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New Brain Tumor Entities Emerge from Molecular Classification of CNS-PNETs
Abstract
Primitive neuroectodermal tumors of the central nervous system (CNS-PNETs) are highly aggressive, poorly differentiated embryonal tumors occurring predominantly in young children but also affecting adolescents and adults. Herein, we demonstrate that a significant proportion of institutionally diagnosed CNS-PNETs display molecular profiles indistinguishable from those of various other well-defined CNS tumor entities, facilitating diagnosis and appropriate therapy for patients with these tumors. From the remaining fraction of CNS-PNETs, we identify four new CNS tumor entities, each associated with a recurrent genetic alteration and distinct histopathological and clinical features. These new molecular entities, designated "CNS neuroblastoma with FOXR2 activation (CNS NB-FOXR2)," "CNS Ewing sarcoma family tumor with CIC alteration (CNS EFT-CIC)," "CNS high-grade neuroepithelial tumor with MN1 alteration (CNS HGNET-MN1)," and "CNS high-grade neuroepithelial tumor with BCOR alteration (CNS HGNET-BCOR)," will enable meaningful clinical trials and the development of therapeutic strategies for patients affected by poorly differentiated CNS tumors.
... Gene profiles of tumors harboring CIC::NUTM1 fusions cluster tightly with those of CIC::DUX4-related sarcomas, regardless of their anatomic location in the central nervous system, kidney, bone, or soft tissues [7,8]. Compared to CIC::DUX4 sarcomas, those with the CIC::NUTM1 fusion tend to lead to a more aggressive clinical course, suggesting that this fusion is likely to be associated with worse prognoses, but additional clinical studies are needed to further investigate this hypothesis. ...
... These characteristics are reminiscent of the myoepithelial-like features of CIC::DUX4 sarcoma, and both exhibit distinct nucleoli [22]. Some cases of CIC::NUTM1 are likely distinct from NC and are better classified as sarcomas, including those of the central nervous system or soft tissues [8]. Moreover, gene expression profiling performed in a large series of round cell sarcomas demonstrated that tumors with the CIC::NUTM1 fusion clustered tightly with CIC::DUX4 and CIC::FOXO4-positive sarcomas and separately from NCs [7], indicating a likely biologic relationship between these CIC-rearranged neoplasms; accordingly, the morphologies of these tumors more closely resemble those of classical CIC-fusion sarcomas. ...
... Meanwhile, cytokeratin or P40 may be occasionally absent in NC [23]. Previous molecular studies have demonstrated that the CIC::NUTM1 fusion (n = 3) clustered tightly with CIC::DUX4 and CIC-FOXO4fusion sarcomas and was split from NCs harboring the BRD3/4::NUTM1 fusion, suggesting that all CIC-rearranged tumors belong to a unique family of tumors [7,8]. Although NUTM1 is positive in both CIC::NUTM1 and NC, the staining pattern is different. ...
Background
CIC-rearranged sarcomas (CRS) represent a new entity of undifferentiated small round cell sarcoma belonging to the Ewing-like sarcomas family. CRS are the most common type. Fusion partners for the CIC gene include DUX4, FOXO4, and the recently recognizedNUTM1. Rare cases of CIC::NUTM1 sarcoma in pediatric patients have recently been reported in brain, kidney, bone, and soft tissues. However, such cases have not been identified in the soft tissues of the limbs.
Case presentation
We reported a case of CIC::NUTM1 sarcoma located in the right upper limb of an 18-year-old man. The tumor displayed morphologic features typical of CIC::DUX4 sarcomas, with small- to medium-sized round cells, a lobular pattern, focal spindling, myxoid stroma, and patchy necrosis. The tumor diffusely expressed NUTM1, was positive for WT1cter at weak to moderate intensity, and was focally positive for CD99, while it was negative for keratins, EMA, P40, MyoD1, myogenin, NKX2.2, BCOR, and pan-TRK. Fluorescence in situ hybridization analyses revealed cleavage of the CIC and NUTM1 genes.
Conclusion
CIC::NUTM1 sarcomas represent a novel molecular variant of CRS with a preference for the central nervous system and younger pediatric persons. Its morphology and phenotype may be mistaken for NUT carcinomas, and the behavior is more progressive than other forms of CRS. For this rare and newly discovered gene fusion variant, it is necessary to integrate molecular and immunohistochemical findings with morphologic features in the diagnosis of undifferentiated neoplasms.
... Embryonal tumors of the CNS are characterized by genetic driving events, which are extremely aggressive, mainly affecting children (Sturm et al., 2016). DNA analysis and gene expression profiling allowed the identification of novel entities, leading to a reclassification of these tumors (Louis et al., 2014). ...
... It typically harbors rearrangements activating the transcription factor FOXR2 (CNS WHO grade 4). It arises in children classically within the supratentorial compartment, rarely with intraventricular location (Sturm et al., 2016). On MRI, this tumor typically manifests as a delineated mass with a cystic and a solid component, exhibiting a mild contrast enhancement (Holsten et al., 2021). ...
... CNS NB-FOXR2 is a recent addition to CNS 2021 WHO, discovered by using DNA methylation analysis, which revealed that several tumors may have belonged to distinct entities. This novel entity has chromosomal rearrangements with overexpression of FOXR2 gene (Sturm et al., 2016;Louis et al., 2020). Next-generation sequencing is required for the discovery of FOXR2 rearrangements, but copy-number analysis may be able to reveal changes to the FOXR2 locus on chromosome Xp11.21. ...
The fifth edition of the WHO Classification of Tumors of the Central Nervous System (CNS), published in 2021, established new approaches to both CNS tumor nomenclature and grading, emphasizing the importance of integrated diagnoses and layered reports. This edition increased the role of molecular diagnostics in CNS tumor classification while still relying on other established approaches such as histology and immunohistochemistry. Moreover, it introduced new tumor types and subtypes based on novel diagnostic technologies such as DNA methylome profiling. Over the past decade, molecular techniques identified numerous key genetic alterations in CSN tumors, with important implications regarding the understanding of pathogenesis but also for prognosis and the development and application of effective molecularly targeted therapies. This review summarizes the major changes in the 2021 fifth edition classification of pediatric CNS tumors, highlighting for each entity the molecular alterations and other information that are relevant for diagnostic, prognostic, or therapeutic purposes and that patients' and oncologists' need from a pathology report.
... Re nement of the process has occurred through recognition of additional tumor-speci c histologic patterns and by integrating testing modalities such as cytogenetics, immunohistochemistry, and nucleic acid sequencing ndings into the classi cation schemes. More recently, it has been recognized that normal and neoplastic tissues have inherent epigenetic signatures encoded in their methylome [2][3][4][5] . The pattern of CpG methylation has been established as a stable and reliable biomarker for tumors and normal tissues 2,6 . ...
... Histologically-de ned tumor types often consist of heterogeneous molecular subtypes with distinct biological and clinical behavior 10,11 . Due to the lack of recurrent de ning mutations, some of these molecular subtypes may only be recognizable by their distinct methylation or transcriptomic signatures 5,12 . ...
... These arrays have favorable characteristics, including relatively low cost, comparable performance on both fresh or formalin-xed and para nembedded (FFPE) tissues, and stability of the methylation mark even in material stored for multiple decades 13 . In addition, the ability to assay FFPE has facilitated the accumulation of large tumor cohorts and allows for easy integration into standard clinical work ows 5,6,14 . ...
As part of the advancement in therapeutic decision-making for brain tumor patients at St. Jude Children’s Research Hospital (SJCRH), we develop and compare the performance of three classification models: a deep learning neural network (NN), an exact bootstrap k-nearest neighbor (kNN), and a random forest classifier (RF) model to predict the 82 molecularly distinct central nervous system (CNS) tumor classes based on DNA-methylation profiles of 2,801 patients. We validate their classification accuracy, precision, and recall with 2,054 samples from two independent cohorts. Although all models perform robustly to missing data, the NN model achieves the highest classification accuracy and maintains better balance between precision and recall than kNN and RF. Average precision and recall of NN reduce to that of RF and kNN only when tumor purity was less than 50%. In conclusion, DNA-methylation based deep learning approach provides the most potential advancement toward precision medicine for brain tumors.
... In recent years, analysis of DNA methylation profiles in central nervous system (CNS) tumors has led to the discovery of new tumor types that exhibit distinct genetic alterations [2]. In 2016, Sturm et al. demonstrated that a portion of tumors, which had been previously diagnosed as primitive neuroectodermal tumors (PNETs), had a distinct DNA methylation profile and were characterized by exon 15 internal tandem (ITD) duplication of BCOR [8]. This new molecular entity was designated CNS high-grade neuroepithelial tumor with BCOR alteration (CNS HGNET-BCOR) [8]. ...
... In 2016, Sturm et al. demonstrated that a portion of tumors, which had been previously diagnosed as primitive neuroectodermal tumors (PNETs), had a distinct DNA methylation profile and were characterized by exon 15 internal tandem (ITD) duplication of BCOR [8]. This new molecular entity was designated CNS high-grade neuroepithelial tumor with BCOR alteration (CNS HGNET-BCOR) [8]. Subsequent studies have confirmed that these tumors are characterized by peculiar histopathological and immunohistochemical features, including circumscribed growth, ependymoma-like perivascular pseudorosettes, delicate branching capillaries, microcystic pattern, focal or absent GFAP immunostaining, focal OLIG2 positivity, variable expression of NeuN, and immunoreactivity for BCOR [1,3,17]. ...
... After the first description of CNS tumor with BCOR ITD [8], other tumors with overlapping morphological and immunohistochemical features, but harboring fusions of BCOR with either EP300 or its paralog CREBBP, instead of BCOR ITD, have been reported in the CNS [7,9,11,12,14,15]. These tumors exhibit a DNA methylation profile close to that of CNS tumors with BCOR ITD, suggesting a similarity between the two [9,11,14,15]. ...
The fifth edition of the World Health Organization (WHO) classification of central nervous system (CNS) tumors introduced the new tumor type CNS tumor with BCOR internal tandem duplication (ITD), characterized by a distinct DNA methylation profile and peculiar histopathological features, including a circumscribed growth pattern, ependymoma-like perivascular pseudorosettes, microcystic pattern, absent or focal GFAP immunostaining, OLIG2 positivity, and BCOR immunoreactivity. We describe a rare case of a CNS tumor in a 45-year-old man with histopathological and immunohistochemical features overlapping the CNS tumor with BCOR internal tandem duplication (ITD) but lacking BCOR immunostaining and BCOR ITD. Instead, the tumor showed CREBBP::BCORL1 fusion and pathogenic mutations in BCOR and CREBBP, along with a DNA methylation profile matching the “CNS tumor with EP300:BCOR(L1) fusion” methylation class. Two CNS tumors with fusions between CREBBP, or its paralog EP300, and BCORL1, and approximately twenty CNS tumors with CREBBP/EP300::BCOR fusions have been reported to date. They exhibited similar ependymoma-like features or a microcystic pattern, along with focal or absent GFAP immunostaining, and shared the same DNA methylation profile. Given their morphological and epigenetic similarities, circumscribed CNS tumors with EP300/CREBBP::BCOR(L1) fusions and CNS tumors with BCOR ITD may represent variants of the same tumor type. The ependymoma-like aspect coupled with the lack of diffuse GFAP immunostaining and the presence of OLIG2 positivity are useful clues for recognizing these tumors in histopathological practice. The diagnosis should be confirmed after testing for BCOR(L1) gene fusions and BCOR ITD.
... Herein, we present for the first time a case harboring pseudorosettes. The variety of histopathology and grading (neurocytic from embryonal morphology with neuropil islands and spongioblastic features) in association with the co-expression of OLIG2 and neuronal markers may morphologically resemble CNS NB-FOXR2 [2,6]. However, in contrast to the aforementioned tumors, DGONC expresses SOX10, which may potentially be important for a diagnosis [7]. ...
... However, in contrast to the aforementioned tumors, DGONC expresses SOX10, which may potentially be important for a diagnosis [7]. A chromosome 1q gain, which is almost always observed in CNS NB-FOXR2, may also be encountered in DGONC (19/35 reported cases, 54%) [1,5,6]. The monosomy of chromosome 14, which is a desirable diagnostic criterion for the current WHO classification, may be exceptionally absent (1/44 reported cases, 2%) [1][2][3][4][5], as was the case in our observation. ...
A novel histomolecular tumor of the central nervous system (CNS), the “diffuse glioneuronal tumor with oligodendroglioma-like features and nuclear clusters (DGONC),” has recently been identified, based on a distinct DNA methylation profile and has been added to the 2021 World Health Organization Classification of CNS Tumors. This glioneuronal tumor mainly affects the supratentorial area in children and recurrently presents with a monosomy of chromosome 14. Herein, we report the case of a DNA-methylation based diagnosis of DGONC having atypical features, such as pseudo-rosettes and the absence of a chromosome 14 monosomy, thus rendering its diagnosis very challenging. Because of the wide variety of morphologies harbored by DGONC, a large range of differential diagnoses may be hypothesized from benign to malignant. Interestingly, the current case, like one previously reported, exhibited a co-expression of OLIG2, synaptophysin and SOX10, without GFAP immunopositivity. This particular immunophenotype seems to be a good indicator for a DGONC diagnosis. The classification of DGONC amongst glioneuronal or embryonal tumors is still debated. The clinical (a pediatric supratentorial tumor), morphological (from a benign oligodendroglioma-like tumor with microcalcifications and possible neuropil-like islands to a malignant embryonal tumor with a possible spongioblastic pattern), and immunohistochemical (co-expression of OLIG2 and synaptophsyin) profiles resemble CNS, neuroblastoma, FOXR2-activated and may potentially bring them together in a future classification. Further comprehensive studies are needed to conclude the cellular origin of DGONC and its prognosis.
... CIC::NUTM1 was first reported in 2016 in a molecular analysis of primitive neuroectodermal tumors of the central nervous system (CNS), where two patients harbored a fusion between CIC exon 16 and NUTM1 exon 4 (Sturm et al., 2016). Since then, CIC:: NUTM1 fusions have been identified in at least an additional one CNS patient and twenty sarcoma patients, where the chimera occurs between CIC exons 16, 17, 18, and 20 and NUTM1 exons 2, 3, 4, 5, and 6 (Mangray et al., 2018;Schaefer et al., 2018;Watson et al., 2018;Le Loarer et al., 2019;Biederman et al., 2022;Yang et al., 2022;Ma et al., 2023;Sievers et al., 2023) ( Figure 1). ...
... Sturm et al., 2016; Mangray et al., 2018;Schaefer et al., 2018; Watson et al., 2018;Le Loarer et al., 2019;Biederman et al., 2022;Yang et al., 2022;Ma et al., 2023; Sievers et al., 2023) ( ...
Capicua (CIC)-rearranged sarcomas are an aggressive subset of undifferentiated round cell sarcomas. CIC::DUX4, the proto-typical CIC fusion oncoprotein is associated with rapid clinical progression and chemotherapy resistance leading to poor clinical outcomes. Recent studies have identified additional CIC fusions (CIC::NUTM1, CIC::FOXO4, and CIC::LEUTX) that largely retain CIC-binding specificity but leverage C-terminal binding partners (NUTM1, FOXO4, and LEUTX) to potentially activate transcriptional programs that drive oncogenesis. Moreover, the recent development of preclinical models to study CIC::DUX4 sarcoma have advanced our understanding of the underlying biological mechanisms and uncovered key dependencies that can be translated into rational therapies. In this review, we will highlight these recent advancements in CIC-rearranged sarcoma biology with a vision for clinical translation to improve patient outcomes.
... The above-noted studies examined the presence of identifying mutations, copy-number abnormalities, or mutagenic effects; yet such approaches have a relatively narrow scope and do not take into account larger-scale changes across the genome that could reflect significant shifts in biology from in vivo to in vitro environments. Genome-wide assessment of DNA methylation patterns by microarray has arguably become the single most powerful tool for diagnosing and subtyping brain tumors [9], creating new diagnostic categories and merging others while revealing some of the limitations of traditional histopathologic diagnosis [10,11]. It interrogates hundreds of thousands of methylation loci throughout the genome to give a highly detailed epigenetic profile for a sample. ...
... (www.preprints.org) | NOT PEER-REVIEWED | Posted: 7 March 2024 doi:10.20944/preprints202403.0432.v110 ...
Diffuse midline gliomas are among the deadliest human cancers and have had little progress in treatment in the last 50 years. Cell cultures of these tumors have been developed recently, but it is unknown the degree to which such cultures retain the characteristics of the source tumors. DNA methylation profiling offers a powerful tool to look at genome-wide epigenetic changes that are biologically meaningful and can help assess the similarity of cultured tumor cells to their in vivo progenitors. Paraffinized diagnostic tissue from three diffuse intrinsic pontine gliomas with H3 K27M mutations was compared with subsequent passages of neurosphere cell cultures from those tumors. Each cell line was passaged 3-4 times and analyzed with DNA methylation arrays and standard algorithms that provided comparison of diagnostic classification and cluster analysis. All samples tested maintained high classifier scores and clustered within the reference group of H3 K27M-mutant diffuse midline gliomas. There was a gain of 1q in all cell lines, with two cell lines initially manifesting the gain of 1q only during culture. In vitro cell cultures of H3 K27M-mutant gliomas maintain high degrees of similarity in DNA methylation profiles to their source tumor, confirming their fidelity even with some chromosomal changes.
... Illumina Infinium BeadChip arrays). Profiling of DNA methylation, a central epigenetic mark, has enabled the discovery and delineation of molecular classes of brain tumors, sarcomas, leukemias, and many other types of cancer (Sturm et al. 2016, Capper et al. 2018, Giacopelli et al. 2021, Kö lsche et al. 2021. A recent machine learning-based patient classification system for brain tumors (often referred to as the "Heidelberg classifier") showcases the high clinical utility of this data type and is integrated in routine workflows worldwide, having classified over 100 000 cases as of December 2022 (Capper et al. 2018, Sturm et al. 2023. ...
... We first presented our approach in studies of pediatric high-grade glioma and medulloblastoma (Sturm et al. 2012, Hovestadt et al. 2013) and made it available to the research community as the conumee Bioconductor package in 2015 (Hovestadt and Zapatka 2015). With nearly 20 000 downloads, conumee is one of the most widely used tools for inferring CNVs from DNA methylation arrays and has been applied in numerous large-scale cancer research projects (Sturm et al. 2016, Northcott et al. 2017. Conumee-derived CNV profiles are frequently displayed in the most recent WHO classification of Tumors of the Central Nervous System and are integrated in automated molecular reports of the Heidelberg classifier, where inferred CNVs may have an impact on tumor classification and clinical decisions (Capper et al. 2018, WHO Classification of Tumours Editorial Board 2021. ...
Motivation
Copy-number variations (CNVs) are common genetic alterations in cancer and their detection may impact tumor classification and therapeutic decisions. However, detection of clinically relevant large and focal CNVs remains challenging when sample material or resources are limited. This has motivated us to create a software tool to infer CNVs from DNA methylation arrays which are often generated as part of clinical routines and in research settings.
Results
We present our R package, conumee 2.0, that combines tangent normalization, an adjustable genomic binning heuristic, and weighted circular binary segmentation to utilize DNA methylation arrays for CNV analysis and mitigate technical biases and batch effects. Segmentation results were validated in a lung squamous cell carcinoma dataset from TCGA (n = 367 samples) by comparison to segmentations derived from genotyping arrays (Pearson’s correlation coefficient of 0.91). We further introduce a segmented block bootstrapping approach to detect focal alternations that achieved 60.9% sensitivity and 98.6% specificity for deletions affecting CDKN2A/B (60.0% and 96.9% for RB1, respectively) in a low-grade glioma cohort from TCGA (n = 239 samples). Finally, our tool provides functionality to detect and summarize CNVs across large sample cohorts.
Availability and implementation
Conumee 2.0 is available under open-source license at: https://github.com/hovestadtlab/conumee2.
Supplementary information
Supplementary data are available at Bioinformatics online.
... When combining the Illumina Infinium Methylation array data with AI methods, several clinical-grade DNAm classifiers were created [5,8], particularly suitable for individualized cancer diagnostics [9][10][11][12]. However, a comprehensive R package specially for it is lacking. ...
... Based on these AI techniques, our package has shown excellent efficacy in overcoming the inter-observer variability of histopathological diagnosis, which poses a big challenge to the neural tumor case study, largely increasing the misclassification rate [10,23,24]. This problem is even more severe for the sarcoma case because approximately half of the sarcoma entities lack morphologic or molecular hallmarks [25]. ...
DNA methylation profiling is a useful tool to increase the accuracy of a cancer diagnosis. However, a comprehensive R package specially for it is lacking. Hence, we developed the R package methylClass for methylation-based classification. Within it, we provide the eSVM (ensemble-based support vector machine) model to achieve much higher accuracy in methylation data classification than the popular random forest model and overcome the time-consuming problem of the traditional SVM. In addition, some novel feature selection methods are included in the package to improve the classification. Furthermore, because methylation data can be converted to other omics, such as copy number variation data, we also provide functions for multi-omics studies. The testing of this package on four datasets shows the accurate performance of our package, especially eSVM, which can be used in both methylation and multi-omics models and outperforms other methods in both cases. methylClass is available at: https://github.com/yuabrahamliu/methylClass.
... A tumorsejtek OLIG2re intenzíven, szinaptofizinre részben pozitívak. Prognózisáról egyelőre csak limitált mennyiségű adat áll rendelkezésre(47).Központi idegrendszeri tumor BCOR internal tandem duplikációval Ez a ritka, bizonytalan hisztogenezisű tumor az esetek döntő többségében a cerebrális és cerebelláris hemiszfériumokban alakul ki. Általában kicsi, monomorf, kerek vagy orsó ala-6. ...
... Hátterében a BCOR gén 15. exonjának internal tandem duplikációja (ITD) mutatható ki. Prognózisáról jelenleg limitált mennyiségű adat áll rendelkezésre(47).A központi idegrendszer embrionális tumora, NEC/NOS (KIR WHO grade 4) Ebbe a diagnosztikai csoportba olyan embrionális tumorok sorolhatók, amelyek sem klinikopatológiai, sem molekuláris sajátosságaik alapján nem illeszthetők egyik fent tárgyalt kategóriába sem. A jövőben várhatóan több, egyelőre ide sorolt tumorcsoport külön entitásként jelenik majd meg.A PINEÁLIS RÉGIÓ TUMORAI Ez a lokalizáció alapján definiált csoport változatos hisztogenezisű, prognózisú és különböző életkorban előforduló tumorokat foglal magában. ...
Considerable changes were introduced into the 5th World Health Organization (WHO) classification of central nervous system (CNS) tumors, published in 2021, including new entities, a clearer classification of previous categories, correlating better with clinical behavior and changes in nomenclature. The number of definitions based on molecular features in addition to histopathology continued to increase. Here, we highlight the basic principles of the 5th CNS WHO classification and discuss glial, glioneuronal, neuronal, choroid plexus, embryonal and pineal tumors, as well as meningiomas in more details. We pay special attention to new entities as well as altered criteria and designations. Our primary goal is to present the "classical" pathological aspects, but the inseparable molecular pathological features are also briefly discussed, to the absolutely necessary extent for comprehension. We aim to provide a guideline to understand the modern classification of CNS tumors for practitioners of neuro-oncology and neuropathology.
... Gene expression data of embryonal brain tumor samples (CEL files) were acquired from the publicly available data sets GSE10327, 14 GSE122077, 6 GSE70678, 15 and GSE73038. 16 Sample annotations were obtained from the respective GEO deposits and the corresponding publications. Sample annotations were checked for redundancy. ...
... 6,14-16 Consensus clustering of the compiled data confirmed robust clustering of tumor entities based on transcriptome profiles (Supplementary Figure 6A), as previously described. 16 Tissue from a subset of embryonal brain tumors of our case series was also available for global DNA methylation analysis (n = 28). As expected, and previously described, 29 consensus clustering based on methylome data showcased robust clustering of samples according to tumor entities (Supplementary Figure 6B). ...
Background
Embryonal tumors with multilayered rosettes (ETMR) are rare malignant embryonal brain tumors. The prognosis of ETMR is poor and novel therapeutic approaches are desperately needed. Comprehension of ETMR tumor biology is currently based on only few previous molecular studies, which mainly focused on the analyses of nucleic acids. In this study, we explored integrated ETMR proteomics.
Methods
Using mass spectrometry, proteome data were acquired from 16 ETMR and the ETMR cell line BT183. Proteome data were integrated with case-matched global DNA methylation data, publicly available transcriptome data, and proteome data of further embryonal and pediatric brain tumors.
Results
Proteome-based cluster analyses grouped ETMR samples according to histomorphology, separating neuropil-rich tumors with neuronal signatures from primitive tumors with signatures relating to stemness and chromosome organization. Integrated proteomics showcased that ETMR and BT183 cells harbor proteasome regulatory proteins in abundancy, implicating their strong dependency on the proteasome machinery to safeguard proteostasis. Indeed, in vitro assays using BT183 highlighted that ETMR tumor cells are highly vulnerable towards treatment with the CNS penetrant proteasome inhibitor Marizomib.
Conclusions
In summary, histomorphology stipulates the proteome signatures of ETMR, and proteasome regulatory proteins are pervasively abundant in these tumors. As validated in vitro, proteasome inhibition poses a promising therapeutic option in ETMR.
... There is a lack of clinical prognosis and correlation with histological features in certain types of tumors, such as in pediatric posterior fossa ependymomas (11) or in diffuse gliomas (12). These factors collectively contribute to a reduced likelihood of achieving accurate diagnoses (8,9,(13)(14)(15). As a result, updates in the classification system led to the incorporation of molecular markers for the first time in 2016 (16,17). ...
Brain tumors and genomics have a long-standing history given that glioblastoma was the first cancer studied by the cancer genome atlas. The numerous and continuous advances through the decades in sequencing technologies have aided in the advanced molecular characterization of brain tumors for diagnosis, prognosis, and treatment. Since the implementation of molecular biomarkers by the WHO CNS in 2016, the genomics of brain tumors has been integrated into diagnostic criteria. Long-read sequencing, also known as third generation sequencing, is an emerging technique that allows for the sequencing of longer DNA segments leading to improved detection of structural variants and epigenetics. These capabilities are opening a way for better characterization of brain tumors. Here, we present a comprehensive summary of the state of the art of third-generation sequencing in the application for brain tumor diagnosis, prognosis, and treatment. We discuss the advantages and potential new implementations of long-read sequencing into clinical paradigms for neuro-oncology patients.
... Consequently, clinically relevant models are needed to understand tumor biology and to develop more effective and less toxic therapeutic options. Additionally, multiple seminal studies have revealed complex biological heterogeneities of brain tumors and identified molecularly distinct subgroups within the classic pathological diagnosis of brain tumors [6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22]. These findings highlight the need to develop a large panel of animal models replicating the full spectrum of molecular subtypes of pediatric brain tumors. ...
Background: Despite multimodality therapies, the prognosis of patients with malignant brain tumors remains extremely poor. One of the major obstacles that hinders development of effective therapies is the limited availability of clinically relevant and biologically accurate (CRBA) mouse models. Methods: We have developed a freehand surgical technique that allows for rapid and safe injection of fresh human brain tumor specimens directly into the matching locations (cerebrum, cerebellum, or brainstem) in the brains of SCID mice. Results: Using this technique, we successfully developed 188 PDOX models from 408 brain tumor patient samples (both high-and low-grade) with a success rate of 72.3% in high-grade glioma, 64.2% in medulloblastoma, 50% in ATRT, 33.8% in ependymoma, and 11.6% in low-grade gliomas. Detailed characterization confirmed their replication of the histopathological and genetic abnormalities of the original patient tumors. Conclusions: The protocol is easy to follow, without a sterotactic frame, in order to generate large cohorts of tumor-bearing mice to meet the needs of biological studies and preclinical drug testing.
... The above-noted studies examined the presence of identifying mutations, copy-number abnormalities, or mutagenic effects; yet such approaches have a relatively narrow scope and do not take into account larger-scale changes across the genome that could reflect significant shifts in biology from in vivo to in vitro environments. The genome-wide assessment of DNA methylation patterns by microarray has arguably become the single most powerful tool for diagnosing and subtyping brain tumors [9], creating new diagnostic categories and merging others while revealing some of the limitations of traditional histopathologic diagnosis [10,11]. It interrogates hundreds of thousands of methylation loci throughout the genome to give a highly detailed epigenetic profile for a sample. ...
Diffuse midline gliomas are among the deadliest human cancers and have had little progress in treatment in the last 50 years. Cell cultures of these tumors have been developed recently, but the degree to which such cultures retain the characteristics of the source tumors is unknown. DNA methylation profiling offers a powerful tool to look at genome-wide epigenetic changes that are biologically meaningful and can help assess the similarity of cultured tumor cells to their in vivo progenitors. Paraffinized diagnostic tissue from three diffuse intrinsic pontine gliomas with H3 K27M mutations was compared with subsequent passages of neurosphere cell cultures from those tumors. Each cell line was passaged 3–4 times and analyzed with DNA methylation arrays and standard algorithms that provided a comparison of diagnostic classification and cluster analysis. All samples tested maintained high classifier scores and clustered within the reference group of H3 K27M-mutant diffuse midline gliomas. There was a gain of 1q in all cell lines, with two cell lines initially manifesting the gain of 1q only during culture. In vitro cell cultures of H3 K27M-mutant gliomas maintain high degrees of similarity in DNA methylation profiles to their source tumor, confirming their fidelity even with some chromosomal changes.
... EWSR1 gene rearrangements have been reported in CNS neoplasms [9,11]; Roosen, Ode, Bunt, & Kool [13],; Sturm et al [15]. Some of these EWSR1 rearrangements are fusions with the PLAGL1 gene, and other PLAGL1 gene rearrangements in CNS neoplasms have also been described [14]. ...
Purpose
Central nervous system (CNS) embryonal tumors are a diverse group of malignant tumors typically affecting pediatric patients that recently have been better defined, and this paper describes evolution of a unique type of embryonal tumor at relapse.
Methods
Two pediatric patients with CNS embryonal tumors with EWSR1-PLAGL1 rearrangements treated at Arkansas Children’s Hospital with histopathologic and molecular data are described.
Results
These two patients at diagnosis were classified as CNS embryonal tumors with EWSR1-PLAGL1 rearrangements based on histologic appearance and molecular data. At relapse both patient’s disease was reclassified as atypical teratoid rhabdoid tumor (ATRT) based on loss of INI-1, presence of SMARCB1 alterations, and methylation profiling results.
Conclusion
CNS embryonal tumors with EWSR1-PLAGL1 rearrangements acquire or include a population of cells with SMARCB1 alterations that are the component that predominate at relapse, suggesting treatment aimed at this disease component at diagnosis should be considered.
... An accurate and detailed diagnosis is vital for treatment decisions and determines patient outcome 4,5 . The current diagnostic procedure of (pediatric) brain tumors requires a tumor tissue biopsy for histopathological investigation [6][7][8] . In this evaluation, inter-observatory variability occasionally results in misdiagnosis 6-8 . ...
Pediatric central nervous system tumors remain challenging to diagnose. Imaging approaches do not provide sufficient detail to discriminate between different tumor types, while the histopathological examination of tumor tissue shows high interobserver variability. Recent studies have demonstrated the accurate classification of central nervous system tumors based on the DNA-methylation profile on a tumor biopsy. However, a brain biopsy holds significant risk of bleeding and damaging the surrounding tissues. Liquid biopsy approaches analyzing circulating tumor DNA show high potential as an alternative and less invasive tool to study the DNA-methylation pattern of tumors. In this study, we explore the potential of classifying pediatric brain tumors based on methylation profiling of the cell-free DNA in cerebrospinal fluid (CSF). For this proof-of-concept study, we collected 20 cerebrospinal fluid samples of pediatric brain cancer patients via a ventricular drain placed for reasons of increased intracranial pressure. Analyses on the circulating cell-free DNA (cfDNA) showed high variability of cfDNA quantities across patients ranging from levels below the limit of quantification to 40 ng cfDNA per milliliter of CSF. Classification based on methylation profiling of cfDNA from CSF was correct for 8 out of 20 samples in our cohort. Accurate results were mostly observed in samples of high quality, more specifically those with limited high-molecular weight DNA contamination. Interestingly, we show that centrifugation of the CSF prior to processing increases the fraction of fragmented cfDNA to high-molecular weight DNA. In addition, classification was mostly correct for samples with high tumoral cfDNA fraction as estimated by computational deconvolution (> 40%). In summary, analysis of cfDNA in the CSF shows potential as a tool for diagnosing pediatric nervous system tumors especially in patients with high levels of tumoral cfDNA in the CSF, however further optimization of the collection procedure, experimental workflow, and bioinformatic approach is required to also allow classification for patients with low tumoral fractions in the CSF.
... DNA methylation arrays greatly improve diagnostic precision and reliability, and are highly effective for the discovery of new tumor types [1]. This is particularly true of central nervous system (CNS) embryonal tumors, a tumor class comprising a growing list of subgroups largely refined by epigenetic studies [2]. These tumors are high-grade malignancies composed of primitive neuroepithelial cells, predominantly seen in childhood. ...
Central nervous system (CNS) embryonal tumors are a heterogeneous group of high-grade malignancies, and the increasing clinical use of methylation profiling and next-generation sequencing has led to the identification of molecularly distinct subtypes. One proposed tumor type, CNS tumor with BRD4::LEUTX fusion, has been described. As only a few CNS tumors with BRD4::LEUTX fusions have been described, we herein characterize a cohort of 9 such cases (4 new, 5 previously published) to further describe their clinicopathologic and molecular features. We demonstrate that CNS embryonal tumor with BRD4::LEUTX fusion comprises a well-defined methylation class/cluster. We find that patients are young (4 years or younger), with large tumors at variable locations, and frequently with evidence of leptomeningeal/cerebrospinal fluid (CSF) dissemination. Histologically, tumors were highly cellular with high-grade embryonal features. Immunohistochemically, 5/5 cases showed synaptophysin and 4/5 showed OLIG2 expression, thus overlapping with CNS neuroblastoma, FOXR2-activated. DNA copy number profiles were generally flat; however, two tumors had chromosome 1q gains. No recurring genomic changes, besides the presence of the fusion, were found. The LEUTX portion of the fusion transcript was constant in all cases assessed, while the BRD4 portion varied but included a domain with proto-oncogenic activity in all cases. Two patients with clinical follow up available had tumors with excellent response to chemotherapy. Two of our patients were alive without evidence of recurrence or progression after gross total resection and chemotherapy at 16 and 33 months. One patient relapsed, and the last of our four patients died of disease one month after diagnosis. Overall, this case series provides additional evidence for this as a distinct tumor type defined by the presence of a specific fusion as well as a distinct DNA methylation signature. Studies on larger series are required to further characterize these tumors.
... A recent research using genome-wide DNA methylation analysis by Sturm et al. highlighted that embryonal tumors of the central nervous system (CNS), formerly called CNS-primitive neuroectodermal tumors (PNET), consisted of various types of tumors including a number of novel tumor entitiessuch as BCOR -altered high-grade neuroepithelial tumors 1) . Considering the clinical and molecular heterogeneities, performing genetic and epigenetic analysis has become essential to reveal the molecular background of the tumor and establish a "true" diagnosis in patients with CNS embryonal tumors, not otherwise specified (NOS). ...
Among intracranial embryonal tumors, genetic/epigenetic analysis can help unveil the molecular background. Herein, we report a case of a cerebellar pontine angular embryonic tumor complicated by Cowden syndrome in an infant. The tumor radiologically lacked evidence of interaction with the cerebellum and appeared to show continuity with the pons. Pathological appearance was similar to that of a medulloblastoma with extensive nodularity. DNA methylation analysis indicated that the tumor was a “medulloblastoma, sonic hedgehog-activated” with a substantial confidence. Although the diagnosis deviated from the definition of medulloblastoma, clinical, pathological and molecular data suggested that it was an “ectopic” medulloblastoma.
... For the knowledge matrix, we used four integrated datasets, namely, bloodCellMarkersIRISDMAP, svmMarkers, canonicalPathways, and oncogenicPathways, which were incorporated into the R package PLIER 22 . The knowledge matrix included 545 biological pathways 23 , 83 cell-and tissue-specific genetic markers 24 , 189 cancerrelated pathways 25 , and 817 gene sets. The methPLIER-CTD was constructed using a gene set for 2545 adenocarcinoma-related chemicals obtained from the Comparative Toxicogenomics Database (CTD). ...
DNA methylation is an epigenetic modification that results in dynamic changes during ontogenesis and cell differentiation. DNA methylation patterns regulate gene expression and have been widely researched. While tools for DNA methylation analysis have been developed, most of them have focused on intergroup comparative analysis within a dataset; therefore, it is difficult to conduct cross-dataset studies, such as rare disease studies or cross-institutional studies. This study describes a novel method for DNA methylation analysis, namely, methPLIER, which enables interdataset comparative analyses. methPLIER combines Pathway Level Information Extractor (PLIER), which is a non-negative matrix factorization (NMF) method, with regularization by a knowledge matrix and transfer learning. methPLIER can be used to perform intersample and interdataset comparative analysis based on latent feature matrices, which are obtained via matrix factorization of large-scale data, and factor-loading matrices, which are obtained through matrix factorization of the data to be analyzed. We used methPLIER to analyze a lung cancer dataset and confirmed that the data decomposition reflected sample characteristics for recurrence-free survival. Moreover, methPLIER can analyze data obtained via different preprocessing methods, thereby reducing distributional bias among datasets due to preprocessing. Furthermore, methPLIER can be employed for comparative analyses of methylation data obtained from different platforms, thereby reducing bias in data distribution due to platform differences. methPLIER is expected to facilitate cross-sectional DNA methylation data analysis and enhance DNA methylation data resources.
... BEND1, BEND5, and RBB are revealed to bind DNA through sequence-specific recognition (Dai et al., 2015(Dai et al., , 2013Xuan et al., 2012), and BEND3 specifically binds heterochromatin (Sathyan et al., 2011a). Additionally, several human BEND2 fusion proteins have been identified in tumors, and the activation of BEND2 is suggested to promote oncogenic activity (Burford et al., 2018;Scarpa et al., 2017;Sturm et al., 2016;Williamson et al., 2019). These data suggest that BEND2 might have a role in regulating chromatin and transcriptional gene expression. ...
Reproductive aging, characterized by a decline in female reproductive potential, is a significant biomedical challenge. A key factor in reproductive aging is the depletion of the ovarian reserve, the pool of primordial follicles in the ovary. Recent studies have implicated BEND2, a BEN domain-containing protein family member, in mammalian spermatogenesis. In the testis, Bend2 expresses two protein isoforms: full-length and truncated. Ablation of both proteins results in an arrested spermatogenesis. Because the Bend2 locus is on the X chromosome, and the Bend2 -/y mutants are sterile, Bend2 ’s role in oogenesis remained elusive.
In this study, we employed a novel Bend2 mutation that completely blocks the expression of the full-length BEND2 protein but allows the expression of the truncated BEND2 isoform. However, this mutation does not confer male sterility, allowing us to investigate BEND2’s role in mice’s oocyte quality, follicular dynamics, and fertility. Our findings demonstrate that full-length BEND2 is dispensable for male fertility, and its ablation leads to impaired oocyte quality, reduced follicular formation, and an accelerated decline in fertility. These results reveal a critical role for BEND2 in oogenesis and provide insights into the mechanisms underlying reproductive aging. Furthermore, these findings hold relevance for the diagnostic landscape of human infertility.
... В 2016 г. опухоль центральной нервной системы (ЦНС) с внутренней тандемной дупликацией (ITD) BCOR была выделена из группы неспецифицированных эмбриональных опухолей ЦНС на основании характерного профиля метилирования ДНК, а также выявления повторяющейся аберрации -ITD в экзоне 15 гена BCOR [1]. ...
In 2022, central nervous system (CNS) tumor with BCOR internal tandem duplication (BCOR ITD) was included in the fifth edition of the World Health Organization Classification of Tumors of the CNS as part of the embryonal tumor group. The identification a distinct DNA methylation profile and the presence of a recurrent genetic aberration – BCOR ITD – made it possible to recognize these tumors as a separate entity. In most cases, these tumors occur in children under 5 years of age and are located in the hemispheres of the cerebellum or brain. Since CNS tumor with BCOR ITD is a rare and relatively new tumor entity, it is not yet widely known and can be misdiagnosed. The most common initial diagnosis in patients referred for a second opinion to our reference center is anaplastic/classical medulloblastoma. In this article, we aimed to summarize the characteristic clinical, morphological and immunophenotypic features of CNS tumors with BCOR ITD based on 8 clinical cases confirmed by molecular genetic testing. The study was approved by the Independent Ethics Committee and the Scientific Council of the Dmitry Rogachev National Medical Research Center of Pediatric Hematology, Oncology and Immunology.
... Prior to 2021, astroblastomas were diagnosed purely on histological features showing tumour cells in characteristic perivascular pseudorosettes [5]. Astroblastoma is now defined by the latest WHO classification as harbouring an alteration in the MN1 gene, typically MN1/BEND2 fusion [6]. Recent reports including the current study have shown astroblastomalike tumours with EWSR1/BEND2 gene fusion without MN1 abnormalities suggesting an epigenetically distinct subtype of astroblastoma. ...
Astroblastomas are rare tumours of the central nervous
system, and only a few cases of brain stem/primary spinal cord
astroblastoma have been reported. It has been suggested
that EWSR1/BEND2 fusion defines an epigenetically
distinct subtype of astroblastoma. We present a rare case
of primary intramedullary spinal cord astroblastoma with
EWSR1/BEND2 fusion. To our knowledge, this is only
the third reported case in the literature with this specific
diagnosis, the first in an adult female, and the first that has
been treated upfront with total surgical resection.
Embryonal tumors of the central nervous system (CNS) are rare and aggressive malignancies accounting for less than 1% of all central nervous system tumors. The occurrence of metastasis to extracranial sites, especially the parotid region, is highly uncommon. We present a rare case of metastatic frontal embryonal tumor (ET) in the parotid region. A 9‐year‐old boy presented with a progressively enlarging left parotid mass. Past history revealed that he was a known case of a frontal lobe embryonal tumor. Fine‐needle aspiration cytology (FNAC) combined with immunocytochemistry from the parotid revealed a metastatic embryonal tumor. This case report highlights the importance of considering metastatic tumors in evaluating parotid masses, even in pediatric patients, and emphasizes the diagnostic potential of FNAC in diagnosing such rare and unusual tumors for prompt and appropriate patient management.
In the most recent fifth edition of the World Health Organization Classification of Tumors of the Central Nervous System, astroblastoma has been defined by molecular rearrangements involving the MN1 gene, with common partners being BEND2 or CXXC5 . Accordingly, this tumor entity is now known as “astroblastoma, MN1 -altered.” However, gliomas with EWSR1::BEND2 fusions, devoid of MN1 fusion alterations, have recently been shown to exhibit astroblastoma-like histomorphologic features and reside in a distinct epigenetic subgroup based on DNA methylation studies similar to high-grade neuroepithelial tumor with MN1 alteration, which includes astroblastoma, MN1 altered tumors. This new epigenetically distinct subtype of astroblastoma containing EWSR1::BEND2 fusions lacks the required MN1 alteration and, thus, does not satisfy the current molecular classification of these lesions. Here, we describe a case of glioma with histologic features and DNA methylation profiling consistent with astroblastoma with a novel YAP1: : BEND2 fusion. This case and others further expand the molecular findings observable in astroblastoma-like tumors outside the constraints of MN1 alteration. Such cases of astroblastoma with EWSR1::BEND2 and YAP1::BEND2 fusions challenge the current molecular classification of astroblastoma based solely on an MN1 alteration.
We report the novel clinical presentation of a primary brain neoplasm in a 30-year-old man with a mass-like area in the anteromedial temporal lobe. Histopathological analysis revealed a low-grade neuroepithelial tumor with cytologically abnormal neurons and atypical glial cells within the cerebral cortex. Molecular analysis showed a previously undescribed FGFR2::DLG5 rearrangement. We discuss the clinical significance and molecular implications of this fusion event, shedding light on its potential impact on tumor development and patient prognosis. Additionally, an extensive review places the finding in this case in the context of protein fusions in brain tumors in general and highlights their diverse manifestations, underlying molecular mechanisms, and therapeutic implications.
With the advancements in precision medicine, the demands on pathological diagnostics have increased, requiring standardized, quantitative, and integrated assessments of histomorphological and molecular pathological data. Great hopes are placed in artificial intelligence (AI) methods, which have demonstrated the ability to analyze complex clinical, histological, and molecular data for disease classification, biomarker quantification, and prognosis estimation. This paper provides an overview of the latest developments in pathology AI, discusses the limitations, particularly concerning the black box character of AI, and describes solutions to make decision processes more transparent using methods of so-called explainable AI (XAI).
Brain disorders represent a significant challenge in medical science due to the formidable blood–brain barrier (BBB), which severely limits the penetration of conventional therapeutics, hindering effective treatment strategies. This review delves into the innovative realm of biomimetic nanodelivery systems, including stem cell‐derived nanoghosts, tumor cell membrane‐coated nanoparticles, and erythrocyte membrane‐based carriers, highlighting their potential to circumvent the BBB's restrictions. By mimicking native cell properties, these nanocarriers emerge as a promising solution for enhancing drug delivery to the brain, offering a strategic advantage in overcoming the barrier's selective permeability. The unique benefits of leveraging cell membranes from various sources is evaluated and advanced technologies for fabricating cell membrane‐encapsulated nanoparticles capable of masquerading as endogenous cells are examined. This enables the targeted delivery of a broad spectrum of therapeutic agents, ranging from small molecule drugs to proteins, thereby providing an innovative approach to neurocare. Further, the review contrasts the capabilities and limitations of these biomimetic nanocarriers with traditional delivery methods, underlining their potential to enable targeted, sustained, and minimally invasive treatment modalities. This review is concluded with a perspective on the clinical translation of these biomimetic systems, underscoring their transformative impact on the therapeutic landscape for intractable brain diseases.
A novel methylation class, “neuroepithelial tumor, with PLAGL1 fusion” (NET-PLAGL1), has recently been described, based on epigenetic features, as a supratentorial pediatric brain tumor with recurrent histopathological features suggesting an ependymal differentiation. Because of the recent identification of this neoplastic entity, few histopathological, radiological and clinical data are available. Herein, we present a detailed series of nine cases of PLAGL1-fused supratentorial tumors, reclassified from a series of supratentorial ependymomas, non-ZFTA/non-YAP1 fusion-positive and subependymomas of the young. This study included extensive clinical, radiological, histopathological, ultrastructural, immunohistochemical, genetic and epigenetic (DNA methylation profiling) data for characterization. An important aim of this work was to evaluate the sensitivity and specificity of a novel fluorescent in situ hybridization (FISH) targeting the PLAGL1 gene. Using histopathology, immunohistochemistry and electron microscopy, we confirmed the ependymal differentiation of this new neoplastic entity. Indeed, the cases histopathologically presented as “mixed subependymomas-ependymomas” with well-circumscribed tumors exhibiting a diffuse immunoreactivity for GFAP, without expression of Olig2 or SOX10. Ultrastructurally, they also harbored features reminiscent of ependymal differentiation, such as cilia. Different gene partners were fused with PLAGL1: FOXO1, EWSR1 and for the first time MAML2. The PLAGL1 FISH presented a 100% sensitivity and specificity according to RNA sequencing and DNA methylation profiling results. This cohort of supratentorial PLAGL1-fused tumors highlights: 1/ the ependymal cell origin of this new neoplastic entity; 2/ benefit of looking for a PLAGL1 fusion in supratentorial cases of non-ZFTA/non-YAP1 ependymomas; and 3/ the usefulness of PLAGL1 FISH.
Cytomorphological features of NUT carcinoma include sheets or discrete nests of primitive, monotonous, round to oval shaped tumour cells with high N/C ratio and brisk mitotic figures. Abrupt squamous differentiation might be a diagnostic hint. More than 50% positivity of NUT immunohistochemistry staining is diagnostic. NUT carcinoma represents a poorly differentiated malignancy by extremely aggressive clinical course and poor prognosis. It frequently manifests in midline organs, notably in the mediastinum and lung. The rising preferences for utilizing the EBUS-FNA procedure in diagnosing thoracic and lung lesions stems from its high diagnostic yield. Hence, recognizing the cytomorphological features of NUT carcinoma is crucial for timely treatment and improved patient survival.
Anaplastic large cell lymphoma (ALCL) is a mature T-cell lymphoma that accounts for for 10–15% of childhood lymphomas. Despite the observation that more than 90% of pediatric cases harbor the anaplastic lymphoma kinase (ALK) rearrangement resulting in aberrant ALK kinase expression, there is significant clinical, morphologic, and biological heterogeneity. To gain insights into the genomic aberrations and molecular heterogeneity within ALK-positive ALCL(ALK+ ALCL), we analyzed 46 pediatric ALK+ ALCLs by whole-exome sequencing, RNA-sequencing, and DNA methylation profiling. Whole-exome sequencing found on average 25 SNV/Indel events per sample with recurring genetic events in regulators of DNA damage (TP53, MDM4), transcription (JUNB), and epigenetic regulators (TET1, KMT2B, KMT2A, KMT2C, KMT2E). Gene expression and methylation profiling consistently subclassified ALK+ ALCLs into two groups characterized by diferential ALK expression levels. The ALK-low group showed enrichment of pathways associated with immune response, cytokine signaling, and a hypermethylated predominant pattern compared to the ALK- high group, which had more frequent copy number changes, and was enriched with pathways associated with cell growth, proliferation, metabolic pathways, and. Taken together, these findings suggest that there is molecular heterogeneity within pediatric ALK+ALCL, predicting distinct biological mechanisms that may provide novel insights into disease pathogenesis and represent prognostic markers
Molecularly defined neoplasms are increasingly recognized, given the broader application and performance of molecular studies. These studies allow us to better characterize these neoplasms and learn about their pathogenesis. In the thorax, molecularly defined neoplasms include tumors such as NUT carcinoma, SMARCA4-deficient undifferentiated tumor (DUT), primary pulmonary myxoid sarcoma with EWSR1::CREB1 fusion, hyalinizing clear cell carcinoma, and SMARCB1-deficient neoplasms. Overall, these tumors are rare but are now more often recognized given more widely available immunostains such as NUT (NUT carcinoma), BRG1 (SMARCA4-DUT), and INI-1 (SMARCB1-deficient neoplasm). Furthermore, cytogenetic studies for EWSR1 to support a hyalinizing clear cell carcinoma or primary pulmonary myxoid sarcoma are, in general, easily accessible. This enables pathologists to recognize and diagnose these tumors. The diagnosis of these tumors is important for clinical management and treatment. For instance, clinical trials are available for patients with NUT carcinoma, SMARCA4-DUT, and SMACRB1-deficient neoplasms. Herein, our current knowledge of clinical, morphologic, immunophenotypic, and molecular features of NUT carcinomas, SMARCA4-DUT, primary pulmonary myxoid sarcomas, hyalinizing clear cell carcinoma, and SMARCB1-deficient neoplasms will be reviewed.
Atypical teratoid/rhabdoid tumors (AT/RTs) are pediatric brain tumors known for their aggressiveness and aberrant but still unresolved epigenetic regulation. To better understand their malignancy, we investigated how AT/RT-specific DNA hypermethylation was associated with gene expression and altered transcription factor binding and how it is linked to upstream regulation. Medulloblastomas, choroid plexus tumors, pluripotent stem cells, and fetal brain were used as references. A part of the genomic regions, which were hypermethylated in AT/RTs similarly as in pluripotent stem cells and demethylated in the fetal brain, were targeted by neural transcriptional regulators. AT/RT-unique DNA hypermethylation was associated with polycomb repressive complex 2 and linked to suppressed genes with a role in neural development and tumorigenesis. Activity of the several NEUROG/NEUROD pioneer factors, which are unable to bind to methylated DNA, was compromised via the suppressed expression or DNA hypermethylation of their target sites, which was also experimentally validated for NEUROD1 in medulloblastomas and AT/RT samples. These results highlight and characterize the role of DNA hypermethylation in AT/RT malignancy and halted neural cell differentiation.
In the 2021 WHO classification, this category covers a group of six relatively circumscribed neoplasms (more solid growth pattern) with glial differentiation: pilocytic astrocytoma, high-grade astrocytoma with piloid features, pleomorphic xanthoastrocytoma, subependymal giant cell astrocytoma, chordoid glioma, and MN1-altered astroblastoma. With a few exceptions (e.g., high-grade astrocytoma with piloid features), they have a natural history that is generally more favorable than the adult-type diffuse gliomas, being attempt of gross total resection the treatment of choice. This chapter reviews the clinicopathologic features and differential diagnosis of these entities as well as of their subtypes.
There is a distinct difference between the clinical and molecular characteristics of diffuse gliomas that primarily occur in adults and those that occur primarily in children (age < 15). Thus, in the 2021 WHO classification, pediatric-type and adult-type gliomas are separated. According to its different behavior, pediatric-type diffuse gliomas are subdivided into two different families: (1) pediatric-type diffuse low-grade gliomas (expected to have good prognosis) and (2) pediatric-type diffuse high-grade gliomas (expected to behave aggressively). Due to the fact that these two new families of tumor types feature diffuse growth in the brain but with sometimes overlapping and less specific histological features, precise classification requires molecular characterization and the integration of histopathological and molecular information (integrated diagnosis). This chapter reviews the clinicopathologic features and differential diagnosis of pediatric-type diffuse low-grade gliomas and pediatric-type diffuse high-grade gliomas.
Although there is significant histological variability, embryonal tumors are grouped together because they are, at least partially, composed of immature neuroepithelial cells resembling the cells that make up the most primitive or embryonal stages of the CNS: the germinal matrix stem cell population. They are tumors mainly, but not exclusively, occurring in children and adolescents (0–14 years) and together comprise 15–20% of all CNS tumors in this age group. Embryonal tumors are characterized by an aggressive behavior with a high frequency of local recurrence, spread through the CSF, and even extraneural metastasis (CNS WHO grade 4). In the 2021 WHO classification, embryonal tumors are classified according to a combination of histopathological and molecular features into two general categories: (1) medulloblastoma, with four molecularly defined groups and four histologically defined groups, and (2) other embryonal tumors, which include atypical teratoid/rhabdoid tumor (AT/RT), embryonal tumor with multilayered rosettes (ETMR), FOXR2-activated CNS neuroblastoma, and CNS tumor with BCOR internal tandem duplication. In addition, cribriform neuroepithelial tumor has been introduced as a provisional entity within this category, whereas the broad designation CNS embryonal tumor is included for embryonal tumors that defy a more specific diagnosis, e.g., that are NOS (diagnostic information necessary to assign a specific diagnosis is not available) or NEC (adequate testing does not reveal signature molecular aberrations). In this chapter, the clinicopathological features and differential diagnosis of these entities are reviewed.
CNS non-meningothelial mesenchymal tumors arise from craniospinal meninges, supporting tissues, vasculature or surrounding osseous structures, representing the homologous of neoplasms encountered far more frequently in the somatic soft tissues or bones. Thus, the 2021 WHO classification of CNS tumors has strived to align its terminology with their counterparts in the WHO Blue Book on Bone and Soft Tissue Tumors. Furthermore, three recently described entities have been included in the group of mesenchymal tumors of uncertain differentiation: CIC-rearranged sarcoma; primary intracranial sarcoma, DICER1-mutant; and intracranial mesenchymal tumor, FET::CREB fusion-positive (a provisional entity). CNS non-meningothelial mesenchymal tumors can occur in patients of any age and may arise anywhere along the neuraxis and its coverings, with tumors arising in meninges more common than ones originating within CNS parenchyma or in choroid plexus. Histologically, they present a broad gradient of malignancy ranging from benign neoplasms to highly malignant sarcomas. This chapter reviews the clinicopathological features and differential diagnosis of solitary fibrous tumor; hemangioblastoma; lipoma; rhabdomyosarcoma; Ewing sarcoma; CIC-rearranged sarcoma; primary intracranial sarcoma, DICER1-mutant; intracranial mesenchymal tumor, FET::CREB fusion-positive; chordoma; chondrosarcoma; mesenchymal chondrosarcoma; and osteosarcoma.
Published in 2021, the fifth edition of the World Health Organization (WHO) classification of tumors of the central nervous system (CNS) introduced new molecular criteria for tumor types that commonly occur in either pediatric or adult age groups. Adolescents and young adults (AYAs) are at the intersection of adult and pediatric care, and both pediatric-type and adult-type CNS tumors occur at that age. Mortality rates for AYAs with CNS tumors have increased by 0.6% per year for males and 1% per year for females from 2007 to 2016. To best serve patients, it is crucial that both pediatric and adult radiologists who interpret neuroimages are familiar with the various pediatric- and adult-type brain tumors and their typical imaging morphologic characteristics. Gliomas account for approximately 80% of all malignant CNS tumors in the AYA age group, with the most common types observed being diffuse astrocytic and glioneuronal tumors. Ependymomas and medulloblastomas also occur in the AYA population but are seen less frequently. Importantly, biologic behavior and progression of distinct molecular subgroups of brain tumors differ across ages. This review discusses newly added or revised gliomas in the fifth edition of the CNS WHO classification, as well as other CNS tumor types common in the AYA population.
A prompt and reliable molecular diagnosis for brain tumors has become crucial in precision medicine. While Comprehensive Genomic Profiling (CGP) has become feasible, there remains room for enhancement in brain tumor diagnosis due to the partial lack of essential genes and limitations in broad copy number analysis. In addition, the long turnaround time of commercially available CGPs poses an additional obstacle to the timely implementation of results in clinics. To address these challenges, we developed a CGP encompassing 113 genes, genome-wide copy number changes, and MGMT promoter methylation. Our CGP incorporates not only diagnostic genes but also supplementary genes valuable for research. Our CGP enables us to simultaneous identification of mutations, gene fusions, focal and broad copy number alterations, and MGMT promoter methylation status, with results delivered within a minimum of 4 days. Validation of our CGP, through comparisons with whole-genome sequencing, RNA sequencing, and pyrosequencing, has certified its accuracy and reliability. We applied our CGP for 23 consecutive cases of intracranial mass lesions, which demonstrated its efficacy in aiding diagnosis and prognostication. Our CGP offers a comprehensive and rapid molecular profiling for gliomas, which could potentially apply to clinical practices and research primarily in the field of brain tumors.
With the advancements in precision medicine, the demands on pathological diagnostics have increased, requiring standardized, quantitative, and integrated assessments of histomorphological and molecular pathological data. Great hopes are placed in artificial intelligence (AI) methods, which have demonstrated the ability to analyze complex clinical, histological, and molecular data for disease classification, biomarker quantification, and prognosis estimation. This paper provides an overview of the latest developments in pathology AI, discusses the limitations, particularly concerning the black box character of AI, and describes solutions to make decision processes more transparent using methods of so-called explainable AI (XAI).
This work was supported by the Wellcome Trust. Y.S.J is supported by a European Molecular Biology Organization long-term fellowship (LTF 1203_2012). J.M.C.T. is supported by Marie Curie Fellowship FP7 PEOPLE-2012-IEF (project number 328264). P.J.C. is a Wellcome Trust Senior Clinical Fellow. Support was provided to A.M.F. by the National Institute for Health Research (NIHR) UCLH Biomedical Research Centre. The ICGC Breast Cancer Consortium was supported by a grant from the European Union (BASIS) and the Wellcome Trust. The ICGC Prostate Cancer Consortium was funded by Cancer Research UK with a grant from the Dallaglio Foundation (grant number C5047/A14835). R.E. is supported by National Institute for Health Research support to the Biomedical Research Centre at The Institute of Cancer Research and Royal Marsden NHS Foundation Trust. We also thank the National Cancer Research Prostate Cancer Mechanisms of Progression and Treatment (PROMPT) collaborative (grant code G0500966/75466) which has funded tissue and urine collections in Cambridge. The authors also acknowledge financial support from the Department of Health via the National Institute for Health Research comprehensive Biomedical Research Centre award to Guy’s and St. Thomas’ NHS Foundation Trust and Breakthrough Breast Cancer Research (ICGC 08/09 and KCL) (A.T.).
The 2016 World Health Organization Classification of Tumors of the Central Nervous System is both a conceptual and practical advance over its 2007 predecessor. For the first time, the WHO classification of CNS tumors uses molecular parameters in addition to histology to define many tumor entities, thus formulating a concept for how CNS tumor diagnoses should be structured in the molecular era. As such, the 2016 CNS WHO presents major restructuring of the diffuse gliomas, medulloblastomas and other embryonal tumors, and incorporates new entities that are defined by both histology and molecular features, including glioblastoma, IDH-wildtype and glioblastoma, IDH-mutant; diffuse midline glioma, H3 K27M-mutant; RELA fusion-positive ependymoma; medulloblastoma, WNT-activated and medulloblastoma, SHH-activated; and embryonal tumour with multilayered rosettes, C19MC-altered. The 2016 edition has added newly recognized neoplasms, and has deleted some entities, variants and patterns that no longer have diagnostic and/or biological relevance. Other notable changes include the addition of brain invasion as a criterion for atypical meningioma and the introduction of a soft tissue-type grading system for the now combined entity of solitary fibrous tumor / hemangiopericytoma-a departure from the manner by which other CNS tumors are graded. Overall, it is hoped that the 2016 CNS WHO will facilitate clinical, experimental and epidemiological studies that will lead to improvements in the lives of patients with brain tumors.
Classification of pediatric tumors into biologically defined subtypes is challenging, and multifaceted approaches are needed. For this aim, we developed a diagnostic classifier based on DNA methylation profiles.
Methylation data generated by the Illumina Infinium HumanMethylation 450 BeadChip arrays were downloaded from the Gene Expression Omnibus (n = 472). Using the data, we built MethPed, which is a multiclass random forest algorithm, based on DNA methylation profiles from nine subgroups of pediatric brain tumors. DNA from 18 regional samples was used to validate MethPed. MethPed was additionally applied to a set of 28 publically available tumors with the heterogeneous diagnosis PNET. MethPed could successfully separate individual histology tumor types at a very high accuracy (κ = 0.98). Analysis of a regional cohort demonstrated the clinical benefit of MethPed, as confirmation of diagnosis of tumors with clear histology but also identified possible differential diagnoses in tumors with complicated and mixed type morphology.
We demonstrate the utility of methylation profiling of pediatric brain tumors and offer MethPed as an easy-to-use toolbox that allows researchers and clinical diagnosticians to test single samples as well as large cohorts for subclass prediction of pediatric brain tumors. This will immediately aid clinical practice and importantly increase our molecular knowledge of these tumors for further therapeutic development.
Clear cell sarcoma of the kidney (CCSK) is one of the major pediatric renal neoplasms, but its associated genetic abnormalities are largely unknown. We identified internal tandem duplications in the BCOR gene (BCL6 corepressor) affecting the C terminus in 100% (20/20) of CCSK tumors but in none (0/193) of the other pediatric renal tumors. CCSK tumors expressed only an aberrant BCOR allele, indicating a close correlation between BCOR aberration and CCSK tumorigenesis.
Mitochondrial genomes are separated from the nuclear genome for most of the cell cycle by the nuclear double membrane, intervening cytoplasm, and the mitochondrial double membrane. Despite these physical barriers, we show that somatically acquired mitochondrial-nuclear genome fusion sequences are present in cancer cells. Most occur in conjunction with intranuclear genomic rearrangements, and the features of the fusion fragments indicate that nonhomologous end joining and/or replication-dependent DNA double-strand break repair are the dominant mechanisms involved. Remarkably, mitochondrial-nuclear genome fusions occur at a similar rate per base pair of DNA as interchromosomal nuclear rearrangements, indicating the presence of a high frequency of contact between mitochondrial and nuclear DNA in some somatic cells. Transmission of mitochondrial DNA to the nuclear genome occurs in neoplastically transformed cells, but we do not exclude the possibility that some mitochondrial-nuclear DNA fusions observed in cancer occurred years earlier in normal somatic cells.
© 2015 Ju et al.; Published by Cold Spring Harbor Laboratory Press.
Major discoveries in the biology of nervous system tumors have raised the question of how non-histological data such as molecular information can be incorporated into the next World Health Organization (WHO) classification of central nervous system tumors. To address this question, a meeting of neuropathologists with expertise in molecular diagnosis was held in Haarlem, The Netherlands, under the sponsorship of the International Society for Neuropathology (ISN). Prior to the meeting, participants solicited input from clinical colleagues in diverse neuro-oncological specialties. The present “white paper” catalogues the recommendations of the meeting, at which a consensus was reached that incorporation of molecular information into the next WHO classification should follow a set of provided “ISN-Haarlem” guidelines. Salient recommendations include that: 1) diagnostic entities should be defined as narrowly as possible to optimize interobserver reproducibility, clinicopathological predictions and therapeutic planning; 2) diagnoses should be “layered” with histologic classification, WHO grade, and molecular information listed below an “integrated diagnosis”; 3) determinations should be made for each tumor entity as to whether molecular information is required, suggested or not needed for its definition; 4) some pediatric entities should be separated from their adult counterparts; 5) input for guiding decisions regarding tumor classification should be solicited from experts in complementary disciplines of neuro-oncology and 6) entity-specific molecular testing and reporting formats should be followed in diagnostic reports. It is hoped that these guidelines will facilitate the forthcoming update of the 4th Edition of the WHO classification of central nervous system tumors.
Amplification of the C19MC oncogenic miRNA cluster and high LIN28 expression has been linked to a distinctly aggressive group of cerebral CNS-PNETs (group 1 CNS-PNETs) arising in young children. In this study, we sought to evaluate the diagnostic specificity of C19MC and LIN28, and the clinical and biological spectra of C19MC amplified and/or LIN28+ CNS-PNETs. We interrogated 450 pediatric brain tumors using FISH and IHC analyses and demonstrate that C19MC alteration is restricted to a sub-group of CNS-PNETs with high LIN28 expression; however, LIN28 immunopositivity was not exclusive to CNS-PNETs but was also detected in a proportion of other malignant pediatric brain tumors including rhabdoid brain tumors and malignant gliomas. C19MC amplified/LIN28+ group 1 CNS-PNETs arose predominantly in children <4 years old; a majority arose in the cerebrum but 24 % (13/54) of tumors had extra-cerebral origins. Notably, group 1 CNS-PNETs encompassed several histologic classes including embryonal tumor with abundant neuropil and true rosettes (ETANTR), medulloepithelioma, ependymoblastoma and CNS-PNETs with variable differentiation. Strikingly, gene expression and methylation profiling analyses revealed a common molecular signature enriched for primitive neural features, high LIN28/LIN28B and DNMT3B expression for all group 1 CNS-PNETs regardless of location or tumor histology. Our collective findings suggest that current known histologic categories of CNS-PNETs which include ETANTRs, medulloepitheliomas, ependymoblastomas in various CNS locations, comprise a common molecular and diagnostic entity and identify inhibitors of the LIN28/let7/PI3K/mTOR axis and DNMT3B as promising therapeutics for this distinct histogenetic entity.
Electronic supplementary material
The online version of this article (doi:10.1007/s00401-014-1291-1) contains supplementary material, which is available to authorized users.
Members of the nuclear factor-kappaB (NF-kappaB) family of transcriptional regulators are central mediators of the cellular inflammatory response. Although constitutive NF-kappaB signalling is present in most human tumours, mutations in pathway members are rare, complicating efforts to understand and block aberrant NF-kappaB activity in cancer. Here we show that more than two-thirds of supratentorial ependymomas contain oncogenic fusions between RELA, the principal effector of canonical NF-kappaB signalling, and an uncharacterized gene, C11orf95. In each case, C11orf95-RELA fusions resulted from chromothripsis involving chromosome 11q13.1. C11orf95-RELA fusion proteins translocated spontaneously to the nucleus to activate NF-kappaB target genes, and rapidly transformed neural stem cells--the cell of origin of ependymoma--to form these tumours in mice. Our data identify a highly recurrent genetic alteration of RELA in human cancer, and the C11orf95-RELA fusion protein as a potential therapeutic target in supratentorial ependymoma.
Diffuse intrinsic pontine glioma (DIPG) is a fatal brain cancer that arises in the brainstem of children, with no effective treatment and near 100% fatality. The failure of most therapies can be attributed to the delicate location of these tumors and to the selection of therapies on the basis of assumptions that DIPGs are molecularly similar to adult disease. Recent studies have unraveled the unique genetic makeup of this brain cancer, with nearly 80% found to harbor a p.Lys27Met histone H3.3 or p.Lys27Met histone H3.1 alteration. However, DIPGs are still thought of as one disease, with limited understanding of the genetic drivers of these tumors. To understand what drives DIPGs, we integrated whole-genome sequencing with methylation, expression and copy number profiling, discovering that DIPGs comprise three molecularly distinct subgroups (H3-K27M, silent and MYCN) and uncovering a new recurrent activating mutation affecting the activin receptor gene ACVR1 in 20% of DIPGs. Mutations in ACVR1 were constitutively activating, leading to SMAD phosphorylation and increased expression of the downstream activin signaling targets ID1 and ID2. Our results highlight distinct molecular subgroups and novel therapeutic targets for this incurable pediatric cancer.
Three histological variants are known within the family of embryonal rosette-forming neuroepithelial brain tumors. These include embryonal tumor with abundant neuropil and true rosettes (ETANTR), ependymoblastoma (EBL), and medulloepithelioma (MEPL). In this study, we performed a comprehensive clinical, pathological, and molecular analysis of 97 cases of these rare brain neoplasms, including genome-wide DNA methylation and copy number profiling of 41 tumors. We identified uniform molecular signatures in all tumors irrespective of histological patterns, indicating that ETANTR, EBL, and MEPL comprise a single biological entity. As such, future WHO classification schemes should consider lumping these variants into a single diagnostic category, such as embryonal tumor with multilayered rosettes (ETMR). We recommend combined LIN28A immunohistochemistry and FISH analysis of the 19q13.42 locus for molecular diagnosis of this tumor category. Recognition of this distinct pediatric brain tumor entity based on the fact that the three histological variants are molecularly and clinically uniform will help to distinguish ETMR from other embryonal CNS tumors and to better understand the biology of these highly aggressive and therapy-resistant pediatric CNS malignancies, possibly leading to alternate treatment strategies.
Malignant peripheral nerve sheath tumors (MPNSTs) are sarcomas of Schwann cell lineage origin that occur sporadically or in association with the inherited syndrome neurofibromatosis type 1. To identify genetic drivers of MPNST development, we used the Sleeping Beauty (SB) transposon-based somatic mutagenesis system in mice with somatic loss of transformation-related protein p53 (Trp53) function and/or overexpression of human epidermal growth factor receptor (EGFR). Common insertion site (CIS) analysis of 269 neurofibromas and 106 MPNSTs identified 695 and 87 sites with a statistically significant number of recurrent transposon insertions, respectively. Comparison to human data sets identified new and known driver genes for MPNST formation at these sites. Pairwise co-occurrence analysis of CIS-associated genes identified many cooperating mutations that are enriched in Wnt/β-catenin, PI3K-AKT-mTOR and growth factor receptor signaling pathways. Lastly, we identified several new proto-oncogenes, including Foxr2 (encoding forkhead box R2), which we functionally validated as a proto-oncogene involved in MPNST maintenance.
Recent studies have identified a Lys 27-to-methionine (K27M) mutation at one allele of H3F3A, one of the two genes encoding histone H3 variant H3.3, in 60% of high-grade pediatric glioma cases. The median survival of this group of patients after diagnosis is ∼1 yr. Here we show that the levels of H3K27 di- and trimethylation (H3K27me2 and H3K27me3) are reduced globally in H3.3K27M patient samples due to the expression of the H3.3K27M mutant allele. Remarkably, we also observed that H3K27me3 and Ezh2 (the catalytic subunit of H3K27 methyltransferase) at chromatin are dramatically increased locally at hundreds of gene loci in H3.3K27M patient cells. Moreover, the gain of H3K27me3 and Ezh2 at gene promoters alters the expression of genes that are associated with various cancer pathways. These results indicate that H3.3K27M mutation reprograms epigenetic landscape and gene expression, which may drive tumorigenesis.
Embryonal tumor with multilayered rosettes (ETMR, previously known as ETANTR) is a highly aggressive embryonal CNS tumor, which almost exclusively affects infants and is associated with a dismal prognosis. Accurate diagnosis is of critical clinical importance because of its poor response to current treatment protocols and its distinct biology. Amplification of the miRNA cluster at 19q13.42 has been identified previously as a genetic hallmark for ETMR, but an immunohistochemistry-based assay for clinical routine diagnostics [such as INI-1 for atypical teratoid rhabdoid tumor (AT/RT)] is still lacking. In this study, we screened for an ETMR-specific marker using a gene-expression profiling dataset of more than 1,400 brain tumors and identified LIN28A as a highly specific marker for ETMR. The encoded protein binds small RNA and has been implicated in stem cell pluripotency, metabolism and tumorigenesis. Using an LIN28A specific antibody, we carried out immunohistochemical analysis of LIN28A in more than 800 childhood brain-tumor samples and confirmed its high specificity for ETMR. Strong LIN28A immunoexpression was found in all 37 ETMR samples tested, whereas focal reactivity was only present in a small (6/50) proportion of AT/RT samples. All other pediatric brain tumors were completely LIN28A-negative. In summary, we established LIN28A immunohistochemistry as a highly sensitive and specific, rapid, inexpensive diagnostic tool for routine pathological verification of ETMR.
Medulloblastoma is an aggressively growing tumour, arising in the cerebellum or medulla/brain stem. It is the most common malignant brain tumour in children, and shows tremendous biological and clinical heterogeneity. Despite recent treatment advances, approximately 40% of children experience tumour recurrence, and 30% will die from their disease. Those who survive often have a significantly reduced quality of life. Four tumour subgroups with distinct clinical, biological and genetic profiles are currently identified. WNT tumours, showing activated wingless pathway signalling, carry a favourable prognosis under current treatment regimens. SHH tumours show hedgehog pathway activation, and have an intermediate prognosis. Group 3 and 4 tumours are molecularly less well characterized, and also present the greatest clinical challenges. The full repertoire of genetic events driving this distinction, however, remains unclear. Here we describe an integrative deep-sequencing analysis of 125 tumour-normal pairs, conducted as part of the International Cancer Genome Consortium (ICGC) PedBrain Tumor Project. Tetraploidy was identified as a frequent early event in Group 3 and 4 tumours, and a positive correlation between patient age and mutation rate was observed. Several recurrent mutations were identified, both in known medulloblastoma-related genes (CTNNB1, PTCH1, MLL2, SMARCA4) and in genes not previously linked to this tumour (DDX3X, CTDNEP1, KDM6A, TBR1), often in subgroup-specific patterns. RNA sequencing confirmed these alterations, and revealed the expression of what are, to our knowledge, the first medulloblastoma fusion genes identified. Chromatin modifiers were frequently altered across all subgroups. These findings enhance our understanding of the genomic complexity and heterogeneity underlying medulloblastoma, and provide several potential targets for new therapeutics, especially for Group 3 and 4 patients.
Medulloblastomas are the most common malignant brain tumours in children. Identifying and understanding the genetic events that drive these tumours is critical for the development of more effective diagnostic, prognostic and therapeutic strategies. Recently, our group and others described distinct molecular subtypes of medulloblastoma on the basis of transcriptional and copy number profiles. Here we use whole-exome hybrid capture and deep sequencing to identify somatic mutations across the coding regions of 92 primary medulloblastoma/normal pairs. Overall, medulloblastomas have low mutation rates consistent with other paediatric tumours, with a median of 0.35 non-silent mutations per megabase. We identified twelve genes mutated at statistically significant frequencies, including previously known mutated genes in medulloblastoma such as CTNNB1, PTCH1, MLL2, SMARCA4 and TP53. Recurrent somatic mutations were newly identified in an RNA helicase gene, DDX3X, often concurrent with CTNNB1 mutations, and in the nuclear co-repressor (N-CoR) complex genes GPS2, BCOR and LDB1. We show that mutant DDX3X potentiates transactivation of a TCF promoter and enhances cell viability in combination with mutant, but not wild-type, β-catenin. Together, our study reveals the alteration of WNT, hedgehog, histone methyltransferase and now N-CoR pathways across medulloblastomas and within specific subtypes of this disease, and nominates the RNA helicase DDX3X as a component of pathogenic β-catenin signalling in medulloblastoma.
Sequence reads for paediatric glioblastoma multiforme (GBM) samples have been deposited in the European Genome Archive under the accession number EGAS00001000226. This has been corrected in the HTML and PDF versions online.
To identify somatic mutations in pediatric diffuse intrinsic pontine glioma (DIPG), we performed whole-genome sequencing of DNA from seven DIPGs and matched germline tissue and targeted sequencing of an additional 43 DIPGs and 36 non-brainstem pediatric glioblastomas (non-BS-PGs). We found that 78% of DIPGs and 22% of non-BS-PGs contained a mutation in H3F3A, encoding histone H3.3, or in the related HIST1H3B, encoding histone H3.1, that caused a p.Lys27Met amino acid substitution in each protein. An additional 14% of non-BS-PGs had somatic mutations in H3F3A causing a p.Gly34Arg alteration.
Glioblastoma multiforme (GBM) is a lethal brain tumour in adults and children. However, DNA copy number and gene expression signatures indicate differences between adult and paediatric cases. To explore the genetic events underlying this distinction, we sequenced the exomes of 48 paediatric GBM samples. Somatic mutations in the H3.3-ATRX-DAXX chromatin remodelling pathway were identified in 44% of tumours (21/48). Recurrent mutations in H3F3A, which encodes the replication-independent histone 3 variant H3.3, were observed in 31% of tumours, and led to amino acid substitutions at two critical positions within the histone tail (K27M, G34R/G34V) involved in key regulatory post-translational modifications. Mutations in ATRX (α-thalassaemia/mental retardation syndrome X-linked) and DAXX (death-domain associated protein), encoding two subunits of a chromatin remodelling complex required for H3.3 incorporation at pericentric heterochromatin and telomeres, were identified in 31% of samples overall, and in 100% of tumours harbouring a G34R or G34V H3.3 mutation. Somatic TP53 mutations were identified in 54% of all cases, and in 86% of samples with H3F3A and/or ATRX mutations. Screening of a large cohort of gliomas of various grades and histologies (n = 784) showed H3F3A mutations to be specific to GBM and highly prevalent in children and young adults. Furthermore, the presence of H3F3A/ATRX-DAXX/TP53 mutations was strongly associated with alternative lengthening of telomeres and specific gene expression profiles. This is, to our knowledge, the first report to highlight recurrent mutations in a regulatory histone in humans, and our data suggest that defects of the chromatin architecture underlie paediatric and young adult GBM pathogenesis.
Neuroblastoma tumors frequently show loss of heterozygosity of chromosome 11q with a shortest region of overlap in the 11q23 region. These deletions are thought to cause inactivation of tumor suppressor genes leading to haploinsufficiency. Alternatively, micro-deletions could lead to gene fusion products that are tumor driving. To identify such events we analyzed a series of neuroblastomas by comparative genomic hybridization and single-nucleotide polymorphism arrays and integrated these data with Affymetrix mRNA profiling data with the bioinformatic tool R2 (http://r2.amc.nl). We identified three neuroblastoma samples with small interstitial deletions at 11q23, upstream of the forkhead-box R1 transcription factor (FOXR1). Genes at the proximal side of the deletion were fused to FOXR1, resulting in fusion transcripts of MLL-FOXR1 and PAFAH1B2-FOXR1. FOXR1 expression has only been detected in early embryogenesis. Affymetrix microarray analysis showed high FOXR1 mRNA expression exclusively in the neuroblastomas with micro-deletions and rare cases of other tumor types, including osteosarcoma cell line HOS. RNAi silencing of FOXR1 strongly inhibited proliferation of HOS cells and triggered apoptosis. Expression profiling of these cells and reporter assays suggested that FOXR1 is a negative regulator of fork-head box factor-mediated transcription. The neural crest stem cell line JoMa1 proliferates in culture conditional to activity of a MYC-ER transgene. Over-expression of the wild-type FOXR1 could functionally replace MYC and drive proliferation of JoMa1. We conclude that FOXR1 is recurrently activated in neuroblastoma by intrachromosomal deletion/fusion events, resulting in overexpression of fusion transcripts. Forkhead-box transcription factors have not been previously implicated in neuroblastoma pathogenesis. Furthermore, this is the first identification of intrachromosomal fusion genes in neuroblastoma.
Functional interpretation of candidate gene lists is an essential task in modern biomedical research. Here, we present the
2011 update of g:Profiler (http://biit.cs.ut.ee/gprofiler/), a popular collection of web tools for functional analysis. g:GOSt and g:Cocoa combine comprehensive methods for interpreting
gene lists, ordered lists and list collections in the context of biomedical ontologies, pathways, transcription factor and
microRNA regulatory motifs and protein–protein interactions. Additional tools, namely the biomolecule ID mapping service (g:Convert),
gene expression similarity searcher (g:Sorter) and gene homology searcher (g:Orth) provide numerous ways for further analysis
and interpretation. In this update, we have implemented several features of interest to the community: (i) functional analysis
of single nucleotide polymorphisms and other DNA polymorphisms is supported by chromosomal queries; (ii) network analysis
identifies enriched protein–protein interaction modules in gene lists; (iii) functional analysis covers human disease genes;
and (iv) improved statistics and filtering provide more concise results. g:Profiler is a regularly updated resource that is
available for a wide range of species, including mammals, plants, fungi and insects.
A recent genomewide mutational analysis of glioblastomas (World Health Organization [WHO] grade IV glioma) revealed somatic mutations of the isocitrate dehydrogenase 1 gene (IDH1) in a fraction of such tumors, most frequently in tumors that were known to have evolved from lower-grade gliomas (secondary glioblastomas).
We determined the sequence of the IDH1 gene and the related IDH2 gene in 445 central nervous system (CNS) tumors and 494 non-CNS tumors. The enzymatic activity of the proteins that were produced from normal and mutant IDH1 and IDH2 genes was determined in cultured glioma cells that were transfected with these genes.
We identified mutations that affected amino acid 132 of IDH1 in more than 70% of WHO grade II and III astrocytomas and oligodendrogliomas and in glioblastomas that developed from these lower-grade lesions. Tumors without mutations in IDH1 often had mutations affecting the analogous amino acid (R172) of the IDH2 gene. Tumors with IDH1 or IDH2 mutations had distinctive genetic and clinical characteristics, and patients with such tumors had a better outcome than those with wild-type IDH genes. Each of four tested IDH1 and IDH2 mutations reduced the enzymatic activity of the encoded protein.
Mutations of NADP(+)-dependent isocitrate dehydrogenases encoded by IDH1 and IDH2 occur in a majority of several types of malignant gliomas.
The Tel gene (or ETV6) is the target of the translocation (12;22)(p13;q11) in myeloid leukemia. TEL is a member of the ETS family of transcription
factors and contains the pointed protein interaction (PNT) domain and an ETS DNA binding domain (DBD). By contrast to other
chimeric proteins that contain TEL's PNT domain, such as TEL–platelet-derived growth factor β receptor in t(5;12)(q33;p13),
MN1-TEL contains the DBD of TEL. The N-terminal MN1 moiety is rich in proline residues and contains two polyglutamine stretches,
suggesting that MN1-TEL may act as a deregulated transcription factor. We now show that MN1-TEL type I, unlike TEL and MN1,
transforms NIH 3T3 cells. The transforming potential depends on both N-terminal MN1 sequences and a functional TEL DBD. Furthermore,
we demonstrate that MN1 has transcription activity and that MN1-TEL acts as a chimeric transcription factor on the Moloney
sarcoma virus long terminal repeat and a synthetic promoter containing TEL binding sites. The transactivating capacity of
MN1-TEL depended on both the DBD of TEL and sequences in MN1. MN1-TEL contributes to leukemogenesis by a mechanism distinct
from that of other chimeric proteins containing TEL.
The translocation t(12;22)(p13;q11) creates an MN1-TEL fusion gene leading to acute myeloid leukemia. MN1 is a transcription coactivator of the retinoic acid and vitamin D receptors, and TEL (ETV6) is a member of the E26-transformation-specific family of transcription factors. In MN1-TEL, the transactivating domains of MN1 are combined with the DNA-binding domain of TEL. We show that MN1-TEL inhibits retinoic acid receptor (RAR)-mediated transcription, counteracts coactivators such as p160 and p300, and acts as a dominant-negative mutant of MN1. Compared to MN1, the same transactivation domains in MN1-TEL are poorly stimulated by p160, p300 or histone deacetylase inhibitors, indicating that the block of RAR-mediated transcription by MN1-TEL is caused by dysfunctional transactivation domains rather than by recruitment of corepressors. The mechanism leading to myeloid leukemia in t(12;22) thus differs from the translocations that involve RAR itself.
The fourth edition of the World Health Organization (WHO) classification of tumours of the central nervous system, published in 2007, lists several new entities, including angiocentric glioma, papillary glioneuronal tumour, rosette-forming glioneuronal tumour of the fourth ventricle, papillary tumour of the pineal region, pituicytoma and spindle cell oncocytoma of the adenohypophysis. Histological variants were added if there was evidence of a different age distribution, location, genetic profile or clinical behaviour; these included pilomyxoid astrocytoma, anaplastic medulloblastoma and medulloblastoma with extensive nodularity. The WHO grading scheme and the sections on genetic profiles were updated and the rhabdoid tumour predisposition syndrome was added to the list of familial tumour syndromes typically involving the nervous system. As in the previous, 2000 edition of the WHO 'Blue Book', the classification is accompanied by a concise commentary on clinico-pathological characteristics of each tumour type. The 2007 WHO classification is based on the consensus of an international Working Group of 25 pathologists and geneticists, as well as contributions from more than 70 international experts overall, and is presented as the standard for the definition of brain tumours to the clinical oncology and cancer research communities world-wide.
We report a previously uncharacterized α-helical module, the BEN domain, in diverse animal proteins such as BANP/SMAR1, NAC1
and the Drosophila mod(mdg4) isoform C, in the chordopoxvirus virosomal protein E5R and in several proteins of polydnaviruses. Contextual analysis
suggests that the BEN domain mediates protein–DNA and protein–protein interactions during chromatin organization and transcription.
The presence of BEN domains in a poxviral early virosomal protein and in polydnaviral proteins also suggests a possible role
for them in organization of viral DNA during replication or transcription.
Contact: aravind{at}ncbi.nlm.nih.gov
Supplementary information: Supplementary data for this study can also be accessed at http://www.ncbi.nlm.nih.gov/CBBresearch/Lakshmin/BEN/
Rapid improvements in sequencing and array-based platforms are resulting in a flood of diverse genome-wide data, including data from exome and whole-genome sequencing, epigenetic surveys, expression profiling of coding and noncoding RNAs, single nucleotide polymorphism (SNP) and copy number profiling, and functional assays. Analysis of these large, diverse data sets holds the promise of a more comprehensive understanding of the genome and its relation to human disease. Experienced and knowledgeable human review is an essential component of this process, complementing computational approaches. This calls for efficient and intuitive visualization tools able to scale to very large data sets and to flexibly integrate multiple data types, including clinical data. However, the sheer volume and scope of data pose a significant challenge to the development of such tools.
Ependymal tumors across age groups are currently classified and graded solely by histopathology. It is, however, commonly accepted that this classification scheme has limited clinical utility based on its lack of reproducibility in predicting patients' outcome. We aimed at establishing a uniform molecular classification using DNA methylation profiling. Nine molecular subgroups were identified in a large cohort of 500 tumors, 3 in each anatomical compartment of the CNS, spine, posterior fossa, supratentorial. Two supratentorial subgroups are characterized by prototypic fusion genes involving RELA and YAP1, respectively. Regarding clinical associations, the molecular classification proposed herein outperforms the current histopathological classification and thus might serve as a basis for the next World Health Organization classification of CNS tumors.
Copyright © 2015 Elsevier Inc. All rights reserved.
NUT midline carcinoma, a squamous cell carcinoma, is one of the most aggressive human cancers, and there is a desperate need for effective therapies for patients with this disease. Will the new bromodomain and extra terminal (BET) inhibitors prove to be one such treatment for this rare and enigmatic cancer?
Supratentorial PNETs (sPNET) are uncommon embryonal malignancies of the central nervous system whose prognosis has historically been poor. We evaluated the outcome and prognostic factors of children with sPNET treated prospectively on a Children's Oncology Group trial.
Following surgery, patients received craniospinal radiotherapy with concurrent carboplatin followed by six months of maintenance chemotherapy with cyclophosphamide and vincristine.
Five-year overall survival (OS) and progression-free survival (PFS) for all patients was 58 ± 7% and 48 ± 7%. For patients with pineoblastoma (n = 23), five-year OS and PFS was 81 ± 9% and 62 ± 11%. Extent of resection but not M-stage was prognostic. Five-year OS and PFS for 37 patients with non-pineal tumors (NPsPNET) was 44 ± 8% and 39 ± 8%, significantly worse than for PB (P = 0.055 and 0.009 respectively). Extent of resection and major radiotherapy deviations were prognostic. Five year OS was 59 +/- 11.4% for those undergoing complete resection versus 10.4 +/- 7% for those who did not (P = 0.017). Central pathologic review called 14 (38%) "classic" sPNET, 8 (22%) "undifferentiated" and 13 (35%) "malignant gliomas." There was no significant difference between the subgroups, although survival distributions approached significance when the combined "classic" and "undifferentiated" group was compared to the "malignant gliomas."
Carboplatin during RT followed by 6 months of non-intensive chemotherapy is a feasible treatment strategy for patients with sPNET. Aggressive surgical resection should be attempted if feasible. The classification of supratentorial small cell malignancies can be difficult. Pediatr Blood Cancer © 2015 Wiley Periodicals, Inc.
© 2015 Wiley Periodicals, Inc.
Malignant rhabdoid tumor (MRT) can occur in the kidney, central nervous system, or extracranial/extrarenal locations and is characterized by alterations in the SMARCB1 gene. The tumors occur in infants and young children and confer a poor prognosis requiring aggressive therapeutic interventions to improve the chances for survival. MRTs pose a diagnostic challenge, as they display heterogeneous histopathologic features and differentiate along multiple lineages. The identification of alterations in the SMARCB1 gene in MRT using immunohistochemical (IHC) staining has lead to improved diagnosis of MRT as well as the discovery of the loss of SMARCB1 expression in some non-MRTs. Whether loss of SMARCB1 plays a pathogenic role in nonrhabdoid tumors remains to be determined; however, most of these tumors lack the clinical and other molecular features of MRT. We review the histopathologic features of MRT and the importance and significance of loss of expression of SMARCB1 in both MRT and nonrhabdoid tumors.
Round cell sarcomas harboring CIC-DUX4 fusions have recently been described as highly aggressive soft tissue tumors of children and young adults. Due to partial morphologic and immunohistochemical overlap with Ewing sarcoma (ES), CIC-DUX4-positive tumors have generally been classified as ES-like and managed similarly; however, a systematic comparison at the molecular and immunohistochemical levels between these two groups has not yet been conducted. Based on an initial observation that CIC-DUX4-positive tumors show nuclear immunoreactivity for WT1 and ETS transcription factors, FLI1 and ERG, we performed a detailed immunohistochemical and molecular analysis including these markers, to further investigate the relationship between CIC-DUX4 tumors and ES. The study group included 21 CIC-DUX4-positive sarcomas and 20 EWSR1-rearranged ES. Immunohistochemically, CIC-DUX4 sarcomas showed membranous CD99 positivity in 18 (86%) cases, but only 5 (24%) with a diffuse pattern, while WT1 and FLI1 were strongly positive in all cases. ERG was positive in 18% of cases. All ES expressed CD99 and FLI1, while ERG positivity was only seen in EWSR1-ERG fusion positive ES. WT1 was negative in all ES. Expression profiling validated by q-PCR revealed a distinct gene signature associated with CIC-DUX4 fusion, with upregulation of ETS transcription factors (ETV4, ETV1, and ETV5) and WT1, among top overexpressed genes compared to ES, other sarcomas and normal tissue. In conclusion, the distinct gene signature and immunoprofile of CIC-DUX4 sarcomas suggest a distinct pathogenesis from ES. The consistent WT1 expression may provide a useful clue in the diagnosis in the context of round cell sarcomas negative for EWSR1 rearrangement. © 2014 Wiley Periodicals, Inc.
A simple classification system for central nervous system neoplasms occurring primarily in infancy and childhood and largely composed of undifferentiated neuroepithelial cells is proposed. Classification is based upon appearance of the tumor as determined by light microscopy, immunocytochemical techniques, and ultrastructural features without consideration for site of origin. This classification is based on the concept that neoplastic transformation of primitive neuroepithelial cells in subependymal zones at all levels of the central nervous system or pineal body may develop into tumors largely composed of similar cells. It therefore seems appropriate to call these neoplasms primitive neuroectodermal tumors and to use descriptive terms to indicate the direction of cellular differentiation, when it has occurred. Proposed terminology for the five subtypes of undifferentiated neuroepithelial round cell tumors is as follows: 1) Primitive neuroectodermal tumor, not otherwise specified (PNET, NOS), 2) PNET with glial differentiation, 3) PNET with ependymal differentiation, 4) PNET with neuronal differentiation, and 5) PNET with multi- or bipotential differentiation. If the tumor is located in the cerebellum, medulloblastoma may be added in parentheses; if in the pineal body, pineal parenchymal neoplasm may be added.
H3F3A mutations are seen in ∼30% of pediatric glioblastoma (GBMs) and involve either the lysine residue at position 27 (K27M) or glycine at position 34 (G34R/V). Sixteen genes encode histone H3, each variant differing in only a few amino acids. Therefore, how mutations in a single H3 gene contribute to carcinogenesis is unknown. H3F3A K27M mutations are predicted to alter methylation of H3K27. H3K27me3 is a repressive mark critical to stem cell maintenance and is mediated by EZH2, a member of the polycomb-group (PcG) family. We evaluated H3K27me3 and EZH2 expression using immunohistochemistry in 76 pediatric brain tumors. H3K27me3 was lowered/absent in tumor cells but preserved in endothelial cells and infiltrating lymphocytes in six out of 20 GBMs. H3K27me3 showed strong immunoreactivity in all other tumor subtypes. Sequencing of GBMs showed H3F3A K27M mutations in all six cases with lowered/absent H3K27me3. EZH2 expression was high in GBMs, but absent/focal in other tumors. However, no significant differences in EZH2 expression were observed between H3F3A K27M mutant and wild type GBMs, suggesting that EZH2 mediated trimethylation of H3K27 is inhibited in GBM harboring K27M mutations. Our results indicate that H3F3A K27M mutant GBMs show decreased H3K27me3 that may be of both diagnostic and biological relevance.
Atypical teratoid/rhabdoid tumor (AT/RT) is a rare malignant pediatric brain tumor characterized by genetic alterations affecting the SMARCB1 (hSNF5/INI1) locus in chromosome band 22q11.2. To identify potential additional genetic alterations, high-resolution genome-wide analysis was performed using a molecular inversion probe single-nucleotide polymorphism (MIP SNP) assay (Affymetrix OncoScan formalin-fixed paraffin-embedded express) on DNA isolated from 18 formalin-fixed paraffin-embedded archival samples. Alterations affecting the SMARCB1 locus could be demonstrated by MIP SNP in 15 out of 16 evaluable cases (94%). These comprised five tumors with homozygous deletions, six tumors with heterozygous deletions, and four tumors with copy number neutral loss of heterozygosity (LOH) involving chromosome band 22q11.2. Remarkably, MIB SNP analysis did not yield any further recurrent chromosomal gains, losses, or copy neutral LOH. On MIP SNP screening for somatic mutations, the presence of a SMARCB1 mutation (c.472C>T p.R158X) was confirmed, but no recurrent mutations of other cancer relevant genes could be identified. Results of fluorescence in situ hybridization, multiplex ligation-dependent probe amplification, and SMARCB1 sequencing were highly congruent with that of the MIP SNP assay. In conclusion, these data further suggest the absence of recurrent genomic alterations other than SMARCB1 in AT/RT. © 2012 Wiley Periodicals, Inc.
Childhood CNS primitive neuro-ectodermal brain tumours (PNETs) are very aggressive brain tumours for which the molecular features and best treatment approaches are unknown. We assessed a large cohort of these rare tumours to identify molecular markers to enhance clinical management of this disease.
We obtained 142 primary hemispheric CNS PNET samples from 20 institutions in nine countries and examined transcriptional profiles for a subset of 51 samples and copy number profiles for a subset of 77 samples. We used clustering, gene, and pathway enrichment analyses to identify tumour subgroups and group-specific molecular markers, and applied immunohistochemical and gene-expression analyses to validate and assess the clinical significance of the subgroup markers.
We identified three molecular subgroups of CNS PNETs that were distinguished by primitive neural (group 1), oligoneural (group 2), and mesenchymal lineage (group 3) gene-expression signatures with differential expression of cell-lineage markers LIN28 and OLIG2. Patients with group 1 tumours were most often female (male:female ratio 0·61 for group 1 vs 1·25 for group 2 and 1·63 for group 3; p=0·043 [group 1 vs groups 2 and 3]), youngest (median age at diagnosis 2·9 years [95% CI 2·4-5·2] for group 1 vs 7·9 years [6·0-9·7] for group 2 and 5·9 years [4·9-7·8] for group 3; p=0·005), and had poorest survival (median survival 0·8 years [95% CI 0·5-1·2] in group 1, 1·8 years [1·4-2·3] in group 2 and 4·3 years [0·8-7·8] in group 3; p=0·019). Patients with group 3 tumours had the highest incidence of metastases at diagnosis (no distant metastasis:metastasis ratio 0·90 for group 3 vs 2·80 for group 1 and 5·67 for group 2; p=0·037).
LIN28 and OLIG2 are promising diagnostic and prognostic molecular markers for CNS PNET that warrant further assessment in prospective clinical trials.
Canadian Institute of Health Research, Brainchild/SickKids Foundation, and the Samantha Dickson Brain Tumour Trust.
To the Editor:
Rapid improvements in sequencing and array-based platforms are resulting in a flood of diverse genome-wide data, including data from exome and whole-genome sequencing, epigenetic surveys, expression profiling of coding and noncoding RNAs, single nucleotide polymorphism (SNP) and copy number profiling, and functional assays. Analysis of these large, diverse data sets holds the promise of a more comprehensive understanding of the genome and its relation to human disease. Experienced and knowledgeable human review is an essential component of this process, complementing computational approaches. This calls for efficient and intuitive visualization tools able to scale to very large data sets and to flexibly integrate multiple data types, including clinical data. However, the sheer volume and scope of data pose a significant challenge to the development of such tools.
Ependymoblastoma (EBL) and embryonal tumor with abundant neuropil and true rosettes (ETANTR) are very aggressive embryonal neoplasms characterized by the presence of ependymoblastic multilayered rosettes typically occurring in children below 6 years of age. It has not been established whether these two tumors really comprise distinct entities. Earlier, using array-CGH, we identified a unique focal amplification at 19q13.42 in a case of ETANTR. In the present study, we investigated this locus by fluorescence in situ hybridization in 41 tumors, which had morphologically been diagnosed as EBL or ETANTR. Strikingly, FISH analysis revealed 19q13.42 amplifications in 37/40 samples (93%). Among tumors harboring the amplification, 19 samples were identified as ETANTR and 18 as EBL. The three remaining tumors showed a polysomy of chromosome 19. Analysis of recurrent/metastatic tumors (n = 7) showed that the proportion of nuclei carrying the amplification was increased (up to 80-100% of nuclei) in comparison to the corresponding primary tumors. In conclusion, we have identified a hallmark cytogenetic aberration occurring in virtually all embryonal brain tumors with ependymoblastic rosettes suggesting that ETANTR and EBL comprise a single biological entity. FISH analysis of the 19q13.42 locus is a very promising diagnostic tool to identify a subset of primitive neuroectodermal tumors with distinct morphology, biology, and clinical behavior.
Rhabdoid tumors of early infancy are highly aggressive with consequent poor prognosis. Most cases show inactivation of the SMARCB1 (also known as INI1 and hSNF5) tumor suppressor, a core member of the ATP-dependent SWI/SNF chromatin-remodeling complex. Familial cases, described as rhabdoid tumor predisposition syndrome (RTPS), have been linked to heterozygous SMARCB1 germline mutations. We identified inactivation of another member of the SWI/SNF chromatin-remodeling complex, its ATPase subunit SMARCA4 (also known as BRG1), due to a SMARCA4/BRG1 germline mutation and loss of heterozygosity by uniparental disomy in the tumor cells of two sisters with rhabdoid tumors lacking SMARCB1 mutations. SMARCA4 is thus a second member of the SWI/SNF complex involved in cancer predisposition. Its general involvement in other tumor entities remains to be established.
We discovered a high-level amplicon involving the chr19q13.41 microRNA (miRNA) cluster (C19MC) in 11/45 ( approximately 25%) primary CNS-PNET, which results in striking overexpression of miR-517c and 520g. Constitutive expression of miR-517c or 520g promotes in vitro and in vivo oncogenicity, modulates cell survival, and robustly enhances growth of untransformed human neural stem cells (hNSCs) in part by upregulating WNT pathway signaling and restricting differentiation of hNSCs. Remarkably, the C19MC amplicon, which is very rare in other brain tumors (1/263), identifies an aggressive subgroup of CNS-PNET with distinct gene-expression profiles, characteristic histology, and dismal survival. Our data implicate miR-517c and 520g as oncogenes and promising biological markers for CNS-PNET and provide important insights into oncogenic properties of the C19MC locus.
Heterozygous point mutations of isocitrate dehydrogenase (IDH)1 codon 132 are frequent in grade II and III gliomas. Recently, we reported an antibody specific for the IDH1R132H mutation. Here we investigate the capability of this antibody to differentiate wild type and mutated IDH1 protein in central nervous system (CNS) tumors by Western blot and immunohistochemistry. Results of protein analysis are correlated to sequencing data. In Western blot, anti-IDH1R132H mouse monoclonal antibody mIDH1R132H detected a specific band only in mutated tumors. Immunohistochemistry of 345 primary brain tumors demonstrated a strong cytoplasmic and weaker nuclear staining in 122 cases. Correlation with direct sequencing of 186 cases resulted in consensus of 177 cases. Genetic retesting of cases with conflicting findings resulted in a match of 186/186 cases, with all discrepancies resolving in favor of immunohistochemistry. Intriguing is the ability of mIDH1R132H to detect single infiltrating tumor cells. The very high frequency and the distribution of this mutation among specific brain tumor entities allow the highly sensitive and specific discrimination of various tumors by immunohistochemistry, such as anaplastic astrocytoma from primary glioblastoma or diffuse astrocytoma World Health Organization (WHO) grade II from pilocytic astrocytoma or ependymoma. Noteworthy is the discrimination of the infiltrating edge of tumors with IDH1 mutation from reactive gliosis.
A simple classification system for central nervous system neoplasms occurring primarily in infancy and childhood and largely composed of undifferentiated neuroepithelial cells is proposed. Classification is based upon appearance of the tumor as determined by light microscopy, immunocytochemical techniques, and ultrastructural features without consideration for site of origin. This classification is based on the concept that neoplastic transformation of primitive neuroepithelial cells in subependymal zones at all levels of the central nervous system or pineal body may develop into tumors largely composed of similar cells. It therefore seems appropriate to call these neoplasms primitive neuroectodermal tumors and to use descriptive terms to indicate the direction of cellular differentiation, when it has occurred. Proposed terminology for the five subtypes of undifferentiated neuroepithelial round cell tumors is as follows: 1) Primitive neuroectodermal tumor, not otherwise specified (PNET, NOS), 2) PNET with glial differentiation, 3) PNET with ependymal differentiation, 4) PNET with neuronal differentiation, and 5) PNET with multi- or bipotential differentiation. If the tumor is located in the cerebellum, medulloblastoma may be added in parentheses; if in the pineal body, pineal parenchymal neoplasm may be added.
Controversial issues relating to the pathobiology and classification of central nervous system primitive neuroectodermal tumors (PNETs) have plagued neuropathologists for more than 70 years. Hypotheses advanced in the mid-1920's have remained as fixed concepts in contemporary literature, largely consequent to repetitious support by a small number of neuropathologists despite a growing body of information discrediting these ideas from neuroembryologists, oncologists, neuroscien-tists and pathologists.
Attention has largely focused upon PNETs arising in the cerebellum (commonly known as medul-loblastomas [MBs]), because about 80% of central nervous system (CNS) PNETs originate in this site. It has been asserted that the 20% which do not are biologically different, although most individuals agree that the histological features of PNETs that occur in different sites throughout the CNS are indistinguishable from those growing in the cerebellum.
The historical aspects of this controversy are examined in the face of evidence that there is, in fact, a unique class of CNS tumors which should appropriately be regarded as primitive neuroectodermal in nature. Specifically, a number of different approaches to the problem have yielded data supporting this hypothesis. These approaches include the identification of patterns of expression among a variety of cellular antigens (demonstrated by the use of immunopathological techniques), molecular analyses of cell lines derived from these tumors, experimental production of PNETs and molecular genetic analyses.
Differences of opinion among surgeons, oncologists and radiotherapists are typically resolved by conducting cooperative studies of patients with these tumors who are diagnosed and treated at multiple centers.
We have encountered a series of seven unusual neuroblastic pediatric central nervous system (CNS) neoplasms with a unique constellation of histologic, immunohistochemical, and ultrastructural features. The tumors presented in five girls and two boys, ages 1 to 3 years. In six cases the lesions involved the frontoparietal region, in one case the tectal plate. The tumors consisted of small to medium-sized, round to oval, hyperchromatic cells with poorly defined cytoplasmic borders. Cells were found in clusters and cords set in a paucicellular fibrillar neuropil matrix. Distinctive, virtually anuclear regions of neuropil were scattered throughout the lesions. True rosettes with well-formed central lumens often filled with granular debris were present, along with perivascular pseudorosettes and occasional Homer-Wright rosettes. Mitoses and apoptosis were frequent, but large regions of confluent necrosis were absent. Immunohistochemically, the neuropil-like areas as well as the perinuclear cytoplasm of many embryonal tumor cells were positive for synaptophysin and neurofilament protein. Ultrastructurally, the tumor cells showed microtubule-containing neuronal processes, some with neurosecretory granules. While the lesions were largely glial fibrillary acidic protein (GFAP) negative, there was focal GFAP positivity consistent with divergent differentiation in one case. The clinical outcome was poor, with five patients dead from their disease 5 to 14 months after initial presentation and one patient with recurrent disease 7 months after resection and chemotherapy. The final patient is alive without recurrent disease 30 months after initial presentation. These lesions present distinctive histological features within the group of primitive neuroectodermal tumors.
Evaluation of histone 3
- S Venneti
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- L M Sullivan
- D Martinez
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