BACKGROUND: Diffuse midline glioma, H3 K27-altered (DMG) is a fatal tumour that arises in the midline structures of the brain. When located in the pons, it is more commonly referred to as diffuse intrinsic pontine glioma (DIPG). DMG/DIPG is usually diagnosed when children are < 10 years, and it has a median overall survival of < 12 months after diagnosis. Radiological imaging is still the gold standard for DIPG diagnosis while the use of biopsy procedures led to our knowledge on its biology, such as with the identification of the canonical histone H3K27M mutation. However, the need to improve survival encourages the development of non-invasive, fast and inexpensive assays on biofluids for optimizing molecular diagnoses in DMG/DIPG. Here, we propose a rapid, new, imaging and epigenetics-based approach to diagnose DMG/DIPG in the plasma of paediatric patients. METHODS: A total of 20 healthy children (mean age: 10.5 years) and 24 children diagnosed with DMG/DIPG (mean age: 8.5 years) were recruited. Individual histones (H2A, H2B, H3, H4, macroH2A1.1 and macroH2A1.2), histone dimers and nucleosomes were assayed in biofluids by means of a new advanced flow cytometry ImageStream(X)-adapted method. RESULTS: We report a significant increase in circulating histone dimers and tetramers (macroH2A1.1/H2B versus control: p value < 0.0001; macroH2A1.2/H2B versus control: p value < 0.0001; H2A/H2B versus control: p value < 0.0001; H3/H4 versus control: p value = 0.008; H2A/H2B/H3/H4 versus control: p value < 0.0001) and a significant downregulation of individual histones (H2B versus control: p value < 0.0001; H3 versus control: p value < 0.0001; H4 versus control: p value < 0.0001). Moreover, histones were also detectable in the cerebrospinal fluid (CSF) of patients with DMG/DIPG and in the supernatant of SF8628, OPBG-DIPG002 and OPBG-DIPG004 DMG/DIPG cell lines, with patterns mostly similar to each other, but distinct compared to blood plasma. CONCLUSIONS: In summary, we identified circulating histone signatures able to detect the presence of DMG/DIPG in biofluids of children, using a rapid and non-invasive ImageStream(X)-based imaging technology, which may improve diagnosis and benefit the patients.

• MeSH
• difuzní intrinsický pontinní gliom * genetika   diagnóza   krev   MeSH
• dítě MeSH
• gliom * genetika   diagnóza   krev   MeSH
• histony * genetika   krev   metabolismus   MeSH
• lidé MeSH
• mladiství MeSH
• mutace MeSH
• nádorové biomarkery krev   MeSH
• nádory mozkového kmene * genetika   diagnóza   krev   MeSH
• nukleozomy metabolismus   MeSH
• předškolní dítě MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• histony * MeSH
• nádorové biomarkery MeSH
• nukleozomy MeSH

Diffuse pediatric-type high-grade gliomas (pedHGG), H3- and IDH-wildtype, encompass three main DNA-methylation-based subtypes: pedHGG-MYCN, pedHGG-RTK1A/B/C, and pedHGG-RTK2A/B. Since their first description in 2017 tumors of pedHGG-RTK2A/B have not been comprehensively characterized and clinical correlates remain elusive. In a recent series of pedHGG with a Gliomatosis cerebri (GC) growth pattern, an increased incidence of pedHGG-RTK2A/B (n = 18) was observed. We added 14 epigenetically defined pedHGG-RTK2A/B tumors to this GC series and provided centrally reviewed radiological, histological, and molecular characterization. The final cohort of 32 pedHGG-RTK2A/B tumors consisted of 25 pedHGG-RTK2A (78%) and seven pedHGG-RTK2B (22%) cases. The median age was 11.6 years (range, 4-17) with a median overall survival of 16.0 months (range 10.9-28.2). Seven of 11 of the newly added cases with imaging available showed a GC phenotype at diagnosis or follow-up. PedHGG-RTK2B tumors exhibited frequent bithalamic involvement (6/7, 86%). Central neuropathology review confirmed a diffuse glial neoplasm in all tumors with prominent angiocentric features in both subclasses. Most tumors (24/27 with available data, 89%) diffusely expressed EGFR with focal angiocentric enhancement. PedHGG-RTK2A tumors lacked OLIG2 expression, whereas 43% (3/7) of pedHGG-RTK2B expressed this glial transcription factor. ATRX loss occurred in 3/6 pedHGG-RTK2B samples with available data (50%). DNA sequencing (pedHGG-RTK2A: n = 18, pedHGG-RTK2B: n = 5) found EGFR alterations (15/23, 65%; predominantly point mutations) in both subclasses. Mutations in BCOR (14/18, 78%), SETD2 (7/18, 39%), and the hTERT promoter (7/19, 37%) occurred exclusively in pedHGG-RTK2A tumors, while pedHGG-RTK2B tumors were enriched for TP53 alterations (4/5, 80%). In conclusion, pedHGG-RTK2A/B tumors are characterized by highly diffuse-infiltrating growth patterns and specific radiological and histo-molecular features. By comprehensively characterizing methylation-based tumors, the chance to develop specific and effective therapy concepts for these detrimental tumors increases.

• Klíčová slova
• Gliomatosis cerebri, Methylation, Pediatric high-grade glioma, RTK2A, RTK2B, Receptor tyrosine kinase, pedHGG,
• MeSH
• dítě MeSH
• fenotyp MeSH
• gliom * genetika   patologie   diagnostické zobrazování   MeSH
• lidé MeSH
• metylace DNA * MeSH
• mladiství MeSH
• nádory mozku * genetika   patologie   diagnostické zobrazování   MeSH
• neuroepitelové nádory * genetika   patologie   diagnostické zobrazování   MeSH
• předškolní dítě MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: The term gliomatosis cerebri (GC), a radiology-defined highly infiltrating diffuse glioma, has been abandoned since molecular GC-associated features could not be established. METHODS: We conducted a multinational retrospective study of 104 children and adolescents with GC providing comprehensive clinical and (epi-)genetic characterization. RESULTS: Median overall survival (OS) was 15.5 months (interquartile range, 10.9-27.7) with a 2-year survival rate of 28%. Histopathological grading correlated significantly with median OS: CNS WHO grade II: 47.8 months (25.2-55.7); grade III: 15.9 months (11.4-26.3); grade IV: 10.4 months (8.8-14.4). By DNA methylation profiling (n = 49), most tumors were classified as pediatric-type diffuse high-grade glioma (pedHGG), H3-/IDH-wild-type (n = 31/49, 63.3%) with enriched subclasses pedHGG_RTK2 (n = 19), pedHGG_A/B (n = 6), and pedHGG_MYCN (n = 5), but only one pedHGG_RTK1 case. Within the pedHGG, H3-/IDH-wild-type subgroup, recurrent alterations in EGFR (n = 10) and BCOR (n = 9) were identified. Additionally, we observed structural aberrations in chromosome 6 in 16/49 tumors (32.7%) across tumor types. In the pedHGG, H3-/IDH-wild-type subgroup TP53 alterations had a significant negative effect on OS. CONCLUSIONS: Contrary to previous studies, our representative pediatric GC study provides evidence that GC has a strong predilection to arise on the background of specific molecular features (especially pedHGG_RTK2, pedHGG_A/B, EGFR and BCOR mutations, chromosome 6 rearrangements).

• Klíčová slova
• H3-wild-type and IDH-wild-type, chromosome 6, gliomatosis cerebri, pedHGG_RTK2, pediatric-type glioma, pediatric-type high-grade glioma,
• MeSH
• dítě MeSH
• fenotyp MeSH
• gliom * genetika   patologie   MeSH
• kojenec MeSH
• lidé MeSH
• metylace DNA MeSH
• míra přežití MeSH
• mladiství MeSH
• mutace MeSH
• nádorové biomarkery genetika   MeSH
• nádory mozku * genetika   patologie   MeSH
• následné studie MeSH
• neuroepitelové nádory * patologie   genetika   MeSH
• předškolní dítě MeSH
• prognóza MeSH
• retrospektivní studie MeSH
• stupeň nádoru MeSH
• Check Tag
• dítě MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• Názvy látek
• nádorové biomarkery MeSH

Pediatric central nervous system (CNS) tumors represent the most common cause of cancer-related death in children aged 0-14 years. They differ from their adult counterparts, showing extensive clinical and molecular heterogeneity as well as a challenging histopathological spectrum that often impairs accurate diagnosis. Here, we use DNA methylation-based CNS tumor classification in combination with copy number, RNA-seq, and ChIP-seq analysis to characterize a newly identified CNS tumor type. In addition, we report histology, patient characteristics, and survival data in this tumor type. We describe a biologically distinct pediatric CNS tumor type (n = 31 cases) that is characterized by focal high-level amplification and resultant overexpression of either PLAGL1 or PLAGL2, and an absence of recurrent genetic alterations characteristic of other pediatric CNS tumor types. Both genes act as transcription factors for a regulatory subset of imprinted genes (IGs), components of the Wnt/β-Catenin pathway, and the potential drug targets RET and CYP2W1, which are also specifically overexpressed in this tumor type. A derived PLAGL-specific gene expression signature indicates dysregulation of imprinting control and differentiation/development. These tumors occurred throughout the neuroaxis including the cerebral hemispheres, cerebellum, and brainstem, and were predominantly composed of primitive embryonal-like cells lacking robust expression of markers of glial or neuronal differentiation (e.g., GFAP, OLIG2, and synaptophysin). Tumors with PLAGL1 amplification were typically diagnosed during adolescence (median age 10.5 years), whereas those with PLAGL2 amplification were diagnosed during early childhood (median age 2 years). The 10-year overall survival was 66% for PLAGL1-amplified tumors, 25% for PLAGL2-amplified tumors, 18% for male patients, and 82% for female patients. In summary, we describe a new type of biologically distinct CNS tumor characterized by PLAGL1/2 amplification that occurs predominantly in infants and toddlers (PLAGL2) or adolescents (PLAGL1) which we consider best classified as a CNS embryonal tumor and which is associated with intermediate survival. The cell of origin and optimal treatment strategies remain to be defined.

• Klíčová slova
• Molecular neuro-oncology, PLAGL1, PLAGL2, Pediatric cancer,
• MeSH
• dítě MeSH
• DNA vazebné proteiny genetika   metabolismus   MeSH
• kojenec MeSH
• lidé MeSH
• metylace DNA MeSH
• nádorové supresorové proteiny genetika   MeSH
• nádory centrálního nervového systému * genetika   MeSH
• předškolní dítě MeSH
• primitivní neuroektodermové nádory * genetika   MeSH
• proteiny buněčného cyklu genetika   MeSH
• proteiny vázající RNA genetika   MeSH
• signální dráha Wnt genetika   MeSH
• transkripční faktory genetika   metabolismus   MeSH
• Check Tag
• dítě MeSH
• kojenec MeSH
• lidé MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA vazebné proteiny MeSH
• nádorové supresorové proteiny MeSH
• PLAGL1 protein, human MeSH Prohlížeč
• PLAGL2 protein, human MeSH Prohlížeč
• proteiny buněčného cyklu MeSH
• proteiny vázající RNA MeSH
• transkripční faktory MeSH

BACKGROUND: Malignant astrocytic gliomas in children show a remarkable biological and clinical diversity. Small in-frame insertions or missense mutations in the epidermal growth factor receptor gene (EGFR) have recently been identified in a distinct subset of pediatric-type bithalamic gliomas with a unique DNA methylation pattern. METHODS: Here, we investigated an epigenetically homogeneous cohort of malignant gliomas (n = 58) distinct from other subtypes and enriched for pediatric cases and thalamic location, in comparison with this recently identified subtype of pediatric bithalamic gliomas. RESULTS: EGFR gene amplification was detected in 16/58 (27%) tumors, and missense mutations or small in-frame insertions in EGFR were found in 20/30 tumors with available sequencing data (67%; 5 of them co-occurring with EGFR amplification). Additionally, 8 of the 30 tumors (27%) harbored an H3.1 or H3.3 K27M mutation (6 of them with a concomitant EGFR alteration). All tumors tested showed loss of H3K27me3 staining, with evidence of overexpression of the EZH inhibitory protein (EZHIP) in the H3 wildtype cases. Although some tumors indeed showed a bithalamic growth pattern, a significant proportion of tumors occurred in the unilateral thalamus or in other (predominantly midline) locations. CONCLUSIONS: Our findings present a distinct molecular class of pediatric-type malignant gliomas largely overlapping with the recently reported bithalamic gliomas characterized by EGFR alteration, but additionally showing a broader spectrum of EGFR alterations and tumor localization. Global H3K27me3 loss in this group appears to be mediated by either H3 K27 mutation or EZHIP overexpression. EGFR inhibition may represent a potential therapeutic strategy in these highly aggressive gliomas.

• Klíčová slova
• EGFR mutation, (bi)thalamic, H3 K27M mutation, K27me3, pediatric-type high-grade glioma,
• MeSH
• dítě MeSH
• erbB receptory genetika   MeSH
• geny erbB-1 MeSH
• gliom * genetika   MeSH
• histony genetika   MeSH
• lidé MeSH
• metylace DNA MeSH
• mutace MeSH
• nádory mozku * genetika   MeSH
• thalamus MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• EGFR protein, human MeSH Prohlížeč
• erbB receptory MeSH
• histony MeSH