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Recently, it was reported that ependymoblastoma and embryonal tumor with abundant neuropil and true rosettes (ETANTR) show 19q13.42 amplification at a high frequency, suggesting that these tumors may constitute a single entity. As ependymoblastic rosettes are the most prominent features in both subtypes, embryonal tumor with multilayered rosettes (ETMR) was proposed, for which 19q13.42 amplification represents a specific molecular hallmark. However, ependymoblastic rosettes are not specific to ependymoblastoma and ETANTR, and are also found in a few other embryonal tumors as well as immature teratomas, and knowledge on 19q13.42 amplification in these tumors is limited. In this study, we performed fluorescence in situ hybridazation (FISH) analysis and differential polymerase chain reaction (PCR), and detected 19q13.42 amplification in three out of four ETANTR, one ependymoblastoma and one medulloepithelioma with ETANTR components, whereas none of the two atypical teratoid/rhabdoid tumors (AT/RT) with ependymoblastic rosettes nor two immature teratomas with developing neuroectodermal structures showed such amplification, suggesting that medulloepitheliomas would possibly be included in ETMR, and ependymoblastic rosettes in AT/RT do not signify that these tumors constitute ETMR. Also, we found C19MC rather than miR-371-373 was amplified in one ETANTR, suggesting that C19MC miRNA cluster seems to be more closely linked to the pathogenesis of ETMR.

Glioneuronal tumor with neuropil-like islands (GTNI) is featured by "neuropil-like islands (NIs)" within dominating astroglial components. Isocitrate dehydrogenase (IDH) mutations, particularly IDH1 R132H (G395A), are found in WHO Grade II and III diffuse gliomas as well as secondary, but not primary, glioblastomas. We reviewed 5 cases of GTNI, and assessed histology and immunohistochemistry with various antibodies, including those for IDH1 R132H, as well as direct DNA sequencing for IDH1 G395A. NIs were variable in morphology, and constantly synaptophysin-positive and glial fibrillary acidic protein-negative. The glioma components were primary glioblastoma in 2 cases, anaplastic astrocytoma in 1 and anaplastic oligoastrocytoma in 2. The IDH1 R132H was expressed in the 2 cases with oligoastrocytoma: In 1, NIs and the astrocytoma-like area as well as the oligodendroglioma-like area were positive. In the other, only the oligodendrogliomalike area was positive. The mutation analysis performed on the latter case with DNA separately sampled from the oligodendroglioma- like area and the astrocytoma-like area detected IDH1 G395A in both areas. We have shown diverse pathologic aspects of GTNI. Also, we have shown that the expression of IDH1 R132H in GTNI is largely concordant with that in diffuse gliomas, and that it can be dependent on each histologic component although the mutant IDH1 gene is ubiquitously present within the tumor.

One of the type VI intermediate filament proteins, nestin, is expressed in neuroepithelial stem cells during neural embryogenesis. Nestin is also expressed in a variety of neoplasms. Its expression in brain tumors has not been thoroughly studied. The objectives of this study were to survey nestin expression in different types of brain tumor, and to evaluate nestin as a marker for diagnosis and prognosis. We used tissue microarrays of 257 brain tumors for an immunohistochemical overview of nestin expression: nestin was frequently expressed in gliomas and schwannomas. Most of the gliomas that expressed high levels of nestin were high-grade gliomas (anaplastic astrocytomas, anaplastic oligodendrogliomas, anaplastic oligoastrocytomas, and glioblastomas). We then focused on high-grade gliomas and performed immunohistochemistry again, using whole-mount slides. As a result, we found (1) significantly different nestin expression between glioblastomas and other high-grade gliomas, and (2) worse overall survival for high-grade gliomas with high nestin expression. Our results suggest that: (1) nestin is a useful marker for diagnosis of high-grade gliomas, (2) nestin is helpful in diagnosis of schwannomas, and (3) nestin expression is related to poor prognosis in high-grade gliomas.

MicroRNAs (miRNAs) regulate a variety of cellular processes via the regulation of multiple target genes. We screened 48 medulloblastomas for mutation, deletion and amplification of nine miRNA genes that were selected on the basis of the presence of potential target sequences within the 3'-untranslated region of the MYCC mRNA. Differential PCR revealed deletions in miR-186 (15%), miR-135a-1 (33%), miR-548d-1 (42%), miR-548d-2 (21%) and miR-512-2 (33%) genes, whereas deletion or amplification was detected in miR-135b (23%) and miR-135a-2 (15%). In miR-33b, deletion, amplification or a mutation at the precursor miRNA were detected in 10% of medulloblastomas. Overall, 35/48 (73%) medulloblastomas had at least one alteration. Real-time RT-PCR revealed MYCC overexpression in 11 of 37 (30%) medulloblastomas, and there was a correlation between MYCC overexpression and miR-512-2 gene deletion (P = 0.0084). Antisense-based knockdown of miR-512-5p (mature sequence of miR-512-2) resulted in significant upregulation of MYCC expression in HeLa and A549 cells, while forced overexpression of miR-512-2 in medulloblastoma/PNET cell lines DAOY, UW-228-2, PFSK resulted in the downregulation of MYCC protein. Furthermore, the results of luciferase reporter assays suggested that miR-512-2 targets the MYCC gene. These results suggest that alterations in the miRNA genes may be an alternative mechanism leading to MYCC overexpression in medulloblastomas.

We recently reported that the vast majority (

Isocitrate dehydrogenase 1 (IDH1) mutations are common in grade II-III diffuse gliomas and secondary glioblastomas. The aim of this study is to investigate the staining pattern of mIDH1R132H, an antibody specific to mutant IDH1 protein, in primary brain tumors and non-neoplastic systemic organs. Eight of 13 diffuse astrocytomas, 1 of 6 anaplastic astrocytomas, 9 of 11 oligodendrogliomas, 15 of 22 anaplastic oligodendrogliomas, 6 of 7 oligoastrocytomas, and 5 of 8 anaplastic oligoastrocytomas showed both cytoplasmic and nuclear positivity. Two of 25 atypical meningiomas and 2 of 42 pituitary adenomas were positive for mIDH1R132H. The following non-neoplastic systemic organs showed positivity in the cytoplasm alone: the myocardium, peribronchial glands, interlobular ducts of the salivary gland, gastric fundic gland, columnar epithelia of the large bowel, hepatocytes, centroacinar cells, the intercalated ducts of the pancreas, renal proximal and distal tubules, adrenocortex, ovarian granulosa cells, and the choroid plexus epithelia. It was concluded that the immunopositivity detected in non-neoplastic systemic organs was due to cross-reactivity, because immunohistochemistry with an anti-mitochondrial antibody revealed that the mIDH1R132H staining pattern was identical to that of the mitochondria. Therefore, mIDH1R132H positivity should only be considered to be validated when a cell shows both cytoplasmic and nuclear staining.

The current World Health Organization classification recognizes three histological types of grade II low-grade diffuse glioma (diffuse astrocytoma, oligoastrocytoma, and oligodendroglioma). However, the diagnostic criteria, in particular for oligoastrocytoma, are highly subjective. The aim of our study was to establish genetic profiles for diffuse gliomas and to estimate their predictive impact. In this study, we screened 360 World Health Organization grade II gliomas for mutations in the IDH1, IDH2, and TP53 genes and for 1p/19q loss and correlated these with clinical outcome. Most tumors (86%) were characterized genetically by TP53 mutation plus IDH1/2 mutation (32%), 1p/19q loss plus IDH1/2 mutation (37%), or IDH1/2 mutation only (17%). TP53 mutations only or 1p/19q loss only was rare (2 and 3%, respectively). The median survival of patients with TP53 mutation ± IDH1/2 mutation was significantly shorter than that of patients with 1p/19q loss ± IDH1/2 mutation (51.8 months vs. 58.7 months, respectively; P = 0.0037). Multivariate analysis with adjustment for age and treatment confirmed these results (P = 0.0087) and also revealed that TP53 mutation is a significant prognostic marker for shorter survival (P = 0.0005) and 1p/19q loss for longer survival (P = 0.0002), while IDH1/2 mutations are not prognostic (P = 0.8737). The molecular classification on the basis of IDH1/2 mutation, TP53 mutation, and 1p/19q loss has power similar to histological classification and avoids the ambiguity inherent to the diagnosis of oligoastrocytoma.

The objective of this study was to evaluate, in a series of 43 pediatric high-grade gliomas (21 anaplastic astrocytoma WHO grade III and 22 glioblastoma WHO grade IV), the prognostic value of histological grading and expression of p53 and YKL-40. Moreover, mutational screening for TP53 and IDH1 was performed in 27 of 43 cases. The prognostic stratification for histological grading showed no difference in overall (OS) and progression-free survival (PFS) between glioblastomas and anaplastic astrocytomas. Overexpression of YKL40 was detected in 25 of 43 (58%) cases, but YKL-40 expression was not prognostic in terms of OS and PFS. p53 protein expression was observed in 13 of 43 (31%) cases but was not prognostic. TP53 mutations were detected in five of 27 (18%) cases (four glioblastomas and one anaplastic astrocytoma). Patients with TP53 mutation had a shorter median OS (9 months) and PFS (8 months) than those without mutations (OS, 17 months; PFS, 16 months), although this trend did not reach statistical significance (p = 0.07). IDH1 mutations were not detected in any of the cases analyzed. Our results suggest that in pediatric high-grade gliomas: (i) histological grading does not have strong prognostic significance, (ii) YKL-40 overexpression is less frequent than adult high-grade gliomas and does not correlate with a more aggressive behavior, (iii) TP53 mutations but not p53 expression may correlate with a more aggressive behavior, and (iv) IDH1 mutations are absent. These observations support the concept that, despite identical histological features, the biology of high-grade gliomas in children differs from that in adults, and therefore different prognostic factors are needed.

Epithelioid glioblastomas (E-GBMs) are rare, highly aggressive tumors consisting of closely packed tumor cells with smooth, round cell borders and abundant eosinophilic cytoplasm. They tend to affect younger patients compared with conventional GBM. BRAF V600E mutation is characteristically found in approximately 50% of all E-GBMs, compared with a low frequency of this mutation in conventional GBM. Here, we report an unusual case of glioma involving the right frontal lobe, basal ganglia and thalamus in an HIV-positive 30-year-old man on antiretroviral therapy. The lesion was composed of abundant discohesive, monotonous epithelioid cells with extensive necrosis, spindle and polyhedral cells, low-grade oligoastrocytoma-like areas, sarcomatous components, and numerous osteoclast-like giant cells (OLGCs) intermingled with epithelioid tumor cells. As the epithelioid cells accounted for more than one-third of the tumor, a pathological diagnosis of E-GBM was made. BRAF V600E mutation was detected in both oligoastrocytoma-like and epithelioid cell components. Similar to previously reported findings on E-GBM associated with low-grade glioma, this case suggested that low-grade astrocytic glioma with BRAF V600E mutation progressed to E-GBM. OLGCs are rarely observed in gliomas, and this is the first case report of E-GBM associated with abundant OLGC infiltration.

The pathogenesis of sarcomatous component in spindle cell carcinoma (SpCC) of the esophagus is unclear. To investigate the involvement of epithelial-mesenchymal transition (EMT) in sarcomatous differentiation, we performed immunohistochemistry for Slug, Twist, ZEB1, and ZEB2, transcription factors associated with EMT and E-cadherin, in 14 cases of SpCC of the esophagus. In order to verify the neoplastic nature of sarcomatous components, TP53 mutation status and protein expression were examined in each case. Nuclear ZEB1 expression was extensive in the sarcomatous component, greater than invasive front of carcinoma components (P0.0001). Membranous E-cadherin expression was mostly lost in sarcomatous cells in all cases (P0.0001). The p53 expression pattern was almost concordant between the two areas in all cases. TP53 mutation analysis revealed that seven cases harbored identical mutations in both components. One case had mutations only in the sarcomatous component. It is noteworthy that none of them harbored mutation in exon 5, unlike conventional esophageal squamous cell carcinoma. These findings show that ZEB1 are widely expressed in the sarcomatous area of SpCC of the esophagus, suggesting the involvement of EMT. The avoidance of exon 5 in terms of TP53 mutation may also be a feature of the tumor.

Epithelioid glioblastomas are one of the rarest histological variants of glioblastomas, which are not formally recognized by the World Health Organization (WHO) classification. Epithelioid glioblastomas usually occur as primary lesions, but there have been several reports of secondary epithelioid glioblastomas or epithelioid glioblastomas with pre- or co-existing lesions to date. The serine/threonine-protein kinase B-Raf (BRAF) V600E mutation has been found at a high frequency of 54% in epithelioid glioblastomas. We present a case of a 26-year-old female patient with an epithelioid glioblastoma with the BRAF V600E mutation in her right frontal lobe. In the present case, a low-grade diffuse astrocytoma component had colocalized with the epithelioid glioblastoma. The component presented prominent calcification on neuroimages as well as by histology, and low-grade diffuse astrocytoma was considered to be a precursor lesion of an epithelioid glioblastoma. However, the BRAF V600E mutation was detected only in epithelioid glioblastoma but not in low-grade diffuse astrocytoma. To the best of our knowledge, this is the first report demonstrating a discrepancy in the BRAF V600E mutation states between epithelioid glioblastoma and colocalized low-grade astrocytoma.

Background There are few previous reports of glioblastoma in young adults, in particular, of the very rare recently proposed rhabdoid or epithelioid types. Case Description We report 2 cases of glioblastoma with rhabdoid features involving a 27-year-old woman and a 41-year-old man. In case 1, the patient presented with generalized seizures, and the initial magnetic resonance imaging showed a very small region of contrast in the left parahippocampal area. After 1 year, the mass suddenly increased in size. Treatment with multiple therapies was administered, but 5 months later, the patient died from multiple systemic metastases. In case 2, the patient presented with a chief complaint of headache for a few weeks. Computed tomography and magnetic resonance imaging showed a left parietal mass with calcification and meningeal dissemination. After undergoing surgical removal, his condition rapidly deteriorated until brain death occurred. Conclusions Glioblastoma with rhabdoid features may represent a specific pattern of clinical progression that emerges from histologic glioblastoma types.

We investigated the fused protein of solute carrier family 44 choline transporter member 1 (SLC44A1) and protein kinase C alpha (PRKCA) in three patients with papillary glioneuronal tumors (PGNT). PGNT and rosette-forming glioneuronal tumors (RGNT) are recently identified, unusual glioneuronal tumor variants which were categorized as novel tumor entities in the 2007 World Health Organization classification system. The molecular background of these tumors remains poorly understood due to the paucity of studies. The SLC44A1-PRKCA fusion was recently detected in three cases of PGNT. We invesitgated for the SLC44A1-PRKCA fusion protein in the three PGNT patients and a further two with RGNT using fluorescence in situ hybridization. Two out of the three PGNT patients had a fused signal (paired red-green signal) representing a rearrangement on chromosomes 9 and 17. A normal signal pattern was observed in the third PGNT patient. Neither of the two RGNT patients demonstrated a fused signal. This suggests that the SLC44A1-PRKCA fusion is a characteristic alteration in PGNT but not RGNT. Therefore, it is a potential biomarker of PGNT. The paired red-green signal that was observed in the PGNT patients implies the presence of a different breakpoint than that previously reported in the 9q31 and 17q24 genes.

Meningiomas show a diverse histopathologic appearance, often referred to as metaplastic changes; however, adenocarcinoma-like metaplasia is an extremely rare condition. Here, we present a novel case. A dura-based bulky mass located in the right frontotemporal region was identified radiologically in an 83-year-old woman. The tumor, yellow to ash-gray in color, was subtotally removed. Histopathological examination revealed robust adenocarcinoma-like structures within a conventional meningothelial neoplasm. Meningioma elements showed a WHO grade I to III histology. Morphological and immunophenotypic transition between meningothelial and columnar epithelial cells was confirmed on detailed observation. It was of note that the adenocarcinomatous components shared an immunophenotype with intestinal epithelium, expressing CDX2, MUC2 and cytokeratin 20. The present case could be differentiated from secretory meningioma based on distinct cellular atypia, lack of intracytoplasmic lumina and pseudosammoma bodies, and the intact status of the KLF4 gene. In addition, the morphological and immunophenotypic transition excluded the possibility of metastatic carcinoma within meningioma. This is the first reported case of meningioma with adenocarcinoma-like metaplasia harboring an intestinal immunophenotype.

Embryonal tumor with abundant neuropil and true rosettes (ETANTR) is a very aggressive embryonal central nervous system (CNS) tumor, histologically featuring ependymoblastic rosettes and neuronal differentiation in a neuropil-like background. 19q13.42 amplification was identified in ETANTR and epndymoblastoma, suggesting that these tumors constitute a single entity, called embryonal tumor with multilayered rosettes (ETMR). Here, we report a case involving a 2-year-old boy with a pontine embryonal tumor composed of clusters of poorly differentiated neuroepithelial cells, and smaller neuroblastic/neurocytic cells in a fibrillary and paucicellular neuropil-like matrix, where clear ependymoblastic rosettes were not detected but only one structure suggestive of an ependymoblastic multilayered rosette was found. Fluorescence in situ hybridazation analysis revealed 19q13.42 amplification, supporting the diagnosis of ETANTR. This report indicates that rare ependymoblasic rosettes found in embryonal tumors, which are otherwise CNS primitive neuroectodermal tumors or medulloblastomas, are significant for considering the examination of 19q13.42 amplification to confirm the diagnosis of ETMR.

Pleomorphic xanthoastrocytoma (PXA) is classified by the World Health Organization as a grade II astrocytic tumor with relatively favorable prognosis among gliomas. A valine-to-glutamic acid substitution at position 600 of the serine/threonine-protein kinase BRAF (BRAF V600E) mutation, which is commonly found in PXA, has recently been detected in approximately 50 % of all epithelioid glioblastoma (GBM) cases. We herein report a case of epithelioid GBM developing at the site of a left temporal PXA 13 years after the treatment of the primary tumor. The BRAF V600E mutation was detected in both tumors. These findings suggest that epithelioid GBM may arise from a PXA with a BRAF V600E mutation.

Epithelioid glioblastoma is among the rarest variants of glioblastoma and is not formally recognized in the World Health Organization classification; it is composed of monotonous, discohesive sheets of small, round cells with eccentric nuclei and eosinophilic cytoplasm devoid of cytoplasmic stellate processes, showing the retention of nuclear staining of INI‐1 protein. Here, we report a case involving a 22‐year‐old man with a right occipital lobe tumor, which comprised mainly epithelioid tumor cells with a small area of diffusely infiltrating less atypical astrocytoma cells showing a lower cell density. Array comparative genomic hybridization separately performed for each histologically distinct component demonstrated eight shared copy number alterations (CNAs) and three CNAs observed only in epithelioid cells; one of the latter was a homozygous deletion of a tumor suppressor gene, LSAMP, at 3q13.31. BRAF V600E mutation was observed both in epithelioid tumor cells and in diffusely infiltrating less atypical astrocytoma cells. Our findings suggest that the regional loss of LSAMP led to the aggressive nature of epithelioid cells in the present case of epithelioid glioblastoma.

Angiomatous meningiomas are rare meningioma subtypes, which are characterized by abundant, well-formed vessels. We encountered two cases of newly diagnosed angiomatous meningiomas exhibiting tumor cells with brown pigments, which were histochemically proven to be iron. In an attempt to understand its pathological significance, we assessed this unusual finding in representatives for each grade of meningiomas and immunoexpression of transferrin receptor (CD71) and the oxidative DNA damage marker, 8-hydroxy-2'-deoxyguanosine (8-OHdG). Iron deposition in the tumor cells was observed in 8/15 (53%) angiomatous meningioma cases, 2/6 (33%) microcystic meningiomas and 2/20 (10%) meningothelial meningiomas, which included clustered microvessels, but not in fibrous, atypical or anaplastic meningiomas (P = 0.001). Cytoplasmic CD71 expression was largely negative in angiomatous meningioma cases, but positive in meningothelial and high-grade meningiomas, suggesting that the transferrin-dependent iron transporter was involved in iron uptake in meningiomas. Nuclear expression of 8-OHdG was observed in ≥ 50% of the tumor cells in all 15 cases of angiomatous meningioma and was associated with the presence of regressive histopathological findings, such as hyalinized vessels and cystic changes. In addition, the fraction of iron-containing tumor cells was correlated to those expressing 8-OHdG (P = 0.005). Our finding indicates that cytoplasmic iron deposition in tumor cells is characteristic of highly vascularized benign meningiomas and related to increased oxidative DNA damage markers.

Angiomatous meningiomas are rare meningioma subtypes, which are characterized by abundant, well-formed vessels. We encountered two cases of newly diagnosed angiomatous meningiomas exhibiting tumor cells with brown pigments, which were histochemically proven to be iron. In an attempt to understand its pathological significance, we assessed this unusual finding in representatives for each grade of meningiomas and immunoexpression of transferrin receptor (CD71) and the oxidative DNA damage marker, 8-hydroxy-2'-deoxyguanosine (8-OHdG). Iron deposition in the tumor cells was observed in 8/15 (53%) angiomatous meningioma cases, 2/6 (33%) microcystic meningiomas and 2/20 (10%) meningothelial meningiomas, which included clustered microvessels, but not in fibrous, atypical or anaplastic meningiomas (P = 0.001). Cytoplasmic CD71 expression was largely negative in angiomatous meningioma cases, but positive in meningothelial and high-grade meningiomas, suggesting that the transferrin-dependent iron transporter was involved in iron uptake in meningiomas. Nuclear expression of 8-OHdG was observed in ≥ 50% of the tumor cells in all 15 cases of angiomatous meningioma and was associated with the presence of regressive histopathological findings, such as hyalinized vessels and cystic changes. In addition, the fraction of iron-containing tumor cells was correlated to those expressing 8-OHdG (P = 0.005). Our finding indicates that cytoplasmic iron deposition in tumor cells is characteristic of highly vascularized benign meningiomas and related to increased oxidative DNA damage markers.

Primary hepatic mucosa-associated lymphoid tissue (MALT) lymphoma is extremely rare, and the etiology of disease is not fully understood. We present herein the case of a primary hepatic MALT lymphoma with Helicobacter pylori and hepatitis C virus infection. A 71-year-old male was admitted to our institution to undergo a precise evaluation of a hepatic tumor incidentally detected during a computed tomography (CT) scan for chest examination. Dynamic CT showed faint enhancement during the arterial phase. The gadoxetate disodium-enhanced magnetic resonance imaging showed a hyper-intensity on the arterial phase and low intensity during the late and hepatocyte phases. Liver biopsy specimen showed small to intermediate size atypical lymphocytes with positive CD20 immunohistochemical staining. It was finally diagnosed as primary hepatic MALT lymphoma. FDG-PET/CT showed faintly increased uptake with a maximum standardized uptake value of 4.6, and did not show other pathological uptake. We present the rare case of primary hepatic MALT lymphoma and discussed the etiology of this disease.

Astroblastoma is a rare glial tumor of unknown origin, usually affecting the cerebral hemispheres of children and young adults. Here we report an unusual cerebral tumor in a 60-year-old woman. On MRI, the tumor appeared as a well circumscribed lesion in the left frontal lobe. Histopathologically, it was composed of rounded eosinophilic cells, and was divisible into two areas. One area was characterized by a collection of GFAP-positive cells around sclerotic blood vessels (astroblastic pseudorosettes and perivascular hyalinization), and had a Ki-67 labeling index of 2.8%. However, the other area was highly cellular, showing many GFAP-negative cells often with a rhabdoid appearance, mitoses and a Ki-67 index of 15.7%. Thus, a final diagnosis of malignant astroblastoma was made. In both areas of the tumor, nearly all the cells were positive for epithelial membrane antigen, and many were positive for oligodendrocyte transcription factor 2 (Olig2). Focal expression of cytokeratin was also evident. With regard to genetic markers, the tumor cells were positive for INI1 and negative for mutant IDH1. The p53 labeling index was1%. Ultrastructurally, the presence of intra- and intercellular lumina with microvilli was a feature. DNA examination of IDH1/2 and TP53 showed no mutations. In conclusion, although ependymal features were evident ultrastructurally in the present tumor, the immunohistochemical expression pattern of Olig2 was that of diffuse astrocytoma. On the other hand, the absence of mutations in both IDH1/2 and TP53 suggested that the present tumor was not a purely astrocytic neoplasm. Further studies, including molecular and genetic analyses, will provide insight into the histogenesis of astroblastoma.

Recently, it was reported that ependymoblastoma and embryonal tumor with abundant neuropil and true rosettes (ETANTR) show 19q13.42 amplification at a high frequency, suggesting that these tumors may constitute a single entity. As ependymoblastic rosettes are the most prominent features in both subtypes, embryonal tumor with multilayered rosettes (ETMR) was proposed, for which 19q13.42 amplification represents a specific molecular hallmark. However, ependymoblastic rosettes are not specific to ependymoblastoma and ETANTR, and are also found in a few other embryonal tumors as well as immature teratomas, and knowledge on 19q13.42 amplification in these tumors is limited. In this study, we performed fluorescence in situ hybridazation (FISH) analysis and differential polymerase chain reaction (PCR), and detected 19q13.42 amplification in three out of four ETANTR, one ependymoblastoma and one medulloepithelioma with ETANTR components, whereas none of the two atypical teratoid/rhabdoid tumors (AT/RT) with ependymoblastic rosettes nor two immature teratomas with developing neuroectodermal structures showed such amplification, suggesting that medulloepitheliomas would possibly be included in ETMR, and ependymoblastic rosettes in AT/RT do not signify that these tumors constitute ETMR. Also, we found C19MC rather than miR‐371‐373 was amplified in one ETANTR, suggesting that C19MC miRNA cluster seems to be more closely linked to the pathogenesis of ETMR.

Papillary tumor of the pineal region (PTPR) is a recently described neoplasm. Several studies have been published on this tumor, but its pathological features and the appropriate treatment remain unclear. PTPR is reported to originate from ependymal cells in the subcommissural organ. Isocitrate dehydrogenase 1 and 2 (IDH1/2) mutations have been a focus area in glioma research as promising predictors. We report a case of PTPR that was characterized by local recurrence, although subtotal removal and radiotherapy seemed effective for many months. Histological examination showed ependymal features in the surgical specimens. As far as we are aware, this case study is the first to show that the IDH1/2 genotypes in PTPR cells are wild-type genotypes, which is consistent with the negative immunoreactivity that was observed for the IDH1 mutant antibody in this study.

Mutations in the genes encoding isocitrate dehydrogenase (IDH) 1/2 have been detected in a significant proportion of diffuse gliomas and in a small fraction of acute myeloid leukemia (AML) cases. Recently, in an examination of various types of mesenchymal tumor, IDH1/2 mutations were only found in cartilaginous tumors including central conventional and periosteal enchondromas/chondrosarcomas. The frequency of IDH1/2 mutations was 56%, and the IDH1 R132C mutation, which is not common in diffuse gliomas or AML, accounted for 40% of these mutations. In this study, we investigated the IDH1/2 mutation status of intracranial chondrosarcomas and chordomas, which are morphologically similar and affect similar regions of the cranial cavity. Of the 13 chondrosarcomas analyzed, six (46.1%) displayed IDH1/2 mutations (the predominant type was IDH1 R132C). Also, an IDH2 mutation (R172S) was observed in one case. Conversely, none of the ten chordomas analyzed displayed any IDH1 or IDH2 mutations. Our data suggest that the IDH1/2 mutation status could be valuable for distinguishing intracranial chondrosarcomas from chordomas.

The NADP+-dependent isocitrate dehydrogenases 1 and 2 (IDH1/2) catalyze the oxidative decarboxylation of isocitrate into α-ketoglutarate (α-KG). IDH1 and IDH2 mutations have been frequently found in some types of gliomas (low-grade diffuse gliomas WHO grade II, anaplastic gliomas WHO grade III, and secondary glioblastomas WHO grade IV), and have received significant attention because of their specificity to single codons. Since the unveiling of IDH1/2 mutations, many studies have investigated their clinical impact on gliomas. While the favorable influence of these mutations in high-grade gliomas has been well established, their prognostic impact on low-grade diffuse gliomas is much less clear. While the mechanism of IDH1/2 mutations in gliomagenesis remains to be clarified, its elucidation might lead to novel therapeutic strategies against gliomas.

Neuronal differentiation of oligodendroglioma has been demonstrated by immunohistochemical and ultrastructural examinations in recent studies. However, oligodendrogliomas displaying a complete neurocytic morphology or even gangliocytic differentiation are rare. We describe a case of anaplastic oligodendroglioma that was characterized by the presence of ganglion cells in a 40-year-old-male. Histologically, the tumor was mainly composed of classical oligodendroglioma cells. The most exceptional finding of this tumor was the presence of ganglion cells and intermediate-sized ganglioid cells. Immunohistochemical analysis revealed that these cells were positive for Olig2 and negative for glial fibrillary acid protein (GFAP). Synaptophysin and microtubule-associated protein 2 (MAP2) were mainly detected in the ganglion cells. Fluorescence in situ hybridization analysis (FISH) revealed the deletion of the 1p and 19q chromosome arms in both the oligodendroglioma cells and ganglion cells. The R132H mutated isocitrate dehydrogenase 1 (IDH1) protein was detected by immunohistochemistry and direct DNA sequencing. The morphological, immunohistochemical, and genetic features of the tumor suggested a diagnosis of anaplastic oligodendroglioma, and this tumor was considered to be a rare form of oligodendroglioma displaying ganglioglioma-like maturation. FISH and mutant IDH1 examinations are useful diagnostic tools for the differential diagnosis of this tumor, i.e., ganglioglioma with anaplastic oligodendroglial features.

MicroRNAs (miRNAs) regulate a variety of cellular processes via the regulation of multiple target genes. We screened 48 medulloblastomas for mutation, deletion and amplification of nine miRNA genes that were selected on the basis of the presence of potential target sequences within the 3′‐untranslated region of the MYCC mRNA. Differential PCR revealed deletions in miR‐186 (15%), miR‐135a‐1 (33%), miR‐548d‐1 (42%), miR‐548d‐2 (21%) and miR‐512‐2 (33%) genes, whereas deletion or amplification was detected in miR‐135b (23%) and miR‐135a‐2 (15%). In miR‐33b, deletion, amplification or a mutation at the precursor miRNA were detected in 10% of medulloblastomas. Overall, 35/48 (73%) medulloblastomas had at least one alteration. Real‐time RT‐PCR revealed MYCC overexpression in 11 of 37 (30%) medulloblastomas, and there was a correlation between MYCC overexpression and miR‐512‐2 gene deletion (P = 0.0084). Antisense‐based knockdown of miR‐512‐5p (mature sequence of miR‐512‐2) resulted in significant upregulation of MYCC expression in HeLa and A549 cells, while forced overexpression of miR‐512‐2 in medulloblastoma/PNET cell lines DAOY, UW‐228‐2, PFSK resulted in the downregulation of MYCC protein. Furthermore, the results of luciferase reporter assays suggested that miR‐512‐2 targets the MYCC gene. These results suggest that alterations in the miRNA genes may be an alternative mechanism leading to MYCC overexpression in medulloblastomas.

We recently reported that the vast majority (90%) of low-grade diffuse gliomas (diffuse astrocytoma, oligoastrocytoma and oligodendroglioma) carry at least one of the following genetic alterations: IDH1/2 mutation, TP53 mutation or 1p/19q loss. Only 7% of cases were triple-negative (ie, lacking any of these alterations). In the present study, array comparative genomic hybridization (CGH) in 15 triple-negative WHO grade II gliomas (eight diffuse astrocytomas and seven oligodendrogliomas) showed loss at 9p21 (p14(ARF) , p15(INK4b) , p16(INK4a) loci) and 13q14-13q32 (containing the RB1 locus) in three and two cases, respectively. Further analyses in 31 triple-negative cases as well as a total of 160 non-triple-negative cases revealed that alterations in the RB1 pathway (homozygous deletion and promoter methylation of the p15(INK4b) , p16(INK4a) and RB1 genes) were significantly more frequent in triple-negative (26%) than in non-triple-negative cases (11%; P = 0.0371). Multivariate analysis after adjustment for age, histology and treatment showed that RB1 pathway alterations were significantly associated with unfavorable outcome for patients with low-grade diffuse glioma [hazard ratio, 3.024 (1.279-6.631); P = 0.0057]. These results suggest that a fraction of low-grade diffuse gliomas lacking common genetic alterations may develop through a distinct genetic pathway, which may include loss of cell-cycle control regulated by the RB1 pathway.

The aim of this study was to establish the frequency of amplification of tyrosine kinase receptor genes PDGFRA, KIT and KDR (VEGFR2) at 4q12 in glioblastomas at a population level, and to assess whether such alterations have any clinical impact. Screening of 390 glioblastomas from a population-based study by differential PCR revealed amplification of the PDGFRA, KIT and KDR genes in 33 (8.5%), 17 (4.4%) and 13 (3.3%) glioblastomas, respectively. None of these alterations was prognostic for overall survival. Patients with glioblastoma showing KIT amplification were significantly younger than those with glioblastoma showing no amplification (51.7 ± 21.7 years vs. 59.3 ± 13.1 years; P=0.0231). Twelve glioblastomas showed concurrent amplification of the PDGFRA, KIT and KDR genes, whereas 18 glioblastomas showed PDGFRA amplification only. A significant inverse association was observed between KIT amplification and EGFR amplification (P=0.0260), whereas a borderline positive association was found between KIT amplification and TP53 mutation (P=0.0579). No significant difference was observed in the frequency of amplification of these genes in primary and secondary glioblastomas or in glioblastomas with and without IDH1 mutations, suggesting that amplification of PDGFRA, KIT and KDR may be implicated in the pathogenesis of a small fraction of both subtypes of glioblastoma.

Glioblastomas are morphologically and genetically heterogeneous, but little is known about the regional patterns of genomic imbalance within glioblastomas. We recently established a reliable whole genome amplification (WGA) method to randomly amplify DNA from paraffin-embedded histological sections with minimum amplification bias [Huang et al (J Mol Diagn 11: 109-116, 2009)]. In this study, chromosomal imbalance was assessed by array comparative genomic hybridization (CGH; Agilent 105K, Agilent Technologies, Santa Clara, CA, USA), using WGA-DNA from two to five separate tumor areas of 14 primary glioblastomas (total, 41 tumor areas). Chromosomal imbalances significantly differed among glioblastomas; the only alterations that were observed inor =6 cases were loss of chromosome 10q, gain at 7p and loss of 10p. Genetic alterations common to all areas analyzed within a single tumor included gains at 1q32.1 (PIK3C2B, MDM4), 4q11-q12 (KIT, PDGFRA), 7p12.1-11.2 (EGFR), 12q13.3-12q14.1 (GLI1, CDK4) and 12q15 (MDM2), and loss at 9p21.1-24.3 (p16(INK4a)/p14(ARF)), 10p15.3-q26.3 (PTEN, etc.) and 13q12.11-q34 (SPRY2, RB1). These are likely to be causative in the pathogenesis of glioblastomas (driver mutations). In addition, there were numerous tumor area-specific genomic imbalances, which may be either nonfunctional (passenger mutations) or functional, but constitute secondary events reflecting progressive genomic instability, a hallmark of glioblastomas.

Purpose: To establish the frequency of IDH1 mutations in glioblastomas at a population level, and to assess whether they allow reliable discrimination between primary (de novo) glioblastomas and secondary glioblastomas that progressed from low-grade or anaplastic astrocytoma. Experimental Design: We screened glioblastomas from a population-based study for IDH1 mutations and correlated them with clinical data and other genetic alterations. Results: IDH1 mutations were detected in 36 of 407 glioblastomas (8.8%). Glioblastoma patients with IDH1 mutations were younger (mean, 47.9 years) than those with EGFR amplification (60.9 years) and were associated with significantly longer survival (mean, 27.1 versus 11.3 months; P < 0.0001). IDH1 mutations were frequent in glioblastomas diagnosed as secondary (22 of 30; 73%), but rare in primary glioblastomas (14 of 377; 3.7%: P < 0.0001). IDH1 mutations as genetic marker of secondary glioblastoma corresponded to the respective clinical diagnosis in 95% of cases. Glioblastomas with IDH1 mutation diagnosed as primary had clinical and genetic profiles similar to those of secondary glioblastomas, suggesting that they may have rapidly progressed from a less malignant precursor lesion that escaped clinical diagnosis and were thus misclassified as primary. Conversely, glioblastomas without IDH1 mutations clinically diagnosed as secondary typically developed from anaplastic rather than low-grade gliomas, suggesting that at least some were actually primary glioblastomas, that may have been misclassified, possibly due to histologic sampling error. Conclusion: IDH1 mutations are a strong predictor of a more favorable prognosis and a highly selective molecular marker of secondary glioblastomas that complements clinical criteria for distinguishing them from primary glioblastomas. (Clin Cancer Res 2009;15(19):6002–7)

We established a line of transgenic rats expressing v-erbB, the viral form of EGFR, under the transcriptional regulation of the S100beta promoter. This is the first transgenic rat model that spontaneously develops brain tumors. The rats maintained under conventional housing conditions until the occurrence of severe neurological symptoms or death. Of the 54 rats analyzed, 52 (96%) developed brain tumors. Histopathologically, they were classified into four subtypes: glioblastoma-like malignant glioma, anaplastic oligodendroglioma, low-grade oligodendroglioma, and low-grade astrocytoma. Eleven transgenic rats developed two different histological types of brain tumor, which were considered to be of multiclonal origin, because of the lack of mutual histological transitions. Almost all brain tumors contained neoplastic cells immunoreactive for S100 and GFAP. Diffuse immunoreactivity for Olig2 and Nkx2.2 was observed in neoplastic cells in all oligodendroglial tumors, but not in the malignant gliomas. Electron microscopy revealed the presence of intermediate filament bundles devoid of side arms, indicating glial differentiation. There was no evidence of cilia, microvilli, cell junction, neurosecretory granules, synaptic structures, or neurofilaments, excluding the possibility of ependymal or neuronal tumors. Reproducible development of particular histologic types of brain tumors in unique localizations might be useful for further study on the histogenesis and molecular mechanisms of development of glial tumors due to disruption of the EGFR pathway.

IDH1 encodes isocitrate dehydrogenase 1, which participates in the citric acid cycle and was recently reported to be mutated in 12% of glioblastomas. We assessed IDH1 mutations in 321 gliomas of various histological types and biological behaviors. A total of 130 IDH1 mutations was detected, and all were located at amino acid residue 132. Of these, 91% were G--A mutations (Arg--His). IDH1 mutations were frequent in low-grade diffuse astrocytomas (88%) and in secondary glioblastomas that developed through progression from low-grade diffuse or anaplastic astrocytoma (82%). Similarly, high frequencies of IDH1 mutations were found in oligodendrogliomas (79%) and oligoastrocytomas (94%). Analyses of multiple biopsies from the same patient (51 cases) showed that there were no cases in which an IDH1 mutation occurred after the acquisition of either a TP53 mutation or loss of 1p/19q, suggesting that IDH1 mutations are very early events in gliomagenesis and may affect a common glial precursor cell population. IDH1 mutations were co-present with TP53 mutations in 63% of low-grade diffuse astrocytomas and with loss of heterozygosity 1p/19q in 64% of oligodendrogliomas; they were rare in pilocytic astrocytomas (10%) and primary glioblastomas (5%) and absent in ependymomas. The frequent presence of IDH1 mutations in secondary glioblastomas and their near-complete absence in primary glioblastomas reinforce the concept that despite their histological similarities, these subtypes are genetically and clinically distinct entities.

Nijmegen breakage syndrome caused by NBS1 germline mutations is a rare autosomal recessive disease with clinical features that include microcephaly, increased radiosensitivity, and predisposition to cancer. NBS1 plays a key role in DNA double-strand break repair and the maintenance of genomic stability. We screened 87 glioblastomas for NBS1 mutations (all 16 exons). Single-strand conformation polymorphism followed by direct DNA sequencing revealed 12 NBS1 mutations (8 missense and 4 intronic mutations) in 9 (32%) of 28 primary (de novo) glioblastomas carrying 2 or more TP53 mutations. None of the NBS1 mutations has been previously reported as a germline mutation in Nijmegen breakage syndrome patients. NBS1 mutations were not detected in 19 primary glioblastomas with 1 TP53 mutation or in 21 primary glioblastomas without TP53 mutations. Secondary glioblastomas that developed through progression from low-grade or anaplastic astrocytoma had TP53 mutations in 16 (84%) of 19 cases, but none contained mutations of the NBS1 gene. These results suggest that multiple TP53 mutations in glioblastomas are due to deficient repair of DNA double-strand breaks caused by mutational inactivation of the NBS1 gene.

Multinodular and vacuolating neuronal tumors (MVNT) have been recently referred to as a distinctive neuronal tumor entity based on histopathological findings. They are characterized by multiple tumor nodules, vacuolar alteration and widespread immunolabeling for human neuronal protein HuC/HuD. Only 13 cases have been reported in the literature to date and little is known about the histopathology of these tumors. Herein, we report a case of MVNT with additional confirmation of immunohistochemical features. A 22-year-old woman presented with a continuous headache. MRI showed a subcortical white matter lesion with multiple satellite nodules in the frontal lobe appearing as T2/fluid-attenuated inversion recovery (FLAIR) hyperintensities. Histological examination of the resected lesion revealed well-defined multiple nodules composed of predominant vacuolating tumor cells. The tumor cells exhibited consistent immunolabeling for doublecortin, as well as HuC/HuD, both representative neuronal biomarkers associated with earlier stages of neuronal development. Immunopositivity for oligodendrocyte transcription factor 2 (Olig2) and S100 was also detected in tumor cells. Additionally, significant overexpression of alpha-internexin was observed in the background neuropil limited to tumor nodules. Neuronal nuclear antigen (NeuN), synaptophysin and neurofilament, markers for mature neurons, were either negative or weakly positive. The expression profile of neuronal biomarkers can be distinguished from that of classic neuronal tumors and is the immunohistochemical hallmark of MVNT. In summary, we identified the characteristic tumoral expression of HuC/HuD and doublecortin and the presence of abundant neuropil localized in MVNT tumor nodules, which exhibited widespread alpha-internexin expression. These results supported the presumption that MVNT is a distinct histopathological entity.

Olig2 is a recently identified transcription factor involved in the phenotype definition of cells in the oligodendroglial lineage. The expression of Olig2 transcript has been demonstrated in human oligodendroglial tumors, although the protein expression has not been studied extensively. We developed a polyclonal antibody to human Olig2 and analyzed it immunohistochemically. The antibody depicted a single distinct band of predicted molecular weight by Western blotting, and did not cross-react with human Olig1. In normal human brain tissue, the nuclei of oligodendrocytes of interfascicular, perivascular, and perineuronal disposition were clearly labeled by the antibody. Similarly, the nuclei of oligodendroglial tumors were labeled. There was no apparent correlation between the staining intensity and histological grade. Astrocytic components within the tumors were generally less or not stained. Astrocytic tumors were also positive with the Olig2 antiserum to a lesser extent, and the difference between oligodendroglial and astrocytic tumors was demonstrated by a statistical analysis. Olig2 and glial fibrillary acidic protein were expressed in a mutually exclusive manner, and Olig2 expression was cell-cycle related. Neither central neurocytoma nor schwannoma cases were stained. Our antibody was demonstrated to be useful in recognizing normal oligodendrocytes on paraffin sections, and applicable in diagnosis of some brain tumors.

Aim: Papillary glioneuronal tumor (PGNT) and rosette-forming glioneuronal tumor (RGNT) were recently established unusual glioneuronal phenotypes and were categorized as a novel tumor entity in the 2007 World Health Organization classification. The molecular background of these glioneuronal tumors remains poorly understood due to a paucity of reports. Recently, the SLC44A1-PRKCA fusion has been identified in 3 cases of PGNT. Methods: The aim of this study was to confirm it in our 3 cases of PGNT and analyze the presence of specific fusion in 2 cases of RGNT, using fluorescence in situ hybridization. Results: Two out of three PGNT cases showed one adjacent red-green signal pair representing the rearrangement on chromosome 9 and 17, but not a complete fused yellow signal. Normal signal pattern was observed in the other PGNT case. Neither of two RGNT cases revealed adjacent red-green signal pair or yellow signal. Conclusions: The SLC44A1-PRKCA fusion is a characteristic alteration in PGNT but not in RGNT, and is suggested as a potential biomarker of PGNT. One adjacent red-green signal pair observed in the PGNTs may imply the presence of different breakpoints from those previously reported in the 9q31 and 17q24 genes. Disclosure: All authors have declared no conflicts of interest.

MicroRNAs (miRNAs) regulate a variety of cellular processes (including differentiation, proliferation and apoptosis) via the regulation of multiple target genes, including oncogenes and tumor suppressors. Abnormal miRNA expression may therefore play a significant role in tumor development. Medulloblastoma is a common malignant brain tumor in children. Several studies have shown abnormal miRNA-expression profiles in medulloblastomas, but little is known as to whether such profiles are caused by alterations in miRNA genes. In the present study, we screened 48 medulloblastomas for mutation, deletion, and amplification of nine miRNA genes (miR-33b, miR-135a-1, miR-135a-2, miR-135b, miR-186, miR-200b, miR-512-2, miR-548d-1, and miR-548d-2). These miRNAs were selected on the basis of the presence of potential target sequences within the 3’-untranslated region of the MYCC messenger RNA, since MYCC is frequently overexpressed without MYCC gene amplification in medulloblastoma. Differential PCR revealed deletions in miR-186 (15%), miR-135a-1 (33%), miR-548d-1 (42%), miR-548d-2 (21%) and miR-512-2 (33%) genes, whereas deletion or amplification was detected in miR-135b (23%) and miR-135a-2 (15%). In miR-33b, deletion, amplification or a mutation at the precursor miRNA were detected in 10% of medulloblastomas. Overall, 35 out of 48 (73%) medulloblastomas had at least one alteration in the miRNA genes analyzed. Real-time RT-PCR revealed MYCC overexpression in 11 out of 37 medulloblastomas analyzed (30%), and there was a significant correlation between MYCC overexpression and miR-512-2 gene deletion (P = 0.0084). These results suggest that deletion or amplification in miRNA genes is frequent, and may play a role in the pathogenesis of medulloblastomas. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3006.

Transgenic rats expressing v-erbB (viral form of the EGF receptor) under transcriptional regulation by the S100beta promoter develop brain tumors (Ohgaki et al. J Neuropathol Experimental Neurol 65: 1111-1117, 2006). In the present study, we carried out detailed immunohistochemical and ultrastructural characterization of the brain tumors that developed in these rats. Of 49 homozygous transgenic rats between 16 and 94 weeks of age (mean, 59 weeks), 31 rats were autopsied because they showed severe neurological symptoms and/or became moribund. Among these, 30 rats had brain tumors, which were classified histologically as malignant glioma, anaplastic oligodendroglioma, and low-grade oligodendroglioma. Six transgenic rats developed two different histologic types of brain tumor, which were considered to be of multiclonal origin, because of the lack of histological transitions. All brain tumors contained neoplastic cells immunoreactive for S100 and GFAP. Diffuse immunoreactivity for Olig2 and Nkx2.2 was observed in neoplastic cells in all seven anaplastic oligodendrogliomas and in all three low-grade oligodendrogliomas analyzed, but in none of 26 malignant gliomas. Electron microscopy, carried out on four malignant gliomas and four anaplastic oligodendrogliomas, revealed the presence of intermediate filament bundles devoid of side arms, indicating glial differentiation. There was no evidence of cilia, microvilli, neurosecretory granules, synaptic structures or neurofilaments, excluding the possibility of ependymal or neuronal tumors. The present study thus provides additional evidence that the brain tumors developing in S100beta-v-erbB transgenic rats are of glial origin, with or without oligodendroglial differentiation. Reproducible development of three distinct histologic types of brain tumor in unique localizations may be explained by activation of the v-erbB transgene driven by the S100beta promoter in specific precursor cells during development of the brain. Thus, S100beta-v-erbB transgenic rats may be useful to study the histogenesis and molecular mechanisms of development of glial tumors due to disruption of the EGFR pathway.