Correlation between DNA Methylation and cell proliferation identifies new candidate predictive markers in meningioma

International audienceMeningiomas are the most common primary tumors of the central nervous system. Based on the 2021 WHO classification, they are classified into three grades reflecting recurrence risk and aggressiveness. However, the WHO’s histopathological criteria defining these grades are somewhat subjective. Together with reliable immunohistochemical proliferation indices, other molecular markers such as those studied with genome-wide epigenetics promise to revamp the current prognostic classification. In this study, 48 meningiomas of various grades were randomly included and explored for DNA methylation with the Infinium MethylationEPIC microarray over 850k CpG sites. We conducted differential and correlative analyses on grade and several proliferation indices and markers, such as mitotic index and Ki-67 or MCM6 immunohistochemistry. We also set up Cox proportional hazard models for extensive associations between CpG methylation and survival. We identified loci highly correlated with cell growth and a targeted methylation signature of regulatory regions persistently associated with proliferation, grade, and survival. Candidate genes under the control of these regions include SMC4, ESRRG, PAX6, DOK7, VAV2, OTX1, and PCDHA-PCDHB-PCDHG, i.e., the protocadherin gene clusters. This study highlights the crucial role played by epigenetic mechanisms in shaping dysregulated cellular proliferation and provides potential biomarkers bearing prognostic and therapeutic value for the clinical management of meningioma

Validation of the Idylla GeneFusion assay to detect fusions and MET exon-skipping in non-small cell lung cancers

Abstract Gene fusions and MET exon skipping drive oncogenesis in 8–9% and 3% of non-small cell lung cancers (NSCLC) respectively. Their detection are essential for the management of patients since they confer sensitivity to specific targeted therapies with significant clinical benefit over conventional chemotherapy. Immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) account for historical reference techniques however molecular-based technologies (RNA-based sequencing and RT-PCR) are emerging as alternative or complementary methods. Here, we evaluated the analytical performance of the fully-automated RT-PCR Idylla GeneFusion assay compared to reference methods using 35 fixed NSCLC samples. Idylla demonstrated overall agreement, sensitivity and specificity of 100% compared to RNASeq. Interestingly, it succeeded in retrieving 10 out of 11 samples with inconclusive results due to insufficient RNA quality for sequencing. Idylla showed an overall agreement, sensitivity and specificity of 90.32%, 91.67% and 89.47% compared to IHC/FISH respectively. Using commercial standards, the limit of detection of the Idylla system for the most frequent fusions and exon skipping ranges between 5 and 10 ng RNA input. These results support that the Idylla assay is a reliable and rapid option for the detection of these alterations, however a particular attention is needed for the interpretation of the expression imbalance

Utilisation du logiciel PELICAN pour la saisie et l’export de comptes rendus standardisés dans les tumeurs du système nerveux central : application aux méningiomes

International audienceIntroduction : L’application PELICAN (« Partager efficacement en laboratoire les informations des comptes rendus anatomopathologiques ») est un outil permettant de générer des comptes rendus standardisés et de renseigner automatiquement une base de données, utilisée au CHRU de Nancy depuis 2014 en neuropathologie tumorale. L’objectif de cet article était d’illustrer l’utilisation de cette application pour les méningiomes, avec une première exploitation statistique.Matériels et méthodes : L’étude inclut les cas de méningiomes enregistrés dans l’application PELICAN jusqu’en juillet 2018. L’application PELICAN est un fichier Microsoft Excel comportant un logiciel écrit en Visual Basic pour Applications, utilisé par le pathologiste pour élaborer le compte rendu. Les principales données cliniques ont été recueillies à partir de la fiche de recensement du Registre de l’Hérault. Le suivi était renseigné de façon systématique pour les méningiomes atypiques.Résultats : Deux cent quatre-vingt-quinze méningiomes ont été analysés dont 250 méningiomes de grade I, 42 méningiomes de grade II et 3 méningiomes de grade III. Dans les méningiomes de grade II, on constatait une proportion significativement plus élevée d’hommes (p = 0,002) et d’infiltration de la dure-mère (p < 0,001), une augmentation significative de l’indice Ki-67 (p < 0,0001) et une diminution significative de l’expression des récepteurs à la progestérone (p < 0,001). Dans les méningiomes atypiques, un indice Ki-67 de plus de 20 % était corrélé significativement à une survie sans progression plus courte (p = 0,032).Conclusion : L’application PELICAN est un outil d’utilisation simple, permettant de générer des comptes rendus standardisés tout en alimentant une base de données, ouvrant de très intéressantes perspectives d’un point de vue épidémiologique et scientifique

Adult diffuse low-grade gliomas: 35-year experience at the Nancy France neurooncology unit

International audienceBackground: To report survival, spontaneous prognostic factors, and treatment efficacy in a French monocentric cohort of diffuse low-grade glioma (DLGG) patients over 35 years of follow-up.Methods: A monocentric retrospective study of 339 patients diagnosed with a new DLGG between 01/01/1982 and 01/01/2017 was created. Inclusion criteria were patient age ≥18 years at diagnosis and histological diagnosis of WHO grade II glioma (according to 1993, 2007, and 2016 WHO classifications). The survival parameters were estimated using the Kaplan-Meier method with a 95% confidence interval. Differences in survival were tested for statistical significance by the log-rank test. Factors were considered significant when p ≤ 0.1 and p ≤ 0.05 in the univariate and multivariate analyses, respectively.Results: A total of 339 patients were included with a median follow-up of 8.7 years. The Kaplan-Meier median overall survival was 15.7 years. At the time of radiological diagnosis, Karnofsky Performance Status score and initial tumor volume were significant independent prognostic factors. Oncological prognostic factors were the extent of resection for patients who underwent surgery and the timing of radiotherapy for those concerned. In this study, patients who had delayed radiotherapy (provided remaining low grade) did not have worse survival compared with patients who had early radiotherapy. The functional capabilities of the patients were preserved enough so that they could remain independent during at least three quarters of the follow-up.Conclusion: This large monocentric series spread over a long time clarifies the effects of different therapeutic strategies and their combination in the management of DLGG

Ki-67 and MCM6 labelling index are correlated with overall survival in anaplastic oligodendroglioma, IDH1-mutant and 1p/19q codeleted: a multicenter study from the French POLA Network

International audienceBackground & Objectives: Anaplastic oligodendroglioma (AO), IDH-mutant and 1p/19q codeleted (IDHmut+/1p19qcodel) are high grade gliomas. Only few prognostic markers were studied in this specific histomolecular subgroup. The primary aim of this study was to evaluate and compare the prognostic value of two proliferation markers, MCM6 and Ki-67, in a French multicenter series of IDHmut+/1p19qcodel AO (POLA network), using immunohistochemistry. Further transcriptomic approaches were implemented to uncover the molecular pathways associated with the overexpression of these markers.Methods: Two hundred and thirty-one IDHmut+/1p19qcodel AO cases were included from the French national POLA network. MCM6 and Ki-67 labelling index (LI) were evaluated using computer color image analyser. Transcriptomic data were analysed in a subset of 68 microarray samples from the French POLA Network.Results: High MCM6 (≥ 50%) and Ki-67 (≥ 15%) LI correlated with shorter overall survival (P=0.013 and P=0.004, respectively). A high proliferation index, defined by MCM6 ≥ 50% and/or Ki-67 ≥ 15%, was independently correlated to a shorter survival (P=0.027; multivariate Cox model including age, mitotic index, MCM6 and Ki-67). Transcriptomic analyses revealed that while the high mRNA level of both MCM6 and Ki-67 were positively associated with clusters enriched in gene functions like cell cycle progression, DNA replication, mitosis, pro-neural phenotype as well as neurogenesis, they were negatively associated with clusters of other functions like microglial cell activation, immune response, positive regulation of myelination, oligodendrocyte development, beta-amyloid binding, and postsynaptic specialization.Conclusion: In conclusion, both MCM6 and Ki-67 LI were correlated to overall survival. Because multivariate analyses showed that overexpression of MCM6 and/or Ki-67 was independently correlated to shorter survival, these two easy-to-use and costless markers could be used in association in daily practice in order to predict clinical outcome. Transcriptomics showed that IDHmut+/1p19qcodel AO are highly proliferative tumours with upregulated pro-neural phenotype associated genes, and downregulated immune response, glial differentiation, and myelin-related function

In vivo characterization of physiological and metabolic changes related to isocitrate dehydrogenase 1 mutation expcression by multiparametric MRI and MRS in a rat model with orthotopically grafted human-derived glioblastoma cell lines

International audienceThe physiological mechanism induced by the isocitrate dehydrogenase 1 (IDH1) mutation, associated with better treatment response in gliomas, remains unknown. The aim of this preclinical study was to characterize the IDH1 mutation through in vivo multiparametric MRI and MRS. Multiparametric MRI, including the measurement of blood flow, vascularity, oxygenation, permeability, and in vivo MRS, was performed on a 4.7 T animal MRI system in rat brains grafted with human‐derived glioblastoma U87 cell lines expressing or not the IDH1 mutation by the CRISPR/Cas9 method, and secondarily characterized with additional ex vivo HR‐MAS and histological analyses. In univariate analyses, compared with IDH1−, IDH1+ tumors exhibited higher vascular density (p < 0.01) and better perfusion (p = 0.02 for cerebral blood flow), but lower vessel permeability (p < 0.01 for time to peak (TTP), p = 0.04 for contrast enhancement) and decreased T1 map values (p = 0.02). Using linear discriminant analysis, vascular density and TTP values were found to be independent MRI parameters for characterizing the IDH1 mutation (p < 0.01). In vivo MRS and ex vivo HR‐MAS analysis showed lower metabolites of tumor aggressiveness for IDH1+ tumors (p < 0.01). Overall, the IDH1 mutation exhibited a higher vascularity on MRI, a lower permeability, and a less aggressive metabolic profile. These MRI features may prove helpful to better pinpoint the physiological mechanisms induced by this mutation

Ki‐67 and MCM6 labelling indices are correlated with overall survival in anaplastic oligodendroglioma, IDH1 ‐mutant and 1p/19q‐codeleted: a multicenter study from the French POLA network

International audienceAnaplastic oligodendroglioma (AO), IDH‐mutant and 1p/19q codeleted (IDHmut+/1p19qcodel), is a high‐grade glioma with only limited prognostic markers. The primary objective of this study was to evaluate, by immunohistochemistry, the prognostic value of two proliferation markers, MCM6 and Ki‐67, in a large series of IDHmut+/1p19qcodel AO included in the POLA (“Prise en charge des Oligodendrogliomes Anaplasiques”) French national multicenter network. We additionally examined the transcriptome obtained from this series to understand the functional pathways dysregulated with the mRNA overexpression of these two markers. The labelling indices (LI) of MCM6 and Ki‐67 were obtained via computer‐assisted color image analyses on immunostained AO tissues of the cohort (n=220). Furthermore, a subgroup of AO (n=68/220) was used to perform transcriptomic analyses. A high LI of either MCM6 (≥50%) or Ki‐67 (≥ 15%) correlated with shorter overall survival, both in univariate (P=0.013 and P=0.004, respectively) and multivariate analyses (P=0.027; multivariate Cox model including age, mitotic index, MCM6 and Ki‐67). MCM6 and Ki‐67 LI also correlated with overall survival in an additional retrospective cohort of 30 grade II IDHmut+/1p19qcodel oligodendrogliomas. The prognostic value of MCM6 mRNA level was confirmed in The Cancer Genome Atlas (TCGA) IDHmut+/1p19qcodel gliomas. The transcriptomic approach revealed that high transcriptional expressions of MCM6 and MKI67 were both linked positively with cell cycle progression, DNA replication, mitosis, pro‐neural phenotype as well as neurogenesis, and negatively with microglial cell activation, immune response, positive regulation of myelination, oligodendrocyte development, beta‐amyloid binding, and postsynaptic specialization.In conclusion, the overexpression of MCM6 and/or Ki‐67 is independently associated to shorter overall survival in IDHmut+/1p19qcodel AO. These two easy‐to‐use and cost‐effective markers could thus be used concurrently in routine pathology practice. Additionally, the transcriptomic analyses showed that AO with high proliferation index have down‐regulated immune response and lower microglial cells activation, and bears pro‐neural phenotype

The dural angioleiomyoma harbors frequent GJA4 mutation and a distinct DNA methylation profile

International audienceThe International Society for the Study of Vascular Anomalies (ISSVA) has defined four vascular lesions in the central nervous system (CNS): arteriovenous malformations, cavernous angiomas (also known as cerebral cavernous malformations), venous malformations, and telangiectasias. From a retrospective central radiological and histopathological review of 202 CNS vascular lesions, we identified three cases of unclassified vascular lesions. Interestingly, they shared the same radiological and histopathological features evoking the cavernous subtype of angioleiomyomas described in the soft tissue. We grouped them together with four additional similar cases from our clinicopathological network and performed combined molecular analyses. In addition, cases were compared with a cohort of 5 soft tissue angioleiomyomas. Three out 6 CNS lesions presented the same p.Gly41Cys GJA4 mutation recently reported in hepatic hemangiomas and cutaneous venous malformations and found in 4/5 soft tissue angioleiomyomas of our cohort with available data. Most DNA methylation profiles were not classifiable using the CNS brain tumor (version 12.5), and sarcoma (version 12.2) classifiers. However, using unsupervised t-SNE analysis and hierarchical clustering analysis, 5 of the 6 lesions grouped together and formed a distinct epigenetic group, separated from the clusters of soft tissue angioleiomyomas, other vascular tumors, inflammatory myofibroblastic tumors and meningiomas. Our extensive literature review identified several cases similar to these lesions, with a wide variety of denominations. Based on radiological and histomolecular findings, we suggest the new terminology of “dural angioleiomyomas” (DALM) to designate these lesions characterized by a distinct DNA methylation pattern and frequent GJA4 mutations

Recurrent fusions in PLAGL1 define a distinct subset of pediatric-type supratentorial neuroepithelial tumors

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