Long-term elevation of complement factors in cerebrospinal fluid of patients with Borna disease virus 1 (BoDV-1) encephalitis.

BACKGROUND: Borna disease virus 1 (BoDV-1) causes rare but severe zoonotic infections in humans, presenting as severe encephalitis. The case-fatality risk is very high and no effective countermeasures have been established so far. An immunopathology is presumed, while data on immune responses in humans are limited. Evidence of a role of the complement system in various neurological disorders and central nervous viral infections is increasing and specific inhibitors are available as therapeutic options. METHODS: In this study, we investigated factors of the complement system in the cerebrospinal fluid (CSF) of patients with BoDV-1 infections (n = 17) in comparison to non-inflammatory control CSF samples (n = 11), using a bead-based multiplex assay. In addition, immunohistochemistry was performed using post-mortem brain tissue samples. RESULTS: We found an intrathecal elevation of complement factors of all complement pathways and an active cascade during human BoDV-1 infections. The increase of certain complement factors such as C1q was persistent and C3 complement deposits were detected in post-mortem brain sections. Intrathecal complement levels were negatively correlated with survival. CONCLUSION: Further investigations are warranted to clarify, whether targeting the complement cascade by specific inhibitors might be beneficial for patients suffering from severe BoDV-1 encephalitis.

Irinotecan and temozolomide in combination with dasatinib and rapamycin versus irinotecan and temozolomide for patients with relapsed or refractory neuroblastoma (RIST-rNB-2011): a multicentre, open-label, randomised, controlled, phase 2 trial.

BACKGROUND: Neuroblastoma is the most common extracranial solid tumour in children. Relapsed or refractory neuroblastoma is associated with a poor outcome. We assessed the combination of irinotecan-temozolomide and dasatinib-rapamycin (RIST) in patients with relapsed or refractory neuroblastoma. METHODS: The multicentre, open-label, randomised, controlled, phase 2, RIST-rNB-2011 trial recruited from 40 paediatric oncology centres in Germany and Austria. Patients aged 1-25 years with high-risk relapsed (defined as recurrence of all stage IV and MYCN amplification stages, after response to treatment) or refractory (progressive disease during primary treatment) neuroblastoma, with Lansky and Karnofsky performance status at least 50%, were assigned (1:1) to RIST (RIST group) or irinotecan-temozolomide (control group) by block randomisation, stratified by MYCN status. We compared RIST (oral rapamycin [loading 3 mg/m2 on day 1, maintenance 1 mg/m2 on days 2-4] and oral dasatinib [2 mg/kg per day] for 4 days with 3 days off, followed by intravenous irinotecan [50 mg/m2 per day] and oral temozolomide [150 mg/m2 per day] for 5 days with 2 days off; one course each of rapamycin-dasatinib and irinotecan-temozolomide for four cycles over 8 weeks, then two courses of rapamycin-dasatinib followed by one course of irinotecan-temozolomide for 12 weeks) with irinotecan-temozolomide alone (with identical dosing as experimental group). The primary endpoint of progression-free survival was analysed in all eligible patients who received at least one course of therapy. The safety population consisted of all patients who received at least one course of therapy and had at least one post-baseline safety assessment. This trial is registered at ClinicalTrials.gov, NCT01467986, and is closed to accrual. FINDINGS: Between Aug 26, 2013, and Sept 21, 2020, 129 patients were randomly assigned to the RIST group (n=63) or control group (n=66). Median age was 5·4 years (IQR 3·7-8·1). 124 patients (78 [63%] male and 46 [37%] female) were included in the efficacy analysis. At a median follow-up of 72 months (IQR 31-88), the median progression-free survival was 11 months (95% CI 7-17) in the RIST group and 5 months (2-8) in the control group (hazard ratio 0·62, one-sided 90% CI 0·81; p=0·019). Median progression-free survival in patients with amplified MYCN (n=48) was 6 months (95% CI 4-24) in the RIST group versus 2 months (2-5) in the control group (HR 0·45 [95% CI 0·24-0·84], p=0·012); median progression-free survival in patients without amplified MYCN (n=76) was 14 months (95% CI 9-7) in the RIST group versus 8 months (4-15) in the control group (HR 0·84 [95% CI 0·51-1·38], p=0·49). The most common grade 3 or worse adverse events were neutropenia (54 [81%] of 67 patients given RIST vs 49 [82%] of 60 patients given control), thrombocytopenia (45 [67%] vs 41 [68%]), and anaemia (39 [58%] vs 38 [63%]). Nine serious treatment-related adverse events were reported (five patients given control and four patients given RIST). There were no treatment-related deaths in the control group and one in the RIST group (multiorgan failure). INTERPRETATION: RIST-rNB-2011 demonstrated that targeting of MYCN-amplified relapsed or refractory neuroblastoma with a pathway-directed metronomic combination of a multkinase inhibitor and an mTOR inhibitor can improve progression-free survival and overall survival. This exclusive efficacy in MYCN-amplified, relapsed neuroblastoma warrants further investigation in the first-line setting. FUNDING: Deutsche Krebshilfe.

Neuroinflammatory disease signatures in SPG11-related hereditary spastic paraplegia patients.

Biallelic loss of SPG11 function constitutes the most frequent cause of complicated autosomal recessive hereditary spastic paraplegia (HSP) with thin corpus callosum, resulting in progressive multisystem neurodegeneration. While the impact of neuroinflammation is an emerging and potentially treatable aspect in neurodegenerative diseases and leukodystrophies, the role of immune cells in SPG11-HSP patients is unknown. Here, we performed a comprehensive immunological characterization of SPG11-HSP, including examination of three human postmortem brain donations, immunophenotyping of patients' peripheral blood cells and patient-specific induced pluripotent stem cell-derived microglia-like cells (iMGL). We delineate a previously unknown role of innate immunity in SPG11-HSP. Neuropathological analysis of SPG11-HSP patient brain tissue revealed profound microgliosis in areas of neurodegeneration, downregulation of homeostatic microglial markers and cell-intrinsic accumulation of lipids and lipofuscin in IBA1+ cells. In a larger cohort of SPG11-HSP patients, the ratio of peripheral classical and intermediate monocytes was increased, along with increased serum levels of IL-6 that correlated with disease severity. Stimulation of patient-specific iMGLs with IFNγ led to increased phagocytic activity compared to control iMGL as well as increased upregulation and release of proinflammatory cytokines and chemokines, such as CXCL10. On a molecular basis, we identified increased STAT1 phosphorylation as mechanism connecting IFNγ-mediated immune hyperactivation and SPG11 loss of function. STAT1 expression was increased both in human postmortem brain tissue and in an Spg11-/- mouse model. Application of an STAT1 inhibitor decreased CXCL10 production in SPG11 iMGL and rescued their toxic effect on SPG11 neurons. Our data establish neuroinflammation as a novel disease mechanism in SPG11-HSP patients and constitute the first description of myeloid cell/ microglia activation in human SPG11-HSP. IFNγ/ STAT1-mediated neurotoxic effects of hyperreactive microglia upon SPG11 loss of function indicate that immunomodulation strategies may slow down disease progression.

Loss of Uhrf1 in neural stem cells leads to activation of retroviral elements and delayed neurodegeneration.

In order to understand whether early epigenetic mechanisms instruct the long-term behavior of neural stem cells (NSCs) and their progeny, we examined Uhrf1 (ubiquitin-like PHD ring finger-1; also known as Np95), as it is highly expressed in NSCs of the developing brain and rapidly down-regulated upon differentiation. Conditional deletion of Uhrf1 in the developing cerebral cortex resulted in rather normal proliferation and neurogenesis but severe postnatal neurodegeneration. During development, deletion of Uhrf1 lead to global DNA hypomethylation with a strong activation of the intracisternal A particle (IAP) family of endogenous retroviral elements, accompanied by an increase in 5-hydroxymethylcytosine. Down-regulation of Tet enzymes rescued the IAP activation in Uhrf1 conditional knockout (cKO) cells, suggesting an antagonistic interplay between Uhrf1 and Tet on IAP regulation. As IAP up-regulation persists into postnatal stages in the Uhrf1 cKO mice, our data show the lack of means to repress IAPs in differentiating neurons that normally never express Uhrf1 The high load of viral proteins and other transcriptional deregulation ultimately led to postnatal neurodegeneration. Taken together, these data show that early developmental NSC factors can have long-term effects in neuronal differentiation and survival. Moreover, they highlight how specific the consequences of widespread changes in DNA methylation are for certain classes of retroviral elements.

Mitochondrial and Cellular Function in Fibroblasts, Induced Neurons, and Astrocytes Derived from Case Study Patients: Insights into Major Depression as a Mitochondria-Associated Disease.

The link between mitochondria and major depressive disorder (MDD) is increasingly evident, underscored both by mitochondria's involvement in many mechanisms identified in depression and the high prevalence of MDD in individuals with mitochondrial disorders. Mitochondrial functions and energy metabolism are increasingly considered to be involved in MDD's pathogenesis. This study focused on cellular and mitochondrial (dys)function in two atypical cases: an antidepressant non-responding MDD patient ("Non-R") and another with an unexplained mitochondrial disorder ("Mito"). Skin biopsies from these patients and controls were used to generate various cell types, including astrocytes and neurons, and cellular and mitochondrial functions were analyzed. Similarities were observed between the Mito patient and a broader MDD cohort, including decreased respiration and mitochondrial function. Conversely, the Non-R patient exhibited increased respiratory rates, mitochondrial calcium, and resting membrane potential. In conclusion, the Non-R patient's data offered a new perspective on MDD, suggesting a detrimental imbalance in mitochondrial and cellular processes, rather than simply reduced functions. Meanwhile, the Mito patient's data revealed the extensive effects of mitochondrial dysfunctions on cellular functions, potentially highlighting new MDD-associated impairments. Together, these case studies enhance our comprehension of MDD.

TSPO PET signal using [18F]GE180 is associated with survival in recurrent gliomas.

PURPOSE: Glioma patients, especially recurrent glioma, suffer from a poor prognosis. While advances to classify glioma on a molecular level improved prognostication at initial diagnosis, markers to prognosticate survival in the recurrent situation are still needed. As 18 kDa translocator protein (TSPO) was previously reported to be associated with aggressive histopathological glioma features, we correlated the TSPO positron emission tomography (PET) signal using [18F]GE180 in a large cohort of recurrent glioma patients with their clinical outcome. METHODS: In patients with [18F]GE180 PET at glioma recurrence, [18F]GE180 PET parameters (e.g., SUVmax) as well as other imaging features (e.g., MRI volume, [18F]FET PET parameters when available) were evaluated together with patient characteristics (age, sex, Karnofsky-Performance score) and neuropathological features (e.g. WHO 2021 grade, IDH-mutation status). Uni- and multivariate Cox regression and Kaplan-Meier survival analyses were performed to identify prognostic factors for post-recurrence survival (PRS) and time to treatment failure (TTF). RESULTS: Eighty-eight consecutive patients were evaluated. TSPO tracer uptake correlated with tumor grade at recurrence (p < 0.05), with no significant differences in IDH-wild-type versus IDH-mutant tumors. Within the subgroup of IDH-mutant glioma (n = 46), patients with low SUVmax (median split, ≤ 1.60) had a significantly longer PRS (median 41.6 vs. 25.3 months, p = 0.031) and TTF (32.2 vs 8.7 months, p = 0.001). Also among IDH-wild-type glioblastoma (n = 42), patients with low SUVmax (≤ 1.89) had a significantly longer PRS (median not reached vs 8.2 months, p = 0.002). SUVmax remained an independent prognostic factor for PRS in the multivariate analysis including CNS WHO 2021 grade, IDH status, and age. Tumor volume defined by [18F]FET PET or contrast-enhanced MRI correlated weakly with TSPO tracer uptake. Treatment regimen did not differ among the median split subgroups. CONCLUSION: Our data suggest that TSPO PET using [18F]GE180 can help to prognosticate recurrent glioma patients even among homogeneous molecular subgroups and may therefore serve as valuable non-invasive biomarker for individualized patient management.

Induced neural progenitor cells and iPS-neurons from major depressive disorder patients show altered bioenergetics and electrophysiological properties.

The molecular pathomechanisms of major depressive disorder (MDD) are still not completely understood. Here, we follow the hypothesis, that mitochondria dysfunction which is inevitably associated with bioenergetic disbalance is a risk factor that contributes to the susceptibility of an individual to develop MDD. Thus, we investigated molecular mechanisms related to mitochondrial function in induced neuronal progenitor cells (NPCs) which were reprogrammed from fibroblasts of eight MDD patients and eight non-depressed controls. We found significantly lower maximal respiration rates, altered cytosolic basal calcium levels, and smaller soma size in NPCs derived from MDD patients. These findings are partially consistent with our earlier observations in MDD patient-derived fibroblasts. Furthermore, we differentiated MDD and control NPCs into iPS-neurons and analyzed their passive biophysical and active electrophysiological properties to investigate whether neuronal function can be related to altered mitochondrial activity and bioenergetics. Interestingly, MDD patient-derived iPS-neurons showed significantly lower membrane capacitance, a less hyperpolarized membrane potential, increased Na+ current density and increased spontaneous electrical activity. Our findings indicate that functional differences evident in fibroblasts derived from MDD patients are partially present after reprogramming to induced-NPCs, could relate to altered function of iPS-neurons and thus might be associated with the aetiology of major depressive disorder.

Update on quality assurance in neuropathology: Summary of the round robin trials on TERT promoter mutation, H3-3A mutation, 1p/19q codeletion, and KIAA1549::BRAF fusion testing in Germany in 2020 and 2021.

We previously reported on the first neuropathological round robin trials operated together with Quality in Pathology (QuIP) GmbH in 2018 and 2019 in Germany, i.e., the trials on IDH mutational testing and MGMT promoter methylation analysis [1]. For 2020 and 2021, the spectrum of round robin trials has been expanded to cover the most commonly used assays in neuropathological institutions. In addition to IDH mutation and MGMT promoter methylation testing, there is a long tradition for 1p/19q codeletion testing relevant in the context of the diagnosis of oligodendroglioma. With the 5th edition of the World Health Organization (WHO) classification of the central nervous system tumors, additional molecular markers came into focus: TERT promoter mutation is often assessed as a molecular diagnostic criterion for IDH-wildtype glioblastoma. Moreover, several molecular diagnostic markers have been introduced for pediatric brain tumors. Here, trials on KIAA1549::BRAF fusions (common in pilocytic astrocytomas) and H3-3A mutations (in diffuse midline gliomas, H3-K27-altered and diffuse hemispheric gliomas, H3-G34-mutant) were most desired by the neuropathological community. In this update, we report on these novel round robin trials. In summary, success rates in all four trials ranged from 75 to 96%, arguing for an overall high quality level in the field of molecular neuropathological diagnostics.

Heterogeneity of Amino Acid Profiles of Proneural and Mesenchymal Brain-Tumor Initiating Cells.

Glioblastomas are highly malignant brain tumors that derive from brain-tumor-initiating cells (BTICs) and can be subdivided into several molecular subtypes. Metformin is an antidiabetic drug currently under investigation as a potential antineoplastic agent. The effects of metformin on glucose metabolism have been extensively studied, but there are only few data on amino acid metabolism. We investigated the basic amino acid profiles of proneural and mesenchymal BTICs to explore a potential distinct utilization and biosynthesis in these subgroups. We further measured extracellular amino acid concentrations of different BTICs at baseline and after treatment with metformin. Effects of metformin on apoptosis and autophagy were determined using Western Blot, annexin V/7-AAD FACS-analyses and a vector containing the human LC3B gene fused to green fluorescent protein. The effects of metformin on BTICs were challenged in an orthotopic BTIC model. The investigated proneural BTICs showed increased activity of the serine and glycine pathway, whereas mesenchymal BTICs in our study preferably metabolized aspartate and glutamate. Metformin treatment led to increased autophagy and strong inhibition of carbon flux from glucose to amino acids in all subtypes. However, oral treatment with metformin at tolerable doses did not significantly inhibit tumor growth in vivo. In conclusion, we found distinct amino acid profiles of proneural and mesenchymal BTICs, and inhibitory effects of metformin on BTICs in vitro. However, further studies are warranted to better understand potential resistance mechanisms against metformin in vivo.

ATRT-SHH comprises three molecular subgroups with characteristic clinical and histopathological features and prognostic significance.

Atypical teratoid/rhabdoid tumor (ATRT) is an aggressive central nervous system tumor characterized by loss of SMARCB1/INI1 protein expression and comprises three distinct molecular groups, ATRT-TYR, ATRT-MYC and ATRT-SHH. ATRT-SHH represents the largest molecular group and is heterogeneous with regard to age, tumor location and epigenetic profile. We, therefore, aimed to investigate if heterogeneity within ATRT-SHH might also have biological and clinical importance. Consensus clustering of DNA methylation profiles and confirmatory t-SNE analysis of 65 ATRT-SHH yielded three robust molecular subgroups, i.e., SHH-1A, SHH-1B and SHH-2. These subgroups differed by median age of onset (SHH-1A: 18 months, SHH-1B: 107 months, SHH-2: 13 months) and tumor location (SHH-1A: 88% supratentorial; SHH-1B: 85% supratentorial; SHH-2: 93% infratentorial, often extending to the pineal region). Subgroups showed comparable SMARCB1 mutational profiles, but pathogenic/likely pathogenic SMARCB1 germline variants were over-represented in SHH-2 (63%) as compared to SHH-1A (20%) and SHH-1B (0%). Protein expression of proneural marker ASCL1 (enriched in SHH-1B) and glial markers OLIG2 and GFAP (absent in SHH-2) as well as global mRNA expression patterns differed, but all subgroups were characterized by overexpression of SHH as well as Notch pathway members. In a Drosophila model, knockdown of Snr1 (the fly homologue of SMARCB1) in hedgehog activated cells not only altered hedgehog signaling, but also caused aberrant Notch signaling and formation of tumor-like structures. Finally, on survival analysis, molecular subgroup and age of onset (but not ASCL1 staining status) were independently associated with overall survival, older patients (> 3 years) harboring SHH-1B experiencing relatively favorable outcome. In conclusion, ATRT-SHH comprises three subgroups characterized by SHH and Notch pathway activation, but divergent molecular and clinical features. Our data suggest that molecular subgrouping of ATRT-SHH has prognostic relevance and might aid to stratify patients within future clinical trials.

Metabolic Heterogeneity of Brain Tumor Cells of Proneural and Mesenchymal Origin.

Brain-tumor-initiating cells (BTICs) of proneural and mesenchymal origin contribute to the highly malignant phenotype of glioblastoma (GB) and resistance to current therapies. BTICs of different subtypes were challenged with oxidative phosphorylation (OXPHOS) inhibition with metformin to assess the differential effects of metabolic intervention on key resistance features. Whereas mesenchymal BTICs varied according to their invasiveness, they were in general more glycolytic and less responsive to metformin. Proneural BTICs were less invasive, catabolized glucose more via the pentose phosphate pathway, and responded better to metformin. Targeting glycolysis may be a promising approach to inhibit tumor cells of mesenchymal origin, whereas proneural cells are more responsive to OXPHOS inhibition. Future clinical trials exploring metabolic interventions should account for metabolic heterogeneity of brain tumors.

Molecular profiling of pediatric meningiomas shows tumor characteristics distinct from adult meningiomas.

In contrast to adults, meningiomas are uncommon tumors in childhood and adolescence. Whether adult and pediatric meningiomas differ on a molecular level is unclear. Here we report detailed genomic analyses of 37 pediatric meningiomas by sequencing and DNA methylation profiling. Histologically, the series was dominated by meningioma subtypes with aggressive behavior, with 70% of patients suffering from WHO grade II or III meningiomas. The most frequent cytogenetic aberrations were loss of chromosomes 22 (23/37 [62%]), 1 (9/37 [24%]), 18 (7/37 [19%]), and 14 (5/37 [14%]). Tumors with NF2 alterations exhibited overall increased chromosomal instability. Unsupervised clustering of DNA methylation profiles revealed separation into three groups: designated group 1 composed of clear cell and papillary meningiomas, whereas group 2A comprised predominantly atypical meningiomas and group 2B enriched for rare high-grade subtypes (rhabdoid, chordoid). Meningiomas from NF2 patients clustered exclusively within groups 1 and 2A. When compared with a dataset of 105 adult meningiomas, the pediatric meningiomas largely grouped separately. Targeted panel DNA sequencing of 34 tumors revealed frequent NF2 alterations, while other typical alterations found in adult non-NF2 tumors were absent. These data demonstrate that pediatric meningiomas are characterized by molecular features distinct from adult tumors.

Intraoperative imaging of brain tumors with fluorescein: confocal laser endomicroscopy in neurosurgery. Clinical and user experience.

OBJECTIVE: Confocal laser endomicroscopy (CLE) is an established tool in basic research for tissue imaging at the level of microstructures. Miniaturization and refinement of the technology have made this modality available for operative imaging with a handheld device. Sufficient image contrast is provided by the preoperative application of fluorescein sodium. The authors report their first experiences in a clinical case series using the new confocal laser endomicroscope. METHODS: Handling, operative workflow, and visualization of the CLE were critically evaluated in 12 cases of different CNS tumors. Three different imaging positions in relation to the tumor were chosen: the tumor border (I), tumor center (II), and perilesional zone (III). Respective diagnostic sampling with H & E staining and matching intraoperative neuronavigation and microscope images are provided. RESULTS: CLE was found to be beneficial in terms of high-quality visualization of fine structures and for displaying hidden anatomical details. The handling of the device was good, and the workflow was easy. CONCLUSIONS: Handling ergonomics and image acquisition are intuitive. The endomicroscope allows excellent additional visualization of microstructures in the surgical field with a minimally invasive technique and could improve safety and clinical outcomes. The new confocal laser endomicroscope is an advanced tool with the potential to change intracranial tumor surgery. Imaging of these microstructures is novel, and research with comparative validation with traditional neuropathological assessments is needed.

LEF1 supports metastatic brain colonization by regulating glutathione metabolism and increasing ROS resistance in breast cancer.

More than half of all brain metastases show infiltrating rather than displacing growth at the macro-metastasis/organ parenchyma interface (MMPI), a finding associated with shorter survival. The lymphoid enhancer-binding factor-1 (LEF1) is an epithelial-mesenchymal transition (EMT) transcription factor that is commonly overexpressed in brain-colonizing cancer cells. Here, we overexpressed LEF1 in an in vivo breast cancer brain colonization model. It shortened survival, albeit without engaging EMT at the MMPI. By differential proteome analysis, we identified a novel function of LEF1 as a regulator of the glutathione (GSH) system, the principal cellular redox buffer. LEF1 overexpression also conferred resistance against therapeutic GSH depletion during brain colonization and improved management of intracellular ROS. We conclude that besides EMT, LEF1 facilitates metastasis by improving the antioxidative capacity of epithelial breast cancer cells, in particular during colonization of the brain parenchyma.

Quality assurance in neuropathology: Experiences from the round robin trials on IDH mutation and MGMT promoter methylation testing launched by the Quality Assurance Initiative Pathology (QuIP) in 2018 and 2019.

We here report on the first neuropathological round robin trials initiated by the Quality Assurance Initiative Pathology (QuIP) in Germany in the years 2018 and 2019. Testing services as external laboratory controls were offered for IDH1-R132H immunohistochemistry in 2018 followed by a molecular trial for IDH1 and IDH2 mutations in 2019 including the rare mutational variants. Also in 2019, a trial on MGMT promoter methylation testing was offered. On a national scale, trial offers were well received with around 40 participating institutions. The international announcement of the molecular IDH1/IDH2 mutational trial achieved only moderate European outspread. Success rates in all three trials were excellent (IDH1-R132H immunohistochemistry 2018: 94%, 18 out of 20 possible points required; IDH1/IDH2 mutational status 2019: 100%, 19 out of 20 possible points required; MGMT promoter methylation 2019: 94%, 19 out of 20 possible points required) indicating that quality standards are high in the broad majority of the institutions. Trial participation also involved filling in a questionnaire asking for background information on local testing procedures. We here present a first assessment of the information collected providing unique insights in the landscape of molecular testing in neuropathology. Derived from this information we identify future challenges and provide an outlook on the development of quality assurance in the field of neuropathology.

Use of metformin and survival of patients with high-grade glioma.

High-grade glioma (HGG) is associated with poor prognosis. Drug repurposing evolves as new modality to improve standard therapy. The antidiabetic drug metformin has been found to inhibit glioma cell growth in vitro and in vivo. The aim of the present retrospective cohort study was to evaluate the survival of patients with HGG with or without treatment with metformin, based on a large cohort of a cancer registry. The analysis included 1,093 patients with HGG diagnosed between 1998 and 2013 from the population-based clinical cancer registry Regensburg (Germany), which covers 2.1 Mio inhabitants and 98% of all cancer diagnoses. We performed multivariable adjusted Cox-regression analyses. Hazard Ratios (HRs) with 95% Confidence Intervals (CIs) for overall survival (OS) and progression-free survival (PFS) of patients with HGG with or without treatment with metformin were obtained. Use of metformin was associated with a significantly better overall and progression-free survival of patients with WHO grade III glioma (HR for OS = 0.30; 95% CI = 0.11-0.81, HR for PFS = 0.29; 95% CI = 0.11-0.78), while there were no significant relations with OS (HR = 0.83; 95% CI = 0.57-1.20) or PFS (HR = 0.85; 95% CI = 0.59-1.22) in patients with WHO grade IV glioma. In conclusion, use of metformin is associated with better overall and progression-free survival of patients with WHO grade III. Possible underlying mechanisms include the higher prevalence of IDH mutations in WHO grade III glioma, which might sensitize to the metabolic drug metformin.

The neuropathology of fatal encephalomyelitis in human Borna virus infection.

After many years of controversy, there is now recent and solid evidence that classical Borna disease virus 1 (BoDV-1) can infect humans. On the basis of six brain autopsies, we provide the first systematic overview on BoDV-1 tissue distribution and the lesion pattern in fatal BoDV-1-induced encephalitis. All brains revealed a non-purulent, lymphocytic sclerosing panencephalomyelitis with detection of BoDV-1-typical eosinophilic, spherical intranuclear Joest-Degen inclusion bodies. While the composition of histopathological changes was constant, the inflammatory distribution pattern varied interindividually, affecting predominantly the basal nuclei in two patients, hippocampus in one patient, whereas two patients showed a more diffuse distribution. By immunohistochemistry and RNA in situ hybridization, BoDV-1 was detected in all examined brain tissue samples. Furthermore, infection of the peripheral nervous system was observed. This study aims at raising awareness to human bornavirus encephalitis as differential diagnosis in lymphocytic sclerosing panencephalomyelitis. A higher attention to human BoDV-1 infection by health professionals may likely increase the detection of more cases and foster a clearer picture of the disease.

PI3K: A master regulator of brain metastasis-promoting macrophages/microglia.

Mutations and activation of the PI3K signaling pathway in breast cancer cells have been linked to brain metastases. However, here we describe that in some breast cancer brain metastases samples the protein expression of PI3K signaling components is restricted to the metastatic microenvironment. In contrast to the therapeutic effects of PI3K inhibition on the breast cancer cells, the reaction of the brain microenvironment is less understood. Therefore we aimed to quantify the PI3K pathway activity in breast cancer brain metastasis and investigate the effects of PI3K inhibition on the central nervous system (CNS) microenvironment. First, to systematically quantify the PI3K pathway activity in breast cancer brain metastases, we performed a prospective biomarker study using a reverse phase protein array (RPPA). The majority, namely 30 out of 48 (62.5%) brain metastatic tissues examined, revealed high PI3K signaling activity that was associated with a median overall survival (OS) of 9.41 months, while that of patients, whose brain metastases showed only moderate or low PI3K activity, amounted to only 1.93 and 6.71 months, respectively. Second, we identified PI3K as a master regulator of metastasis-promoting macrophages/microglia during CNS colonization; and treatment with buparlisib (BKM120), a pan-PI3K Class I inhibitor with a good blood-brain-barrier penetrance, reduced their metastasis-promoting features. In conclusion, PI3K signaling is active in the majority of breast cancer brain metastases. Since PI3K inhibition does not only affect the metastatic cells but also re-educates the metastasis-promoting macrophages/microglia, PI3K inhibition may hold considerable promise in the treatment of brain metastasis and the respective microenvironment.

Chronophin regulates active vitamin B6 levels and transcriptomic features of glioblastoma cell lines cultured under non-adherent, serum-free conditions.

BACKGROUND: The phosphatase chronophin (CIN/PDXP) has been shown to be an important regulator of glioma cell migration and invasion. It has two known substrates: p-Ser3-cofilin, the phosphorylated form of the actin binding protein cofilin, and pyridoxal 5'-phosphate, the active form of vitamin B6. Phosphoregulation of cofilin, among other functions, plays an important role in cell migration, whereas active vitamin B6 is a cofactor for more than one hundred enzymatic reactions. The role of CIN has yet only been examined in glioblastoma cell line models derived under serum culture conditions. RESULTS: We found that CIN is highly expressed in cells cultured under non-adherent, serum-free conditions that are thought to better mimic the in vivo situation. Furthermore, the substrates of CIN, p-Ser3-cofilin and active vitamin B6, were significantly reduced as compared to cell lines cultured in serum-containing medium. To further examine its molecular role we stably knocked down the CIN protein with two different shRNA hairpins in the glioblastoma cell lines NCH421k and NCH644. Both cell lines did not show any significant alterations in proliferation but expression of differentiation markers (such as GFAP or TUBB3) was increased in the knockdown cell lines. In addition, colony formation was significantly impaired in NCH644. Of note, in both cell lines CIN knockdown increased active vitamin B6 levels with vitamin B6 being known to be important for S-adenosylmethionine biosynthesis. Nevertheless, global histone and DNA methylation remained unaltered as was chemoresistance towards temozolomide. To further elucidate the role of phosphocofilin in glioblastoma cells we applied inhibitors for ROCK1/2 and LIMK1/2 to our model. LIMK- and ROCK-inhibitor treatment alone was not toxic for glioblastoma cells. However, it had profound, but antagonistic effects in NCH421k and NCH644 under chemotherapy. CONCLUSION: In non-adherent glioblastoma cell lines cultured in serum-free medium, chronophin knockdown induces phenotypic changes, e.g. in colony formation and transcription, but these are highly dependent on the cellular background. The same is true for phenotypes observed after treatment with inhibitors for kinases regulating cofilin phosphorylation (ROCKs and LIMKs). Targeting the cofilin phosphorylation pathway might therefore not be a straightforward therapeutic option in glioblastoma.

Information needs and requirements in patients with brain tumours and their relatives.

Patients with brain tumours face a number of medical and social challenges. Previous studies have shown that these patients and their relatives need a high level of patient-oriented information and counselling. However, these needs are often underestimated. In this single-centre cross-sectional study, we evaluated, for the first time, the information needs of patients with brain tumours and their relatives depending on diagnosis, age and level of education. The participants were interviewed using pre-specified questionnaires. Answers were evaluated descriptively using standard statistical methods. A total of 888 questionnaires were sent out. The return rate was 50.7%. The majority of patients (nP = 103; 59.9%) and a higher proportion of relatives (nR = 103; 72.5%; p = 0.019) wished to receive a maximum of information. The majority (79.7% of patients; 83.1% of relatives) also stated that they preferred a personal, face-to-face meeting as primary source of information. The need for information increased with education (p = 0.015), and decreased with tumour grade (p = 0.025) and age (p = 0.118). Our data indicate that patients with brain tumours and their relatives have high information needs throughout their disease and continuously require information and counselling. Optimal provision of information is based on personal preferences, which needs to be evaluated appropriately. Patient-oriented information and counselling are parts of a successful communication strategy that can improve cancer care significantly.