Pediatric Ependymoma: A Proteomics Perspective.

BACKGROUND/AIM: Proteomics based on high-resolution mass spectrometry (MS) is the tool of choice for the analysis of protein presence, modifications and interactions, with increasing emphasis on the examination of tumor tissues. Application of MS-based proteomics offers a detailed picture of tumor tissue characteristics, facilitating the appreciation of different tumor entities, whilst providing reliable and fast results for therapeutic marker targeting and prognostic factor assessment. Through use of the high analytical resolution of nano-high-pressure liquid chromatography (nanoHPLC) and the high resolution of an Orbitrap Elite mass spectrometer, the present study aimed to provide knowledge on the proteome of the generally unknown entity of pediatric ependymal tumors. MATERIALS AND METHODS: Ten resected specimens of childhood ependymoma were analyzed through a one-dimensional (1D) nanoLC-MS/MS approach. Method optimization steps were undertaken for both the sample preparation/protein extraction procedure and LC parameters, aiming to achieve the highest possible identification rates. RESULTS: Following method optimization, each nanoLC-MS/MS run resulted in identification of more than 5,000 proteins and more than 25,000 peptides for every analyzed sample, thus detailing the greater part of the ependymoma proteome. Identified proteins were found to spread throughout all known tumor categories regarding their molecular function and subcellular localization. CONCLUSION: Through the proposed nanoLC-MS/MS method herein we report, for the firs time, the ependymoma proteome database. A large number of similarities regarding proteome content are revealed compared to other two pediatric brain tumor entities; astrocytomas and medulloblastomas. Furthermore, through our approach, the majority of currently proposed markers for ependymoma (e.g. nucleolin, nestin, Ki67 and laminin subunit A2) as well as all major key players of the phosphoinositide 3-kinase pathway (seemingly implicated in ependymoma), were definitely detected.

Giant Atypical Intraventricular Meningioma in a Child.

We present the case of a 14-year-old boy with a giant tumor of the lateral ventricle. The patient was operated upon. Histopathology showed the presence of an atypical meningioma. Postoperative imaging confirmed the complete tumor excision. Meningiomas although frequent in adults are rare in children. Intraventricular meningiomas are exceedingly rare. Complete surgical excision should be the goal of treatment and is usually associated with a favorable outcome.

The Role of Fast Cell Cycle Analysis in Pediatric Brain Tumors.

Cell cycle analysis by flow cytometry has not been adequately studied in pediatric brain tumors. We investigated the value of a modified rapid (within 6 min) cell cycle analysis protocol for the characterization of malignancy of pediatric brain tumors and for the differentiation of neoplastic from nonneoplastic tissue for possible intraoperative application. We retrospectively studied brain tumor specimens from patients treated at our institute over a 5-year period. All tumor samples were histopathologically verified before flow-cytometric analysis. The histopathological examination of permanent tissue sections was the gold standard. There were 68 brain tumor cases. All tumors had significantly lower G0/G1 and significantly higher S phase and mitosis fractions than normal brain tissue. Furthermore low-grade tumors could be differentiated from high-grade tumors. DNA aneuploidy was detected in 35 tumors. A correlation between S phase fraction and Ki-67 index was found in medulloblastomas and anaplastic ependymomas. Rapid cell cycle analysis by flow cytometry is a promising method for the identification of neoplastic tissue intraoperatively. Low-grade tumors could be differentiated from high-grade tumors. Thus, cell cycle analysis can be a valuable adjunct to the histopathological evaluation of pediatric brain tumors, whereas its intraoperative application warrants further investigation.

The value of cell cycle analysis by propidium-iodine staining of CD56+ cells in pediatric brain tumors.

PURPOSE: Cell cycle analysis by flow cytometry has not been adequately studied in pediatric brain tumors. We investigated the value of cell cycle analysis by propidium-iodine (PI) staining of CD56+ (gated) cells for the characterization of pediatric brain tumor's malignancy. METHODS: Patients that underwent surgery for an intracranial lesion and tissue sample was available were included in the study. All tumor samples were histopathologically verified before flow cytometric analysis. RESULTS: There were 46 pediatric brain tumor cases. As control we used samples from three cases of brain tissue obtained during surgery for epilepsy. All tumors had significant lower G0/G1 and significant higher S-phase, G2/M fraction, S+G2/M and S+G2/M/G0/G1-phase fraction than normal brain tissue. Low-grade tumors had significant lower S-phase than high grade tumors. Grade IV tumors had significant lower G0/G1 fraction and higher S+G2/M/G0/G1 index than grade III tumors. DNA diploidy was detected in 24 tumors and DNA aneuploidy was detected in 23 tumors. There was a significant correlation between Ki-67 index and both S+G2/M and S+G2/M/G0/G1 phase fraction. CONCLUSIONS: Cell cycle analysis by PI staining of CD56+ cells is a promising method for the assessment of pediatric brain tumors malignancy and could be a valuable adjunct to histopathological evaluation.

Proteomic studies of pediatric medulloblastoma tumors with 17p deletion.

CNS tumors are the leading cause of cancer-related death in children. Medulloblastoma is the commonest pediatric CNS malignancy, wherein, despite multimodal therapy with surgery, radiation, and chemotherapy, 5 year survival rates merely approach 60%. Until present, gene expression and cytogenetic studies have produced contradicting findings regarding the molecular background of the specific disease. Through integration of genomics, bioinformatics, and proteomics, the current study aims to shed light at the proteomic-related molecular events responsible for MBL pathophysiology, as well as to provide molecular/protein/pathway answers concerning tumor-onset. Experiments were performed on tissues collected at surgery. With 17p loss being the commonest chromosomal aberrance observed in our sample set, array-CGH were employed to first distinguish for 17p-positive cases. 2-DE coupled to mass spectrometry identification exposed the MBL-specific protein profile. Protein profiles of malignant tissues were compared against profiles of normal cerebellar tissues, and quantitative protein differences were determined. Bioinformatics, functional and database analyses, characterization, and subnetwork profiling generated information on MBL protein interactions. Key molecules of the PI3K/mTOR signaling network were identified via the techniques applied herein. Among the findings IGF2, PI3K, Rictor, MAPKAP1, S6K1, 4EBP1, and ELF4A, as part of the IGF network (implicating PI3K/mTOR), were founded to be deregulated.

Hernia de disco lumbar en un niño con fibrosis quística: caso clínico / Lumbar disc herniation in a child with cystic fibrosis: case report

Presentamos el caso de un niño con fibrosis quística y hernia de disco lumbar. Un niño de 8 años de edad consultó por dolor lumbar que aumentaba con la tos, al sentarse, caminar o inclinarse, y disminuía al acostarse. La prueba de levantamiento de la pierna en extensión fue positiva cuando el miembro inferior derecho llegó a 60 grados. La prueba contralateral fue negativa. La resonancia magnética nuclear mostró una protrusión central del disco intervertebral entre L5-S1. El tratamiento conservador no fue efectivo, por lo cual se efectuó tratamiento quirúrgico, que hizo desaparecer el dolor. Según nuestro conocimiento, este es el primer caso comunicado de hernia de disco lumbar en un niño con fibrosis quística. Aunque este caso podría ser una coincidencia, se debe realizar una investigación detallada ante el dolor de espalda, síntoma frecuente en pacientes con fibrosis quística.

We report a case of child with cystic fibrosis and lumbar disc herniation. An 8-year-old boy presented with low back pain that exacerbated on coughing, sitting, walking, or bending and diminished when lying down. The straight leg raising test was positive when the right leg was lifted at 60 degrees. Crossed leg raising test was negative. Lumbar MRI revealed a L5-S1central disc protrusion. Conservative treatment was not effective and the patient underwent surgery. Postoperatively the patient experienced regression of the pain. To the best of our knowledge this is the first reported case of lumbar disc herniation in a child with cystic fibrosis. Although this case might be coincidental, thorough investigation of back pain, which is frequent in patients with cystic fibrosis, should be performed.

Hernia de disco lumbar en un niño con fibrosis quística: caso clínico / Lumbar disc herniation in a child with cystic fibrosis: case report

Presentamos el caso de un niño con fibrosis quística y hernia de disco lumbar. Un niño de 8 años de edad consultó por dolor lumbar que aumentaba con la tos, al sentarse, caminar o inclinarse, y disminuía al acostarse. La prueba de levantamiento de la pierna en extensión fue positiva cuando el miembro inferior derecho llegó a 60 grados. La prueba contralateral fue negativa. La resonancia magnética nuclear mostró una protrusión central del disco intervertebral entre L5-S1. El tratamiento conservador no fue efectivo, por lo cual se efectuó tratamiento quirúrgico, que hizo desaparecer el dolor. Según nuestro conocimiento, este es el primer caso comunicado de hernia de disco lumbar en un niño con fibrosis quística. Aunque este caso podría ser una coincidencia, se debe realizar una investigación detallada ante el dolor de espalda, síntoma frecuente en pacientes con fibrosis quística.(AU)

We report a case of child with cystic fibrosis and lumbar disc herniation. An 8-year-old boy presented with low back pain that exacerbated on coughing, sitting, walking, or bending and diminished when lying down. The straight leg raising test was positive when the right leg was lifted at 60 degrees. Crossed leg raising test was negative. Lumbar MRI revealed a L5-S1central disc protrusion. Conservative treatment was not effective and the patient underwent surgery. Postoperatively the patient experienced regression of the pain. To the best of our knowledge this is the first reported case of lumbar disc herniation in a child with cystic fibrosis. Although this case might be coincidental, thorough investigation of back pain, which is frequent in patients with cystic fibrosis, should be performed.(AU)

Lumbar disc herniation in a child with cystic fibrosis: case report.

We report a case of child with cystic fibrosis and lumbar disc herniation. An 8-year-old boy presented with low back pain that exacerbated on coughing, sitting, walking, or bending and diminished when lying down. The straight leg raising test was positive when the right leg was lifted at 60 degrees. Crossed leg raising test was negative. Lumbar MRI revealed a L5-S1central disc protrusion. Conservative treatment was not effective and the patient underwent surgery. Postoperatively the patient experienced regression of the pain. To the best of our knowledge this is the first reported case of lumbar disc herniation in a child with cystic fibrosis. Although this case might be coincidental, thorough investigation of back pain, which is frequent in patients with cystic fibrosis, should be performed.

Lumbar disc herniation in a child with cystic fibrosis: case report. / Lumbar disc herniation in a child with cystic fibrosis: case report.

We report a case of child with cystic fibrosis and lumbar disc herniation. An 8-year-old boy presented with low back pain that exacerbated on coughing, sitting, walking, or bending and diminished when lying down. The straight leg raising test was positive when the right leg was lifted at 60 degrees. Crossed leg raising test was negative. Lumbar MRI revealed a L5-S1central disc protrusion. Conservative treatment was not effective and the patient underwent surgery. Postoperatively the patient experienced regression of the pain. To the best of our knowledge this is the first reported case of lumbar disc herniation in a child with cystic fibrosis. Although this case might be coincidental, thorough investigation of back pain, which is frequent in patients with cystic fibrosis, should be performed.

Expression of heat shock proteins in medulloblastoma.

OBJECT: Medulloblastoma (MB) is the most common malignant brain tumor in children. Heat shock proteins (HSPs) comprise a superfamily of proteins that serve as molecular chaperones and are overexpressed in a wide range of human cancers. The purpose of the present study was to investigate the expression of HSP27 (pSer(82)), HSP27 (pSer(15)), HSP40, HSP60, HSP70, HSP90-α, Akt, and phospho-Akt by multiplex bead array assay of MBs. The results of HSP and Akt expression were correlated with MB subtype; immunohistochemical expression of Ki-67 index, bcl-2, and p53; and patients' prognosis. METHODS: The authors retrospectively evaluated 25 children with MB who underwent surgery. Immunohistochemical analysis of Ki-67, p53, and bcl-2 expression was performed in all cases. By using multiplex bead array assay, a simultaneous detection of HSP27 (pSer(82)), HSP27 (pSer(15)), HSP40, HSP60, HSP70, HSP90-α, Akt, and phospho-Akt was performed. RESULTS: Medulloblastoma with extensive nodularity had significantly lower HSP27 (pSer(15)) expression (p = 0.039) but significantly higher HSP60 expression (p = 0.021) than classic MB. Large-cell MB had significantly higher HSP70 expression (p = 0.028) than classic MB. No significant difference was found between HSP27 (pSer(82)), HSP40, HSP90-α, Akt, or phospho-Akt expression and MB subtype. Large-cell MBs had significantly higher Ki-67 index compared with classic MBs (p = 0.033). When analyzing all MBs, there was a significant negative correlation between HSP27 (pSer(15)) and Ki-67 index (r = -0.475, p = 0.016); a significant positive correlation between HSP70 expression and Ki-67 index (r = 0.407, p = 0.043); and a significant positive correlation between HSP70 expression and bcl-2 index (r = 0.491, p = 0.023). Patients with large-cell MB had a worse survival than those with classic MB, but the difference did not reach statistical significance (p = 0.076). CONCLUSIONS: A substantial expression of several HSPs in MB was observed. Given that HSPs represent an attractive strategy for anticancer therapy, further studies, involving larger series of patients, are obviously necessary to clarify the relationship of HSPs with tumor aggressiveness and prognosis.

Supratentorial ependymomas in children: Analysis of nine cases.

BACKGROUND: Intracranial ependymomas are the third most common primary brain tumor in children. In the present study, we set out to investigate the expression of p-53, p-27, bcl-2, epidermal growth factor receptor (EGFR) and of neuronal markers in pediatric supratentorial ependymomas, in correlation with Ki-67/MIB-1 proliferation index and prognosis. MATERIALS AND METHODS: Nine children with supratentorial ependymomas that were treated surgically in our institute over the last seven years were identified and included in the study. The extent of resection was classified as gross total and subtotal, and was determined by MRI scans. The ependymal tumors were classified according to WHO classification. RESULTS: Headache and seizures were the most common presenting symptoms and papilledema the most common sign. In seven cases, gross total excision was performed, and in two cases, the resection was subtotal. All ependymomas were anaplastic. Ki-67/MIB-1 was detected in 20-40% of the nuclei in all tumors. There was also increased expression of p-53, bcl-2, p-27, and EGFR. There was expression of neuronal markers in three cases. After a mean follow-up period of 32.1 months (range 16-74 months), eight children were alive. Five children suffered from tumor recurrence. CONCLUSIONS: Complete surgical excision should be the goal of surgery. The prognostic role of Ki-67, p-53, p-27, bcl-2, EGFR, and neuronal markers expression needs to be determined in multi-institutional studies due to tumor's rarity.

BRAF alterations in pediatric low grade gliomas and mixed neuronal-glial tumors.

Low grade astrocytomas are the most common brain tumor in children. Recent studies have identified alterations in the BRAF serine/threonine kinase gene that result in mitogen activated protein kinase pathway activation. Herewith, we investigated the genetic changes of BRAF in pediatric low grade gliomas and their relation to pathological findings and Ki-67 proliferation index. The results showed gene fusions between KIAA1549 and BRAF in 66.7 % of tumors. The majority involved the KIAA1549-BRAF exon 16-exon 9 variant. Fusion junction between KIAA1549 exon 15 and BRAF exon 9 was found in five tumors, in which the myxoid component was the predominant. This has not been previously reported. No significant correlation was found between specific KIAA1549 and BRAF fusion junctions and Ki-67 index. All of the samples included in this study were tested for the presence of the BRAF(V600E) mutation, and no positive sample was found.

Embryonal tumor with abundant neuropil and true rosettes: a systematic literature review and report of 2 new cases.

Embryonal tumor with abundant neuropil and true rosettes has been recently defined as a distinct central nervous system embryonal neoplasm, although it was initially regarded as a subtype of central nervous system primitive neuroectodermal tumor. To date 70 cases have been reported. We have performed a literature review and we present 2 new cases. Analysis of the reported data revealed that radiotherapy, tumor excision and high-dose adjuvant chemotherapy with sequential autologous hematopoietic stem cell rescue have a prognostic significance.