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Ependymomas encompass multiple, clinically relevant tumor types based on localization, genetic alterations, and epigenetic and transcriptomic profiles. Tumors belonging to the methylation class of spinal ependymoma (SP-EPN) represent the most common intramedullary neoplasms in children and adults. However, molecular data of SP-EPN are scarce, and clear treatment recommendations are lacking. The only known recurrent genetic events in SP-EPN are loss of chromosome 22q and NF2 mutations. Yet, it remains unclear whether SP-EPN with germline or sporadic NF2 mutations or with NF2 wild type status differ clinically or molecularly. To provide a comprehensive molecular profile of SP-EPN, we integrated epigenetic, genomic, transcriptomic, and histological analyses of up to 237 cases. Clustering of methylation data revealed two distinct molecular SP-EPN subtypes. The distribution of NF2 mutated cases differed significantly across these subtypes (p

PURPOSE Fluorescence imaging is an important surgical adjunct in malignant glioma surgery. 5-aminolevulinic acid (5-ALA) has been proven effective for radical tumor resection and extended progression-free survival in a phase III randomized trial and therefore integrated into surgery for malignant glioma. Importantly, however, some limitations still exist in its use, which include false positivity and false negativity as well as inability of re-administration. In this study, we aimed to develop a novel, spray-type fluorescent probe using hydroxymethyl rhodamine green (HMRG) as a fluorescent scaffold. METHODS We have previously established a fluorescent probe library comprised of more than 320 kinds of HMRG probes. They have HMRG as a fluorescent scaffold with various types of dipeptides attached to it. Primary probe screening was performed using the homogenized tumor samples from patients with glioblastoma operated at our institution. Secondary screening followed using the selected probes and fresh tumor samples obtained from patients with glioblastoma operated from 2016 until 2018. Diced electrophoresis gel (DEG) assay, two-dimensional gel electrophoresis followed by a multi-well plate-based fluorometric assay, was performed to identify responsible enzymes for the selected probe. Further experiments with inhibitors, real-time PCR, immunohistochemistry, and western blotting were performed for confirmation. RESULTS Proline-arginine-HMRG (PR-HMRG) was selected as a candidate probe based upon the above two-step screenings. It achieved 79.4% accuracy in receiver operating characteristic curve analysis. Calpain-1 was found to be responsible to cleave PR-HMRG probe by DEG-proteome analysis. Calpain-1 protein was expressed at significantly higher level in tumors that were fluoresced by PR-HMRG than in those that were not. CONCLUSIONS Our innovative screening method was able to find PR-HMRG as a novel fluorescent probe effective for rapid detection of glioblastoma. A preclinical study is planned to assess the efficacy and safety of the selected probe.

BACKGROUND Ependymomas are currently classified into 9 subgroups by DNA methylation profiles. Although spinal cord ependymoma (SP-EPN) is distinct from other tumors, diversity within SP-EPN is still unclear. Here, we used transcriptomic and epigenomic profiles to investigate the diversity among Japanese SP-EPN cases. MATERIALS AND METHODS We analyzed 57 SP-EPN patients (32 males and 25 females, aged from 18 to 78 years, median: 52), including two cases of neurofibromatosis type 2, five cases of grade 3 (WHO grade). We obtained transcriptome (RNA-seq) and DNA methylation (Infinium Methylation EPIC array) data from fresh frozen specimens of SP-EPN resected at the University of Tokyo Hospital and our collaborative groups. RESULTS Three cases had a previous intracranial ependymoma operation. Hierarchical clustering of the DNA methylation data showed that these three cases of intracranial origin as a different cluster from spinal origin. The 45 grade 2 spinal ependymoma showed a relatively homogenous methylation pattern. However, the methylation status of HOX gene cluster regions is compatible with the segment of origin, which reflects the cells of origins are derived after the determination of segment identity. RNA sequencing of 57 cases revealed two subgroups within grade 2. Gene ontology analysis of differentially expressed genes suggested the difference in metabolic state such as rRNA translation and mitochondrial respiration between the two expression subgroups. CONCLUSION Epigenetic analysis indicated the accurate body segment origin of SP-EPN. We observed that metabolic states could divide grade 2 spinal cord ependymoma into 2 subgroups and will present the relationship to clinicopathological information.

BACKGROUND convolutional neural network (CNN) model using MRI imaging are likely to be effective for grading glioma. However, interpretation of the judgment basis of CNN is difficult. The purpose of this study is twofold. One is to create a high accuracy machine learning model that grading of glioma (Glioma grade II, between III and IV). The other is to visualize the judgment basis of the model. METHODS We targeted cases that were imaged at our Hospital during the period from August 2014 to January 2018. Five types of MRI are used. The five types are two types of DWI (b1000DWI and b2000DWI, respectively, with a value of 1000 and 2000), apparent diffusion coefficient (ADC), fractional anisotropy (FA), and mean kurtosis (MK). The images were input to CNN wtihout ROI. Seven types of CNN were prepared, and 100 epochs of learning were performed. RESULTS 55 cases were included. There were 14 grade II, 12 grade III, and 29 grade IV). Of these, 44 cases up to July 2017 in chronological order were used as a dataset for learning, and 11 cases from August 2017 to January 2018 were taken as independent test dataset. The best results were obtained using MK as input, with a percentage of correct cases of test data of 0.82. Subsequently, we applied Grad-CAM to the model. Grad-CAM is visualized technique that shows where CNN focused on. The model focused on the tumor part of the image. In addition, in the cases of glioma and meningioma coexisting, grading was performed focusing on the glioma area not on meningioma area. CONCLUSION When grading glioma, it was considered that CNN decide glioma’s grade focusing on tumor part of the image, as well as humans.

INTRODUCTION Neuroendoscopic surgery is useful for intraventricular tumors accompanied by ventriculomegaly. However, it is often challenging for cases with small ventricles. Our institution is actively performing surgeries for pediatric intraventricular tumors without frank ventriculomegaly. METHODS Seven cases of intraventricular tumors without ventriculomegaly (5 cases of subependymal giant cell astrocytoma (SEGA) and 2 cases of germ cell tumors (GCTs)) were analyzed. The age ranged between 3 and 14 years (median 5 years). The sizes of SEGA were between 10-27mm, and all the tumors showed an enlargement around the foramen of Monro, which was the indication for surgery. Biopsy and third ventriculostomy were performed for GCTs. For resection, after making a small craniotomy of 2 x 3 cm, ellipse-cone-like sheath with a diameter of 12mm or 17mm was inserted through it to the lateral ventricle, which enabled a wide surgical view. Under a rigid endoscope of 4mm diameter, 2 types of surgical instruments were employed, making the microsurgical procedure like under a microscope, with a wider view, possible. For the cases of tumor resection, septostomy and placement of a drain in the ventricle were performed at the end of surgery. RESULTS The lesions were safely approached in all the cases. For resection, endoscopic microsurgery was possible, and tumor was totally removed in all the cases. No postoperative complication was observed in any of them. CONCLUSIONS Our experience shows that tumor resection can be safely achieved with the aid of endoscope even for cases without ventriculomegaly.

INTRODUCTION Diffuse midline glioma (DMG) mostly affects young children. The newly-introduced disease entity DMG, H3K27M-mutant uniformly portends poor prognosis, and therefore that in the pons is usually treated based upon radiological diagnosis without histological confirmation. DMG is rarer in adolescents and young adults (AYA), and remains poorly characterized. In this study, we sought to investigate the clinical, pathological, and molecular profiles of DMG in AYA generation. METHODS Patients of age between 16 and 39 undergoing biopsy at the University of Tokyo Hospital between 2003 and 2019 were included in the study. Clinical data and images were retrospectively reviewed. Genetic analyses were performed in cases with abundant tissues. RESULTS Ten patients included 8 brainstem and 2 thalamic DMG. The median age was 25 years (range, 19–38). Pathological diagnosis was DMG, H3K27M-mutant in 3 patients, glioblastoma, IDH-mutant in 1, anaplastic astrocytoma, IDH-wildtype in 4, diffuse astrocytoma, IDH-mutant in 1, and diffuse astrocytoma, IDH-wildtype in 1. Genetic analyses detected H3F3A-K27M mutation in 2, HIST1H3B-K27M mutation in 1, IDH1-R132H mutation in 1, and IDH1-R132S mutation in 1. With a median follow-up of 23 months (range, 2–61), only 3 patients died 29–61 months after diagnosis, and the remaining 7 patients survived for 2–59 months. Neither IDH1 mutation nor H3K27M mutation was associated with survival in this series. CONCLUSION Survival of AYA patients with DMG was seemingly variable with some long survivors. H3K27M mutation was present in a subset of patients. A further study is warranted to correlate molecular profile with clinical pictures including patient survival.

BACKGROUND Ependymomas commonly occur in the fourth ventricle and the spinal cord. Gross total resection, age and WHO grade are known prognostic factors. Ependymomas are currently classified into 9 distinct subgroups by DNA methylation profile analysis. Spinal cord ependymoma is distinct from other subgroups. To investigate heterogeneity within spinal cord ependymoma, we examined DNA methylation profiles. MATERIALS AND METHODS We used Infinium MethylationEPIC array (illumina) to obtain DNA methylation data from frozen specimens of spinal ependymoma resected at the University of Tokyo, Osaka City University, and Tokyo Metropolitan Neurological Hospital. Japan Pediatric Molecular Neuro-Oncology Group provided methylation data for 11 reported cases. Cluster analysis was performed using Cluster3.0. RESULTS We analyzed 34 patients, 21 male and 13 female, aged from 18 to 76 years (median 50.5 years), including 2 cases with neurofibromatosis type 2. WHO grade was grade_3 in 2 cases and grade_2 in others. Clustering of the DNA methylation data showed that WHO grade_3 cases tended to be classified into a subgroup distinct from other cases. CONCLUSION This is the largest DNA methylation profiling study on spinal cord ependymoma to date. The study may suggest a new subgroup correlated with higher WHO grade.

The systems that can objectively predict the future trends of a particular research field are always anticipated while conducting medical research. Such systems also provide a considerable aid to researchers while determining and acquiring appropriate research budgets. This study intended to establish a novel and versatile algorithm that can predict the latest trends in neuro-oncology. Seventy-nine neuro-oncological research fields were selected using computational sorting methods, such as text-mining analyses, along with 30 journals that represent the recent trends in the neuro-oncology field. Further, the annual impact (AI) for each year with respect to each journal and field (number of articles published in the journal × the impact factor of the journal) was calculated as a novel concept. Subsequently, the AI index (AII) for the year was defined as the sum of the AIs for the aforementioned 30 journals. With respect to the aforementioned neuro-oncological research fields, the AII trends from 2008 to 2017 were subjected to machine learning predicting analyses. The prediction accuracy of the latest trends in neuro-oncology was validated using actual data obtained from previous studies. In particular, the linear prediction model achieved a relatively good accuracy. The most notable and latest predicted fields in neuro-oncology included some interesting emerging fields, such as microenvironment and anti-mitosis, as well as the already renowned fields, such as immunology and epigenetics. Furthermore, we retrospectively attempted an analysis of the fields different from neuro-oncology. Interestingly, as of 2008, the future emergence of the CRISPR-Cas9 gene editing system has been predicted using this system. Overall, the presented algorithm displays potential to be an effective and versatile tool for the prediction of future trends in a particular medical field.

PURPOSE Diffusion tensor-based tractography (DTT) is a method to estimate the direction of white matter fibers, but it is difficult to verify the relationship with brain function spatially with high accuracy. We developed a registration method to fuse the real space (brain surface photograph) and preoperative fused 3D image (virtual space) using the landmark method and thin plate spline method. In our previous study, this method was able to achieve highly accurate alignment registration error 0.7±0.1mm (mean±SE) even after brain shift due to craniotomy. In this study, we proposed a method to examine spatial errors of DTT and direct cortical stimulation (DCS) and verify its accuracy. METHODS We included 7 gliomas performed awake surgery. We created the fused three – dimensional image before surgery and acquired the brain surface photograph immediately after craniotomy, then we aligned them using the proposed method. Sites that showed speech arrest by DCS were plotted on the fused image. A circle with a radius of 15 mm centered on the same site was taken as the range over which the current spreads. The surface area of each of the circles was calculated to make it true if there was arcuate fasciculus drawn with DTT in the circle, and false if it did not exist. By using this method, the accuracy of the DTT was verified. RESULT: In 7 cases, speech arrest was shown at 21 DCS plots. The probability of the presence of DTT within the current spread of DCS was 64.4%. CONCLUSION The proposed method indicates that DTT does not necessarily match the DCS results by verification using real space and virtual space. We present some illustrative cases.

Background Although inactivation of the von Hippel-Lindau gene (VHL), located on chromosome 3p25, is considered to be a major cause of hemangioblastomas (HBs), the incidence of biallelic inactivation of VHL is reportedly low. The aim of this study was to determine the prevalence of VHL alterations in HBs, as well as to identify additional molecular aberrations. Methods Genetic and epigenetic alterations were comprehensively and comparatively analyzed in 11 VHL-related and 21 sporadic HBs. Results VHL alterations detected by sequencing and multiplex ligation-dependent probe amplification (MLPA) analysis were more frequent in VHL-related HBs than in sporadic HBs (100% vs 62%; P = 0.029). VHL alterations were found only in 4 sporadic HBs by direct sequencing; however, targeted deep sequencing detected 9 additional alterations. Loss of heterozygosity (LOH) on chromosome 3 was found in 64% and 57% of VHL-related and sporadic HBs, respectively, by single nucleotide polymorphism (SNP) array analysis. Among 19 tumors with chromosome 3 LOH, 5 were classified as copy-neutral LOH. VHL promoter hypermethylation was detected only in sporadic HBs (33%), indicating that epigenetic suppression of VHL is a common mechanism in sporadic HBs. The rate of biallelic VHL inactivation among VHL-related and sporadic HBs was 64% and 52%, respectively. LOH on either chromosome 6 or 10 was detected only in sporadic HBs (43%). Conclusion Although biallelic inactivation of VHL is a dominant mechanistic cause of the pathogenesis of HB, other unknown mechanisms may also be involved, and such mechanisms may be different between VHL-related and sporadic HB.

Hemangioblastoma (HB) is a rare benign tumor of the central nervous system that usually occurs in the posterior fossa.1 About 20% of cases of intracranial HB are associated with von Hippel–Lindau disease (VHL),2 and ∼80% of cases of spinal cord HB develop in patients with VHL.3 For intracranial HB, surgical resection is an indispensable modality for lesions causing higher intracranial pressure and various associated symptoms. On the other hand, for patients with VHL who generally harbor multiple intracranial lesions, repeated surgical procedures are less agreeable and surgery should be reserved as a salvage modality. Recently, clinical studies with a large number of patients with VHL have been reported from the National Institutes of Health.4–7 In those reports, all the silent lesions were principally recommended to be observed conservatively, since HB usually shows a slow and stuttering pattern of growth.4 In addition, based on these results, the published reports at that time regarding outcomes of stereotactic radiosurgery (SRS) showed preferable results, but the follow-up periods reported were insufficient to declare “tumor control.”4,7 In fact, we have previously reported early results of SRS for 13 patients with 38 HBs and concluded that SRS would be safe and effective, although a limited number of the patients were observed over 10 years and long-term results were still unclear.8 From this point of view, the present study reconsidered the outcomes of SRS for intracranial HB, factors associated with tumor control, and adverse events with a larger number of the patients and the lesions in the longer follow-up periods.

Mutations in H3F3A, which encodes histone H3.3, commonly occur in pediatric glioblastoma. Additionally, H3F3A K27M substitutions occur in gliomas that arise at midline locations (eg, pons, thalamus, spine); moreover, this substitution occurs mainly in tumors in children and adolescents. Here, we sought to determine the association between H3F3A mutations and adult thalamic glioma.Genomic H3F3A was sequenced from 20 separate thalamic gliomas. Additionally, for 14 of the 20 gliomas, 639 genes--including cancer-related genes and chromatin-modifier genes--were sequenced, and the Infinium HumanMethylation450K BeadChip was used to examine DNA methylation across the genome.Of the 20 tumors, 18 were high-grade thalamic gliomas, and of these 18, 11 were from patients under 50 years of age (median age, 38 y; range, 17-46), and 7 were from patients over 50 years of age. The H3F3A K27M mutation was present in 10 of the 11 (91%) younger patients and absent from all 7 older patients. Additionally, H3F3A K27M was not detected in the 2 diffuse astrocytomas. Further sequencing revealed recurrent mutations in TP53, ATRX, NF1, and EGFR. Gliomas with H3F3A K27M from pediatric or young adult patients had similar, characteristic DNA methylation profiles. In contrast, thalamic gliomas with wild-type H3F3A had DNA methylation profiles similar to those of hemispheric glioblastomas.We found that high-grade thalamic gliomas from young adults, like those from children and adolescents, frequently had H3F3A K27M.

OBJECTIVE: Creating a machine learning model using diffusion tensor and kurtosis images are effective for malignancy diagnosing of glioma. However, it is unknown whether an IDH mutation, which is the most important mutation in glioma, can be predicted using diffusion tensor and kurtosis images. The purpose of our research was to predict the IDH mutation by using diffusion tensor and kurtosis images based on a machine learning. METHODS: Between August 2014 and July 2017, 38 consecutive patients with suspected glioma and preoperative MRI including diffusion coefficient (ADC), fractional anisotropy (FA), mean kurtosis (MK), were included in the study. All IDH mutations were identified by Sanger sequencing. A volume of interest (VOI) was created manually and applied to T2-weighted images, diffusion-weighted images (DWI), ADC, FA, MK. 476 per imaging sequence and 2856 features were extracted. RESULTS: 39 datasets were obtained. The number of cases for the IDH mutation and wild-type was 21 cases of the IDH1 mutation (3 diffuse astrocytoma, 8 anaplastic astrocytoma, 4 oligodendroglioma, 3 anaplastic oligodendroglioma, 3 glioblastoma, 18 cases of wild-type(2 anaplastic astrocytoma, 16 glioblastoma). An IDH2 mutation was not observed in our cases. Features were selected by using recursive feature elimination (RFE). The most accurate machine learning model was created using the extracted eight features from ADC, Fa, and MK. The best AUC was 0.95 ± 0.02 in a support vector machine (SVM). Discussion: Imaging sequences useful for the IDH1 mutation identification are ADC, MK, FA, similar to malignancy diagnosing of glioma. However, useful features were different. The machine learning models using diffusion images may be useful for identifying the IDH1 mutations.

BACKGROUND: Mutations in the H3F3A gene, which encodes histone H3.3, were recently reported in cases of pediatric glioblastoma. H3F3A K27M mutations occur in gliomas that arise at midline locations, including the pons, thalamus, and spine; moreover, this particular mutation is found mainly in tumors in children and adolescents. In this study, we aimed to determine the association between H3F3A mutations and adult thalamic glioma. METHODS: Genomic H3F3A was sequenced from 20 separate thalamic gliomas. Additionally, for 14 of the 20 gliomas, 639 genes—including cancer-related genes and chromatin-modifier genes—were sequenced, and the Infinium HumanMethylation450K BeadChip was used to examine DNA methylation across the genome. RESULTS: Of the 20 tumors, 18 were high-grade thalamic gliomas, and of these 18, 11 were from patients under 50 years of age (median age, 38 years; range, 17–46), and seven were from patients more than 50 years of age. Among the 18 high-grade thalamic gliomas, the H3F3A K27M mutation was present in 10 (91%) of the 11 tumors from patients under 50 years of age, and was absent in all seven tumors from patients over 50 years of age. The H3F3A K27M mutation was not detected in two diffuse astrocytomas. By additional sequencing, recurrent mutations were identified in TP53, ATRX, NF1 and EGFR. In addition, a KDM6A mutation was found in one case of diffuse astrocytoma and a CREBBP mutation was identified in one case of glioblastoma with the H3F3A K27M mutation. Gliomas with H3F3A K27M from pediatric or young-adult patients had similar, characteristic DNA methylation profiles. In contrast, thalamic gliomas with wild-type H3F3A had DNA methylation profiles similar to those of hemispheric glioblastomas. The patients with high-grade thalamic gliomas harboring the H3F3A K27M mutation that occur frequently in young adults had a poor prognosis similar to those with wild-type H3F3A tumors that occur mainly in older adults. CONCLUSIONS: We found that high-grade thalamic gliomas from young adults, like those from children and adolescents, frequently had H3F3A K27M. SECONDARY CATEGORY: Neuropathology & Tumor Biomarkers.