Posterior Fossa Ependymoma Research Articles

Cell Ecosystem and Signaling Pathways of Primary and Metastatic Pediatric Posterior Fossa Ependymoma

Pediatric ependymoma is a devastating brain cancer marked by its relapsing pattern and lack of effective chemotherapies. This shortage of treatments is partially due to limited knowledge about ependymoma tumorigenic mechanisms. Although there is evidence that ependymoma originates in radial glia, the specific pathways underlying the progression and metastasis of these tumors are unknown. By means of single-cell transcriptomics, immunofluorescence, and in situ hybridization, we show that the expression profile of tumor cells from pediatric ependymomas in the posterior fossa is consistent with an origin in LGR5 + stem cells. Tumor stem cells recapitulate the developmental lineages of radial glia in neurogenic niches, promote an inflammatory microenvironment in cooperation with microglia, and upon metastatic progression initiate a mesenchymal program driven by reactive gliosis and hypoxia-related genes. Our results uncover the cell ecosystem of pediatric posterior fossa ependymoma and identify WNT/s-catenin and TGF-s signaling as major drivers of tumorigenesis for this cancer.

Spatial Agreement of Brainstem Dose Distributions Depending on Biological Model in Proton Therapy for Pediatric Brain Tumors

PurposeDuring radiation therapy for pediatric brain tumors, the brainstem is a critical organ at risk, possibly with different radio-sensitivity across its substructures. In proton therapy, treatment planning is currently performed using a constant relative biological effectiveness (RBE) of 1.1 (RBE1.1), whereas preclinical studies point toward spatial variability of this factor. To shed light on this biological uncertainty, we investigated the spatial agreement between isodose maps produced by different RBE models, with emphasis on (smaller) substructures of the brainstem. Methods and MaterialsProton plans were recalculated using Monte Carlo simulations in 3 anonymized pediatric patients with brain tumors (a craniopharyngioma, a low-grade glioma, and a posterior fossa ependymoma) to obtain dose and linear energy transfer distributions. Doses and volume metrics for the brainstem and its substructures were calculated using a constant RBE1.1, 4 phenomenological RBE models with varying (α/β)x parameters, and with a simpler linear energy transfer-dependent model. The spatial agreement between the dose distributions of constant RBE1.1 versus the variable RBE models was compared using the Dice similarity coefficient. ResultsThe spatial agreement between the variable RBE dose distributions and RBE1.1 decreased with increasing isodose levels in all patient cases. The patient with ependymoma showed the greatest variation in dose and dose volumes, where V50Gy(RBE) in the brainstem increased from 32% (RBE1.1) to 35% to 49% depending on the applied model, corresponding to a spatial agreement (Dice similarity coefficient) between 0.79 and 0.95. The remaining patients showed similar trends, however, with lower absolute values due to lower brainstem doses. ConclusionsAll phenomenological RBE models fully enclosed the isodose volumes of the constant RBE1.1, and the volumes based on variable RBE spatially agreed. The spatial agreement was dependent on the isodose level, where higher isodose levels showed larger expansions and less agreement between the variable RBE models and RBE1.1.

CIMPACT-NOW update 7: advancing the molecular classification of ependymal tumors.

Advances in our understanding of the biological basis and molecular characteristics of ependymal tumors since the latest iteration of the World Health Organization (WHO) classification of CNS tumors (2016) have prompted the cIMPACT-NOW group to recommend a new classification. Separation of ependymal tumors by anatomic site is an important principle of the new classification and was prompted by methylome profiling data to indicate that molecular groups of ependymal tumors in the posterior fossa and supratentorial and spinal compartments are distinct. Common recurrent genetic or epigenetic alterations found in tumors belonging to the main molecular groups have been used to define tumor types at intracranial sites; C11orf95 and YAP1 fusion genes for supratentorial tumors and two types of posterior fossa ependymoma defined by methylation group, PFA and PFB. A recently described type of aggressive spinal ependymoma with MYCN amplification has also been included. Myxopapillary ependymoma and subependymoma have been retained as histopathologically defined tumor types, but the classification has dropped the distinction between classic and anaplastic ependymoma. While the cIMPACT-NOW group considered that data to inform assignment of grade to molecularly defined ependymomas are insufficiently mature, it recommends assigning WHO grade 2 to myxopapillary ependymoma and allows grade 2 or grade 3 to be assigned to ependymomas not defined by molecular status.

Ependymomas in infancy: underlying genetic alterations, histological features, and clinical outcome

IntroductionYoung age is an adverse prognostic factor in children with ependymomas. Treatment of these infants is challenging since beneficial therapeutic options are limited. As ependymomas are considered a biologically heterogeneous group, we aimed to characterize infant ependymomas with regard to their histological and genetic features.Materials and methodsWe analyzed 28 ependymomas occurring in children younger than 18 months at diagnosis enrolled into the HIT2000-E protocols with the aim to postpone irradiation until the age of 18 months if possible. All cases underwent neuropathological review, including immunohistochemical characterization. Genome-wide copy number alterations (CNA) were assessed by molecular inversion probe assays, and RELA and YAP1 fusions were detected by RT-PCR and sequencing.ResultsAll infant ependymomas were anaplastic (WHO grade III). Twenty-one (75%) cases were located in the posterior fossa. Gross total resection was accomplished in 12 (57%) of these cases. All posterior fossa tumors showed loss of H3-K27me3 characteristic of PFA ependymomas. CNA analysis showed a stable genome in all cases with lack of chromosome 1q gain, an adverse prognostic marker in PFA ependymomas of older children. However, after a median follow-up of 5.4 years, 15 (71%) relapsed, and 9 (43%) died. Seven ependymomas (25%) occurred in the supratentorial region. Gross total resection could be achieved in only two of these cases. Four tumors carried C11orf95-RELA fusions, and two cases had typical YAP1-MAMLD1 fusions (one case was not analyzable). The RELA-fused cases did not display CDKN2A loss as an adverse indicator of prognosis in this disease entity. Although three infants (43%) with supratentorial ependymomas relapsed, all patients survived (median follow-up, 8.0 years).ConclusionInfant ependymomas seem to fall into three biological entities, with supratentorial tumors carrying RELA or YAP fusions and PFA posterior fossa ependymomas. The latter showed a poor outcome even though chromosome 1q gain was absent.

Radiology Profile as a Potential Instrument to Differentiate Between Posterior Fossa Ependymoma (PF-EPN) Group A and B

Posterior fossa ependymoma (PF-EPN) was categorized into PF-EPN-A and PF-EPN-B subgroups based on the DNA methylation profiling. PF-EPN-A was reported to have poorer prognosis compared with PF-EPN-B. In this study, we particularly evaluated preoperative imaging to distinguish PF-EPN-A from PF-EPN-B. Sixteen cases of PF-EPN were treated in our institution from 1999 to 2018. The patients were divided into PF-EPN-A and PF-EPN-B groups based on H3K27me3 immunostaining positivity. We evaluated progression-free survival, overall survival, as well as preoperative magnetic resonance imaging and computed tomography scan images in both groups. Based on T1WI and Gd-T1WI magnetic resonance images, the tumor contrast rate was determined from dividing the volume of gadolinium enhanced tumor by the overall tumor volume. Nine cases (4 male, 5 female) were grouped as PF-EPN-A, and 7 (4 male, 3 female) as PF-EPN-B. The median age of PF-EPN-A and PF-EPN-B were 4 and 43 years old, respectively. In the PF-EPN-A group, the progression-free survival median value was 32.6 months, and the overall survival median was 96.9 months. In contrast, PFS in PF-EPN-B did not reach a median value (P < 0.05) and all the patients were alive (P < 0.05) at the end of the study. With imaging, tumor contrast rate in PF-EPN-B was more than 50% and significantly different from PF-EPN-A (P= 0.0294). Calcification was mainly observed in PF-EPN-A, whereas cystic formation was only seen in PF-EPN-B. Contrast rate less than 50%, based on the magnetic resonance images, was characteristic in the PF-EPN-A group. Comparatively, cystic component and absence of calcification were more characteristic in the PF-EPN-B group.

Chondro-Osseous Metaplasia in Ependymoma: A Rare Histopathological Finding

Ependymoma is a circumscribed glioma composed of uniform glial cells with bland nuclei in a fibrillary matrix. It is characterized by the presence of perivascular pseudorosettes. Unusual histopathological findings have rarely been reported in ependymomas, 0.5% of all diagnosed cases. Such unusual and exceedingly rare histological findings include osseous or chondroid metaplasia. To the best of our knowledge, only 15 cases of osseocartilaginous ependymomas have been reported in English literature. We report a 3-year-old boy who presented with ataxia, vomiting, and headache for three months. Radiological imaging revealed a posterior fossa lesion. Histopathological examination of the lesion confirmed a posterior fossa ependymoma with chondro-osseous metaplasia. The present case outlines the clinical presentation, histopathological findings, and outcome of chondro-osseous metaplasia in ependymomas. To date, the etiology of chondro-osseous metaplasia in ependymomas remains uncertain. Further research exploring such phenomenon is of paramount importance to explain how these tumors develop.

Adult outcome after treatment of pediatric posterior fossa ependymoma: long-term follow-up of a single consecutive institutional series of 22 patients with more than 5 years of survival.

Ependymoma is the third most common posterior fossa tumor in children; however, there is a lack of long-term follow-up data on outcomes after surgical treatment of posterior fossa ependymoma (PFE) in pediatric patients. Therefore, the authors sought to investigate the long-term outcomes of children treated for PFE at their institution. The authors performed a retrospective analysis of outcome data from children who underwent treatment for PFE and survived for at least 5 years. The authors identified 22 children (median age at the time of surgery 3 years, range 0-18 years) who underwent primary tumor resection of PFE during the period from 1945 to 2014 and who had at least 5 years of observed survival. None of these 22 patients were lost to follow-up, and they represent the long-term survivors (38%) from a total of 58 pediatric PFE patients treated. Nine (26%) of the 34 children treated during the pre-MRI era (1945-1986) were long-term survivors, while the observed 5-year survival rate in the children treated during the MRI era (1987-2014) was 13 (54%) of 24 patients. The majority of patients (n = 16) received adjuvant radiotherapy, and 4 of these received proton-beam irradiation. Six children had either no adjuvant treatment (n = 3) or only chemotherapy as adjuvant treatment (n = 3). Fourteen patients were alive at the time of this report. According to MRI findings, all of these patients were tumor free except 1 patient (age 78 years) with a known residual tumor after 65 years of event-free survival.Repeat resections for residual or recurrent tumor were performed in 9 patients, mostly for local residual disease with progressive clinical symptoms; 4 patients underwent only 1 repeated resection, whereas 5 patients each had 3 or more resections within 15 years after their initial surgery. At further follow-up, 5 of the patients who underwent a second surgery were found to be dead from the disease with or without undergoing additional resections, which were performed from 6 to 13 years after the second procedure. The other 4 patients, however, were tumor free on the latest follow-up MRI, performed from 6 to 27 years after the last resection. Hence, repeated surgery appears to increase the chance of tumor control in some patients, along with modern (proton-beam) radiotherapy. Six of 8 patients with more than 20 years of survival are in a good clinical condition, 5 of them in full-time work and 1 in part-time work. Pediatric PFE occurs mostly in young children, and there is marked risk for local recurrence among 5-year survivors even after gross-total resection and postoperative radiotherapy. Repeated resections are therefore an important part of treatment and may lead to persistent tumor control. Even though the majority of children with PFE die from their tumor disease, some patients survive for more than 50 years with excellent functional outcome and working capacity.

Ependymoma Pediatric Brain Tumor Protein Fingerprinting by Integrated Mass Spectrometry Platforms: A Pilot Investigation.

Ependymoma pediatric brain tumor occurs at approximate frequencies of 10–15% in supratentorial and 20–30% in posterior fossa regions. These tumors have an almost selective response to surgery and relative and confirmed resistance to radiotherapy and chemotherapic agents, respectively. Alongside histopathological grading, clinical and treatment evaluation of ependymomas currently consider the tumor localization and the genomic outlined associated molecular subgroups, with the supratentorial and the posterior fossa ependymomas nowadays considered diverse diseases. On these grounds and in trying to better understand the molecular features of these tumors, the present investigation aimed to originally investigate the proteomic profile of pediatric ependymoma tissues of different grade and localization by mass spectrometry platforms to disclose potential distinct protein phenotypes. To this purpose, acid-soluble and acid-insoluble fractions of ependymoma tumor tissues homogenates were analyzed by LC-MS following both the top-down and the shotgun proteomic approaches, respectively, to either investigate the intact proteome or its digested form. The two approaches were complementary in profiling the ependymoma tumor tissues and showed distinguished profiles for supratentorial and posterior fossa ependymomas and for WHO II and III tumor grades. Top-down proteomic analysis revealed statistically significant higher levels of thymosin beta 4, 10 kDa heat shock protein, non-histone chromosomal protein HMG-17, and mono-/uncitrullinated forms ratio of the glial fibrillary acidic protein (GFAP) fragment 388–432 in supratentorial ependymomas—the same GFAP fragment as well as the hemoglobin alpha- and the beta-chain marked grade II with respect to grade III posterior fossa ependymomas. Gene ontology classification of shotgun data of the identified cancer and the non-cancer related proteins disclosed protein elements exclusively marking tumor localization and pathways that were selectively overrepresented. These results, although preliminary, seem consistent with different protein profiles of ependymomas of diverse grade of aggressiveness and brain region development and contributed to enlarging the molecular knowledge of this still enigmatic tumor.

Is H3K27me3 status really a strong prognostic indicator for pediatric posterior fossa ependymomas? A single surgeon, single center experience.

Posterior fossa ependymomas (PFE) are among the most frequently occurring solid tumors in children. Their definitive treatment is surgical excision and adjuvant radio-chemotherapy. This study aimed to investigate prognostic effects of age, H3K27me3 status, extent of resection, radiation treatment (RT), Ki67 index, WHO grade, and ATRX and H3K27M mutations in PFE patients. This retrospective study included 42 pediatric patients with PFE who had undergone operation at our institution between 1996 and 2018. Patient demographics and treatment information were obtained from patient notes. Information on radiological location of tumors (median vs paramedian), extent of tumor resection, and recurrence was obtained from preoperative and postoperative magnetic resonance imaging. Formalin-fixed paraffin-embedded tumor samples were evaluated for H3K27me3 immunostaining, Ki67 index, WHO grades, and ATRX and H3K27M mutations. Tumor samples with global reduction in H3K27me3 were grouped as posterior fossa ependymoma group A (PFA) and those with H3K27me3 nuclear immunopositivity as posterior fossa ependymoma group B (PFB). We evaluated the cohort's 5-year progression-free survival (PFS) and overall survival (OS). There were 20 (47.6%) female and 22 (52.4%) male patients in the cohort. The mean age of patients was 4.4 (range, 0.71-14.51) years. Overall, tumors in 31 (73.8%) and 11 (26.2%) patients were found to be PFA and PFB, respectively. There was no statistically significant age or sex difference between PFA and PFB. All patients received chemotherapy, whereas only 28 (66.6%) received RT. The WHO grades of PFA were statistically higher than those of PFB. There was no significant difference between PFA and PFB in terms of extent of resection, disease recurrence, and survival parameters. Nine of 42 tumor samples had ATRX mutations. One patient with PFA showed H3K27M mutation. Age, WHO grade, H3K27me3 status, and RT had no effect on patients' PFS and OS. Patients with total surgical excisions had significantly better PFS and OS rates. Those with Ki67 < 50% also had better OS rates. Determining H3K27me3 status by immunohistochemistry is a widely accepted method for molecular subgrouping of PFEs. Most of the reports in the literature state that molecular subgroups of PFEs have significantly different clinical outcomes. However, in our present series, we have shown that the extent of surgical excision is still the most important prognostic indicator in PFEs. We also conclude that the prognostic effect of H3K27me3 status-based molecular subgrouping may be minimized with a more aggressive surgical strategy followed in PFAs.

Characterization of global 5-hydroxymethylcytosine in pediatric posterior fossa ependymoma

Background5-Hydroxymethylcytosine (5hmC) is a novel epigenetic mark and may be involved in the mechanisms of tumorigenesis and malignant transformation. However, the role of 5hmC in ependymoma, the third most common brain tumor in children, remains unclear. The aim of this study sought to identify the characterization of 5hmC levels in pediatric posterior fossa ependymoma and to evaluate whether 5hmC levels could be a potential factor to predict clinical outcomes.ResultsOur results showed that 5hmC levels were globally decreased in posterior fossa ependymoma compared with normal cerebellum tissues (P < 0.001). Group A posterior fossa ependymomas had higher 5hmC levels than group B tumors (P = 0.007). Moreover, 5hmC levels positively correlated with Ki-67 index in posterior fossa ependymoma (r = 0.428, P = 0.003). Multivariate Cox hazards model revealed that patients with high 5hmC levels (> 0.102%) had worse PFS and OS than patients with lower 5hmC levels (< 0.102%) (PFS: HR = 3.014; 95% CI, 1.040–8.738; P = 0.042; OS: HR = 2.788; 95% CI, 0.974–7.982; P = 0.047).ConclusionsOur findings suggest that loss of 5hmC is an epigenetic hallmark for pediatric posterior fossa ependymoma. 5hmC levels may represent a potential biomarker to predict prognosis in children with posterior fossa ependymoma.

TET2 promotor methylation and TET2 protein expression in pediatric posterior fossa ependymoma.

Pediatric posterior fossa ependymoma (PF) is one of the most common brain tumors in children. Recently, two subtypes of PF were identified. PF-A has a dismal prognosis and shows a hypermethylation phenotype, whereas PF-B shows a great genomic instability. The ten-eleven translocation methylcytosine dioxygenase 2 (TET2) gene (TET2) has been linked to the regulation of DNA methylation. We analyzed TET2 promotor methylation and protein expression to assess the role of TET2 in PF. Medical records of all PF cases treated in our institution between 1993 and 2015 were evaluated regarding tumor histology, grade, tumor location, gender, age, tumor recurrence, distant metastasis, survival and time to progression. Subsequently, we analyzed TET2 promotor methylation using methylation-specific polymerase chain reaction. TET2 protein expression was assessed using immunohistochemistry. Low TET2 expression was detected in seven of 17 cases. There was an association between low TET2 expression and tumor recurrence (P = 0.049). A TET2 promotor methylation was detected in five of 10 cases. There was no association between the TET2 promotor methylation with recurrence, tumor grade or gender. TET2 promotor methylation and low TET2 expression was detected in a subgroup of PF. Our data show an association between low TET2 expression and tumor recurrence in PF.

Improved risk-stratification for posterior fossa ependymoma of childhood considering clinical, histological and genetic features \u2013 a retrospective analysis of the HIT ependymoma trial cohort

IntroductionRisk stratification of children with ependymomas of the posterior fossa in current therapeutic protocols is mainly based on clinical criteria. We aimed to identify independent outcome predictors for this disease entity by a systematic integrated analysis of clinical, histological and genetic information in a defined cohort of patients treated according to the German HIT protocols.MethodsTumor samples of 134 patients aged 0.2–15.9 years treated between 1999 and 2010 according to HIT protocols were analyzed for histological features including mitotic activity, necrosis and vascular proliferation and genomic alterations by SNP and molecular inversion probe analysis. Survival analysis was performed by Kaplan-Meier method with log rank test and multivariate Cox regression analysis.ResultsResidual tumor after surgery, chromosome 1q gain and structural genomic alterations were identified as predictors of significantly shorter event-free (EFS) and overall survival (OS). Furthermore, specific histological features including vascular proliferation, necrosis and high mitotic activity were predictive for shorter OS. Multivariate Cox regression revealed residual tumor, chromosome 1q gain and mitotic activity as independent predictors of both EFS and OS. Using these independent predictors of outcome, we were able to build a 3-tiered risk stratification model that separates patients with standard, intermediate and high risk, and which outperforms current stratification procedures.ConclusionThe integration of defined clinical, histological and genetic parameters led to an improved risk-stratification model for posterior fossa ependymoma of childhood. After validation in independent cohorts this model may provide the basis for risk-adapted treatment of children with ependymomas of the posterior fossa.

PATH-03. DIFFERENCES IN CLINICAL COURSE OF GROUP-A AND GROUP-B POSTERIOR FOSSA EPENDYMOMA (PFA, PFB) AS DEFINED BY H3K27ME3 IMMUNOHISTOCHEMICAL ANALYSIS

Abstract PURPOSE Posterior fossa ependymoma is classified as PFA and PFB based on underlying genetic/mutational characteristics, and PFA is reported to have poor prognosis compared to PFB. Recently, immunohistochemical findings of negative H3K27me3 have been reported to suggest PFA and attracted the attention as a surrogate marker. We classified our cases into PFA and PFB guided by immunohistochemical staining results of H3K27me3 and examined their clinical course. MATERIALS AND METHODS Between 1999 and 2018, 17 cases of posterior fossa ependymoma were treated at our institute. Negative immunostaining for H3K27me3 was classified as PFA, and positive as PFB. We evaluated age, gender, pathological diagnosis, progression free survival (PFS) and overall survival (OS) in both groups. RESULTS Nine cases were classified as PFA and eight as PFB. The median age of the PFA group was 4 years-old, 4 males, 5 females, pathological diagnosis was ependymoma in 5, and anaplastic ependymoma in 4 cases. The median follow-up period was 50 months, recurrence was confirmed in 5 cases (56%), PFS was 27.6 months. Three of the 5 patients who relapsed died (33%), and the OS was 50 months. The median age of the PFB group was 50 years, 4 males and 4 females, the pathological diagnosis was ependymoma in 6, and the anaplastic ependymoma in 2 cases. The median follow-up period was 111 months, recurrence was observed in 2 cases (25%), PFS was 98 months and all patients were alive during the period of this research. CONCLUSION Immunohistochemical results of H3K27me3 correlated with prognosis, implying the possibility of it being a surrogate marker of posterior fossa ependymoma.

PATH-16. HISTOPATHOLOGICAL EPENDYMOMA VARIANTS ARE ASSOCIATED WITH DISTINCT CLINICAL PARAMETERS AND DNA METHYLATION PATTERNS

Abstract According to the current WHO classification, ependymal tumors are classified as subependymomas, myxopapillary ependymomas, classic ependymomas, anaplastic ependymomas and RELA-fusion positive ependymoma (RELA-EPN). Among classic ependymomas, the WHO defines rare histological variants, i.e. the clear-cell, papillary, and tanycytic ependymoma. In parallel to this WHO classification scheme, DNA methylation patterns can distinguish nine distinct molecular ependymoma subgroups, some of which tightly overlap with certain histopathological subgroups, e.g. subependyomas or myxopapillary ependymomas. Since very little is known about the molecular background of histological classic ependymoma variants, we analyzed histomorphology, clinical parameters and global DNA methylation patterns of diagnosed tanycytic ependymomas (n=12), clear-cell ependymomas (n=14) and papillary ependymomas (n=19). Surprisingly, up to 42% of these variants did not match to ependymomas using a previously published DNA methylation-based classifier for brain tumors. Among the tumors with a match to one of the nine known ependymoma methylation classes, tanycytic ependymomas were predominantly located in the spine, but showed diverse molecular methylation patterns. Most clear-cell ependymomas showed a common histomorphology, were found supratentorially and fell into the methylation class of RELA-EPN. Papillary ependymomas showed a “papillary”, “trabecular” or “pseudo-papillary” growth pattern. Interestingly, a true papillary growth pattern was strongly associated with the molecular class B of posterior fossa ependymoma (PFB), but tumors displayed DNA methylation sites that were significantly different when compared to PFB ependymomas without papillary growth. Our results show that the diagnosis of classic histological ependymoma variants can be challenging. While clear-cell and papillary ependymomas harbor common molecular features, tanycytic ependymoma may not represent a molecularly distinct subgroup.

Molecular characterization of histopathological ependymoma variants.

According to the WHO classification, ependymal tumors are classified as subependymomas, myxopapillary ependymomas, classic ependymomas, anaplastic ependymomas, and RELA-fusion-positive ependymomas (RELA-EPN). Among classic ependymomas, the WHO defines rare histological variants, i.e., the clear cell, papillary, and tanycytic ependymoma. In parallel, global DNA methylation patterns distinguish nine molecular groups, some of which tightly overlap with histopathological subgroups. However, the match of the aforementioned histological variants to DNA methylation classes remains unclear. We analyzed histomorphology, clinical parameters, and global DNA methylation of tumors with the initial histological diagnoses of tanycytic (n = 12), clear cell (n = 14), or papillary ependymoma (n = 19). Forty percent of these tumors did not match to the epigenetic profile of ependymomas, using a previously published DNA methylation-based classifier for brain tumors. Instead, they were classified as low-grade glioma (n = 3), plexus tumor (n = 2), CNS high-grade neuroepithelial tumor with MN1 alteration (n = 2), papillary tumor of the pineal region (n = 2), neurocytoma (n = 1), or did not match to any known brain tumor methylation class (n = 8). Overall, integrated diagnosis had to be changed in 35.6% of cases as compared to the initial diagnosis. Among the tumors molecularly classified as ependymoma (27/45 cases), tanycytic ependymomas were mostly located in the spine (5/7 cases) and matched to spinal or myxopapillary ependymoma. 6/8 clear cell ependymomas were found supratentorially and fell into the methylation class of RELA-EPN. Papillary ependymomas with a positive ependymoma match (12/19 cases) showed either a "papillary" (n = 5), a "trabecular" (n = 1), or a "pseudo-papillary" (n = 6) growth pattern. The papillary growth pattern was strongly associated with the methylation class B of posterior fossa ependymoma (PFB, 5/5 cases) and tumors displayed DNA methylation sites that were significantly different when compared to PFB ependymomas without papillary growth. Tumors with pseudo-papillary histology matched to the methylation class of myxopapillary ependymoma (4/6 cases), whereas the trabecular case was anatomically and molecularly a spinal ependymoma. Our results show that the diagnosis of histological ependymoma variants is challenging and epigenetic profiles may improve diagnostic accuracy of these cases. Whereas clear cell and papillary ependymomas display correlations between localization, histology, and methylation, tanycytic ependymoma does not represent a molecularly distinct subgroup.

P12.06 Clinicopathological evaluation of PD-L1 expression and cytotoxic T-lymphocyte infiltrates across intracranial molecular subgroups of ependymomas

Abstract BACKGROUND Immune check-point blockade targeting programmed cell death ligand-1 (PD-L1)/programmed death-1 (PD-1) axis has created paradigm shift in the treatment of advanced cancer. The prognosis of the ‘ST-RELA’ and ‘PF-A’ molecular subgroups of ependymomas (EPN) remains poor. This study aims to understand the potential candidature of EPNs for ICB. MATERIAL AND METHODS All supratentorial (ST) Grade II/III EPNs were classified into ST-RELA, ST-YAP and ST-not otherwise specified (NOS), based on L1CAM protein expression and/or presence of RELA/YAP1 fusion transcripts by RT-PCR. All posterior fossa (PF) BEPNs were classified into PF-A and PF-B based on H3K27me3 protein expression and chromosome 1q gain. Immunohistochemistry for PD-L1 (SP263, Ventana) and CD8 was performed. RelA protein enrichment at PDL1 promoter site was analysed by chromatin immunoprecipitation-qPCR (ChIP-qPCR) in three ST-RELA samples. RESULTS A total of 83 intracranial EPNs were included as follows: ST-RELA (n=32), ST-YAP (n=1), ST-NOS (n=11), ST-subependymoma (n=1), PF-A (n=25) and PF-B (n=13). Median tumor infiltrating CD8+ cytotoxic T-lymphocyte (CTL) density was 6/mm2, and was higher in ST EPNs (median 10/mm2) as compared to PF EPNs (median 3/mm2). PD-L1 protein expression in ≥1% tumor cells was noted in 17/83 (20%) of all EPNs, including 12/32 (37%) of ST-RELA and rare ST-NOS (2/11), PF-A (2/25) and PF-B (1/13) tumors. Twelve intracranial EPNs (14%, 12/83) showed high CTL density and concurrent PD-L1 positivity, of which majority (83%, 10/12) were ST-RELA EPNs. Enrichment of RelA protein was seen at PD-L1 promoter. CONCLUSION Increased CTL densities and upregulation of PD-L1 in ST-RELA ependymomas suggests potential candidature for immunotherapy.

Medulloblastoma: Clinical presentation

Medulloblastomas present generally a-specifically as a fast-evolving posterior fossa tumor. Medical literature is poor concerning clinical features of medulloblastomas and their potential significance. In the present study, we reviewed 91 pediatric observations of medulloblastomas treated in Lille between 1997 and 2017. Clinical and epidemiological variables were collected and intercorrelated. They were also compared with anatomical and pathological findings, and outcome, with the aim of defining clinical-pathological entities. We also compared the group with 32 cases of posterior fossa ependymoma and 130 cases of cerebellar astrocytoma treated during the same period. We found that in medulloblastomas, the M/F ratio was higher and diagnostic delay was shorter than in astrocytomas. Also, the mean age was older than in ependymomas. Intracranial hypertension was constant; we further observed that altered general status was common (16.5%) and correlated with a metastatic tumor. We delineated two clusters: the “nodular” cluster, which associates young age, cerebello-pontine angle tumor, herniation, desmoplastic tumor, and tumor predisposition syndrome; and the “metastatic” cluster, which associates altered status, initial metastases, hydrocephalus, and diagnostic delay. Meticulous collection of clinical data at the initial phase is integral part of the oncological evaluation, with a search for genetic and prognostic risk factors, which then permits us to define clinical-pathological entities.

Expression and Clinical Significance of Translation Regulatory Long Non-Coding RNA 1 (TRERNA1) in Ependymomas.

Long noncoding RNAs (lncRNA) have emerged as vital molecules governing epithelial-to-mesenchymal transition (EMT) in cancers. Translation regulatory RNA 1 (TRERNA1) is one such lncRNA known to enhance the transcriptional activity of the EMT-transcription factor, Snail. We have previously demonstrated differential upregulation of EMT-transcription factors and cadherin switching across various clinico-pathologic-molecular subclasses of ependymomas (EPN). With an aim to analyze the correlation between the expression of TRERNA1 in EPNs, we performed gene expression analysis for TRERNA1 on 75 Grade II/III EPNs and correlated with tumor site, C11orf95-RELA fusions, age, MIB-1 proliferative indices, and outcome wherever available. Upregulation of gene expression levels of TRERNA1 was seen in intracranial EPNs, with highest expression levels in pediatric posterior fossa EPNs. High TRERNA1 expression was found associated with higher proliferative indices (p = 0.034) and shorter progression free survival (p = 0.002). Our study, for the first time, demonstrates an association between TRERNA1 expressions and pediatric posterior fossa EPNs. Further in-vivo and in-vitro studies are required to confirm these findings and evaluate TRERNA1 as a novel biomarker and potential therapeutic target in childhood PF-EPNs.

MYCN amplification drives an aggressive form of spinal ependymoma

Spinal ependymal tumors form a histologically and molecularly heterogeneous group of tumors with generally good prognosis. However, their treatment can be challenging if infiltration of the spinal cord or dissemination throughout the central nervous system (CNS) occurs and, in these cases, clinical outcome remains poor. Here, we describe a new and relatively rare subgroup of spinal ependymal tumors identified using DNA methylation profiling that is distinct from other molecular subgroups of ependymoma. Copy number variation plots derived from DNA methylation arrays showed MYCN amplification as a characteristic genetic alteration in all cases of our cohort (n = 13), which was subsequently validated using fluorescence in situ hybridization. The histological diagnosis was anaplastic ependymoma (WHO Grade III) in ten cases and classic ependymoma (WHO Grade II) in three cases. Histological re-evaluation in five primary tumors and seven relapses showed characteristic histological features of ependymoma, namely pseudorosettes, GFAP- and EMA positivity. Electron microscopy revealed cilia, complex intercellular junctions and intermediate filaments in a representative sample. Taking these findings into account, we suggest to designate this molecular subgroup spinal ependymoma with MYCN amplification, SP-EPN-MYCN. SP-EPN-MYCN tumors showed distinct growth patterns with intradural, extramedullary localization mostly within the thoracic and cervical spine, diffuse leptomeningeal spread throughout the whole CNS and infiltrative invasion of the spinal cord. Dissemination was observed in 100% of cases. Despite high-intensity treatment, SP-EPN-MYCN showed significantly worse median progression free survival (PFS) (17 months) and median overall survival (OS) (87 months) than all other previously described molecular spinal ependymoma subgroups. OS and PFS were similar to supratentorial ependymoma with RELA-fusion (ST-EPN-RELA) and posterior fossa ependymoma A (PF-EPN-A), further highlighting the aggressiveness of this distinct new subgroup. We, therefore, propose to establish SP-EPN-MYCN as a new molecular subgroup in ependymoma and advocate for testing newly diagnosed spinal ependymal tumors for MYCN amplification.

Radiation doses to brain substructures associated with cognition in radiotherapy of pediatric brain tumors

Background: Several brain substructures associated with cognition (BSCs) are located close to typical pediatric brain tumors. Pediatric patients therefore have considerable risks of neurocognitive impairment after brain radiotherapy. In this study, we investigated the radiation doses received by BSCs for three common locations of pediatric brain tumor entities.Material and methods: For ten patients in each group [posterior fossa ependymoma (PFE), craniopharyngioma (CP), and hemispheric ependymoma (HE)], the cumulative fraction of BSCs volumes receiving various dose levels were analyzed. We subsequently explored the differences in dose pattern between the three groups and used available dose response models from the literature to estimate treatment-induced intelligence quotient (IQ) decline.Results: Doses to BSCs were found to differ considerably between the groups, depending on their position relative to the tumor. Large inter-patient variations were observed in the ipsilateral structures of the HE groups, and at low doses for all three groups. IQ decline estimates differed depending on the model applied, presenting larger variations in the HE group.Conclusion: While there were notable differences in the dose patterns between the groups, the extent of estimated IQ decline depended more on the model applied. This inter-model variability should be considered in dose–effect assessments on cognitive outcomes of pediatric patients.