to clinical diagnosis and molecular analysis of tumor tissue, if available. RESULTS: 110/129 samples were technically successful, and 50 of these contained detectable circulating tumor DNA (ctDNA) by CNV or methylation profiling. ctDNA was detected in samples from patients with progressive disease but also from patients without known residual disease. CNV plots showed diagnostic and prognostic alterations, such as C19MC amplifications in embryonal tumors with multilayered rosettes or Chr.1q gains and Chr.6q losses in posterior fossa group A ependymoma, respectively. Most CNV profiles mirrored the profiles of the respective tumor tissue. DNA methylation allowed exact classification of the tumor in 22/110 cases and led to incorrect classification in 2/110 cases. Only 5/50 samples with detected ctDNA contained tumor cells detectable through microscopy. CONCLUSIONS: Our results suggest that Nanopore sequencing data of cfDNA from CSF samples may be a promising approach for initial brain tumor diagnostics and an important tool for disease monitoring.

Pituitary neuroendocrine tumors with PIT1/SF1 co-expression show distinct clinicopathological and molecular features.

Pituitary neuroendocrine tumors (PitNETs) are classified according to cell lineage, which requires immunohistochemistry for adenohypophyseal hormones and the transcription factors (TFs) PIT1, SF1, and TPIT. According to the current WHO 2022 classification, PitNETs with co-expression of multiple TFs are termed "plurihormonal". Previously, PIT1/SF1 co-expression was prevailingly reported in PitNETs, which otherwise correspond to the somatotroph lineage. However, little is known about such tumors and the WHO classification has not recognized their significance. We compiled an in-house case series of 100 tumors, previously diagnosed as somatotroph PitNETs. Following TF staining, histopathological features associated with PIT1/SF1 co-expression were assessed. Integration of in-house and publicly available sample data allowed for a meta-analysis of SF1-associated clinicopathological and molecular features across a total of 270 somatotroph PitNETs. The majority (74%, 52/70) of our densely granulated somatotroph PitNETs (DGST) unequivocally co-expressed PIT1 and SF1 (DGST-PIT1/SF1). None (0%, 0/30) of our sparsely granulated somatotroph PitNETs (SGST) stained positive for SF1 (SGST-PIT1). Among DGST, PIT1/SF1 co-expression was significantly associated with scarce FSH/LH expression and fewer fibrous bodies compared to DGST-PIT1. Integrated molecular analyses including publicly available samples confirmed that DGST-PIT1/SF1, DGST-PIT1 and SGST-PIT1 represent distinct tumor subtypes. Clinicopathological meta-analyses indicated that DGST-PIT1 respond more favorably towards treatment with somatostatin analogs compared to DGST-PIT1/SF1, while both these subtypes show an overall less aggressive clinical course than SGST-PIT1. In this study, we spotlight that DGST with co-expression of PIT1 and SF1 represent a common, yet underrecognized, distinct PitNET subtype. Our study questions the rationale of generally classifying such tumors as "plurihormonal", and calls for a refinement of the WHO classification. We propose the term "somatogonadotroph PitNET".

Temporal change of DNA methylation subclasses between matched newly diagnosed and recurrent glioblastoma.

The longitudinal transition of phenotypes is pivotal in glioblastoma treatment resistance and DNA methylation emerged as an important tool for classifying glioblastoma phenotypes. We aimed to characterize DNA methylation subclass heterogeneity during progression and assess its clinical impact. Matched tissues from 47 glioblastoma patients were subjected to DNA methylation profiling, including CpG-site alterations, tissue and serum deconvolution, mass spectrometry, and immunoassay. Effects of clinical characteristics on temporal changes and outcomes were studied. Among 47 patients, 8 (17.0%) had non-matching classifications at recurrence. In the remaining 39 cases, 28.2% showed dominant DNA methylation subclass transitions, with 72.7% being a mesenchymal subclass. In general, glioblastomas with a subclass transition showed upregulated metabolic processes. Newly diagnosed glioblastomas with mesenchymal transition displayed increased stem cell-like states and decreased immune components at diagnosis and exhibited elevated immune signatures and cytokine levels in serum. In contrast, tissue of recurrent glioblastomas with mesenchymal transition showed increased immune components but decreased stem cell-like states. Survival analyses revealed comparable outcomes for patients with and without subclass transitions. This study demonstrates a temporal heterogeneity of DNA methylation subclasses in 28.2% of glioblastomas, not impacting patient survival. Changes in cell state composition associated with subclass transition may be crucial for recurrent glioblastoma targeted therapies.

PerSurge (NOA-30) phase II trial of perampanel treatment around surgery in patients with progressive glioblastoma.

BACKGROUND: Glioblastoma is the most frequent and a particularly malignant primary brain tumor with no efficacy-proven standard therapy for recurrence. It has recently been discovered that excitatory synapses of the AMPA-receptor subtype form between non-malignant brain neurons and tumor cells. This neuron-tumor network connectivity contributed to glioma progression and could be efficiently targeted with the EMA/FDA approved antiepileptic AMPA receptor inhibitor perampanel in preclinical studies. The PerSurge trial was designed to test the clinical potential of perampanel to reduce tumor cell network connectivity and tumor growth with an extended window-of-opportunity concept. METHODS: PerSurge is a phase IIa clinical and translational treatment study around surgical resection of progressive or recurrent glioblastoma. In this multicenter, 2-arm parallel-group, double-blind superiority trial, patients are 1:1 randomized to either receive placebo or perampanel (n = 66 in total). It consists of a treatment and observation period of 60 days per patient, starting 30 days before a planned surgical resection, which itself is not part of the study interventions. Only patients with an expected safe waiting interval are included, and a safety MRI is performed. Tumor cell network connectivity from resected tumor tissue on single cell transcriptome level as well as AI-based assessment of tumor growth dynamics in T2/FLAIR MRI scans before resection will be analyzed as the co-primary endpoints. Secondary endpoints will include further imaging parameters such as pre- and postsurgical contrast enhanced MRI scans, postsurgical T2/FLAIR MRI scans, quality of life, cognitive testing, overall and progression-free survival as well as frequency of epileptic seizures. Further translational research will focus on additional biological aspects of neuron-tumor connectivity. DISCUSSION: This trial is set up to assess first indications of clinical efficacy and tolerability of perampanel in recurrent glioblastoma, a repurposed drug which inhibits neuron-glioma synapses and thereby glioblastoma growth in preclinical models. If perampanel proved to be successful in the clinical setting, it would provide the first evidence that interference with neuron-cancer interactions may indeed lead to a benefit for patients, which would lay the foundation for a larger confirmatory trial in the future. TRIAL REGISTRATION: EU-CT number: 2023-503938-52-00 30.11.2023.

Plasma extracellular vesicles in meningioma patients following radiotherapy as liquid biopsy- a prospective explorative biomarker study (ARO 2023-05/AG-NRO-07).

BACKGROUND: While surgical resection remains the primary treatment approach for symptomatic or growing meningiomas, radiotherapy represents an auspicious alternative in patients with meningiomas not safely amenable to surgery. Biopsies are often omitted in light of potential postoperative neurological deficits, resulting in a lack of histological grading and (molecular) risk stratification. In this prospective explorative biomarker study, extracellular vesicles in the bloodstream will be investigated in patients with macroscopic meningiomas to identify a biomarker for molecular risk stratification and disease monitoring. METHODS: In total, 60 patients with meningiomas and an indication of radiotherapy (RT) and macroscopic tumor on the planning MRI will be enrolled. Blood samples will be obtained before the start, during, and after radiotherapy, as well as during clinical follow-up every 6 months. Extracellular vesicles will be isolated from the blood samples, quantified and correlated with the clinical treatment response or progression. Further, nanopore sequencing-based DNA methylation profiles of plasma EV-DNA will be generated for methylation-based meningioma classification. DISCUSSION: This study will explore the dynamic of plasma EVs in meningioma patients under/after radiotherapy, with the objective of identifying potential biomarkers of (early) tumor progression. DNA methylation profiling of plasma EVs in meningioma patients may enable molecular risk stratification, facilitating a molecularly-guided target volume delineation and adjusted dose prescription during RT treatment planning.

Defining benchmark outcomes for mesial temporal lobe epilepsy surgery: A global multicenter analysis of 1119 cases.

OBJECTIVE: Benchmarking has been proposed to reflect surgical quality and represents the highest standard reference values for desirable results. We sought to determine benchmark outcomes in patients after surgery for drug-resistant mesial temporal lobe epilepsy (MTLE). METHODS: This retrospective multicenter study included patients who underwent MTLE surgery at 19 expert centers on five continents. Benchmarks were defined for 15 endpoints covering surgery and epilepsy outcome at discharge, 1 year after surgery, and the last available follow-up. Patients were risk-stratified by applying outcome-relevant comorbidities, and benchmarks were calculated for low-risk ("benchmark") cases. Respective measures were derived from the median value at each center, and the 75th percentile was considered the benchmark cutoff. RESULTS: A total of 1119 patients with a mean age (range) of 36.7 (1-74) years and a male-to-female ratio of 1:1.1 were included. Most patients (59.2%) underwent anterior temporal lobe resection with amygdalohippocampectomy. The overall rate of complications or neurological deficits was 14.4%, with no in-hospital death. After risk stratification, 377 (33.7%) benchmark cases of 1119 patients were identified, representing 13.6%-72.9% of cases per center and leaving 742 patients in the high-risk cohort. Benchmark cutoffs for any complication, clinically apparent stroke, and reoperation rate at discharge were ≤24.6%, ≤.5%, and ≤3.9%, respectively. A favorable seizure outcome (defined as International League Against Epilepsy class I and II) was reached in 83.6% at 1 year and 79.0% at the last follow-up in benchmark cases, leading to benchmark cutoffs of ≥75.2% (1-year follow-up) and ≥69.5% (mean follow-up of 39.0 months). SIGNIFICANCE: This study presents internationally applicable benchmark outcomes for the efficacy and safety of MTLE surgery. It may allow for comparison between centers, patient registries, and novel surgical and interventional techniques.

Multiple craniotomies in a single surgery - the resection of scattered brain metastases.

Patients with brain metastases (BM), who can benefit from resection of multiple scattered lesions, often will not be offered a procedure involving multiple craniotomies in one session due to the overall poor prognosis. However, carefully selected candidates may well benefit from the resection of multiple lesions using multiple craniotomies through a significantly shortened hospital stay, aggressive decompression, and rapid eligibility for adjuvant therapies. In this retrospective analysis, the records of patients, who were treated for multiple BM using one surgical session involving multiple craniotomies, were reviewed. A group of patients with multiple BM, whose surgery only involved one craniotomy, were assigned to a control group. Clinical and surgical characteristics, preoperative and postoperative Karnofsky Performance Scale (KPS), complication rate, preoperative tumor size, number of lesions, number of craniotomies, skin incisions, and intraoperative repositioning of patients were recorded. Thirty-three patients were included in the multiple-craniotomy group. Thirty patients underwent two craniotomies, while three cases involved three craniotomies. Seven patients (21%) were intraoperatively repositioned from a prone to a supine position, which required an average of 23.3 ± 9.3 min from wound closure to the following skin incision. Thirty-six patients with multiple BM and matching characteristics, who received only one craniotomy for the dominant lesion, served as the control group. No difference was detected in postoperative KPS (p = 0.269), complication rate (p = 0.612), rate of new postoperative neurological deficits (p = 0.278), length of intensive care unit (ICU) (p = 0.991), and hospital stay (p = 0.913). There was a significant difference in average preoperative tumor size (p = 0.002), duration of surgery (p < 0.001), and extent of resection (p = 0.002). In the age of personalized medicine, selected patient may benefit from a single surgery for BM using multiple craniotomies. This study shows no significant increase of the perioperative complication rate for surgeries with multiple craniotomies.

Association of the classification of intraoperative adverse events (ClassIntra) with complications and neurological outcome after neurosurgical procedures: a prospective cohort study.

PURPOSE: To analyze the reliability of the classification of intraoperative adverse events (ClassIntra) to reflect intraoperative complications of neurosurgical procedures and the potential to predict the postoperative outcome including the neurological performance. The ClassIntra classification was recently introduced and found to be reliable for assessing intraoperative adverse events and predicting postoperative complications across different surgical disciplines. Nevertheless, its potential role for neurosurgical procedures remains elusive. METHODS: This is a prospective, monocentric cohort study assessing the ClassIntra in 422 adult patients who underwent a neurosurgical procedure and were hospitalized between July 1, 2021, to December 31, 2021. The primary outcome was the occurrence of intraoperative complications graded according to ClassIntra and the association with postoperative outcome reflected by the Clavien-Dindo classification and comprehensive complication index (CCI). The ClassIntra is defined as intraoperative adverse events as any deviation from the ideal course on a grading scale from grade 0 (no deviation) to grade V (intraoperative death) and was set at sign-out in agreement between neurosurgeon and anesthesiologist. Secondary outcomes were the neurological outcome after surgery as defined by Glasgow Coma Scale (GCS), modified Rankin scale (mRS), Neurologic Assessment in Neuro-Oncology (NANO) scale, National Institute Health of Strokes Scale (NIHSS), and Karnofsky Performance Score (KPS), and need for unscheduled brain scan. RESULTS: Of 442 patients (mean [SD] age, 56.1 [16.2]; 235 [55.7%] women and 187 [44.3%] men) who underwent a neurosurgical procedure, 169 (40.0%) patients had an intraoperative adverse event (iAE) classified as ClassIntra I or higher. The NIHSS score at admission (OR, 1.29; 95% CI, 1.03-1.63, female gender (OR, 0.44; 95% CI, 0.23-0.84), extracranial procedures (OR, 0.17; 95% CI, 0.08-0.61), and emergency cases (OR, 2.84; 95% CI, 1.53-3.78) were independent risk factors for a more severe iAE. A ClassIntra ≥ II was associated with increased odds of postoperative complications classified as Clavien-Dindo (p < 0.01), neurological deterioration at discharge (p < 0.01), prolonged hospital (p < 0.01), and ICU stay (p < 0.01). For elective craniotomies, severity of ClassIntra was associated with the CCI (p < 0.01) and need for unscheduled CT or MRI scan (p < 0.01). The proportion of a ClassIntra ≥ II was significantly higher for emergent craniotomies (56.2%) and associated with in-hospital mortality, and an unfavorable neurological outcome (p < 0.01). CONCLUSION: Findings of this study suggest that the ClassIntra is sensitive for assessing intraoperative adverse events and sufficient to identify patients with a higher risk for developing postoperative complications after a neurosurgical procedure.

High-Resolution Imaging Flow Cytometry Reveals Impact of Incubation Temperature on Labeling of Extracellular Vesicles with Antibodies.

Extracellular vesicles (EVs) are released from basically all cells. Over the last decade, small EVs (sEVs; 50-150 nm) have gained enormous attention in diagnostics and therapy. However, methodological limitations coupled to the lack of EV standards leave many questions in this quickly evolving field unresolved. Recently, by using enhanced green fluorescent protein (eGFP)-labeled sEVs as biological reference material, we systematically optimized imaging flow cytometry for single sEV analysis. Furthermore, we showed that sEVs stained with different fluorescent antibodies can be analyzed in a multiparametric manner. However, many parameters potentially affecting the sEV staining procedure still require further evaluation and optimization. Here, we present a concise, systematic evaluation of the impact of the incubation temperature (4°C, room temperature and 37°C) during sEV antibody staining on the outcome of experiments involving the staining of EVs with fluorescence-conjugated antibodies. We provide evidence that both the staining intensity and the sample recovery can vary depending on the incubation temperature applied, and that observed differences are less pronounced following prolonged incubation times. In addition, this study can serve as an application-specific example of parameter evaluation in EV flow cytometry. © 2020 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry.

Mass Spectrometric Lipid Profiles of Picosecond Infrared Laser-Generated Tissue Aerosols Discriminate Different Brain Tissues.

BACKGROUND AND OBJECTIVES: A picosecond infrared laser (PIRL) has recently been demonstrated to cut biological tissue without scar formation based on the minimal destructive action on the surrounding cells. During cutting with PIRL, the irradiated tissue is ablated by a cold vaporization process termed desorption by impulsive vibrational excitation. In the resulting aerosol, all molecules are dissolved in small droplets and even labile biomolecules like proteins remain intact after ablation. It is hypothesized that these properties enable the PIRL in combination with mass spectrometry as an intelligent laser scalpel for guided surgery. In this study, it was tested if PIRL-generated tissue aerosols are applicable for direct analysis with mass spectrometry, and if the acquired mass spectra can be used to discriminate different brain areas. MATERIALS AND METHODS: Brain tissues were irradiated with PIRL. The aerosols were collected and directly infused into a mass spectrometer via electrospray ionization without any sample preparation or lipid extraction. RESULTS: The laser produced clear cuts with no marks of burning. Lipids from five different classes were identified in the mass spectra of all samples. By principal component analysis the different brain areas were clearly distinguishable from each other. CONCLUSIONS: The results demonstrate the potential for real-time analysis of lipids with a PIRL-based laser scalpel, coupled to a mass spectrometer, for the discrimination of tissues during surgeries. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc.

Decision-making in temporal lobe epilepsy surgery based on invasive stereo-electroencephalography (sEEG).

In medical refractory temporal lobe epilepsy (TLE), the epileptogenic zone can be difficult to identify and therefore difficult to treat, especially in the absence of clear MRI pathologies and specific results from presurgical evaluation. Invasive monitoring with stereo-electroencephalography (sEEG) is a tool for a better determination of the epileptogenic zone. Here, we investigate the impact of sEEG on decision-making in temporal lobe epilepsy surgery. We reviewed patients with TLE who underwent further investigation with sEEG in our epilepsy unit. We examined specifically how sEEG findings influenced our decision regarding indication for a surgical procedure and resection volume. From 2013 to 2017, we performed 152 temporal resections in epilepsy patients. Twenty-one of these patients were designated for further preoperative investigation with sEEG due to incongruent findings in presurgical evaluation. Six patients were implanted bitemporally. In five cases, the hypothesis for the epileptogenic zone and localization had to be changed due to sEEG findings and resulted in a different tailored resection than intended. In three cases, sEEG findings led to the cancelation of the originally intended temporal resection as the epileptogenic zone was not definable or bilateral. In another three cases, the prognosis for reduction of seizures postoperatively had to be reduced due to the sEEG findings. However, the resection was performed after interdisciplinary discussion and informed consent of the patient. The examination by sEEG led to a change of plan for further treatment in 13 patients (61.9%) suffering TLE in total. Invasive monitoring with sEEG electrodes had a strong impact on decision-making for further treatment in patients suffering from temporal lobe epilepsy with incongruent findings in presurgical examination designated for epilepsy surgery. This applies to resection volumes as well as to prediction of seizure outcome.

FASN Is a Biomarker Enriched in Malignant Glioma-Derived Extracellular Vesicles.

Extracellular vesicles (EVs) are known for their important role in cancer progression and hold considerable potential as a source for tumor biomarkers. However, purification of tumor-specific EVs from patient plasma is still an urgent unmet need due to contamination by normal host cell-derived EVs, that results in compromised analytical sensitivity. Here we identified fatty acid synthase (FASN), a key lipogenic enzyme which is highly expressed in malignant glioma cells, to be elevated in CD63- and CD81-positive EVs in glioma patient plasma samples, opening vital opportunities to sort brain tumor-specific EVs.

Seizures as presenting symptom in patients with glioblastoma.

OBJECTIVE: The clinical course and underlying molecular causes in patients with glioblastoma presenting with seizures are poorly understood. Here we investigated clinical features and carrier systems as well as a transaminase relevant in glutamate homeostasis in patients with glioblastoma. METHODS: We performed a retrospective analysis of our clinical glioma database for clinical data during a 2-year period. Patients with glioblastoma were divided into 2 groups: symptomatic and asymptomatic for seizures. Magnetic resonance imaging (MRI) scans and tissue samples from both groups were investigated. A Cox regression analysis was performed for survival and clinical and molecular features. RESULTS: One hundred three patients diagnosed with glioblastoma in this period were identified. Twenty-three patients were symptomatic with seizures (22.3%). All were IDH-1/2 wild-type. We found no significant difference in the tumor localization between the groups. Patients with seizures from glioblastoma had significantly smaller tumors, which caused less edema compared to nonepileptogenic tumors. A significantly increased up-regulation of glutamate carrier systems was evident in symptomatic tumors compared to asymptomatic tumors. Moreover, there seems to be an oversupply of glutamate in symptomatic tumors due to dysregulation in glutamate synthesis. SIGNIFICANCE: Glioblastoma presenting with seizures is morphologically different from asymptomatic tumors. Furthermore, we were able to show that the molecular profile of these tumors, particularly glutamate homeostasis controlling systems, is significantly different.

Relapse of a group 4 medulloblastoma after 18 years as proven by histology and DNA methylation profiling.

BACKGROUND: Recent studies on medulloblastomas (MB) suggest that a large fraction of tumors appearing as late recurrence turn out to be secondary malignancies, e.g., malignant gliomas, after thorough molecular investigation. RESULTS: Here, we report of a patient with a group 4 MB that developed a distant recurrence after more than 18 years. The recurrent tumor was confirmed by histology and genome-wide DNA methylation profiling. CONCLUSION: Our case not only illustrates the potential of very late recurrences after seemingly cured group 4 MB, but also illustrates that detailed molecular analyses are indispensable in patients with a history of a previous malignancy.

The simplified acute physiology score II to predict hospital mortality in aneurysmal subarachnoid hemorrhage.

BACKGROUND: Early prediction of increased morbidity and mortality in aneurysmal subarachnoid hemorrhage (aSAH) remains crucial to improving patient management. Most prediction models lack external validation and focus on disease-specific items without considering physiological parameters and the past medical history. The aim was to assess the validity of the established Simplified Acute Physiology Score II (SAPS-II) in an aSAH cohort for the prediction of hospital mortality and to identify additional physiological and clinical predictors. METHODS: The predictive value of SAPS-II for hospital mortality was assessed in a retrospective analysis of 263 consecutive patients with aSAH. Additional physiological and clinical parameters including the past medical history were analyzed by forward selection multivariate analysis to identify independent predictors of hospital mortality and to improve the prediction model. RESULTS: The SAPS-II predicted hospital mortality with an area under the curve (AUC) of 0.834 with an odds ratio (OR) of 1.097 [95 % confidence interval 1.067-1.128) for each additional point. Forward selection multivariate analysis identified the Glasgow Coma Scale score (P < 0.001), history of chronic headache (P = 0.01) and medication with anticoagulants (P = 0.04) as independent predictors of hospital mortality. Adding these parameters to the SAPS-II, the AUC increased to 0.86. CONCLUSION: This study validates the predictive accuracy of SAPS-II for hospital mortality in aSAH patients. Additional parameters from the past medical history increase its predictive power. From a practical viewpoint, SAPS-II alone already represents a sufficient and powerful score to predict hospital mortality at an early time point and may help to improve patient management.

Impact of intraventricular hemorrhage measured by Graeb and LeRoux score on case fatality risk and chronic hydrocephalus in aneurysmal subarachnoid hemorrhage.

BACKGROUND: Reliable prognostic tools to estimate the case fatality rate (CFR) and the development of chronic hydrocephalus (CHC) in aneurysmal subarachnoid hemorrhage (SAH) are not well defined. This study aims to investigate the practicability and reliability of Fisher, Graeb, and LeRoux scores for SAH patient prognosis. METHODS: A total of 206 patients with aneurysmal SAH were retrospectively analyzed in prediction of CFR and CHC. Clinical data was evaluated and grading was performed using Fisher, Graeb, and LeRoux scores. Univariate and multivariate analyses were performed to identify relevant predictive parameters. RESULTS: CFR was 17.0 % and was associated with higher age, higher Hunt & Hess (H&H) grade, lower Glasgow Coma Scale (GCS) at admission, as well as a higher Fisher, Graeb, and LeRoux score (p < 0.001). There were 19.9 % that developed CHC requiring permanent cerebrospinal fluid diversion. Low initial GCS (p = 0.003), high H&H (p < 0.001), intracerebral hematoma (p = 0.003), high Fisher (p = 0.047), Graeb and LeRoux scores (p < 0.001) were associated with a higher rate of ventricular-peritoneal shunting (VPS) in surviving patients. In multivariate analyses, Graeb score (odds ratio (OR) 1.183 [1.027, 1.363], p = 0.020), LeRoux score (OR 1.120 [1.013-1.239, p = 0.027), and H&H (OR 2.715 [1.496, 4.927], p = 0.001) remained independent prognostic factors for VPS. CONCLUSIONS: Graeb or LeRoux scores improve the prediction of shunt dependency and in parts of CFR in aneurysmal SAH patients therefore confirming the relevance of the extent and distribution of intraventricular blood for the clinical course in SAH.

Unclassifiable CNS tumors in DNA methylation-based classification: clinical challenges and prognostic impact.

DNA methylation analysis has become a powerful tool in neuropathology. Although DNA methylation-based classification usually shows high accuracy, certain samples cannot be classified and remain clinically challenging. We aimed to gain insight into these cases from a clinical perspective. To address, central nervous system (CNS) tumors were subjected to DNA methylation profiling and classified according to their calibrated score using the DKFZ brain tumor classifier (V11.4) as "≥ 0.84" (score ≥ 0.84), "0.3-0.84" (score 0.3-0.84), or "< 0.3" (score < 0.3). Histopathology, patient characteristics, DNA input amount, and tumor purity were correlated. Clinical outcome parameters were time to treatment decision, progression-free, and overall survival. In 1481 patients, the classifier identified 69 (4.6%) tumors with an unreliable score as "< 0.3". Younger age (P < 0.01) and lower tumor purity (P < 0.01) compromised accurate classification. A clinical impact was demonstrated as unclassifiable cases ("< 0.3") had a longer time to treatment decision (P < 0.0001). In a subset of glioblastomas, these cases experienced an increased time to adjuvant treatment start (P < 0.001) and unfavorable survival (P < 0.025). Although DNA methylation profiling adds an important contribution to CNS tumor diagnostics, clinicians should be aware of a potentially longer time to treatment initiation, especially in malignant brain tumors.

Mean global DNA methylation serves as independent prognostic marker in IDH-wildtype glioblastoma.

BACKGROUND: The IDH-wildtype glioblastoma (GBM) patients have a devastating prognosis. Here, we analyzed the potential prognostic value of global DNA methylation of the tumors. METHODS: DNA methylation of 492 primary samples and 31 relapsed samples, each treated with combination therapy, and of 148 primary samples treated with radiation alone were compared with patient survival. We determined the mean methylation values and estimated the immune cell infiltration from the methylation data. Moreover, the mean global DNA methylation of 23 GBM cell lines was profiled and correlated to their cellular radiosensitivity as measured by colony formation assay. RESULTS: High mean DNA methylation levels correlated with improved survival, which was independent from known risk factors (MGMT promoter methylation, age, extent of resection; P = 0.009) and methylation subgroups. Notably, this correlation was also independent of immune cell infiltration, as higher number of immune cells indeed was associated with significantly better OS but lower mean methylation. Radiosensitive GBM cell lines had a significantly higher mean methylation than resistant lines (P = 0.007), and improved OS of patients treated with radiotherapy alone was also associated with higher DNA methylation (P = 0.002). Furthermore, specimens of relapsed GBM revealed a significantly lower mean DNA methylation compared to the matching primary tumor samples (P = 0.041). CONCLUSIONS: Our results indicate that mean global DNA methylation is independently associated with outcome in glioblastoma. The data also suggest that a higher DNA methylation is associated with better radiotherapy response and less aggressive phenotype, both of which presumably contribute to the observed correlation with OS.