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Background The ability to identify genetic alterations in cancers is essential for precision medicine; however, surgical approaches to obtain brain tumor tissue are invasive. Profiling circulating tumor DNA (ctDNA) in liquid biopsies has emerged as a promising approach to avoid invasive procedures. Here, we systematically evaluated the feasibility of profiling pediatric brain tumors using ctDNA obtained from plasma, cerebrospinal fluid (CSF), and urine. Methods We prospectively collected 564 specimens (257 blood, 240 urine, and 67 CSF samples) from 258 patients across all histopathologies. We performed ultra-low-pass whole-genome sequencing (ULP-WGS) to assess copy number variations and estimate tumor fraction and developed a pediatric CNS tumor hybrid capture panel for deep sequencing of specific mutations and fusions. Results ULP-WGS detected copy number alterations in 9/46 (20%) CSF, 3/230 (1.3%) plasma, and 0/153 urine samples. Sequencing detected alterations in 3/10 (30%) CSF, 2/74 (2.7%) plasma, and 0/2 urine samples. The only positive results were in high-grade tumors. However, most samples had insufficient somatic mutations (median 1, range 0-39) discoverable by the sequencing panel to provide sufficient power to detect tumor fractions of greater than 0.1%. Conclusions Children with brain tumors harbor very low levels of ctDNA in blood, CSF, and urine, with CSF having the most DNA detectable. Molecular profiling is feasible in a small subset of high-grade tumors. The level of clonal aberrations per genome is low in most of the tumors, posing a challenge for detection using whole-genome or even targeted sequencing methods. Substantial challenges therefore remain to genetically characterize pediatric brain tumors from liquid biopsies.

Ependymomas encompass a heterogeneous group of central nervous system (CNS) neoplasms that occur along the entire neuroaxis. In recent years, extensive (epi-)genomic profiling efforts have identified several molecular groups of ependymoma that are characterized by distinct molecular alterations and/or patterns. Based on unsupervised visualization of a large cohort of genome-wide DNA methylation data, we identified a highly distinct group of pediatric-type tumors (n = 40) forming a cluster separate from all established CNS tumor types, of which a high proportion were histopathologically diagnosed as ependymoma. RNA sequencing revealed recurrent fusions involving the pleomorphic adenoma gene-like 1 (PLAGL1) gene in 19 of 20 of the samples analyzed, with the most common fusion being EWSR1:PLAGL1 (n = 13). Five tumors showed a PLAGL1:FOXO1 fusion and one a PLAGL1:EP300 fusion. High transcript levels of PLAGL1 were noted in these tumors, with concurrent overexpression of the imprinted genes H19 and IGF2, which are regulated by PLAGL1. Histopathological review of cases with sufficient material (n = 16) demonstrated a broad morphological spectrum of tumors with predominant ependymoma-like features. Immunohistochemically, tumors were GFAP positive and OLIG2- and SOX10 negative. In 3/16 of the cases, a dot-like positivity for EMA was detected. All tumors in our series were located in the supratentorial compartment. Median age of the patients at the time of diagnosis was 6.2 years. Median progression-free survival was 35 months (for 11 patients with data available). In summary, our findings suggest the existence of a novel group of supratentorial neuroepithelial tumors that are characterized by recurrent PLAGL1 fusions and enriched for pediatric patients.

Medulloblastomas (MBs) are the most frequent childhood malignant brain tumor. Four histopathologic variants and 4 genetic subgroups have been defined in the World Health Organization (WHO) 2016 Classification and constitute major risk stratification items directly affecting the patient management. Although MB subgroups have been molecularly defined, immunohistochemical surrogates are needed. The aim of our retrospective study was to evaluate the concordance between immunohistochemistry, using 4 antibodies (YAP1, GAB1, OTX2, and β-catenin), and DNA-methylation profiling in MB subgrouping. From a series of 155 MBs, the κ coefficient of concordance was almost perfect (0.90), with only 8/152 discrepant cases (no DNA-methylation analysis was available in 3 cases). Interestingly, the discrepancies mostly concerned (7/8 cases) MBs with divergent differentiations (myogenic, melanotic, and others) with all of those classified into group 3 (n=6) and group 4 (n=1) by DNA-methylation profiling. Another discrepant case concerned a WNT-activated MB (showing only 1% of immunopositive tumor cell nuclei), highlighting the difficulties of determining an appropriate β-catenin immunostaining cutoff. The high concordance of the routine immunohistochemical panel (YAP1, GAB1, OTX2, and β-catenin) and DNA-methylation profiling confirm its utility as a reliable predictive marker of molecular subtype in MBs. We analyzed the accuracy of 10 different IHC combinations for the determination of MB subtype and found that a combination of 2 antibodies (YAP1 and OTX2) allows for the successful characterization of 144 cases of 152 cases. Finally, our series extends the molecular data of the rare morphologic variant of MBs with melanotic/myogenic differentiations.

Fusions involving neurotrophic tyrosine receptor kinase (NTRK) genes are detected in ≤2% of gliomas and can promote gliomagenesis. The remarkable therapeutic efficacy of TRK inhibitors, which are among the first Food and Drug Administration-approved targeted therapies for NTRK-fused gliomas, has generated significant clinical interest in characterizing these tumors. In this multi-institutional retrospective study of 42 gliomas with NTRK fusions, next generation DNA sequencing (n = 41), next generation RNA sequencing (n = 1), RNA-sequencing fusion panel (n = 16), methylation profile analysis (n = 18), and histologic evaluation (n = 42) were performed. All infantile NTRK-fused gliomas (n = 7) had high-grade histology and, with one exception, no other significant genetic alterations. Pediatric NTRK-fused gliomas (n = 13) typically involved NTRK2, ranged from low- to high-histologic grade, and demonstrated histologic overlap with desmoplastic infantile ganglioglioma, pilocytic astrocytoma, ganglioglioma, and glioblastoma, among other entities, but they rarely matched with high confidence to known methylation class families or with each other; alterations involving ATRX, PTEN, and CDKN2A/2B were present in a subset of cases. Adult NTRK-fused gliomas (n = 22) typically involved NTRK1 and had predominantly high-grade histology; genetic alterations involving IDH1, ATRX, TP53, PTEN, TERT promoter, RB1, CDKN2A/2B, NF1, and polysomy 7 were common. Unsupervised principal component analysis of methylation profiles demonstrated no obvious grouping by histologic grade, NTRK gene involved, or age group. KEGG pathway analysis detected methylation differences in genes involved in PI3K/AKT, MAPK, and other pathways. In summary, the study highlights the clinical, histologic, and molecular heterogeneity of NTRK-fused gliomas, particularly when stratified by age group.

The cIMPACT-NOW Update 7 has replaced the WHO nosology of “ependymoma, RELA fusion positive” by “Supratentorial-ependymoma, C11orf95-fusion positive”. This modification reinforces the idea that supratentorial-ependymomas exhibiting fusion that implicates the C11orf95 (now called ZFTA) gene with or without the RELA gene, represent the same histomolecular entity. A hot off the press molecular study has identified distinct clusters of the DNA methylation class of ZFTA fusion-positive tumors. Interestingly, clusters 2 and 4 comprised tumors of different morphologies, with various ZFTA fusions without involvement of RELA. In this paper, we present a detailed series of thirteen cases of non-RELA ZFTA-fused supratentorial tumors with extensive clinical, radiological, histopathological, immunohistochemical, genetic and epigenetic (DNA methylation profiling) characterization. Contrary to the age of onset and MRI aspects similar to RELA fusion-positive EPN, we noted significant histopathological heterogeneity (pleomorphic xanthoastrocytoma-like, astroblastoma-like, ependymoma-like, and even sarcoma-like patterns) in this cohort. Immunophenotypically, these NFκB immunonegative tumors expressed GFAP variably, but EMA constantly and L1CAM frequently. Different gene partners were fused with ZFTA: NCOA1/2, MAML2 and for the first time MN1. These tumors had epigenetic homologies within the DNA methylation class of ependymomas-RELA and were classified as satellite clusters 2 and 4. Cluster 2 (n = 9) corresponded to tumors with classic ependymal histological features (n = 4) but also had astroblastic features (n = 5). Various types of ZFTA fusions were associated with cluster 2, but as in the original report, ZFTA:MAML2 fusion was frequent. Cluster 4 was enriched with sarcoma-like tumors. Moreover, we reported a novel anatomy of three ZFTA:NCOA1/2 fusions with only 1 ZFTA zinc finger domain in the putative fusion protein, whereas all previously reported non-RELA ZFTA fusions have 4 ZFTA zinc fingers. All three cases presented a sarcoma-like morphology. This genotype/phenotype association requires further studies for confirmation. Our series is the first to extensively characterize this new subset of supratentorial ZFTA-fused ependymomas and highlights the usefulness of ZFTA FISH analysis to confirm the existence of a rearrangement without RELA abnormality.

Ependymomas encompass a heterogeneous group of central nervous system (CNS) neoplasms that occur along the entire neuroaxis. In recent years, extensive (epi-)genomic profiling efforts have identified several molecular groups of ependymoma that are characterized by distinct molecular alterations and/or patterns. Based on unsupervised visualization of a large cohort of genome-wide DNA methylation data, we identified a highly distinct group of pediatric-type tumors (n = 40) forming a cluster separate from all established CNS tumor types, of which a high proportion were histopathologically diagnosed as ependymoma. RNA sequencing revealed recurrent fusions involving the pleomorphic adenoma gene-like 1 (PLAGL1) gene in 19 of 20 of the samples analyzed, with the most common fusion being EWSR1:PLAGL1 (n = 13). Five tumors showed a PLAGL1:FOXO1 fusion and one a PLAGL1:EP300 fusion. High transcript levels of PLAGL1 were noted in these tumors, with concurrent overexpression of the imprinted genes H19 and IGF2, which are regulated by PLAGL1. Histopathological review of cases with sufficient material (n = 16) demonstrated a broad morphological spectrum of largely ependymoma-like tumors. Immunohistochemically, tumors were GFAP-positive and OLIG2- and SOX10-negative. In 3/16 of the cases, a dot-like positivity for EMA was detected. Consistent with other fusion-positive ependymal groups, all tumors in our series were located in the supratentorial compartment. Median age of the patients at the time of diagnosis was 6.2 years. Analysis of time to progression or recurrence revealed survival times comparable to those of patients with ZFTA:RELA-fused ependymoma. In summary, our findings suggest the existence of a novel group of supratentorial ependymomas that are characterized by recurrent PLAGL1 fusions and enriched for pediatric patients.

BET-bromodomain inhibition (BETi) has shown pre-clinical promise for MYC-amplified medulloblastoma. However, the mechanisms for its action, and ultimately for resistance, have not been fully defined. Here, using a combination of expression profiling, genome-scale CRISPR/Cas9-mediated loss of function and ORF/cDNA driven rescue screens, and cell-based models of spontaneous resistance, we identify bHLH/homeobox transcription factors and cell-cycle regulators as key genes mediating BETi’s response and resistance. Cells that acquire drug tolerance exhibit a more neuronally differentiated cell-state and expression of lineage-specific bHLH/homeobox transcription factors. However, they do not terminally differentiate, maintain expression of CCND2, and continue to cycle through S-phase. Moreover, CDK4/CDK6 inhibition delays acquisition of resistance. Therefore, our data provide insights about the mechanisms underlying BETi effects and the appearance of resistance and support the therapeutic use of combined cell-cycle inhibitors with BETi in MYC-amplified medulloblastoma., BET-bromodomain inhibitors could be used to treat medulloblastoma tumors with Myc amplifications. Here, the authors show that both the response and resistance to BET inhibitors in mice is mediated by bHLH/homeobox transcription factors.

Ependymoma with RELA fusion has been defined as a novel entity of the revised World Health Organization 2016 classification of tumors of the central nervous system (CNS), characterized by fusion transcripts of the RELA gene and consequent pathological activation of the NFkB pathway. These tumors represent the majority of supratentorial ependymomas in children. The validation of diagnostic tools to identify this clinically relevant ependymoma entity is essential. Here, we have used interphase fluorescent in situ hybridization (FISH) for C11orf95 and RELA, immunohistochemistry (IHC) for p65‐RelA and the recently developed DNA methylation‐based classification besides conventional histopathology, and compared the precision of the methods in 40 supratentorial pediatric brain tumors diagnosed as ependymomas in the past years. Reverse transcription PCR (RT‐PCR) and RNA sequencing were performed to explore discordant cases. Furthermore, we integrated imaging and clinical features as additional layers of information. The concordance between nuclear RelA expression by IHC and RELA FISH was 100%. Concordance between IHC and DNA methylation profiling, and between FISH and DNA methylation profiling was also high (96.4% and 95.2%, respectively). Thirty‐four out of 40 (85%) cases were confirmed by integrated diagnoses as ependymal tumors, including 22 RELA‐fused ependymomas (71% of ependymal tumors), two YAP1‐fused ependymomas (6%), six non‐RELA/non‐YAP1 ependymomas (18%) and four ependymal/subependymal mixed tumors (12%). Ependymal/subependymal mixed tumors had an excellent clinical outcome despite the presence of histopathological signs of malignancy, suggesting that these tumors should not be diagnosed as classic ependymomas. DNA methylation profiling helped in the differential diagnosis of RELA‐fused ependymomas. IHC and FISH, which are available in the majority of pathology laboratories, are valuable tools to identify RELA‐fused ependymomas.

Purpose of review This review aims to give an update on histopathological, molecular and clinical features of central nervous system (CNS) 'embryonal' tumors. Recent findings The taxonomy of previously called 'CNS primitive neuroectodermal tumor' (CNS PNET) has been deeply modified since the discovery of specific molecular profiles for each various sub-entity of these rare, mainly pediatric, tumors. The term 'embryonal tumors' now refers to medulloblastomas, atypical teratoid rhabdoid tumors (AT/RT) and other rare entities, defined by their specific histopathological features together with expression-based or methylation-based profiling; specific gene mutations or fusions characterize some tumor types. In addition, the compilation of large series of molecular data has allowed to dissecting several of these tumor types in molecular subgroups, increasing the number of tumor entities, and leading to an amazingly complex nosology of rare-to-extremely rare malignancies. This rarity precludes from having strong evidence-based therapeutic recommendations, although international efforts are conducted to define the best treatment strategies. Summary Embryonal tumors now correspond to molecularly well defined entities, which deserve further international collaborations to specify their biology and the appropriate burden of treatment, in order to minimize the long-term side-effects of treatment of these overall rare and severe diseases of childhood.

Background Considering that pediatric high-grade gliomas (HGGs) are biologically distinct from their adult counterparts, the objective of this study was to define the landscape of HGGs in adolescents and young adults (AYAs). Methods We performed a multicentric retrospective study of 112 AYAs from adult and pediatric Ile-de-France neurosurgical units, treated between 1998 and 2013 to analyze their clinicoradiological and histomolecular profiles. The inclusion criteria were age between 15 and 25 years, histopathological HGG diagnosis, available clinical data, and preoperative and follow-up MRI. MRI and tumoral samples were centrally reviewed. Immunohistochemistry and complementary molecular techniques such as targeted/next-generation sequencing, whole exome sequencing, and DNA-methylation analyses were performed to achieve an integrated diagnosis according to the 2016 World Health Organization (WHO) classification. Results Based on 80 documented AYA patients, HGGs constitute heterogeneous clinicopathological and molecular groups, with a predominant representation of pediatric subtypes (histone H3-mutants, 40%) but also adult subtypes (isocitrate dehydrogenase [IDH] mutants, 28%) characterized by the rarity of oligodendrogliomas, IDH mutants, and 1p/19q codeletion and the relative high frequency of “rare adult IDH mutations” (20%). H3G34-mutants (14%) represent the most specific subgroup in AYAs. In the H3K27-mutant subgroup, non-brainstem diffuse midline gliomas are more frequent (66.7%) than diffuse intrinsic pontine gliomas (23.8%), contrary to what is observed in children. We found that WHO grade has no prognostic value, but molecular subgrouping has major prognostic importance. Conclusions HGGs in AYAs could benefit from a specific classification, driven by molecular subtyping rather than age group. Collaborative efforts are needed from pediatric and adult neuro-oncology teams to improve the management of HGGs in AYAs.

Diffuse leptomeningeal glioneuronal tumor (DLGNT) was introduced, for the first time, as a provisional entity in the 2016 WHO classification of central nervous system tumors. DLGNT mainly occur in children and characterized by a widespread leptomeningeal growth occasionally associated with intraspinal tumor nodules, an oligodendroglial-like cytology, glioneuronal differentiation and MAP-Kinase activation associated with either solitary 1p deletion or 1p/19q codeletion in the absence of IDH mutation.We report here two unexpected DLGNTs adult cases, characterized by a unique supratentorial circumscribed intraparenchymal tumor without leptomeningeal involvement in spite of long follow-up. In both cases, the diagnosis of DLGNT was made after DNA-methylation profiling which demonstrated that one case belonged to the DLGNT class whereas the other remained not classifiable but showed on CNV the characteristic genetic findings recorded in DLGNT. Both cases harbored 1p/19q codeletion associated with KIAA1549:BRAF fusion in one case and with BRAF V600E and PIK3CA E545A mutations, in the other.Our study enlarges the clinical and molecular spectrum of DLGNTs, and points out that the terminology of DLGNTs is not fully appropriate since some cases could have neither diffuse growth nor leptomeningeal dissemination. This suggests that DLGNTs encompass a wide spectrum of tumors that has yet to be fully clarified.

The department of neuropathology of Sainte-Anne Hospital uses zinc-formalin as the fixative agent for its samples. No publication referenced in Pubmed has proven the validity of this fixative agent. In the context of the accreditation of our standard staining (HPS for Hemalun-Phloxin-Saffron), we started a file for the validation of this method in which the fixative agent constitutes an « interfering » substance which can modify the quality of the technique. The aim of this study was to prove that the use of zinc-formalin as a fixative agent is as suitable as the fixation with 4 % buffered formalin.A cohort of samples fixed by zinc-formalin and by 4 % buffered formalin was performed on fresh samples, then cut and stained by HPS. The slides were interpreted by three pathologists (one of them was outside our centre) ``blind '' to the fixative agent and they evaluated four criteria (general quality of the staining, components of the extracellular matrix, cytoplasmic details, and nuclear details) and scored them (from 0 to 3) according to the Association française en assurance qualité (AFAQAP) recommendations.The cohort included 43 samples. The results of the analysis showed that for samples fixed by zinc-formalin, three of the four criteria obtained significantly a better score than the samples fixed by classical formalin.Our results show that the zinc-formalin fixative does not constitute an ``interfering '' agent for the quality of the HPS staining for neuropathological samples.

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Accurate pathological diagnosis is crucial for optimal management of patients with cancer. For the approximately 100 known tumour types of the central nervous system, standardization of the diagnostic process has been shown to be particularly challenging - with substantial inter-observer variability in the histopathological diagnosis of many tumour types. Here we present a comprehensive approach for the DNA methylation-based classification of central nervous system tumours across all entities and age groups, and demonstrate its application in a routine diagnostic setting. We show that the availability of this method may have a substantial impact on diagnostic precision compared to standard methods, resulting in a change of diagnosis in up to 12% of prospective cases. For broader accessibility, we have designed a free online classifier tool, the use of which does not require any additional onsite data processing. Our results provide a blueprint for the generation of machine-learning-based tumour classifiers across other cancer entities, with the potential to fundamentally transform tumour pathology.

BACKGROUND Considering that paediatric high-grade gliomas (HGGs) are biologically distinct from their adult counterparts, the objective of this study was to define the landscape of HGGs in adolescents and young adults (AYAs). METHODS We performed a multicentric retrospective study of 112 AYAs from adult and paediatric Ile-de-France neurosurgical units, treated between 1998 and 2013 to analyse their clinicoradiological and histomolecular profiles. The inclusion criteria were age between 15 and 25-years, histopathological HGG diagnosis, available clinical data, pre-operative and follow-up MRI. MRI and tumoral samples were centrally reviewed. Immunohistochemistry and complementary molecular techniques such as targeted/next generation sequencing, whole exome sequencing and DNA-methylation analyses were performed to achieve an integrated diagnosis according to the 2016 WHO classification. RESULTS Based on 80 documented AYA patients, HGGs constitute heterogeneous clinicopathological and molecular groups, with a predominant representation of paediatric-subtypes (Histone H3-mutants, 40%) but also adult-subtypes (IDH-mutants, 28%) characterized by the rarity of oligodendrogliomas, IDH-mutant and 1p/19q co-deleted and the relative high frequency of “rare adult IDH mutations” (20%). H3G34-mutants (14%) represent the most specific subgroup in AYAs. In the H3K27-mutant subgroup, the non-brainstem diffuse midline gliomas are more frequent (66.7%) than diffuse intrinsic pontine gliomas (23.8%), contrary to children. We found that WHO grade has no prognostic value, but molecular subgrouping has major prognostic importance. CONCLUSIONS HGGs in AYAs could benefit from a more personalized neuro-oncological management, driven by molecular subtyping rather than age group. Collaborative efforts are needed from paediatric and adult neuro-oncology teams to improve the management of HGGs in AYAs.

Ganglioglioma (GG) is a grade I tumor characterized by alterations in the MAPK pathway, including BRAF V600E mutation. Recently, diffuse midline glioma with an H3 K27M mutation was added to the WHO 2016 classification as a new grade IV entity. As co-occurrence of H3 K27M and BRAF V600E mutations has been reported in midline tumors and anaplastic GG, we searched for BRAF V600E and H3 K27M mutations in a series of 54 paediatric midline grade I GG (midline GG) to determine the frequency of double mutations and its relevance for prognosis. Twenty-seven patients (50%) possessed the BRAF V600E mutation. The frequency of the co-occurrence of H3F3A/BRAF mutations at diagnosis was 9.3%. No H3 K27M mutation was detected in the absence of the BRAF V600E mutation. Double-immunostaining revealed that BRAF V600E and H3 K27M mutant proteins were present in both the glial and neuronal components. Immunopositivity for the BRAF V600E mutant protein correlated with BRAF mutation status as detected by massARRAY or digital droplet PCR. The median follow-up of patients with double mutation was 4 years. One patient died of progressive disease 8 years after diagnosis, whereas the four other patients were all alive with stable disease at the last clinical follow-up (at 9 months, 1 year and 7 years) without adjuvant therapy. We demonstrate in this first series of midline GGs that the H3 K27M mutation can occur in association with the BRAF V600E mutation in grade I glioneuronal tumors. Despite the presence of H3 K27M mutations, these cases should not be graded and treated as grade IV tumors because they have a better spontaneous outcome than classic diffuse midline H3 K27M-mutant glioma. These data suggest that H3 K27M cannot be considered a specific hallmark of grade IV diffuse gliomas and highlight the importance of integrated histomolecular diagnosis in paediatric brain tumors.

Papillary glioneuronal tumor (PGNT) is a WHO-defined brain tumor entity that poses a major diagnostic challenge. Recently, SLC44A1-PRKCA fusions have been described in PGNT. We subjected 28 brain tumors from different institutions histologically diagnosed as PGNT to molecular and morphological analysis. Array-based methylation analysis revealed that 17/28 tumors exhibited methylation profiles typical for other tumor entities, mostly dysembryoplastic neuroepithelial tumor and hemispheric pilocytic astrocytoma. Conversely, 11/28 tumors exhibited a unique profile, thus constituting a distinct methylation class PGNT. By screening the extended Heidelberg cohort containing over 25,000 CNS tumors, we identified three additional tumors belonging to this methylation cluster but originally histologically diagnosed otherwise. RNA sequencing for the detection of SLC44A1-PRKCA fusions could be performed on 19 of the tumors, 10 of them belonging to the methylation class PGNT. In two additional cases, SLC44A1-PRKCA fusions were confirmed by FISH. We detected fusions involving PRKCA in all cases of this methylation class with material available for analyses: the canonical SLC44A1-PRKCA fusion was observed in 11/12 tumors, while the remaining case exhibited a NOTCH1-PRKCA fusion. Neither of the fusions was found in the tumors belonging to other methylation classes. Our results point towards a high misclassification rate of the morphological diagnosis PGNT and clearly demonstrate the necessity of molecular analyses. PRKCA fusions are highly diagnostic for PGNT, and detection by RNA sequencing enables the identification of rare fusion partners. Methylation analysis recognizes a unique methylation class PGNT irrespective of the nature of the PRKCA fusion.

© Springer-Verlag GmbH Germany, part of Springer Nature 2019., The "isomorphic subtype of diffuse astrocytoma" was identified histologically in 2004 as a supratentorial, highly differentiated glioma with low cellularity, low proliferation and focal diffuse brain infiltration. Patients typically had seizures since childhood and all were operated on as adults. To define the position of these lesions among brain tumours, we histologically, molecularly and clinically analysed 26 histologically prototypical isomorphic diffuse gliomas. Immunohistochemically, they were GFAP-positive, MAP2-, OLIG2- and CD34-negative, nuclear ATRX-expression was retained and proliferation was low. All 24 cases sequenced were IDH-wildtype. In cluster analyses of DNA methylation data, isomorphic diffuse gliomas formed a group clearly distinct from other glial/glio-neuronal brain tumours and normal hemispheric tissue, most closely related to paediatric MYB/MYBL1-altered diffuse astrocytomas and angiocentric gliomas. Half of the isomorphic diffuse gliomas had copy number alterations of MYBL1 or MYB (13/25, 52%). Gene fusions of MYBL1 or MYB with various gene partners were identified in 11/22 (50%) and were associated with an increased RNA-expression of the respective MYB-family gene. Integrating copy number alterations and available RNA sequencing data, 20/26 (77%) of isomorphic diffuse gliomas demonstrated MYBL1 (54%) or MYB (23%) alterations. Clinically, 89% of patients were seizure-free after surgery and all had a good outcome. In summary, we here define a distinct benign tumour class belonging to the family of MYB/MYBL1-altered gliomas. Isomorphic diffuse glioma occurs both in children and adults, has a concise morphology, frequent MYBL1 and MYB alterations and a specific DNA methylation profile. As an exclusively histological diagnosis may be very challenging and as paediatric MYB/MYBL1-altered diffuse astrocytomas may have the same gene fusions, we consider DNA methylation profiling very helpful for their identification., This study was supported by an International League against Epilepsy (ILAE) Grant to D. Capper. I. Blumcke was supported by the European Union (FP6 DESIRE Grant Agreement #602531). M. Mittelbronn would like to thank the Luxembourg National Research Fund (FNR) for the support (FNR PEARL P16/BM/11192868 grant). K. Ligon is funded by the Paediatric Low Grade Astrocytoma Foundation and NCI R01CA215489. T. Jacques is funded by The Brain Tumour Charity, Children with Cancer UK, Great Ormond Street Hospital Children’s Charity, Cancer Research UK, the Olivia Hodson Cancer Fund and the National Institute of Health Research. T. Jacques’s work is partly funded by the NIHR GOSH BRC. The views expressed are those of the authors and not necessarily those of the NHS, the NIHR or the Department of Health. We thank the biomedical scientists in the Division of Neuropathology, the National Hospital for Neurology and Neurosurgery (NHNN) for excellent technical assistance. We also thank clinicians and neuropathologists for referring cases for molecular analysis. Part of the study was funded by the National Institute for Health Research to UCLH Biomedical research centre (BRC399/NS/RB/101410). S. Brandner and Z. Jaunmuktane are also supported by the Department of Health’s NIHR Biomedical Research Centre’s funding scheme. Additional funding was provided by the Brain Tumour Charity (UK) for the Everest Centre for Paediatric Low-Grade Brain Tumour Research

Prevalence and predictors of epileptic seizures are unknown in the malignant variant of ganglioglioma.In a retrospective exploratory dataset of 18 supratentorial anaplastic World Health Organization grade III gangliogliomas, we studied: (i) the prevalence and predictors of epileptic seizures at diagnosis; (ii) the evolution of seizures during tumor evolution; (iii) seizure control rates and predictors of epilepsy control after oncological treatments.Epileptic seizures prevalence progresses throughout the natural course of anaplastic gangliogliomas: 44% at imaging discovery, 67% at histopathological diagnosis, 69% following oncological treatment, 86% at tumor progression, and 100% at the end-of-life phase. The medical control of seizures and their refractory status worsened during the tumor's natural course: 25% of uncontrolled seizures at histopathological diagnosis, 40% following oncological treatment, 45.5% at tumor progression, and 45.5% at the end-of-life phase. Predictors of seizures at diagnosis appeared related to the tumor location (i.e. temporal and/or cortical involvement). Prognostic parameters of seizure control after first-line oncological treatment were temporal tumor location, eosinophilic granular bodies, TP53 mutation, and extent of resection. Prognostic parameters of seizure control at tumor progression were a history of epileptic seizures at diagnosis, seizure control after first-line oncological treatment, eosinophilic granular bodies, and TP53 mutation.Epileptic seizures are frequently observed in anaplastic gangliogliomas and both prevalence and medically refractory status worsen during the tumor's natural course. Both oncological and antiepileptic treatments should be employed to improve the control of epileptic seizures and the quality of life of patients harboring an anaplastic ganglioglioma.

Anaplastic ganglioglioma (AGG) is a rare and malignant variant of ganglioglioma. According to the World Health Organization classification version 2016, their histopathological grading criteria are still ill-defined. The aim of the present study was to assess the clinical, imaging, histopathological, and molecular characteristics and outcomes of AGGs in a large consecutive and retrospective adult and pediatric case series. Eighteen patients with AGGs (13 adults and 5 children) were identified (14 de novo and 4 secondary) from a cohort of 222 gangliogliomas (GG) (8%) treated at our institution between 2000 and 2015. AGGs represented a very aggressive disease with poor outcome (median progression-free survival, 10 months; median overall survival, 27 months). They were located in the temporal lobe only in 22% and presented with seizures (44%) or increased intracranial pressure (44%) at diagnosis. Concerning histopathological and molecular data, they shared morphological characteristics and BRAF V600E mutation (39%) with their benign counterparts but also showed hTERT promoter mutation (61%), p53 accumulation (39%), ATRX loss (17%), or p.K27M H3F3A mutation (17%). AGGs are malignant neoplasms requiring aggressive oncological treatment. In the perspective of targeted therapies, AGGs should be screened for BRAF V600E, hTERT, ATRX, and mutations of histone genes.

Numerous molecular alterations have been described in supratentorial high-grade gliomas (1p19q co-deletion, IDH1/2, histone H3, hTERT promotor mutations, loss of ATRX) which have led to a new histomolecular classification of diffuse gliomas. We aimed at describing these alterations in a series of 19 adults with pure cerebellar high-grade gliomas. Systematic immunohistochemical analyses, including that of IDH1R132H, ATRX, p53, PTEN, EGFR, p16, FGFR3, BRAFV600E, mismatch repair proteins, H3K27me3, H3K36me3, and H3K27M; molecular analyses of IDH1/2, hTERT, BRAF, H3F3A, and HIST1H3B mutation hotspots; and EGFR, PTEN FISH were retrospectively performed in a multicentric study. We histopathologically identified 14 glioblastomas, 4 grade III astrocytomas and 1 gliosarcoma. Two cases showed a H3F3A K27M mutation. Only one case harbored a classical profile of glioblastoma with hTERT mutation, EGFR gain and 10q loss. The most frequent alteration was the absence of p16 immunoexpression. We report a histomolecular analysis of pure cerebellar high grade gliomas. The histomolecular profile appears to be different from that of supratentorial gliomas, with no IDH1/2 gene mutations and only 1 case with a classic profile of de novo glioblastoma. In 2 cases, we identified H3F3A K27M mutation, classically described in pediatric midline gliomas. .

Summary The current consensus recognizes four main medulloblastoma subgroups (wingless, Sonic hedgehog, group 3 and group 4). While medulloblastoma subgroups have been characterized extensively at the (epi-)genomic and transcriptomic levels, the proteome and phosphoproteome landscape remain to be comprehensively elucidated. Using quantitative (phospho)-proteomics in primary human medulloblastomas, we unravel distinct posttranscriptional regulation leading to highly divergent oncogenic signaling and kinase activity profiles in groups 3 and 4 medulloblastomas. Specifically, proteomic and phosphoproteomic analyses identify aberrant ERBB4-SRC signaling in group 4. Hence, enforced expression of an activated SRC combined with p53 inactivation induces murine tumors that resemble group 4 medulloblastoma. Therefore, our integrative proteogenomics approach unveils an oncogenic pathway and potential therapeutic vulnerability in the most common medulloblastoma subgroup.

OBJECTIVEThe goal of this study was to provide insight into the influence of gliomas on gestational outcomes, the impact of pregnancy on gliomas, and the identification of patients at risk.METHODSIn this multiinstitutional retrospective study, the authors identified 52 pregnancies in 50 women diagnosed with a glioma.RESULTSFor gliomas known prior to pregnancy (n = 24), we found the following: 1) An increase in the quantified imaging growth rates occurred during pregnancy in 87% of cases. 2) Clinical deterioration occurred in 38% of cases, with seizures alone resolving after delivery in 57.2% of cases. 3) Oncological treatments were immediately performed after delivery in 25% of cases. For gliomas diagnosed during pregnancy (n = 28), we demonstrated the following: 1) The tumor was discovered during the second and third trimesters in 29% and 54% of cases, respectively, with seizures being the presenting symptom in 68% of cases. 2) The quantified imaging growth rates did not significantly decrease after delivery and before oncological treatment. 3) Clinical deterioration resolved after delivery in 21.4% of cases. 4) Oncological treatments were immediately performed after delivery in 70% of cases. Gliomas with a high grade of malignancy, negative immunoexpression of alpha-internexin, or positive immunoexpression for p53 were more likely to be associated with tumor progression during pregnancy. Deliveries were all uneventful (cesarean section in 54.5% of cases and vaginal delivery in 45.5%), and the infants were developmentally normal.CONCLUSIONSWhen a woman harboring a glioma envisions a pregnancy, or when a glioma is discovered in a pregnant patient, the authors suggest informing her and her partner that pregnancy may impact the evolution of the glioma clinically and radiologically. They strongly advise a multidisciplinary approach to management.■ CLASSIFICATION OF EVIDENCE Type of question: association; study design: case series; evidence: Class IV.

Diffuse intrinsic pontine glioma (DIPG) is the most severe paediatric solid tumour, with no significant therapeutic progress made in the past 50 years. Recent studies suggest that diffuse midline glioma, H3-K27M mutant, may comprise more than one biological entity. The aim of the study was to determine the clinical and biological variables that most impact their prognosis. Ninety-one patients with classically defined DIPG underwent a systematic stereotactic biopsy and were included in this observational retrospective study. Histone H3 genes mutations were assessed by immunochemistry and direct sequencing, whilst global gene expression profiling and chromosomal imbalances were determined by microarrays. A full description of the MRI findings at diagnosis and at relapse was integrated with the molecular profiling data and clinical outcome. All DIPG but one were found to harbour either a somatic H3-K27M mutation and/or loss of H3K27 trimethylation. We also discovered a novel K27M mutation in HIST2H3C, and a lysine-to-isoleucine substitution (K27I) in H3F3A, also creating a loss of trimethylation. Patients with tumours harbouring a K27M mutation in H3.3 (H3F3A) did not respond clinically to radiotherapy as well, relapsed significantly earlier and exhibited more metastatic recurrences than those in H3.1 (HIST1H3B/C). H3.3-K27M-mutated DIPG have a proneural/oligodendroglial phenotype and a pro-metastatic gene expression signature with PDGFRA activation, while H3.1-K27M-mutated tumours exhibit a mesenchymal/astrocytic phenotype and a pro-angiogenic/hypoxic signature supported by expression profiling and radiological findings. H3K27 alterations appear as the founding event in DIPG and the mutations in the two main histone H3 variants drive two distinct oncogenic programmes with potential specific therapeutic targets. Electronic supplementary material The online version of this article (doi:10.1007/s00401-015-1478-0) contains supplementary material, which is available to authorized users.

The aim was to localize the interictal cerebral perfusion abnormalities of focal cortical dysplasia (FCD) in children with Arterial Spin Labeling MRI (ASL) in a retrospective study of nine consecutive children explored with multimodal investigation during interictal periods. We analyzed brain morphology with a 1.5T MRI and a dedicated protocol for epilepsy. Brain perfusion was quantified with pseudo continuous ASL. Brain metabolism was imaged with (18)FDG-PET in six patients. Microvessel histology was studied in five children who underwent epilepsy surgery with CD34 immunostaining on FCD and control samples. Localized decrease of cerebral blood flow (CBF) was found on visual analysis in all patients with ASL. It was co-localized with the structural MRI abnormalities in every case, with PET hypo-metabolism in 5/6 cases, and with histologically proven FCD type IIb in 5/5 cases (all seizure free after surgery). CBF was lower (Kruskal-Wallis test, p=0.001) in FCD than in normal cortex. The total count of CD34+ microvessels was similar in FCD and control cases, but microvasculature showed disorganized architecture. Interictal ASL is a non-invasive method that may help to localize the epileptogenic zone showing hypo-perfusion in FCD. Whether this finding could be generalized to MRI-negative FCD needs to be further studied.

Grade I or III ganglioglioma (GG) is characterized by alterations in the MAPK pathway, including BRAF mutation. Diffuse midline glioma H3 K27M mutated was added to the WHO 2016 classification as a new grade IV entity. A co-occurrence of H3F3A/BRAF mutations has been reported in midline tumors. Firstly, we searched for BRAF V600E and H3 K27M mutations in a series of 54 pediatric midline grade I GG to determine the frequency of double mutations in grade I GG. We then studied the clinico-radiological and histo-molecular features of 12 grade I to III double-mutated tumors to explore the therapeutic and prognostic relevance. The frequency of the co-occurrence of H3F3A/BRAF mutations in grade I midline GG at diagnosis was 9.3%. Double-immunostaining revealed that BRAF V600E and H3 K27M mutant proteins colocalized in both the glial and neuronal components, The series of double-mutated GG included 8 grade I, and 4 grade III at diagnosis. The median survival was 8 years. Three grade I tumors showed a malignant transformation at 0.5, 3.5 and 7 years and two of them showed a loss of the BRAF mutation in recurrent tumor samples. Five patients with grade I were alive with stable disease without adjuvant therapy at the last follow-up. One grade III showed a paradoxical response under BRAF inhibitor. In this work we stress that H3F3A mutation can occur, in association with the BRAF mutation, in grade I GG, and that these have a better spontaneous outcome than classic diffuse midline H3 K27M-mutant gliomas. These data highlight the importance of an integrated histomolecular diagnosis especially in the context of targeted therapy.

Background and purpose Molecular alterations were recently added to the World Health Organization (WHO) 2016 classification of central nervous system (CNS) tumors. We correlated the histological and radiological features of G34R mutant high-grade gliomas, a recently described hemispheric and supratentorial glioma of children and young adults. Materials and methods We performed a retrospective multicenter study on the histopathological and MRI results of 12 patients. Results All tumors were supratentorial. Several radiological aspects were observed. Height over 12 were bulky and well delineated tumors, without visible peritumoral infiltration on MRI and pathologically characterized by highly cellular tissue associated with a moderate peritumoral infiltrative component. Two tumors were ill-defined and hyperintense on T2 sequences and pathologically characterized by diffuse tumoral infiltration. Two tumors were bulky and well delineated with an infiltrative component, both radiologically and histopathologically. Conclusions These different patterns may correspond to different pathological mechanisms and a potential link with prognosis should be assessed in further studies.

Purpose: Despite multimodal therapy, prognosis of pediatric intracranial ependymomas remains poor with a 5-year survival rate below 70% and frequent late deaths. Experimental design: This multicentric European study evaluated putative prognostic biomarkers. Tenascin-C (TNC) immunohistochemical expression and copy number status of 1q25 were retained for a pooled analysis of 5 independent cohorts. The prognostic value of TNC and 1q25 on the overall survival (OS) was assessed using a Cox model adjusted to age at diagnosis, tumor location, WHO grade, extent of resection, radiotherapy and stratified by cohort. Stratification on a predictor that did not satisfy the proportional hazards assumption was considered. Model performance was evaluated and an internal-external cross validation was performed. Results: Among complete cases with 5-year median follow-up (n = 470; 131 deaths), TNC and 1q25 gain were significantly associated with age at diagnosis and posterior fossa tumor location. 1q25 status added independent prognostic value for death beyond the classical variables with a hazard ratio (HR) = 2.19 95%CI = [1.29; 3.76] (p = 0.004), while TNC prognostic relation was tumor location-dependent with HR = 2.19 95%CI = [1.29; 3.76] (p = 0.004) in posterior fossa and HR = 0.64 [0.28; 1.48] (p = 0.295) in supratentorial (interaction p value = 0.015). The derived prognostic score identified 3 different robust risk groups. The omission of upfront RT was not associated with OS for good and intermediate prognostic groups while the absence of upfront RT was negatively associated with OS in the poor risk group. Conclusion: Integrated TNC expression and 1q25 status are useful to better stratify patients and to eventually adapt treatment regimens in pediatric intracranial ependymoma.

Purpose To compare arterial spin labeling (ASL) data between low- and high-grade brain tumors in children to establish a cutoff to distinguish low- from high-grade neoplasms and to assess potential correlations between cerebral blood flow (CBF) and quantitative histologic microvascular data. Materials and Methods Approval was obtained from the regional review board. ASL data obtained in 129 children between 2011 and 2015 were retrospectively analyzed. CBF and relative CBF in the most perfused area of each neoplasm and contrast enhancement were quantified with a semiquantitative ratio. The correlation between CBF and microvascular density was analyzed in specimens stained with anti-CD34. Results were controlled in two validation cohorts with 1.5- and 3.0-T magnetic resonance (MR) imaging. Results Mean CBF was significantly higher for high-grade than for low-grade hemispheric (116 mL/min/100 g [interquartile range {IQR}, 73-131 mL/min/100 g] vs 29 mL/min/100 g [IQR, 23-35 29 mL/min/100 g], P < .001), thalamic (87 mL/min/100 g [IQR, 73-100 mL/min/100 g] vs 36 mL/min/100 g [IQR, 30-40 mL/min/100 g], P = .016), and posterior fossa (59 mL/min/100 g [IQR, 45-91 mL/min/100 g] vs 33 mL/min/100 g [IQR, 25-40 mL/min/100 g], P < .001) tumors. With a cutoff of 50 mL/min/100 g, sensitivity and specificity were 90% (95% confidence interval [CI]: 68, 100) and 93% (95% CI: 66, 100), respectively, for hemispheric tumors; 100% (95% CI: 48, 100) and 80% (95% CI: 28, 100), respectively, for thalamic tumors; and 65% (95% CI: 51, 78) and 94% (95% CI: 80, 99), respectively, for posterior fossa tumors. In posterior fossa tumors, additional use of the CBF-to-contrast enhancement ratio yielded sensitivity and specificity of 96% (95% CI: 87, 100) and 97% (95% CI: 84, 100), respectively. Use of a simple algorithm based on these values yielded an accuracy of 93% (95% CI: 87, 97). Validation sets yielded similar results, with grading accuracy of 88% (95% CI: 62, 98) with 1.5-T MR imaging and 77% (95% CI: 46, 95) with 3.0-T MR imaging. CBF was strongly correlated with microvascular density (R = 0.66, P < .001). Conclusion High-grade pediatric brain tumors display higher CBF than do low-grade tumors, and they may be accurately graded by using these values. CBF is correlated with tumor microvascular density. © RSNA, 2016 Online supplemental material is available for this article.

INTRODUCTION: BIOMEDE is a multicentre, phase-II trial comparing efficacy of erlotinib, everolimus, and dasatinib in combination with radiotherapy, in newly diagnosed diffuse intrinsic pontine glioma (DIPG) (NCT02233049). Randomisation is based on biomarkers determined in upfront stereotactic biopsies: mTOR pathway activation detected by the loss of PTEN expression and EGFR overexpression. METHODS: A central pathology review is mandatory in BIOMEDE for DIPG diagnosis confirmation, assessment of H3K27M immunostatus, PDGFRA amplification by FISH and biomarkers evaluation for treatment allocation (no erlotinib if no EGFR overexpression; no everolimus if no PTEN loss, no specific marker for dasatinib allocation). We evaluated the process feasibility and the agreement between H3.3 immunoprofil and genotyping on the first 120 patients. RESULTS: DIPG was centrally confirmed in 94% of cases (median time=1 day; range 0–6). The differential diagnoses were HGNET-MYCN (n=1) and angiocentric glioma MYB-QKI (n=1). No tumoral infiltration was found in 3 cases and extensive necrosis in 1 case. Overall, 38/118 evaluable cases showed EGFR overexpression (32%) and 90/118 had PTEN loss (76%). 15/105 evaluable cases had PDGFRA amplification (14%). Trimethylation loss at H3K27 was detected in all samples but 4 corresponding to HGNET-MYCN, angiocentric glioma and to 2 H3 wild-type gliomas. Agreement between genotyping (H3.3 and/or H3.1) and H3K27M immunostatus (loss of trimethylation and/or H3K27M) is substantial (Kappa coefficient=0.79, 95%-confidence interval, 0.41 to 1). CONCLUSIONS: These data document the feasibility of real-time molecular pathology and biomarkers screening in international multicentre trials.

OBJECTIVE There are no guidelines for the management of postoperative lateral sinus thrombosis following posterior fossa surgery. Introducing treatment-dose anticoagulant therapy during the immediate postoperative period increases the risk of intracranial bleeding. This study assessed the incidence of and risk factors associated with postoperative lateral sinus thrombosis and the complications related to thrombosis and/or anticoagulation. METHODS This study was a retrospective monocentric analysis of adult patients who underwent surgical removal of a posterior fossa space-occupying lesion with available postoperative imaging. Postoperative lateral sinus thrombosis was defined as a T2* hypointensity within the venous sinus and/or a filling defect on postcontrast MRI or CT scan. RESULTS Among 180 patients, 12 (6.7%; 95% CI 3.0–10.4) were found to have lateral sinus thrombosis on postoperative imaging, none of whom were symptomatic. Unadjusted risk factors for postoperative lateral sinus thrombosis were a history of deep venous thrombosis (p = 0.016), oral contraceptive pill (p = 0.004), midline surgical approach (p = 0.035), and surgical exposure of the sinus (p < 0.001). Seven of the patients (58.3%) with a postoperative lateral sinus thrombosis received immediate treatment-dose anticoagulant therapy. Lateral sinus recanalization occurred radiologically at a mean time of 272 ± 23 days in 85.7% of patients (6 of 7) undergoing treatment-dose anticoagulant therapy and in 20% of patients (1 of 5) not receiving treatment-dose anticoagulant therapy. Postoperative complications occurred in 56.2% of patients (9 of 16) who received treatment-dose curative anticoagulant therapy and in 27% of patients (45 of 164) who did not. CONCLUSIONS Incidental radiological lateral sinus thrombosis following posterior fossa surgery has an incidence of 6.7%. To further define the benefit-to-risk ratio of a treatment-dose anticoagulant therapy, a prospective trial should be considered.

Angiocentric gliomas are pediatric low-grade gliomas (PLGGs) without known recurrent genetic drivers. We performed genomic analysis of new and published data from 249 PLGGs, including 19 angiocentric gliomas. We identified MYB-QKI fusions as a specific and single candidate driver event in angiocentric gliomas. In vitro and in vivo functional studies show that MYB-QKI rearrangements promote tumorigenesis through three mechanisms: MYB activation by truncation, enhancer translocation driving aberrant MYB-QKI expression and hemizygous loss of the tumor suppressor QKI. To our knowledge, this represents the first example of a single driver rearrangement simultaneously transforming cells via three genetic and epigenetic mechanisms in a tumor.

International audience; INTRODUCTION:Papillary Glioneuronal Tumor (PGNT) is a grade I tumor which was classified as a separate entity in the World Health Organization Classification of the Central Nervous System 2007 in the group of mixed glioneuronal tumors. This tumor is rare and subclassifying PGNT represents a challenge. Recently, a fusion between SLC44A1 and PRKCA which encodes a protein kinase C involved in MAPK signaling pathway has been described in two studies (five cases). The current study aimed at raising the cytogenetic, histological and molecular profiles of PGNT and to determine if SLC44A1-PRKCA fusion represented a specific diagnostic marker to distinguish it from other glioneuronal tumors.RESULTS:We report on four pediatric cases of PGNT, along with clinico-radiologic and immunohistological features for which SLC44A1-PRKCA fusion assessment by fluorescence in situ hybridization, BRAF V600E and FGFR1 mutation by immunohistochemistry and direct DNA sequencing and KIAA1549-BRAF fusion by RT-PCR were performed. MAPK signaling pathway activation was investigated using phospho-ERK immunohistochemistry and western blot. We analyzed fifteen cases of tumors with challenging histological or clinical differential diagnoses showing respectively a papillary architecture or periventricular location (PGNT mimics). fluorescence in situ hybridization analysis revealed a constant SLC44A1-PRKCA fusion signal in all PGNTs. None of PGNT mimics showed the SLC44A1-PRKCA fusion signal pattern. All PGNTs were negative for BRAF V600E and FGFR1 mutation, and KIAA1549-BRAF fusion. Phospho-ERK analysis provides arguments for the activation of the MAPK signaling pathway in these tumors.CONCLUSIONS:Here we confirmed and extended the molecular data on PGNT. These results suggest that PGNT belong to low grade glioma with MAPK signaling pathway deregulation. SLC44A1-PRKCA fusion seems to be a specific characteristic of PGNT with a high diagnostic value and detectable by FISH.

Primary leptomeningeal gliomatosis (PLG) is a poorly recognized tumor of the central nervous system. To describe the histopathological, immunohistochemical, and molecular features of PLG. Results of our multicentric retrospective study of 6 PLG cases (3 pediatric and 3 adult) were compared with literature data. The mean age was 54.7 years for adults and 8.7 years for children, with 3 males and 3 females. Clinical symptoms were nonspecific. Cerebrospinal fluid analyses showed a high protein level often associated with pleocytosis but without neoplastic cells. On neuroimaging, diffuse leptomeningeal enhancement and hydrocephalus were observed, except in 1 case. PLG was mostly misinterpreted as infectious or tumoral meningitis. The first biopsy was negative in 50% of cases. Histopathologically, PLG cases corresponded to 1 oligodendroglioma without 1p19q codeletion and 5 astrocytomas without expression of p53. No immunostaining for IDH1R132H and no mutations of IDH1/2 and H3F3A genes were found. Overall survival was highly variable (2-82 months) but seems to be increased in children treated with chemotherapy. This study shows the difficulties of PLG diagnosis. The challenge is to achieve an early biopsy to establish a diagnosis and to begin a treatment, but the prognosis remains poor. PLG seems to have a different molecular and immunohistochemical pattern compared with intraparenchymal malignant gliomas. OS, overall survivalPLG, primary leptomeningeal gliomatosis.

Meningioma is the most frequent primary central nervous system tumor. The risk of recurrence and the prognosis are correlated with the extent of the resection that ideally encompasses the infiltrated dura mater and, if required, the infiltrated bone. No device can deliver real-time intraoperative histopathological information on the tumor environment to help the neurosurgeon to achieve a gross total removal. This study assessed the abilities of nonlinear microscopy to provide relevant and real-time data to help resection of meningiomas. Nine human meningioma samples (four World Health Organization Grade I, five Grade II) were analyzed using different optical modalities: spectral analysis and imaging, lifetime measurements, fluorescence lifetime imaging microscopy, fluorescence emitted under one- and two-photon excitation and the second-harmonic generation signal imaging using a multimodal setup. Nonlinear microscopy produced images close to histopathology as a gold standard. The second-harmonic generation signal delineated the collagen background and two-photon fluorescence underlined cell cytoplasm. The matching between fluorescence images and Hematoxylin and Eosin staining was possible in all cases. Grade I meningioma emitted less autofluorescence than Grade II meningioma and Grade II meningioma exhibited a distinct lifetime value. Autofluorescence was correlated with the proliferation rates and seemed to explain the observed differences between Grade I and II meningiomas. This preliminary multimodal study focused on human meningioma samples confirms the potential of tissue autofluorescence analysis and nonlinear microscopy in helping intraoperatively neurosurgeons to reach the actual boundaries of the tumor infiltration. Correspondence between H&E staining (top pictures) and the two-photon fluorescence imaging (bottom pictures).

Rosette-forming glioneuronal tumors (RGNTs) that do not involve the fourth ventricle are rare. RGNTs were originally thought to be exclusively localized into the fourth ventricle but were found in various anatomical localizations.We review the literature and found 32 cases of this particular RGNT. The outcome was excellent with no mortality reported after surgical treatment. Only two patients had received adjuvant therapy for progression. We added one case of a RGNT located in the cerebellar hemisphere.She underwent a subtotal removal with no evidence of progression after. This WHO grade I tumor with a specific biphasic histopathology is of a good oncological outcome after surgical treatment. A long follow-up is needed as recurrence or metastatic progressions exist.

Objectif Les ependymomes supratentoriels pediatriques (ESTP) sont des tumeurs rares dont le grading histologique est controverse. Des marqueurs moleculaires ont recemment ete identifies, notamment le rearrangement du gene RELA codant pour une proteine du complexe NFκB [1] , [2] . L’objectif etait de comparer les techniques de FISH, immuno-histochimie (IHC) et methylome pour la recherche du rearrangement RELA et les integrer aux donnees histologiques. Methodes Une serie de 41 ESTP a ete constituee. Des sondes break apart ciblant C11orf95 et RELA ont ete concues. L’activation de la voie NFκB a ete etudiee en IHC. Une analyse du methylome a ete realisee et les resultats compares. Les resultats ont ete confrontes aux donnees histologiques et de survie. Resultats L’IHC et la FISH etaient positives dans respectivement 56 % (23/41) et 68 % (21/31). La concordance etait de 96 % (25/26). En methylome 63 % des cas (24/38) etaient RELA + . La concordance avec l’IHC et la FISH etait de 87 % (respectivement 33/38 et 27/31). Vingt-six tumeurs ont ete classees en ESTP dont 92 % (24/26) etaient RELA + . Deux cas presentaient un rearrangement YAP et avaient une evolution clinique favorable. En methylome, 12 tumeurs clusterisaient dans des groupes differents : 4 subependymomes, 6 PNET, 1 gliome, 1 ependymome infratentoriel. Histologiquement, les tumeurs reclassees comme subependymomes presentaient des criteres de malignite mais avaient une evolution clinique favorable. Les 6 tumeurs classees en PNET ne presentaient pas les caracteristiques histologiques habituelles et se caracterisaient par des anomalies chromosomiques en 16q et 22q. Conclusion Le rearrangement RELA et l’activation de la voie NFκB peuvent etre recherches en diagnostic par FISH ou IHC et sont caracteristiques des ESTP. De rares ESTP presentent un rearrangement YAP semblant etre de meilleur pronostic. L’analyse du methylome individualise un groupe de tumeurs a differenciation ependymaire presentant des anomalies chromosomiques specifiques.

Objet Dans la leucemie lymphoide chronique (LLC), le dysfonctionnement des telomeres est associe a l’instabilite chromosomique et au pronostic pejoratif. La reponse physiologique aux telomeres dysfonctionnels est dependante de TP53, et son inactivation dans les cellules leucemiques pourrait conduire a un taux d’instabilite telomerique et chromosomique accru. Notre objectif etait d’evaluer l’impact des alterations de TP53 sur les telomeres et l’instabilite chromosomique dans une serie de patients atteints de LLC. Materiels et methodes Nous avons compare un groupe de patients presentant des anomalies de TP53 (deletions et/ou mutations) (n = 24) a un groupe de patients TP53 WT (n = 91). Le statut mutationnel de TP53 a ete determine par sequencage des exons 2–11. Les analyses cytogenetiques classiques et moleculaires ont ete realisees pour evaluer le niveau d’instabilite chromosomique et detecter les signes de fragilite telomerique (FISH pantelomerique). La longueur des telomeres et l’expression des genes du complexe shelterin et de la telomerase ont ete evaluees par PCR quantitative. Resultats Les patients presentant une mutation de TP53 sans deletion 17p et les patients porteurs d’une deletion 17p associes ou non a une mutation TP53 semblent avoir le meme degre de raccourcissement telomerique et d’activation de la telomerase. Par ailleurs, les alterations de TP53 ont ete correlees avec une diminution de l’expression des genes du complexe shelterin. Les analyses cytogenetiques sequentielles ont montre une forte augmentation de l’instabilite chromosomique chez les patients presentant des alterations de TP53. De facon interessante, les patients TP53 WT dont l’expression des genes du complexe shelterin etait diminuee ont aussi montre une apparition d’anomalies cytogenetiques et une survie sans progression plus courte. Conclusion Ces resultats suggerent que les alterations de TP53 jouent un role majeur dans l’instabilite telomerique et chromosomique dans la LLC. En absence d’alteration de TP53, les anomalies d’expression des genes du complexe shelterin peuvent favoriser l’instabilite chromosomique.