Familial glioma

Glioma is the most common brain tumor, characterized by several histological and malignancy grade. The majority of gliomas are sporadic, but some familial cases have been reported (< 5%). Despite hereditary predisposition to gliomas has been associated to rare inherited cancer syndromes, such as Li-Fraumeni and Turcot's syndromes, neurofibromatosis and tuberous sclerosis, not all familial gliomas can be explained by these syndromes. Most familial gliomas seem to be characterized by cluster of two cases, suggesting the involvment of low penetrance factor risks. Moreover, no sex-linked disorders or SNPs on the X chromosome have been associated with increased glioma risk, except for ATRX gene, whose loss-of-function has been observed in 20 % of adult oligodendrogliomas and in 80 % of grade 2 and 3 astrocytomas. Finally, the risk to inherit tumors such as glioma could also be related to combinations of multiple risk variants: besides GWAS analysis identified many SNPs involved in familial gliomas at 5p15.33 (TERT), 7p11.2 (EGFR), 8q24.21 (CCDC26), 9p21.3 (CDKN2A/CDKN2B), 11q23.3 (PHLDB1) and 20q13.33 (RTEL1), mutatio could be associated with the risk of glioma ns in POT1 gene and rare variants in SPAG9 and RUNDC1 genes could be associated with the risk of glioma

Glioblastoma multiforme: a multidisciplinary approach to overcome chemoresistance and find new therapeutic strategies

Objectives: Glioblastoma multiforme is the most frequent malignant brain tumor. Patients die within 15 months after diagnosis. The failure of current therapies is ascribed to a subpopulation of cells with stem-like properties, called glioma stem cells (GSCs). The aim of this study is to develop new effective therapies. Moreover, we want to better characterize the orthotopic xenograft model established by GSCs injection into NOD/SCID mice. Materials and methods: We tested Temolomide and Valproic acid treatments, alone and in combination, on seven GSC lines by MTT assay and we sequenced p53. Moreover, we characterized our xenograft model investigating the expression of stemness and differentiation markers by immunohistochemistry on FFPE tissues and by immunofluorescence on the correspondent cell line. Finally, we performed aCGH on the DNA extracted from the cell line and from FFPE tissues. Results: GSCs were resistant to Temozolomide and slightly sensitive to Valproic acid. The two drugs exerted a synergistic effect when combined performing a pre-conditioning with Valproic acid. Furthermore, several cell lines carry p53 mutations. IF and IHC showed a perfect correspondence for stemness markers expression, but discordant data for the others. aCGH analysis evidenced numerous alterations specific for the ex vivo sample, suggesting the presence of an in vivo clonal selection. Discussion: This work shows the importance of murine microenvironment in GSCs phenotype in vivo and suggests the possibility to use our combined treatment for therapeutic purposes. Conclusions: Orthotopic models from GSCs and in vitro grown cell lines represent good models for the development of GSC-targeted therapies

Biological heterogeneity of putative bladder cancer stem-like cell populations from human bladder transitional cell carcinoma samples.

Transitional cell carcinoma (TCC) is the most common type of bladder cancer. Emerging evidence has suggested that the capability of a tumor to grow and propagate is dependent on a small subset of cells, the cancer stem-like cells (CSCs) or tumor initiating cells. We report on the isolation and biological characterization of putative bladder CSC populations from primary TCCs. Isolated cells were induced to proliferate in stem cell culture conditions (serum-free medium containing mitogenic growth factors). The proliferating cells formed spheroids (urospheres) and their abilities for extensive proliferation and self-renewal were assayed. Their positivity for several stem cell markers (CD133, Oct-3/4, nestin, and cytokeratins) was also assessed by immunofluorescence tests and they could have the potential to differentiate in the presence of serum. In stem cell culture conditions they gradually showed loss of proliferation, adherence to the substrate, and morphological changes, which might reflect their progressive acquisition of differentiative capacity and loss of self-renewal ability. To evaluate if effective cell selection occurred after isolation, conventional cytogenetic studies on fresh chromosome spreads immediately after isolation and after culture were carried out. In addition, a molecular cytogenetic study by UroVysion assay was carried out on paraffin-embedded tissue sections and on fresh and after culture nuclei preparations. The data collected indicated important karyotype changes and a positive selection for hypo- or near-diploid cells, losing the complexity present in fresh tumors

Cytogenetics of Premature Ovarian Failure: An Investigation on 269 Affected Women

The importance of X chromosome in the aetiology of premature ovarian failure (POF) is well-known but in many cases POF still remains idiopathic. Chromosome aneuploidy increase is a physiological phenomenon related to aging, but the role of low-level sex chromosome mosaicism in ovarian function is still undiscovered. Standard cytogenetic analysis was carried out in a total of 269 patients affected by POF: 27 chromosomal abnormalities were identified, including X chromosome and autosomal structural and numerical abnormalities. In 47 patients with 46,XX karyotype we performed interphase FISH using X alpha-satellite probe in order to identify X chromosome mosaicism rate. Aneuploidy rate in the patient group was significantly higher than the general population group. These findings underline the importance of X chromosome in the aetiology of POF and highlight the potential role of low-level sex chromosome mosaicism in ovarian aging that may lead to a premature onset of menopause

Thyrospheres from B-CPAP cell line with BRAF and TERT promoter mutations have different functional and molecular features than parental cells

Human thyroid cancer derived cell lines are widely used to study the mechanisms involved in thyroid carcinogenesis. However, there is limited availability of non-cross-contaminated cancer cell lines derived from papillary thyroid carcinoma (PTC), and the B-CPAP cell line is one of the few such lines. B-CPAP cells have been genetically and cytogenetically well-characterized, but details of their stemness features remain uncertain. Considering that this cell line is extensively used for in vitro studies on thyroid tumorigenesis, we broaden its functional and molecular profiles as well as the tumorigenic capacity. We used functional assays (sphere-forming capacity and efficiency), assessed self-renewal and propagation efficiency and tested in vivo tumorigenicity in Hsd:Athymic Nude-Foxn1nu mice. Expression of markers of stemness, differentiation, and epithelial-mesenchymal transition were estimated at RNA and protein levels in adherent parental cells and sphere-forming cells. Functional aspects and stemness features were compared with normal thyrocytes. Protein expression of xenograft tumors was evaluated by immunohistochemistry. B-CPAP sphere-forming cells were able to form thyrospheres theoretically indefinitely in an appropriate serum-free medium, reverting to the adherent parental cell phenotype when cultured in differentiation medium. Different expression of ALDH1-A1 and CD44 stemness markers and TTF-1 and CK19 differentiation markers allowed discrimination between isolated sphere-forming cells and adherent parental cells, indicating that sphere-forming cells retained stem-like features. In keeping with these observations, tumorigenicity assays confirmed that, relative to parental adherent cells, thyrospheres had enhanced capacity to initiate xenograft tumors. Thyrospheres from normal cell line retained very low functional capacity, as well as different stemness markers expression compared to tumor thyrospheres. Our findings may constitute a useful background to develop an in vitro model for assessing the origin and progression of papillary thyroid carcinoma bearing BRAFV600E and TERT promoter mutations

Genomic and epigenomic profile of uterine smooth muscle tumors of uncertain malignant potential (Stumps) revealed similarities and differences with leiomyomas and leiomyosarcomas

Uterine smooth muscle tumors of uncertain malignant potential (STUMPs) represent a heterogeneous group of tumors that cannot be histologically diagnosed as unequivocally benign or malignant. For this reason, many authors are working to obtain a better definition of diagnostic and prognostic criteria. In this work, we analyzed the genomic and epigenomic profile of uterine smooth muscle tumors (USMTs) in order to find similarities and differences between STUMPs, leiomyosarcomas (LMSs) and leiomyomas (LMs), and possibly identify prognostic factors in this group of tumors. Array-CGH data on 23 USMTs demonstrated the presence of a more similar genomic profile between STUMPs and LMSs. Some genes, such as PRKDC and PUM2, with a potential prognostic value, were never previously associated with STUMP. The methylation data appears to be very promising, especially with regards to the divergent profile found in the sample that relapsed, characterized by an overall CGI hypomethylation. Finally, the Gene Ontology analysis highlighted some cancer genes that could play a pivotal role in the unexpected aggressive behavior that can be found in some of these tumors. These genes could prove to be prognostic markers in the future

Cytogenetics of Premature Ovarian Failure: An Investigation on 269 Affected Women

The importance of X chromosome in the aetiology of premature ovarian failure (POF) is well-known but in many cases POF still remains idiopathic. Chromosome aneuploidy increase is a physiological phenomenon related to aging, but the role of low-level sex chromosome mosaicism in ovarian function is still undiscovered. Standard cytogenetic analysis was carried out in a total of 269 patients affected by POF: 27 chromosomal abnormalities were identified, including X chromosome and autosomal structural and numerical abnormalities. In 47 patients with 46,XX karyotype we performed interphase FISH using X alpha-satellite probe in order to identify X chromosome mosaicism rate. Aneuploidy rate in the patient group was significantly higher than the general population group. These findings underline the importance of X chromosome in the aetiology of POF and highlight the potential role of low-level sex chromosome mosaicism in ovarian aging that may lead to a premature onset of menopause

Notch Signaling and MicroRNA: The Dynamic Duo Steering Between Neurogenesis and Glioblastomas.

Notch signaling is an evolutionary conserved pathway that plays a central role in development and differentiation of eukaryotic cells. It has been well documented that Notch signaling is inevitable for neuronal cell growth and homeostasis. It regulates process of differentiation from early embryonic stages to fully developed brain. To achieve this streamlined development of neuronal cells, a number of cellular processes are being orchestrated by the Notch signaling. Abrogated Notch signaling is related to several brain tumors, including glioblastomas. On the other hand, microRNAs are small molecules that play decisive role in mediating and modulating Notch signaling. This review discusses the crucial role of Notch signaling in development of nervous system and how this versatile pathway interplay with microRNAs in glioblastoma. This review sheds light on interplay between abrogated Notch signaling and miRNAs in the regulation of neuronal differentiation with special focus on miRNAs mediated regulation of tumorigenesis in glioblastoma. Furthermore, it discusses different aspects of neurogenesis modulated by the Notch signaling that could be exploited for the identification of new diagnostic tools and therapies for the treatment of glioblastoma

Chromosomal Aberrations in Bladder Cancer: Fresh versus Formalin Fixed Paraffin Embedded Tissue and Targeted FISH versus Wide Microarray-Based CGH Analysis

Bladder carcinogenesis is believed to follow two alternative pathways driven by the loss of chromosome 9 and the gain of chromosome 7, albeit other nonrandom copy number alterations (CNAs) were identified. However, confirmation studies are needed since many aspects of this model remain unclear and considerable heterogeneity among cases has emerged. One of the purposes of this study was to evaluate the performance of a targeted test (UroVysion assay) widely used for the detection of Transitional Cell Carcinoma (TCC) of the bladder, in two different types of material derived from the same tumor. We compared the results of UroVysion test performed on Freshly Isolated interphasic Nuclei (FIN) and on Formalin Fixed Paraffin Embedded (FFPE) tissues from 22 TCCs and we didn't find substantial differences. A second goal was to assess the concordance between array-CGH profiles and the targeted chromosomal profiles of UroVysion assay on an additional set of 10 TCCs, in order to evaluate whether UroVysion is an adequately sensitive method for the identification of selected aneuploidies and nonrandom CNAs in TCCs. Our results confirmed the importance of global genomic screening methods, that is array based CGH, to comprehensively determine the genomic profiles of large series of TCCs tumors. However, this technique has yet some limitations, such as not being able to detect low level mosaicism, or not detecting any change in the number of copies for a kind of compensatory effect due to the presence of high cellular heterogeneity. Thus, it is still advisable to use complementary techniques such as array-CGH and FISH, as the former is able to detect alterations at the genome level not excluding any chromosome, but the latter is able to maintain the individual data at the level of single cells, even if it focuses on few genomic regions

Rubinstein-Taybi Syndrome: spectrum of CREBBP mutations in Italian patients

BACKGROUND: Rubinstein-Taybi Syndrome (RSTS, MIM 180849) is a rare congenital disorder characterized by mental and growth retardation, broad and duplicated distal phalanges of thumbs and halluces, facial dysmorphisms and increased risk of tumors. RSTS is caused by chromosomal rearrangements and point mutations in one copy of the CREB-binding protein gene (CREBBP or CBP) in 16p13.3. To date mutations in CREBBP have been reported in 56.6% of RSTS patients and an average figure of 10% has ascribed to deletions. METHODS: Our study is based on the mutation analysis of CREBBP in 31 Italian RSTS patients using segregation analysis of intragenic microsatellites, BAC FISH and direct sequencing of PCR and RT-PCR fragments. RESULTS: We identified a total of five deletions, two of the entire gene and three, all in a mosaic condition, involving either the 5' or the 3' region. By direct sequencing a total of 14 de novo mutations were identified: 10 truncating (5 frameshift and 5 nonsense), one splice site, and three novel missense mutations. Two of the latter affect the HAT domain, while one maps within the conserved nuclear receptor binding of (aa 1–170) and will probably destroy a Nuclear Localization Signal. Identification of the p.Asn1978Ser in the healthy mother of a patient also carrying a de novo frameshift mutation, questions the pathogenetic significance of the missense change reported as recurrent mutation. Thirteen additional polymorphisms, three as of yet unreported, were also detected. CONCLUSION: A high detection rate (61.3%) of mutations is confirmed by this Italian study which also attests one of the highest microdeletion rate (16%) documented so far