Epigenetic principles and mechanisms underlying nervous system functions in health and disease.

Frequent Amplification of a chr19q13.41 MicroRNA Polycistron in Aggressive Primitive Neuroectodermal Brain Tumors

Survival rates of pediatric brain tumor patients have significantly improved over the years due to developments in diagnostic techniques, neurosurgery, chemotherapy, radiotherapy, and supportive care. However, brain tumors are still an important cause of cancer-related deaths in children. Prognosis is still highly dependent on clinical characteristics, such as the age of the patient, tumor type, stage, and localization, but increased knowledge about the genetic and biological features of these tumors is being obtained and might be useful to further improve outcome for these patients. It has become clear that the deregulation of signaling pathways essential in brain development, for example, sonic hedgehog (SHH), Wnt, and Notch pathways, plays an important role in pathogenesis and biological behavior, especially for medulloblastomas. More recently, data have become available about the cells of origin of brain tumors and the possible existence of brain tumor stem cells. Newly developed array-based techniques for studying gene expression, protein expression, copy number aberrations, and epigenetic events have led to the identification of other potentially important biological abnormalities in pediatric medulloblastomas and ependymomas.

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Biological background of pediatric medulloblastoma and ependymoma: a review from a translational research perspective.

Bmi-1 and Mel-18 are structural homologues that belong to the Polycomb group of transcriptional regulators and are believed to stably maintain repression of gene expression by altering the state of chromatin at specific promoters. While a number of clinical and experimental observations have implicated Bmi-1 in human tumorigenesis, the role of Mel-18 in cancer cell growth has not been investigated. We report here that short hairpin RNA-mediated knockdown of either Bmi-1 or Mel-18 in human medulloblastoma DAOY cells results in the inhibition of proliferation, loss of clonogenic survival, anchorage-independent growth, and suppression of tumor formation in nude mice. Furthermore, overexpression of both Bmi-1 and Mel-18 significantly increases the clonogenic survival of Rat1 fibroblasts. In contrast, stable downregulation of Bmi-1 or Mel-18 alone does not affect the growth of normal human WI38 fibroblasts. Proteomics-based characterization of Bmi-1 and Mel-18 protein complexes isolated from cancer cells revealed substantial similarities in their respective compositions. Finally, gene expression analysis identified a number of cancer-relevant pathways that may be controlled by Bmi-1 and Mel-18 and also showed that these Polycomb proteins regulate a set of common gene targets. Taken together, these results suggest that Bmi-1 and Mel-18 may have overlapping functions in cancer cell growth.

Contribution of polycomb homologues Bmi-1 and Mel-18 to medulloblastoma pathogenesis.

Sox2 is a key transcription factor that maintains the proliferation of neuroglial stem cells and inhibits neuronal fate commitment. Moreover, it was recently found that brain tumors contain stem cells that resemble normal neuroglial stem cells in many respects. This study was undertaken to describe Sox2 expression in various brain tumors, and to determine whether Sox2 expression is a universal feature of brain tumors, or whether its expression is limited to a specific lineage of brain tumors. Sox2 immunohistochemistry was performed on 194 brain tumor tissues of various kinds. Fetal and adult normal brain tissues obtained by autopsy and brain tissues of epilepsy patients with cortical dysplasia were used as controls. Semiquantitative reverse transcription polymerase chain reaction was used to confirm the immunohistochemical results. Double immunofluorescence was performed to characterize the lineage of Sox2-positive cells. Sox2 was found to be expressed in various glial tumors, including those with astroglial, oligodendroglial, and ependymal lineages, and in the glial components of mixed neuroglial tumors, regardless of pathologic grade. In brain tumors of embryonal origin, supratentorial primitive neuroectodermal tumors showed robust Sox2 expression, whereas medulloblastomas and pineoblastomas did not. The majority of Sox2-positive tumor cells coexpressed glial fibrillary acidic protein, and most Sox2-negative cells in medulloblastomas and pineoblastomas showed neuronal differentiation. This study suggest that Sox2 may be a tumor marker of glial lineages rather than a universal brain tumor stem cell marker, because its expression pattern was found to correspond to differentiation pathways. On the other hand, the aberrant coexpressions of Sox2 and of a neuronal marker were widely observed in glioblastomas, which reflects a disorganized differentiation pattern that characterizes highly malignant tumors.

Sox2 expression in brain tumors: a reflection of the neuroglial differentiation pathway.

Aims : Medulloblastoma (MB), a kind of infratentorial primitive neuroectodermal tumour (PNET), is the most frequent malignant brain tumour in childhood. In contrast, supratentorial PNET (sPNET) are very infrequent tumours, but they are histologically similar to MB, although they present a worse clinical outcome. We investigated the differences in genetic abnormalities between sPNET and MB.



Methods and results : We analysed 20 central PNET (14 MB and six sPNET) by conventional comparative genomic hybridization (CGH) in order to determine whether a different genetic profile for each tumour exists. Isochromosome 17q was detected in four of the 14 MB cases, but not in any sPNET. Gains at 17q and 7 happened more frequently in MB, and those at 1q in sPNET. Losses at chromosome 10 were detected only in MB, while losses at 16p and 19p happened more frequently in sPNET. A new amplification site, on 4q12, was detected in two MB.



Conclusions : Central PNET are a heterogeneous group of tumours from the genetic point of view. The present and previous data, together with further results from larger series, might contribute to the establishment of specific treatments for supratentorial and infratentorial PNET.

Genetic heterogeneity in supratentorial and infratentorial primitive neuroectodermal tumours of the central nervous system

Aims : Medulloblastoma (MB) is the most common primitive neuroectodermal tumour (PNET) of the central nervous system. Although supratentorial PNET (sPNET) and MB are histologically similar, their clinical behaviour differs, sPNET being more aggressive than MB. The aim of this study was to determine whether sPNET and MB are genetically different entities.



Methods and results : We investigated 32 PNET primary tumour samples (23 MB and nine sPNET) and four PNET cell lines, for the presence of CDKN2A homozygous deletions at exon 1-α of p16/INK4 and exon 1-β of p14/ARF, and promoter hypermethylation of both genes. No homozygous deletion of either p16/INK4 or p14/ARF was demonstrated in any of the PNET primary tumour samples. Methylation of p16/INK4 was found in one of six sPNET and in one of 23 MB, while p14/ARF methylation was observed in three of six sPNET and in three of 21 MB. No methylation of p16/INK4 or p14/ARF was found in any of the PNET cell lines analysed. The three MB cell lines did not show p16/INK4 expression, and only the MB Daoy cell line (homozygously deleted at CDKN2A) presented loss of p14/ARF expression.



Conclusions : Our results in this limited series of central PNET show that p14/ARF is frequently involved in PNET carcinogenesis, with a higher frequency, but not statistically significant, for sPNET than for MB.

M. M. Inda

Papers

Genetic heterogeneity in supratentorial and infratentorial primitive neuroectodermal tumours of the central nervous system

High promoter hypermethylation frequency of p14/ARF in supratentorial PNET but not in medulloblastoma.