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Immunocytochemical and ultrastructural studies of the histogenesis of Ewing's sarcoma and putatively related tumors.

The histogenesis of Ewing's sarcoma (EW) and extraskeletal Ewing's sarcoma (EEW) is still disputable. Their relationship to the so-called Askin's tumor, neuroectodermal tumor of bone, and peripheral neuroblastoma remains to be established. In an attempt to clarify these points, immunocytochemical and ultrastructural studies were done on tissues from 14 cases of EW, 4 cases of EEW, and 9 cases of primitive neuroectodermal tumor (PNET) and compared with neuroblastoma and olfactory neuroblastoma. Six tumors categorized initially as EW and EEW on biopsy, turned out to be PNET by extensive histologic and/or ultrastructural observations. Abundant glycogen was recognized not only in 16 of 18 cases of EW and EEW, but also in seven of nine cases of PNET. Fine fibrillar cell processes were seen between tumor cells, at least in limited areas even in cases of EW and EEW. Immunocytochemically, neuron-specific enolase (NSE), neuroblastoma cell surface antigen (NBCA), neuron cell surface antigen (NCSA), and neurofilament (NF) were demonstrated not only in neuroblastoma, but also frequently in cases of EW, EEW, and PNET. The results seem to suggest that EW and EEW represent the most immature forms of neuroectodermal tumor. Electron microscopic study showed predominantly primitive cells with occasional areas of cell processes, neurosecretory granules, and microtubules, suggesting a neuroectodermal origin.

Adolescent↗

Telomere maintenance in childhood primitive neuroectodermal brain tumors.

Primitive neuroectodermal tumors (PNETs), including medulloblastoma (PNET/MB) and supratentorial PNET (sPNET), are the most common malignant brain tumors of childhood. The stabilization of telomere lengths by telomerase activation is an important step in carcinogenesis and cell immortalization. Epigallocatechin gallate (EGCG), the major polyphenol in green tea, is a telomerase inhibitor with antiproliferative and anticarcinogenic effects against different types of cancer. In this study, we used real-time reverse transcriptase-polymerase chain reaction to measure the mRNA expression of the human telomerase reverse transcriptase (hTERT) in 50 primary PNET samples (43 PNET/MB, 7 sPNET), 14 normal human brain samples, and 6 human PNET cell lines. Compared to normal human cerebellum, 38/50 (76%) primary PNET samples had >or= 5-fold upregulated hTERT mRNA expression. We then examined PNET cell lines for telomerase activity using a quantitative telomeric repeat amplification protocol (TRAP), and for telomere length using terminal restriction fragment analysis. While a positive correlation between hTERT mRNA expression and telomerase activity was detected in PNET cell lines, no correlation was found between telomerase activity and telomere length. Treatment of PNET cell lines with EGCG resulted in a dose-dependent inhibition of telomerase activity at micromolar levels. Although EGCG displayed strong proliferation inhibitory effects against TRAP-positive PNET cell lines, it had no significant effect against TRAP-negative D425 cells. These results provide evidence for a possible role of telomerase in the pathogenesis of most PNETs and indicate that subsets of PNETs maintain telomere length by alternative mechanisms. Inhibition of telomerase function represents a novel experimental therapeutic strategy in childhood PNETs that warrants further investigation.

Adolescent↗

Glial differentiation predicts poor clinical outcome in primitive neuroectodermal brain tumors.

Primitive neuroectodermal tumors (PNETs) of the central nervous system, including medulloblastomas (PNET/MB), are the most common malignant brain tumor of childhood. These tumors often express proteins characteristic of glial differentiation (glial fibrillary acidic protein, GFAP), neuronal differentiation (neurofilament proteins, NFPs), and/or photoreceptor differentiation (retinal-S antigen). To identify biological factors of prognostic significance in PNETs, the expression of glial, neuronal, or photoreceptor antigens was evaluated in the tumor specimens of 86 patients with PNETs by immunohistochemistry after microwave antigen enhancement. Patterns of differentiation were then compared with patient relapse-free survival. Multivariate analysis of PNET immunohistochemistry and clinical variables indicated GFAP expression conferred a 6.7-fold greater risk of relapse than tumors that did not express GFAP or NFPs. Increased risk of relapse was directly related to the amount of GFAP expression. Tumors exhibiting clumps or sheets of GFAP-staining cells were associated with a 3.0-fold increased risk of relapse compared with tumors that did not express GFAP, irrespective of immunohistochemical evidence of other differentiation, while scattered GFAP staining was not associated with increased risk of relapse. These findings indicate that expression of GFAP in PNETs has prognostic power comparable with the most significant clinical factors currently used to predict clinical outcome.

Adolescent↗

[The forensic medical significance of brain tumors].

Thirty-three cases of death because of brain tumors are described. The age and sex of dead subjects are analyzed, as are the place and causes of death and morphologic features of the tumors. Neuroectodermal tumors of the glioblastoma or neurinoma types were the most incident. Seven patients were hospitalized and in none of the cases a correct diagnosis was made in life. Forensic medical significance of brain tumors is demonstrated.

Adult↗

Proton MR spectroscopy of pediatric cerebellar tumors.

PURPOSE: To investigate the role of proton MR spectroscopy in pediatric cerebellar tumor diagnosis. METHODS: Single voxel pulse sequences with long echo time (135 or 270 milliseconds, voxel size 8 to 19 cm3), were used to obtain proton spectra of primary pediatric cerebellar tumors. Eleven primitive neuroectodermal tumors (patient age, 2 to 12 years; mean, 7 years), 11 low-grade astrocytomas (age, 2 to 16 years; mean, 9 years), 4 ependymomas (age, 1 to 6 years; mean, 4 years), 1 mixed glioma ependymo-astrocytoma (age, 11 years), 1 anaplastic ependymoma (age, 7 years), 1 ganglioglioma (age, 14 years), and 1 malignant teratoma (age, 6 days) were studied. Control cerebellum spectra were acquired from five patients without abnormality in cerebellum (age, 2 to 15 years; mean, 8 years). The signal intensities from choline-containing compounds (Cho), creatine/phosphocreatine (Cr), N-acetyl-aspartate (NAA), and lactate (Lac) were quantified. The mean and standard deviation of metabolite ratios were calculated. RESULTS: The control spectra ratios (NAA:Cho = 1.49 +/- 0.36, Cr:Cho = 1.13 +/- 0.23) were distinct from the tumor spectra (NAA:Cho = 0.41 +/- 0.27 and Cr:Cho = 0.37 +/- 0.23). Most of primitive neuroectodermal tumors had low NAA:Cho (0.17 +/- 0.09) and Cr:Cho (0.32 +/- 0.19). Compared with primitive neuroectodermal tumors, low-grade astrocytomas and ependymomas had higher NAA:Cho ratio (0.63 +/- 0.19 and 0.39 +/- 0.12). The Cr:Cho ratio was higher for ependymomas (0.60 +/- 0.20) than for astrocytomas (0.27 +/- 0.12) and primitive neuroectodermal tumors. No NAA was found in the malignant teratoma. Lac:Cho ratio was 0.66 +/- 0.40, 0.58 +/- 0.30, and 0.08 +/- 0.12 for astrocytoma, ependymoma, and primitive neuroectodermal tumor, respectively. Lactate was elevated in the mixed glioma ependymo-astrocytoma, ganglioglioma, and teratoma. The NAA and lactate signals were sometimes obscured by lipids in the spectra. Discriminant analysis was carried out using NAA:Cho and Cr:Cho ratios to differentiate the three major tumor types. The sensitivity/specificity values for diagnosing astrocytoma, ependymoma, and primitive neuroectodermal tumor were found to be 0.91/0.84, 0.75/0.92, and 0.82/0.89, respectively, based on this study. CONCLUSION: In many cases, proton MR spectroscopy can be used to help differentiate cerebellar primitive neuroectodermal tumor, low-grade astrocytoma, and ependymoma.

Adolescent↗

Immunocytochemical studies in canine neuroectodermal brain tumors.

Seventy-four canine neuroectodermal tumors were examined immunocytochemically for the presence of glial fibrillary acidic protein (GFAP). Eleven oligodendrogliomas were examined for the presence of myelin basic protein (MBP) and myelin-associated glycoprotein (MAG). Twenty-three tumors, including ten astrocytomas, one ependymoma, two glioblastomas, one case of gliomatosis, and nine poorly differentiated gliomas were positive for GFAP. Two astrocytomas, eleven oligodendrogliomas, eight ependymomas, four choroid plexus papillomas, two medulloblastomas, one glioblastoma, nine poorly differentiated gliomas, six cases of gliomatosis, and three unclassified tumors were GFAP-negative. In six tumors (including four that were classified as astrocytoma) GFAP staining was equivocal. All oligodendrogliomas were MBP-negative but three expressed MAG. It was concluded that many canine gliomas are not only morphologically but also immunocytochemically similar to human gliomas, but that a larger proportion of canine neuroectodermal growths are undifferentiated tumors.

Animals↗

Expression of SV40 T antigen gene in the oligodendroglia induced primitive neuroectodermal tumor-like tumors in the mouse brain.

Primitive neuroectodermal tumors (PNET) are classified as the embryonal tumors developed in the brain, except for the cerebellum. Although many studies have been reported, the origin and pathogenesis of PNET are still unclear. In this study, we observed the development of undifferentiated tumors indistinguishable from PNET in the transgenic mice which expressed simian virus 40 T antigen (SV40-Tag) selectively in the oligodendroglia under the control of mouse myelin basic protein gene promoter. These PNET-like tumors reproducibly developed in the brain stem of the founder mice and the transgenic progeny derived from one founder mouse. Oligodendroglia-specific expression of SV40-Tag in these transgenic mice was observed by immunohistochemical analysis. Furthermore, expression of the oligodendroglia-specific marker genes was decreased in the tumors as well as in the transgenic brains. These findings suggested that tumors developed in transgenic mice were indistinguishable from PNET, and one of them showed oligodendroglia-like characteristics. Consequently, this transgenic line is a useful animal model to study the pathogenesis of undifferentiated tumor.

Animals↗

Melanotic primitive neuroectodermal (neuroepithelial) tumor of medulla.

A 69-year-old man had a melanotic primitive neuroectodermal tumor of the medulla displaying various neuroepithelial elements including undifferentiated neuroepithelial cells forming Homer Wright's rosettes as well as neoplastic neuroglia resembling those seen in medulloblastoma. The neuroglial tumor cells were verified by demonstrating glial fibrillary acidic protein (GFAP) in the cells. These findings support the concept that the primitive neuroectodermal tumor and medulloblastoma are similar neoplasms. They have been described by such diverse names as melanotic medulloblastomas and progonomas. Review of 18 reported cases of intracranial melanotic primitive neuroectodermal tumors, including the present one, reveals that they have common pathologic features, are most frequent in the cerebellum and fourth ventricle, often metastasize widely within the neuraxis or even systemically, occur more frequently in children than adults, and strike males more often than females.

Aged↗

Immunodetection of endogenous opioid peptides in human brain tumors and associated cyst fluids.

The antitumorigenic effects of endogenous opioid peptides and their presence in extracerebral tumors are well documented. In this study, methionine-enkephaline (met-enkephalin) was measured by radioimmunoassay in 108 glial and nonglial brain tumors and in 44 associated cyst fluids. By immunohistochemistry, the distribution of the peptide and its precursor, preproenkephalin A, was also analyzed. Met-enkephalin and preproenkephalin were detected in the cytoplasm and cell processes of all tumors. Moreover, for neuroectodermal tumors (i.e., gliomas, gangliogliomas, and dysembryoplastic neuroepithelial tumors), a strong inverse correlation (P < 0.0001) was observed between the met-enkephalin levels and the degree of malignancy (242.9, 148.3, 55.3, and 30.3 pg/mg protein for grade 1, 2, 3, and 4, respectively). When compared to normal tissue, this differential expression mainly results from a decrease in the opioid peptide content in high-grade neuroectodermal tumors. Meningiomas and cerebral metastases displayed low met-enkephalin levels, similar to those of grade 4 neuroectodermal tumors. Large amounts of met-enkephalin were found in all cyst fluids. These data suggest that the endogenous opioid system is an integral component of brain tumors and that met-enkephalin may represent a useful malignancy marker in neuroectodermal tumors.

Brain Neoplasms↗

[Soft tissue malignancies in childhood and adolescence. Pathology and clinical relevance based on data from the kiel pediatric tumor registry].

Soft tissue malignancies in childhood and adolescence encompass a wide variety of histologically and genetically different tumor entities. In the files of the Kiel Pediatric Tumor Registry, 4,272 soft tissue malignancies were collected since 1977. Rhabdomyosarcomas are by far the most frequent sarcomas (44.6 % of the cases), followed in decreasing order of frequency by the family of Ewing tumors (peripheral primitive neuroectodermal tumors and extraosseous Ewing's sarcomas; altogether 22.3 %), malignant peripheral nerve sheath tumors (8.1 %), synovial sarcomas (5.0 %), leiomyosarcomas (3.2 %), fibrosarcomas (2.4 %), extrarenal malignant rhabdoid tumors (2.0 %), and alveolar soft tissue sarcomas (1.1 %). A further group (11.3 %) includes rare tumors, intermediate fibrohistiocytic tumors, and unclassified sarcomas. Embryonal rhabdomyosarcomas are 2.5 times more frequent than the alveolar rhabdomyosarcomas, which are prognostically unfavorable and located predominantly in the extremities and the trunk. With regard to clinical findings, histology, molecular biology and prognosis, embryonal and alveolar rhabdomyosarcomas have to be considered as two different tumor types. The family of Ewing tumors includes extraosseous Ewing's sarcoma and peripheral primitive neuroectodermal tumors (synonym: malignant peripheral neuroectodermal tumors), the former tumors without and the latter with neural differentiation. Many cases of infantile malignant peripheral nerve sheath tumors and infantile fibrosarcomas are low-grade malignancies and prognostically more favorable than their "adult" counterparts.

Adolescent↗

Elevation of cerebrospinal fluid catecholamine metabolites in patients with intracranial tumors of neuroectodermal origin.

The concentrations of homovanillic acid (HVA), hydroxymethoxyphenylethyleneglycol (HMPG), and vanillylmandelic acid (VMA) were determined in lumbar cerebrospinal fluid (CSF) specimens. The study population consisted of the following groups: control subjects with malignancies of nonneuroectodermal origin (mostly leukemia in remission), neuroblastoma (extracranial and intracranial or cranial metastases), brain tumors (neuroectodermal and glial), and retinoblastoma. A significant increase in the CSF concentration of HVA was observed in patients with brain tumors (neuroectodermal), neuroblastoma (intracranial or cranial metastases), and retinoblastoma when compared with age-matched controls. In contrast, HMPG and VMA concentrations did not differ from controls except in patients with neuroblastoma (intracranial or cranial metastases) and brain tumors (neuroectodermal) who had significant elevations in HMPG and VMA, respectively. An inverse correlation was noted between the CSF concentration of HVA and clinical response to therapy. Nonresponding patients exhibited increases in HVA when compared with pretreatment values. These data suggest that the quantitative determination of catecholamine metabolites in lumbar CSF is an effective method for diagnosing intracranial tumors of neuroectodermal origin and assessing their response to therapy.

Brain Neoplasms↗

Chlorotoxin, a scorpion-derived peptide, specifically binds to gliomas and tumors of neuroectodermal origin.

Highly migratory neuroectodermal cells share a common embryonic origin with cells of the central nervous system (CNS). They include enteric, parasympathetic, sympathoadrenal, and sensory neurons of the peripheral nervous system, Schwann cells, melanocytes, endocrine cells, and cells forming connective tissue of the face and neck. Because of their common embryologic origin, these cells and the tumors that derive from them can share genetic and antigenic phenotypes with gliomas, tumors derived from CNS glia. We recently discovered that chlorotoxin (ClTx), a 4-kD peptide purified from Leiurus quinquestriatus scorpion, is a highly specific marker for glioma cells in biopsy tissues (Soroceanu et al. Cancer Res 58:4871-4879, 1998) that can target tumors in animal models. We report on the specificity of ClTx as a marker for tumors of neuroectodermal origin that include peripheral neuroectodermal tumors (PNET) and gliomas. Specifically, we histochemically stained frozen and paraffin tissue sections of human biopsy tissues from 262 patients with a synthetically manufactured and biologically active ClTx bearing an N-terminal biotin. The vast majority (74 of 79) of primary human brain tumors investigated showed abundant binding of ClTx with greater than 90% ClTx-positive cells in each section. By comparison, 32 biopsies of uninvolved brain used for comparison were largely ClTx-negative, with only a few isolated reactive astrocytes showing some ClTx binding. However, as with gliomas, the vast majority of PNETs examined showed specific ClTx binding (31 of 34). These include medulloblastomas (4 of 4), neuroblastomas (6 of 7), ganglioneuromas (4 of 4), melanomas (7 of 7), adrenal pheochromocytomas (5 of 6), primitive PNET (1), small cell lung carcinoma (2 of 3), and Ewing's sarcoma (2 of 2). Under identical staining conditions, normal tissues from brain, skin, kidney, and lung were consistently negative for ClTx. These results suggest that chlorotoxin is a reliable and specific histopathological marker for tumors of neuroectodermal origin and that chlorotoxin derivatives with cytolytic activity may have therapeutic potential for these cancers.

Adult↗

In vivo and in vitro models of medulloblastomas and other primitive neuroectodermal brain tumors of childhood.

Recent advances in understanding the basic biology of the neoplastic cells that populate childhood primitive neuroectodermal tumors (PNET) of the central nervous system (CNS) underline several unique properties of these common pediatric brain neoplasms. For example, studies of posterior fossa cerebellar medulloblastomas (MB), a prototypical group of brain tumors that comprise the largest class of PNET, suggest that the molecular phenotype of subpopulations of neoplastic cells in MB partially recapitulates stages in the acquisition of the neuronal phenotype by normal developing human CNS progenitor cells. However, as reviewed here, it appears that the neoplastic cells in MB exhibit one or more molecular defects in the sequence of normal maturational events that enable CNS progenitor cells to exit the cell cycle, become committed to the neuronal lineage, and undergo terminal differentiation into fully mature, permanently postmitotic CNS neurons. Indeed, since PNET emerge almost exclusively in early childhood, the induction of PNET may result from genetic lesions that arise in developing CNS progenitor cells thereby preventing these neural precursors from executing normal programs of lineage commitment and differentiation in the CNS. Clarification of how lineage commitment and maturation in PNET comprised of neuron-like tumor cells deviate from normal CNS development may clarify how oncogenes and tumor suppressor genes exert their effects in a cell type specific manner at different stages in the normal maturation of CNS cells. Recently, a number of potentially effective in vitro and in vivo model systems of PNET have been developed. Since these model systems could facilitate efforts to elucidate mechanisms of neoplastic transformation and tumor progression in the CNS, we review the potential utility of several recently described in vitro (e.g., MB cell lines) and in vivo (e.g., transgenic mice) experimental systems as models of authentic childhood CNS neoplasms.

Animals↗

Clinical and prognostic significance of Ki-67 and proliferating cell nuclear antigen expression in childhood primitive neuroectodermal brain tumors.

The cell proliferation activity of sixteen childhood primitive neuroectodermal tumors (PNETs) was observed immunocytochemically, to determine the cell kinetics and cell proliferation activity of these relatively undifferentiated, malignant brain tumors. Two mouse anti-human monoclonal antibodies (MoABs) were employed for the detection of the nuclear antigen (Ki-67) present during proliferation in frozen sections and proliferating cell nuclear antigen (PCNA) expression in formalin fixed, paraffin embedded tissue sections. A sensitive four step, indirect, streptavidin-biotin, alkaline phosphatase (AP) conjugated, immunocytochemical experimental technique, was used. The anti-Ki-67 MoAB which binds to a special nuclear antigen, identified its expression only in proliferating cells. This nuclear antigen was detectable during the whole mitotic cycle of all malignant and fast proliferating cells, only absent between and during phases G0 (resting phase) and the first gap phase (G1). The mean labeling index (MLI) was defined as the percentage of Ki-67 and PCNA antigen positive cells of the total number counted. The MLI for the PNETs ranged between 1.4% and 11.6%, with a mean MLI of 6.2% for Ki-67; and between 3.2% and 16.8%, with a mean of 9.74% for PCNA. All observed PNETs demonstrated heterogeneous nuclear stainings, but the highest MLIs were found among the poorly differentiated classic medulloblastomas (over 30% for the Ki-67 antigen and 46% for PCNA). MLIs were low in 5/13 PNETs (under 4% for antigen Ki-67 and 9.4% for PCNA) and in this group we defined our lowest (1.4%) MLI, suggesting the presence of in vivo neuritogenesis of these undifferentiated, embryonal tumors. MLIs in 6/13 PNETs were intermediate in magnitude. The MLIs were higher in two PNETs with clear cellular differentiation towards an ependymal (10.4%) and melanocytic (8.8%) direction. Formation of astrocytes within the tumor mass did not affect the intermediate character of the MLI (7.8%). The prognosis of every intracranial tumor is obviously correlated with its proliferation activity and cell kinetics. The clinical significance of these parameters is great since they provide direct information concerning the growth characteristics of an intracranial tumor.

Adolescent↗

TrkC expression predicts good clinical outcome in primitive neuroectodermal brain tumors.

PURPOSE: To identify biologic prognostic factors in childhood primitive neuroectodermal tumors (PNET), including medulloblastoma, that accurately define patient groups with sufficiently good prognosis to permit a reduction in treatment intensity. PATIENTS AND METHODS: We determined expression levels of the neurotrophin receptor TrkC mRNA in formalin-fixed tumor samples from 87 well characterized PNET patients using in situ hybridization. Comparison of TrkC mRNA expression levels with clinical and other laboratory variables was performed using univariate and multivariate Cox regression analysis. RESULTS: High TrkC mRNA expression was found to be associated more with higher 5-year cumulative survival rate than was low TrkC mRNA expression (89% v 46%, respectively). When compared with established clinical prognostic factors and laboratory variables of potential prognostic significance, TrkC mRNA expression, by univariate analysis, was found to be the single most powerful predictor of outcome (hazards ratio, 4.81; P <.00005), exceeding all clinical prognostic factors. In multivariate analysis, the hazards ratio remained significant (P <.00005). CONCLUSION: High TrkC mRNA expression in PNET is a powerful independent predictor of favorable clinical outcome. Assessment of TrkC mRNA levels may aid in treatment planning for patients with PNETs and should be incorporated prospectively into PNET clinical trials.

Adolescent↗

Primitive neuroectodermal kidney tumor: 2 case reports and review of the literature.

Peripheral neuroectodermal tumors are uncommon cancers arising from outside the central nervous system. The urinary system is rarely involved. The differentiation of these tumors from other small cell cancer and neuroblastoma is based on immunohistochemical differences. We report 2 cases of such tumors arising from the renal parenchyma. Tumor behavior and treatment modalities are discussed.

Adult↗

Intraperitoneal dissemination of Ad12-induced undifferentiated neuroectodermal hamster tumors: de novo methylation and transcription patterns of integrated viral and of cellular genes.

The intramuscular (i.m.) injection of human adenovirus type 12 (Ad12) into newborn Syrian hamsters caused widespread dissemination of up to 15 tumors over the entire peritoneal cavity in 70-90% of the animals within 30-50 days. Subcutaneous (s.c.) injections led to local tumor formation only. Independent of location, tumor histology revealed Homer-Wright rosette-like structures typical for primitive neuroectodermal tumors (PNET). All tumor cells showed markers indicative of neuroectodermal and mesenchymal derivations. Each Ad12-induced tumor cell carried multiple copies of integrated Ad12 genomes at one chromosomal site which was different for each tumor. For Ad12 tumor induction in hamsters, the patterns of Ad12 viral and cellular gene expression were important and were affected by changes in DNA methylation, both in the integrated Ad12 DNA and the cellular genome. By applying the bisulfite protocol, the de novo DNA methylation in the integrated Ad12 genomes was determined. These patterns were complex, characterized by regional initiation and by excluding genome segments in the E1A and E1B promoters. In all tumors, the Ad12 segments E1A, E1B, E2A, parts of E3 and E4 were similarly transcribed, as shown by the RT-PCR and DNA microarray methods. Changes in the transcription of a large number of cellular genes was assessed by using mouse gene microarrays encompassing about 1980 different mouse genes with 87-96% homology to hamster genes. Similarities and differences existed in the transcription of cellular genes of different functional classes among the different Ad12-induced tumors. These alterations in cellular gene transcription may be an important parameter in the oncogenic transformation by Ad12.

Adenoviruses, Human↗