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Due to its peculiar histopathological findings, pleomorphic xanthoastrocytoma (PXA), a rare cerebral tumor of young adults with a slow growth and a good prognosis, resembles to the lytic phase of progressive multifocal leukoencephalopathy, a fatal neurodegenerative disease caused by JC polyomavirus (JCPyV). Therefore, the presence of JCPyV DNA was examined in an 11-year-old child with xanthoastrocytoma, WHO grade 3, by quantitative PCR (qPCR) and nested PCR (nPCR) using primers amplifying sequences encoding the N- and C-terminal region of large T antigen (LTAg), the non-coding control region (NCCR), and viral protein 1 (VP1) DNA. The expression of transcripts from LTAg and VP1 genes was also evaluated. In addition, viral microRNAs’ (miRNAs) expression was investigated. Cellular p53 was also searched at both DNA and RNA level. qPCR revealed the presence of JCPyV DNA with a mean value of 6.0 × 104 gEq/mL. nPCR gave a positive result for the 5ʹ region of the LTAg gene and the NCCR, whereas 3ʹ end LTAg and VP1 DNA sequences were not amplifiable. Only LTAg transcripts of 5ʹ end were found whereas VP1 gene transcript was undetectable. Although in most cases, either Mad-1 or Mad-4 NCCRs have been identified in association with JCPyV-positive human brain neoplasms, the archetype NCCR structure was observed in the patient’s sample. Neither viral miRNA miR-J1-5p nor p53 DNA and RNA were detected. Although the expression of LTAg supports the possible role of JCPyV in PXA, further studies are warranted to better understand whether the genesis of xanthoastrocytoma could depend on the transformation capacity of LTAg by Rb sequestration.

Objectives Cerebral hyaline protoplasmic astrocytopathy (HPA) is a clinicopathologic entity characterized by eosinophilic cytoplasmic inclusions within astrocytes. It has been observed in a subset of patients with early-onset epilepsy, brain malformations, and developmental delay. The exact association of this entity with epilepsy is still unknown. This report, with its review of the literature, aims to summarize HPA features to raise awareness regarding this entity. Methods We report on 2 HPA cases and critically review the literature. Results Approximately 42 cases of HPA have been reported, including the 2 cases presented here, consisting of 23 female and 19 male patients. Patient age ranged from 3 to 39 years. All patients had early-onset seizures (3-20 months of age), ranging from partial to generalized, that were refractory despite treatment with antiepileptic drugs. Postoperative follow-up intervals ranged from 2 to 93 months, and the clinical outcome was graded according to the Engel classification, showing variable results. Conclusions Clinicians should consider HPA in differential diagnosis in patients with intractable seizures, especially when they are associated with developmental delay and brain malformations. Increasing awareness of this entity among pathologists may promote better understanding of this condition as well as better diagnosis and treatment for these patients.

Background. First-line therapies for medulloblastoma(MBL) are obtaining higher survival-rates while decreasing late-effects, but treatment at relapse is not standardized. We report the experience with MBL re-irradiation(re-RT), its timing and outcome in different clinical settings and tumor groups. Methods. Patient’s staging/treatment at diagnosis, histotypes/molecular subgroups, relapse site/s, re-treatments outcome are reported. Results. Patients were 25, median age 11.4 years, 8 had metastases, three LCA histotype. According to 2016-2021 WHO-classification, 14 had SHH subgroup tumors(6 TP53 mutated,1 + MYC and 1 + NMYC amplification), 11 non-WNT/non-SHH (2 with MYC/MYCN amplification).Thirteen had received HART-CSI, 11 standard-CSI, one HFRT; all post-radiation chemotherapy(CT), 16 also pre-RT. Median time to relapse (local-LR in 9, distant-DR in 14, LR+DR in two) was 26 months. Fourteen patients were re-operated, in 5 excising single DR-sites, thereafter 3 received CT, two after re-RT; out of 11 not re-operated patients, 4 had re-RT as first treatment and 7 after CT. Re-RT was administered at median 32 months after first RT: focally in 20 cases, CSI in 5, never resulting in radionecrosis. Median post-relapse-PFS/after re-RT were 16.7/8.2 months, while overall survival-OS were 35.1/23.9 months, respectively. Metastatic status both at diagnosis/relapse negatively affected outcome and re-surgery was prognostically favorable.MYC,MYCN,P53 status and molecular subgroups, RT extension/fractionation, gender and age were not statistically prognostic; in the multivariable model, OSs were positively influenced by longer intervals before re-RT, re-surgery and not-SHH subgroups (P=0.019 from recurrence and 0.004 from second RT). Conclusions. Re-surgery+reRT can prolong survival; a substantial fraction of patients with worse outcome belongs to SHH-subgroup.

Atypical teratoid/rhabdoid tumor (AT/RT) is a highly aggressive embryonal CNS neoplasm, characterized by inactivation of SMARCB1 (INI1) or rarely of SMARCA4 (BRG1). While it is predominantly a childhood tumor, AT/RT is rare in adults.We provide a comprehensive systematic review of literature with meta-analysis; 92 adult cases were found from 74 articles. We additionally present 4 cases of adult AT/RTs (age ranging from 19 to 29 years), located to cerebellum in 2 cases, to ponto-cerebellar angle in 1 case and to spinal cord in the remaining case.Microscopic features of our 4 cases showed a highly cellular tumor with rhabdoid morphology and high mitotic activity. All tumor cells lacked nuclear SMARCB1/INI1 protein expression. In case no. 3 we also performed methylation profiling which clustered the tumor with pediatric AT/RT-MYC subgroup. Prognosis remains poor in both pediatric and adult population with a median overall survival of 11 months. Our review demonstrated median overall survival of 15 months among the adult populations. However, consistent with a recent review, adult AT/RT seems to have highly variable prognosis and some patients reach long term survival with 22.9% of 5-year survival without evidence of disease and mean follow up time of 35.9 months (SD = 36.5). 27.1% of dissemination was also reported among the adult population.Adult AT/RTs predominantly arise in female patients and in supratentorial location. Midline structures, including the sellar region, are the most affected sites, especially among females aged40 years. Male gender is more prevalent between the age of 18 and 40 years and more frequently associated with non-midline tumors. Factors significantly associated with better prognosis are patient's age ( 40 years), combined radio-chemotherapy adjuvant approach and Ki-67 score 40%.

Adult-type diffuse gliomas represent a group of highly infiltrative central nervous system tumors with a prognosis that significantly varies depending on the specific subtype and histological grade. Traditionally, adult-type diffuse gliomas have been classified based on their morphological features with a great interobserver variability and discrepancy in patient survival even within the same histological grade. Over the last few decades, advances in molecular profiling have drastically changed the diagnostic approach and classification of brain tumors leading to the development of an integrated morphological and molecular classification endowed with a more clinically relevant value. These concepts were largely anticipated in the revised fourth-edition of WHO classification of central nervous system tumors published in 2016. The fifth-edition (WHO 2021) moved molecular diagnostics forward into a full integration of molecular parameters with the histological features into an integrative diagnostic approach. Diagnosis of adult type diffuse gliomas, IDH mutant and IDH-wildtype has been simplified by introducing revised diagnostic and grading criteria. In this review, we will discuss the most recent updates to the classification of adult-type diffuse gliomas and summarize the essential diagnostic keys providing a practical guidance to pathologists.

Recurrence incidence for paediatric/adolescent high-grade glioma (HGG) exceeds 80%. Reirradiation (reRT) palliates symptoms and delays further progression. Strategies for reRT are scarce: we retrospectively analysed our series to develop rational future approaches.We re-evaluated MRI + RT plans of 21 relapsed HGG-patients, accrued 2010-2021, aged under 18 years. All underwent surgery and RT + chemotherapy at diagnosis. Pathologic/molecular re-evaluation allowed classification based on WHO 2021 criteria in 20/21 patients. Survival analyses and association with clinical parameters were performed.Relapse after 1st RT was local in 12 (7 marginal), 4 disseminated, 5 local + disseminated. Re-RT obtained 8 SD, 1 PR, 1PsPD, 1 mixed response, 10 PD; neurological signs/symptoms improved in 8. Local reRT was given to 12, followed again by 6 local (2 marginal) and 4 local + disseminated second relapses in 10/12 re-evaluated. The 4 with dissemination had 1 whole brain, 2 craniospinal irradiation (CSI), 1 spine reRT and further relapsed with dissemination and local + dissemination in 3/four assessed. Five local + disseminated tumours had 3 CSI, 1 spine reRT, further progressing locally (2), disseminated (1), n.a. (1). Three had a third RT; three were alive at 19.4, 29, 50.3 months after diagnosis. Median times to progression/survival after re-RT were 3.7 months (0.6-16.2 months)/6.9 months (0.6-17.9 months), improved for longer interval between 1st RT and re-RT (P = 0.017) and for non-PD after reRT (P 0.001). First marginal relapse showed potential association with dissemination after re-RT (P = 0.081).This is the biggest series of re-RT in paediatric HGG. Considering the dissemination observed at relapse, our results could prompt the investigation of different first RT fields in a randomized trial.

Background: Brain metastases (BMs) are the most common intracranial tumors in adults and the brain is one of the most frequent metastatic sites for non-small-cell lung cancer (NSCLC). Some studies have hypothesized that EGFR expression in the primary tumor may result in clinically radiological and prognostic changes in related brain metastases, comparable to EGFR's correlation with prognosis in primary brain tumors. There are no current studies that clinically and radiologically demonstrate a difference between BMs from EGFR-mutated and wild-type NSCLC.Methods: We performed a retrospective study to identify prognostic factors for the survival of patients with NSCLC-BMs by exploring the role of driving mutations in NSCLC, focusing on EGFR mutated status, and comparing all morphological, radiological, and clinical features of NSCLC-BMs with their outcome parameters in a cohort of surgically treated patients. Results: The final cohort consisted of 81 patients. The overall survival of the cohort was 15±17 months. The tumor-related edema was associated with neurological symptoms at the clinical onset of the disease (p=.048). We found that EGFR and ALK mutation status did not reach significant associations with age, sex, and the morphology of the lesions. EGFR mutation in the primary tumor is positively associated with higher edema and tumor volume (respectively 22.38±21.35cm3 versus 7.68±8.44cm3 and 72.44±60.71cm3 versus 31.92cm3 p=.046 and p=.028). Moreover, EGFR mutation is associated with a clinical debut of seizures (p=.004).Conclusions: The role of EGFR and ALK mutations of NSCLC on prognostic characteristics of BMs is still to be fully clarified; however the results retrieved from the present study suggest that the presence of EGFR mutations significantly correlates with bigger edema volume and higher incidence of seizures. In recent studies, while EGFR mutation status did not affect the immune pathway scores of primary lung tumors, the overall immune pathway scores in related BMs indicate a peculiar immunogenic phenotype that could explain a large amount of edema volume. This study could be considered the first clinical and radiological demonstration of this immunological phenotype.

Background A prospective 2002–2014 study stratified 160 patients by resection extent and histological grade, reporting results in 2016. We re-analyzed the series after a median of 119 months, adding retrospectively patients’ molecular features. Methods Follow-up of all patients was updated. DNA copy number analysis and gene-fusion detection could be completed for 94/160 patients, methylation classification for 68. Results Progression-free survival (PFS) and overall survival (OS) at 5/10 years were 66/58%, and 80/73%. Ten patients had late relapses (range 66–126 mo), surviving after relapse no longer than those relapsing earlier (0–5 y). On multivariable analysis a better PFS was associated with grade II tumor and complete surgery at diagnosis and/or at radiotherapy; female sex and complete resection showed a positive association with OS. Posterior fossa (PF) tumors scoring ≥0.80 on DNA methylation analysis were classified as PFA (n = 41) and PFB (n = 9). PFB patients had better PFS and OS. Eighteen/32 supratentorial tumors were classified as RELA, and 3 as other molecular entities (anaplastic PXA, LGG MYB, HGNET). RELA had no prognostic impact. Patients with 1q gain or cyclin-dependent kinase inhibitor 2A (CDKN2A) loss had worse outcomes, included significantly more patients >3 years old (P = 0.050) and cases of dissemination at relapse (P = 0.007). Conclusions Previously described prognostic factors were confirmed at 10-year follow-up. Late relapses occurred in 6.2% of patients. Specific molecular features may affect outcome: PFB patients had a very good prognosis; 1q gain and CDKN2A loss were associated with dissemination. To draw reliable conclusions, modern ependymoma trials need to combine diagnostics with molecular risk stratification and long-term follow-up.

Astroblastomas are neuroepithelial tumours defined by the presence of MN1 rearrangement and are typically located in the cerebral hemispheres. Rare cases of astroblastoma-like tumours carrying an EWSR1-BEND2 fusion have been recently described in the brain stem and spinal cord. We report a paediatric case of neuroepithelial astroblastoma-like tumour occurring in the spine and carrying a novel MAMLD1-BEND2 fusion. We believe that our case aligns with the rare astroblastoma-like tumours with EWSR1-BEND2 fusion, in terms of non-hemispheric location, pathology, methylation profile and activation of BEND2 transcription. Whether they may represent a distinct entity or a variant of MN1-altered astroblastoma is not clear.

Background More than 40% of patients with intracranial ependymoma need a salvage treatment within 5 years after diagnosis, and no standard treatment is available as yet. We report the outcome after first relapse of 64 patients treated within the 2nd AIEOP protocol. Methods We considered relapse sites and treatments, that is, various combinations of complete/incomplete surgery, if followed by standard or hypofractionated radiotherapy (RT) ± chemotherapy (CT). Molecular analyses were available for 38/64 samples obtained at first diagnosis. Of the 64 cases, 55 were suitable for subsequent analyses. Results The median follow-up was 147 months after diagnosis, 84 months after first relapse, 5-year EFS/OS were 26.2%/30.8% (median EFS/OS 13/32 months) after relapse. For patients with a local relapse (LR), the 5-year cumulative incidence of second LRs was 51.6%, with a 5-year event-specific probability of being LR-free of 40.0%. Tumor site/grade, need for shunting, age above/below 3 years, molecular subgroup at diagnosis, had no influence on outcomes. Due to variation in the RT dose/fractionation used and the subgroup sizes, it was not possible to assess the impact of the different RT modalities. Multivariable analyses identified completion of surgery, the absence of symptoms at relapse, and female sex as prognostically favorable. Tumors with a 1q gain carried a higher cumulative incidence of dissemination after first relapse. Conclusions Survival after recurrence was significantly influenced by symptoms and completeness of surgery. Only a homogeneous protocol with well-posed, randomized questions could clarify the numerous issues, orient salvage treatment, and ameliorate prognosis for this group of patients.

Astroblastoma is a rare glial neoplasm arising more frequently in young, predominantly female, patients and with unclear clinical behavior and outcome. The diagnostic molecular alteration is the rearrangement of meningioma 1 (MN1) gene.However, little is known about the specific mechanism of tumor development driven by such genetic change. microRNAs (miRNAs) are important gene expression regulators with strong implications in several biological processes. In this study we investigated the microRNAs’ expression and regulation in MN1 altered neoplasms. We collected a cohort of 14 formalin-fixed, paraffin-embedded (FFPE) tumor samples histologically defined classified as astroblastoma. The DNA methylation analysis showed that only 8 cases harbored the MN1 rearrangement characteristic of astroblastoma. The 8 MN1 altered tumors were analyzed for their expression pattern of miRNAs by Nanostring technology. Thirty-nine deregulated miRNAs were found in the 8 astroblastomas compared to normal brain tissue. In order to understand the underlying mechanisms of the miRNAs aberrant expression, we first investigated the methylation status of themicroRNA promoters. Thirty-two out 39 deregulated miRNA resulted epigenetically regulated. with methylation status coherent with microRNA expression in 14/32 miRNAs.. Secondly, we investigated the hypothesis of a genomic alteration as a reason for the abnormal expression of the remaining 18/32 deregulated miRNAs by analyzing the Copy Number Variation (CNV) of tumor samples. but no alteration was found on miRNAs chromosome loci. Finally, we identified validated targets of the 32 deregulated miRNAs and uncovered biological processes putatively correlated to miRNA target genes, clinically and pathologically relevant in MN1-altered astroblastomas. Our findings shed light on the biology of this rare disease with potential implications on prognostic markers and therapy.

Cerebellar liponeurocytoma (CL) is an unusual tumor, histologically composed of a mixture of small to medium-sized, rounded neurocytic cells and a variable lipomatous component. Although CL was originally considered as a subtype of medulloblastoma, subsequent molecular studies demonstrated that this tumor was a distinct entity, exhibiting the tumor protein p53 gene (TP53) missense mutations in 20% of cases, chromosome 17 deletion, and the absence of mutations in the adenomatous polyposis coli gene (APC), the protein patched homolog gene (PTCH), the kinase insert domain receptor gene (KDR), and the β-catenin gene (CTNNB). Apart from these molecular features, little is known about the pathogenesis and the genetic landscape of CL to date. In order to characterize the mutational landscape of CL and identify alterations that are driving tumorigenesis, we report a series of three cases, including one recurrent tumor, analysed by next-generation sequencing (NGS), which identified a total of 22 variants, of which four were missense mutations, nine were synonymous variants, and nine were located on intronic regions. In particular, DNA sequencing identified missense mutations in APC, KDR, and TP53 that could be implicated in promoting tumor progression and angiogenesis of CL. Furthermore, the NGS analysis revealed that recurrent CL did not have additional genetic changes compared with the primary tumor. Moreover, the high frequencies of detected mutations suggested that the identified alterations are germline variants. Indeed, an additional NGS on the genomic DNA obtained from one of the three patients confirmed the presence of the variants in the germline DNA. In conclusion, the obtained data support the hypothesis that CL is a distinct pathological entity that does not show specific somatic alterations driving tumorigenesis.

Neoplastic cells acquire the ability to proliferate endlessly by maintaining telomeres via telomerase, or alternative lengthening of telomeres (ALT). The role of telomere maintenance in pituitary neuroendocrine tumors (PitNETs) has yet to be thoroughly investigated. We analyzed surgical samples of 24 adult recurrent PitNETs (including onset and relapses for 14 of them) and 12 pediatric primary PitNETs. The presence of ALT was assessed using telomere-specific fluorescence in situ hybridization, methylation of telomerase reverse transcriptase promoter (TERTp) by methylation-specific PCR, and ATRX expression by immunohistochemistry. Among the adult recurrent PitNETs, we identified 3/24 (12.5%) ALT-positive cases. ALT was present from the onset and maintained in subsequent relapses, suggesting that this mechanism occurs early in tumorigenesis and is stable during progression. ATRX loss was only seen in one ALT-positive case. Noteworthy, ALT was observed in 3 out of 5 aggressive PitNETs, including two aggressive corticotroph tumors, eventually leading to patient's death. ALT-negative tumors (87.5%) were classified according to their low (29.2%), medium (50%), and high (8.3%) telomere fluorescence intensity, with no significant differences emerging in their molecular, clinical, or pathological characteristics. TERTp methylation was found in 6/24 cases (25%), with a total concordance in methylation status between onset and recurrences, suggesting that this mechanism remains stable throughout disease progression. TERTp methylation did not influence telomere length. In the pediatric cohort of PitNETs, TERTp methylation was also observed in 4/12 cases (33.3%), but no case of ALT activation was observed. In conclusion, ALT is triggered at onset and maintained during tumor progression in a subset of adult PitNETs, suggesting that it could be used for clinical purposes, as a potential predictor of aggressive behavior.

As a relevant element of novelty, the fifth CNS WHO Classification highlights the distinctive pathobiology underlying gliomas arising primarily in children by recognizing for the first time the families of paediatric-type diffuse gliomas, both high-grade and low-grade. This review will focus on the family of paediatric-type diffuse high-grade gliomas, which includes four tumour types: 1) Diffuse midline glioma H3 K27-altered; 2) Diffuse hemispheric glioma H3 G34-mutant; 3) Diffuse paediatric-type high-grade glioma H3-wildtype and IDH-wildtype; and 4) Infant-type hemispheric glioma. The essential and desirable diagnostic criteria as well as the entities entering in the differential will be discussed for each tumour type. A special focus will be given on the issues encountered in the daily practice, especially regarding the diagnosis of the diffuse paediatric-type high-grade glioma H3-wildtype and IDH-wildtype. The advantages and the limits of the multiple molecular tests which may be utilised to define the entities of this tumour family will be evaluated in each diagnostic context.

BACKGROUND: The purposes of this trial were to evaluate the feasibility, response, PFS/OS of a randomized study comparing two different RT schedules for DIPG while administering the same systemic treatment. METHODS: Patients: 2-21 years-old with a not-pretreated radiologically verified DIPG (MRI blindly reviewed at diagnosis and every 12 weeks thereafter) and symptoms duration below 6 months. Biopsy was required if suggested by atypical imaging. Vinorelbine 20 mg/m2+nimotuzumab 150 mg/m2 were administered weekly for 12 weeks; thereafter every other week until tumor progression or for up to 2 years. Standard(ST) arm included focal RT at total dose of 54Gy (1.8Gy/day); for local progression re-irradiation was proposed at 19.8Gy, in case of dissemination craniospinal irradiation(CSI) at 36Gy was adopted. Experimental(SP) arm included three elective courses of RT at defined timepoints at 36Gy, 19.8Gy and 19.8Gy with possible reirradiation for relapse at 9 Gy. Incidences of local(L) and distant(D) progression were assessed in a competing risk setting. RESULTS: Aggregated preliminary results are given for 4 Italian centers. 54 pts were screened and 51 included, 27 in ST, 28 males, median age 7 years (range 3-17). Median time of observation was 17.9 months. Twelve patients needed a shunt, 10 during treatment; 20 were biopsied, in 18 cases according local protocols. 19/20 tumors had H3.3 K27 mutation. 41 relapsed, 28 locally, 13 with a component of dissemination. 36 died, one for tracheotomy bleeding. SP irradiation was feasible and never produced significant radionecrosis. Median EFS/OS were 7.3/12.9 months, respectively; EFS/OS at 1 year were 19.0%/57.3%, not differing between patients with local vs. disseminated relapses. Patients submitted to biopsies had more dissemination (P=0.04) and less local progression (P=0.077). CONCLUSIONS: Treatment was feasible and OS confirmed previous results obtained in a single center. Randomization results will be later reported.

BACKGROUND: Gliomatosis cerebri (GC), a radiologically defined diffusely infiltrating glioma, is no longer considered a distinct entity since the 2016 WHO classification for tumors of the CNS. Due to its rarity and dismal prognosis treatment recommendations in children remain ambiguous. Using central neuroradiological review, we performed a multi-institutional, retrospective study of GC providing comprehensive radiological, clinical, and (epi)genetic characterization. RESULTS: We included 104 patients between 1-19 years. Within a median follow-up of 15.5 months (range, 2.3-138.8), 93 patients (89.4 %) had died, 4 (3.8 %) were lost to follow-up and 7 (6.8 %) were alive with stable/progressive disease. Median progression-free- (PFS) and overall survival (OS) were 8.6 months (interquartile range, 4.3-14.0) and 15.5 months (10.9-27.7), respectively. Former WHO grading correlated significantly with median OS: WHO °II: 47.8 months (25.2-55.7); WHO °III: 15.9 months (11.4-26.3); WHO °IV: 10.4 months (8.8-14.4) (p

Background There is a paucity of knowledge regarding pediatric biomarkers, including the relevance of ErbB pathway aberrations in pediatric tumors. We investigated the occurrence of ErbB receptor aberrations across different pediatric malignancies, to identify patterns of ErbB dysregulation and define biomarkers suitable for patient enrichment in clinical studies. Procedure Tissue samples from 297 patients with nervous system tumors and rhabdomyosarcoma were analyzed for immunohistochemical expression or gene amplification of epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2). Exploratory analyses of HER3/HER4 expression, and mRNA expression of ErbB receptors/ligands (NanoString) were performed. Assay validation followed general procedures, with additional validation to address Clinical Laboratory Improvement Amendments (CLIA) requirements. Results In most tumor types, samples with high ErbB receptor expression were found with heterogeneous distribution. We considered increased/aberrant ErbB pathway activation when greater than or equal to two EGFR/HER2 markers were simultaneously upregulated. ErbB pathway dysregulation was identified in ∼20%-30% of samples for most tumor types (medulloblastoma/primitive neuroectodermal tumors 31.1%, high-grade glioma 27.1%, neuroblastoma 22.7%, rhabdomyosarcoma 23.1%, ependymoma 18.8%), 4.2% of diffuse intrinsic pontine gliomas, and no recurrent or refractory low-grade astrocytomas. In medulloblastoma/primitive neuroectodermal tumors and neuroblastoma, this was attributed mainly to high EGFR polysomy/HER2 amplification, whereas EGFR gene amplification was observed in some high-grade glioma samples. EGFR/HER2 overexpression was most prevalent in ependymoma. Conclusions Overexpression and/or amplification of EGFR/HER2 were identified as potential enrichment biomarkers for clinical trials of ErbB-targeted drugs.

Medulloblastoma with extensive nodularity (MBEN) is one of the few central nervous system (CNS) tumor entities occurring in infants which is traditionally associated with good to excellent prognosis. Some MBEN, however, have been reported with an unfavorable clinical course. We performed an integrated DNA/RNA-based molecular analysis of a multi-institutional MBEN cohort (n = 41) to identify molecular events which might be responsible for variability in patients' clinical outcomes. RNA sequencing analysis of this MBEN cohort disclosed two clear transcriptome clusters (TCL) of these CNS tumors: "TCL1 MBEN" and "TCL2 MBEN" which were associated with various gene expression signatures, mutational landscapes and, importantly, prognosis. Thus, the clinically unfavorable "TCL1 MBEN" subset revealed transcriptome signatures composed of cancer-associated signaling pathways and disclosed a high frequency of clinically relevant germline PTCH1/SUFU alterations. In contrast, gene expression profiles of tumors from the clinically favorable "TCL2 MBEN" subgroup were associated with activation of various neurometabolic and neurotransmission signaling pathways, and germline SHH-pathway gene mutations were extremely rare in this transcriptome cluster. "TCL2 MBEN" also revealed strong and ubiquitous expression of VSNL1 (visinin-like protein 1) both at the mRNA and protein level, which was correlated with a favorable clinical course. Thus, combining mutational and epigenetic profiling with transcriptome analysis including VSNL1 immunohistochemistry, MBEN patients could be stratified into clinical risk groups of potential value for subsequent treatment planning.

Introduction: The H3K27M-mutant diffuse midline glioma (DMG) was first included in the World Health Organization (WHO) Classification of central nervous system (CNS) tumors in 2016, and confirmed in its fifth edition. The biological behavior and dismal prognosis of this tumor resemble diffuse intrinsic pontine gliomas (DIPG). Homogeneously-treated series are rarely reported. Methods: From 2016 onwards, we treated patients with DMG with radiotherapy and concomitant/adjuvant nimotuzumab/vinorelbine, plus re-irradiation at relapse, as already done for DIPG. Results: We treated nine patients, seven females, with a median age at diagnosis of 13 years. Tumor sites were: thalamic in five cases, pontocerebellar in two, pineal in one, and paratrigonal with nodular/leptomeningeal dissemination in one. Three patients were biopsied, and six had partial tumor resections. Central pathological review was always performed. The median time to local progression was 12.7 months, and the median overall survival was 17.8 months. Six patients died of tumor progression, one of cerebral bleeding at progression. Two were alive, one in continuous remission, the other after relapsing, at 38.6 and 46.3 months after diagnosis. Progression-free survival was 33.3% at one year. Overall survival was 88.9%, 33.3% and 22.2% at 1, 2 and 3 years, respectively. Conclusions: This is a small series of homogeneously-treated DMG patients. The results obtained are comparable with those of DIPG patients. Given the phenotypically- and molecularly-defined setting of DMG and severe outcome in this orphan population, they should be treated and included in registries and protocols of DIPG.

Enrolling medulloblastoma(MB) patients in the PNET5 protocol is a daily problem in Italy; since June 2015, 59 cases have been enrolled in 13 centres. So far, 44 of the 103 patients claiming for eligibility did not enter the protocol: 13 metastases, 5 for residual, 20 having exclusion criteria, 4 insufficient frozen material, 2 failure to comply with the correct procedures. No case was lost due to delayed centralization, which is respected even with committing weekends; review of the radiation plan was performed on Saturday for 2 cases, and radiotherapy began on the same day. We made some procedural changes to meet expected deadlines; each local centre notifies the national coordinator of a possible case's existence at MRI diagnosis, of the expected surgery date as well as its realization. MRI imaging is reviewed within 2 days after centralization. Paediatricians notify the national coordinator and pathology/biology reference centre of the MB diagnosis; the shipment of frozen tissue, blood and FFPE is booked. A slot is reserved to priority perform the central pathology review, as well as central molecular diagnosis of genetically defined subgroup (WHO classification) upon receipt of the frozen material. Upon receipt of the FFPE and frozen material, the national reference centre undertakes a double-check with the national coordinator and the local treatment centre to validate the eligibility. Within the 7th day from the receipt of the material: IHC, MYC/MYCN, Monosomy 6, beta-catenin mutation and methylation array are performed. Priority execution of somatic (blood control) sequencing of the PTCH, SUFU, and TP53 genes is also triggered for SHH-activated MB, with the deadline on the 15th day. So far we have had 99% agreement between molecular subgrouping and methylation array. CONCLUSIONS: PNET5 requirements are multiple and changing over time; difficulties may and must be overcome by mutual fast collaboration.

BACKGROUND: The purposes of this trial were evaluating the feasibility, response and PFS/OS of patients receiving upfront radiotherapy and reirradiation at progression with concomitant nimotuzumab/vinorelbine chemotherapy as the standard arm of the ongoing protocol for DIPG. METHODS: Patients aged 2-21 years with not-pretreated DMG (evidence of H3.3K27M mutation at immunohistochemistry) received vinorelbine 20 mg/m2+nimotuzumab 150 mg/m2 weekly for 12 weeks; thereafter every other week until tumor progression or for up to 2 years. Focal photons irradiation was delivered within the 3rd week after first nimotuzumab+vinorelbine administration with a total dose of 54 Gy in 1.8 Gy daily fractions. For local progression re-irradiation was proposed at 19.8 Gy; in case of dissemination craniospinal-irradiation at 36 Gy was adopted. MRI were blindly centrally reviewed at diagnosis and every 12 weeks thereafter. RESULTS: Aggregated preliminary results are available for 20 patients from 4 Italian centers: 12 males, median age 11 years (range 3-17). Median time of observation was 12.5 months; 8 had thalamic/basal ganglia disease, in 5 pons was involved (2 pontobulbar, 3 pontomesencephalic), 6 spinal, 2 cerebellar. 13 patients had only biopsy, the others partial or near-total resection.14 relapsed: 5 locally, 4 with dissemination, 5 with both; 12 died, one was lost at follow-up. Two patients received reirradiation, one of them was irradiated three times without evidence of radionecrosis, still alive at 26 months from diagnosis. Median EFS/OS were 8.3/10.2 months, respectively; EFS/OS at 1 year were 25.8/36.7%. Survival curves between spinal and cerebral locations showed no difference. Patients after partial/near-total resection vs biopsied seemed to have earlier relapses (P 0.017) with EFS at 6 months of 34.3/75.0% respectively. CONCLUSIONS: This is one of the first series of DMG homogeneously and prospectively treated; treatment was feasible. A potential role of reirradiation emerge as in DIPGs.

We report two cases of pediatric low-grade glioma (pLGG) treated with vemurafenib, an oral BRAF-inhibitor: a 12-year-old girl with involvement of basal ganglia, hypothalamus, and diencephalic junction; a 3-year-old boy, with an optic pathway/hypothalamic glioma extending along the left optic tract and basal ganglia. Both received a biopsy and molecular analysis was performed in the girl’s tumor, showing BRAF V600E mutation. Therefore, instead of surgical removal planned by neurosurgeons, first-line treatment with vemurafenib was started: after one month 45% reduction of the mass according to RANO criteria was found, as well as better balance control and strong reduction of the right arm paresis; five months later, a 70% shrinkage was detected, stabilized to 76% after a year. The young boy first started chemotherapy with vincristine and carboplatin, but at the end of the induction phase the tumor had increased and ascites, hydrocephalus, and visual impairment occurred. Molecular testing showing BRAF V600E mutation on the initial tumor biopsy was obtained; therefore, the surgical option was postponed and therapy with vemurafenib started. After only three days, visual acuity and muscle tone improved; brain MRI showed a 34% reduction of the mass after one week, increased up to 65% after six months. Therefore, no ulterior surgery was necessary. In pLGG, the neurosurgical biopsy is essential to let an early and rapid molecular diagnosis of BRAF mutations and guide subsequent targeted therapies. Our cases demonstrate how a prompt radiological response to vemurafenib and the related clinical improvement can influence both therapeutic and surgical decisions, hopefully reducing the occurrence of second neurosurgery with associated risks of neurological sequelae. To our knowledge, this is the first report assessing such a quick shrinkage in pLGG treated with vemurafenib, highlighting the importance of an early investigation of BRAF status in all cases of LGG in children.

Marburg type is a rare variant of Multiple Sclerosis (MS) characterized by a severe and progressive course/evolution leading to exitus in few months. Its radiological presentation commonly raises suspicion of a malignancy, often requiring a brain biopsy for histological confirmation. Hereby we report an atypical radiological presentation and progression of a pathologically confirmed case of Marburg MS Keywords: Multiple Sclerosis; Marburg variant; Magnetic resonance imaging.

Purpose Our purpose was to better understand the pathogenesis of seizures associated with angiocentric glioma. Angiocentric glioma is an indolent and rare low-grade glioma. Its typical clinical presentation is with epileptic seizures. The pathogenesis of tumor-associated seizures is poorly understood. Among the possible pathomechanisms, the increased neurotoxic concentrations of the glutamate has been proposed. Glutamate transporters, pyruvate carboxylase and glutamine synthetase are involved in maintaining the physiological concentration of glutamate in the inter synaptic spaces. Methods We evaluated the immunohistochemical expression of EAAT2 (the most important glutamate transporter), pyruvate carboxylase and glutamine synthetase in 17 angiocentric gliomas. Results EAAT2 was never expressed (0%) in the neoplastic cells in none of the cases studied. Pyruvate carboxylase was expressed in the cytoplasm of the neoplastic cells in 16/17 cases (94 %). Glutamine synthetase was expressed in the cytoplasm of the neoplastic cells in 15/17 cases (88 %). Conclusion The net result of this enzymatic expression, in particular considering the loss of EAAT2, could be an increased glutamate concentration in the synaptic clef, which might increase local network excitability initially involving intratumoral neurons. The observation that the angiocentric glioma-associated epilepsy might be at least in part related to EAAT2 deficiency opens up interesting therapeutic perspectives.

Aim Diffuse midline glioma (DMG) H3 K27M-mutant is a specific entity that, as the name indicates, tends to occur in midline structures including the thalamus, brainstem, and spinal cord. DMG predominates in children, is an aggressive tumor with poor prognosis, and is considered a WHO grade IV tumor regardless of histological features. The exact frequency of these mutations in adults diagnosed with glioma in the midline is unknown. Materials and methods We report a series of 6 more adult cases, and we critically review the current literature on adults with DMG H3 K27M-mutant. Results There were 5 males and 1 female. The age ranged from 26 to 52 years (median 39 years). All cases showed astrocytic differentiation, with positive staining for H3 K27M protein, and loss of H3 K27me in the tumor cells confirming the diagnosis. Conclusion H3 K27M-mutant midline glioma can occur in adults, affecting midline structures. Increasing awareness of the reporting pathologists of this entity might help in a better determination of the frequency of mutant DMG in adults as well as better diagnosis and patient counseling of the outcome.

Purpose The maintenance of telomere length prevents cancer cell senescence and occurs via two mutually exclusive mechanisms: (a) reactivation of telomerase expression and (b) activation of alternative lengthening of telomeres (ALT). ALT is frequently related to alterations on ATRX, a chromatin-remodelling protein. Recent data have identified different molecular subgroups of paediatric high-grade glioma (pHGG) with mutations of H3F3A, TERTp and ATRX; however, differences in telomere length among these molecular subgroups were not thoroughly examined. Methods We investigated which genetic alterations trigger the ALT mechanism in 52 IDH-wildtype, 1p/19q-wildtype pHGG. Samples were analysed for telomere length using Tel-FISH. ATRX nuclear loss of expression was assessed by IHC, H3F3A and TERTp mutations by DNA sequencing, and TERTp methylation by MS-PCR. Results Mutant H3.3 was found in 21 cases (40.3%): 19.2% with K27M mutation and 21.1% with G34R mutation. All H3.3G34R-mutated cases showed the ALT phenotype (100%); on the opposite, only 40% of the H3.3K27M-mutated showed ALT activation. ATRX nuclear loss was seen in 16 cases (30.7%), associated sometimes with the G34R mutation, and never with the K27M mutation. ATRX nuclear loss was always related to telomere elongation. TERTp C250T mutations were rare (5.4%) and were not associated with high intensity Tel-FISH signals, as TERTp hyper-methylation detected in 21% of the cases. H3.3/ATRX/TERTp-wildtype pHGG revealed all basal levels of telomere length. Conclusion Our results show a strong association between H3.3 mutations and ALT, and highlight the different telomeric profiles in histone-defined subgroups: H3.3-G34R mutants always trigger ALT to maintain telomere length, irrespective of ATRX status, whereas only some H3.3-K27M tumours activate ALT. These findings suggest that acquiring the gly34 mutation on H3.3 might suffice to trigger the ALT mechanism.

Introduction: Medulloblastoma is the most common malignant brain tumor in children, but accounts for only 1% of brain cancers in adults. For standard-risk pediatric medulloblastoma, current therapy includes craniospinal irradiation (CSI) at reduced doses (23.4Gy) associated with chemotherapy. Whereas most same-stage adult patients are still given CSI at 36Gy, with or without chemotherapy, we report here on our use of reduced-dose CSI associated with chemotherapy for older patients. Methods: We gathered non-metastatic patients over 18years old (median age 28years, range 18–48) with minimal or no residual disease after surgery, no negative histological subtypes, treated between 1996–2018 at the Centre Léon Bérard (Lyon) and the INT (Milano). A series of 54 children with similar tumors treated in Milano was used for comparison. Results: Forty-four adults were considered (median follow-up 101months): 36 had 23.4Gy of CSI, and 8 had 30.6Gy, plus a boost to the posterior fossa/tumor bed; 43 had chemotherapy as all 54 children, who had a median 83-month follow-up. The PFS and OS were 82.2 ± 6.1% and 89 ± 5.2% at 5 years, and 78.5 ± 6.9% and 75.2 ± 7.8% at ten, not significantly different from those of the children. CSI doses higher than 23.4Gy did not influence PFS. Female adult patients tended to have a better outcome than males. Conclusion: The results obtained in our combined series are comparable with, or even better than those obtained after high CSI doses, underscoring the need to reconsider this treatment in adults.

BACKGROUND This 2002–2014 Italian prospective study stratified 160 patients by surgical resection (complete=NED/incomplete=ED) and centrally-reviewed grade. Grade2/NED patients received focal radiotherapy (RT) up to 59.4Gy, Grade3/NED received 4 courses of VEC(vincristine,etoposide,cyclophosphamide) after RT.ED patients received 1–4 VEC courses, second-look surgery, 59.4 Gy+8Gy boost on measurable residue. METHODS We re-analyzed data at 115 months follow-up including methylation profile on available samples. RESULTS Global PFS/OS at 5/10 years were 66/59% and 80/74%, respectively. Of the 64 relapsers at median 20 months, 53 died at median 37/18 months after diagnosis/relapse, respectively.10/64 relapsed after 5 years (66–126 months); 4 died, relapse was local in 8/10, metastatic 1, combined 1;5/10 patients were below age 3, 5 females, 8 PF tumors. Their survival post-relapse was not longer than earlier relapsers’. At univariable analysis, age over 3 years, female sex, complete surgery, grade 2, no shunt confirmed better PFS/OS. 66/95 analyzed tumors received a score >0.80 through the DNA methylation-based central nervous system tumor classifier: 41/8 as PFA/PFB, respectively,14/17 ST as RELA-positive (3 scored for other molecular entities i.e. anaplastic PXA, LGG MYB, HGNET). Prognostic factors were equally distributed among PFA/PFB groups,1 only group B patient relapsed locally at 96 months. CONCLUSIONS Already published prognostic factors remained at long-term follow-up;6.2% patients had late relapses. OS after relapse was not better in late relapsers. Group B confirmed better prognosis but all patients had received «at least» adjuvant RT. Modern ependymoma trials need long follow-up to draw firm conclusions.

Mechanical stimuli are a fundamental player in the pathophysiology of the brain influencing its physiological development and contributing to the onset and progression of many diseases. In some pathological states, the involvement of mechanical and physical stimuli might be extremely subtle; in others, it is more evident and particularly relevant. Among the latter pathologies, one of the most serious life-threatening condition is the brain abscess (BA), a focal infection localized in the brain parenchyma, which causes large brain mechanical deformations, giving rise to a wide range of neurological impairments. In this paper, we present the first nano-mechanical characterization of surgically removed human brain abscess tissues by means of atomic force microscopy (AFM) in the spectroscopy mode. Consistently with previous histological findings, we modeled the brain abscess as a multilayered structure, composed of three main layers: the cerebritis layer, the collagen capsule, and the internal inflammatory border. We probed the viscoelastic behavior of each layer separately through the measure of the apparent Young’s modulus (E), that gives information about the sample stiffness, and the AFM hysteresis (H), that estimates the contribution of viscous and dissipative forces. Our experimental findings provide a full mechanical characterization of the abscess, showing an average E of (94 ± 5) kPa and H of 0.37 ± 0.01 for the cerebritis layer, an average E = (1.04 ± 0.05) MPa and H = 0.10 ± 0.01 for the collagen capsule and an average E = (9.8 ± 0.4) kPa and H = 0.57 ± 0.01 for the internal border. The results here presented have the potential to contribute to the development of novel surgical instruments dedicated to the treatment of the pathology and to stimulate the implementation of novel constitutive mechanical models for the estimation of brain compression and damage during BA progression.

High-grade gliomas (HGGs) in pediatric age have the same bad prognosis as those arising in adults. Approximately one-half of HGGs in children occur in the brain stem, most frequently within the pons as diffuse intrinsic pontine glioma or other midline structures. Although they have the same histological appearance of adult malignant gliomas, in recent years, the extensive use of molecular profiling techniques has demonstrated significant molecular differences between the two age groups. These data have led to a major reclassification of pediatric HGG (pHGG) based on molecular subgrouping with significant clinical correlations in terms of age at presentation, anatomical location, and prognosis. The most important molecular groups are: (1) the histone mutations related pHGG, that is, H3.K27-mutated midline and H3.G34-mutated hemispheric pHGG; (2) the rare isocitrate dehydrogenase (IDH)-mutated pHGG occurring mainly in adolescents; and (3) the H3-/IDH wild type, a heterogenous group of pHGG still object of further molecular stratification. Another important group of pHGG is that occurring in patients with cancer predisposition syndromes such as Li-Fraumeni syndrome, constitutional mismatch repair deficiency, and neurofibromatosis-1 (NF1). In this review, the different subgroups of pHGG and their major driver molecular alterations will be discussed.