miR148b is a major coordinator of breast cancer progression in a relapse-associated microRNA signature by targeting ITGA5, ROCK1, PIK3CA, NRAS, and CSF1.

Breast cancer is often fatal during its metastatic dissemination. To unravel the role of microRNAs (miRs) during malignancy, we analyzed miR expression in 77 primary breast carcinomas and identified 16 relapse-associated miRs that correlate with survival and/or distinguish tumor subtypes in different datasets. Among them, miR-148b, down-regulated in aggressive breast tumors, was found to be a major coordinator of malignancy. In fact, it is able to oppose various steps of tumor progression when overexpressed in cell lines by influencing invasion, survival to anoikis, extravasation, lung metastasis formation, and chemotherapy response. miR-148b controls malignancy by coordinating a novel pathway involving over 130 genes and, in particular, it directly targets players of the integrin signaling, such as ITGA5, ROCK1, PIK3CA/p110α, and NRAS, as well as CSF1, a growth factor for stroma cells. Our findings reveal the importance of the identified 16 miRs for disease outcome predictions and suggest a critical role for miR-148b in the control of breast cancer progression.

The cell block technique revisited for cells cultured in adherence and as "spheres".

A simple modification of the cell block technique for cultured cells grown in different conditions and suitable for the construction of tissue micro arrays (TMA) is described. The application of mechanical stirring during clot formation allows the uniform dispersion of cells in the fibrin mesh, thus increasing the final volume of the embedded material of evenly distributed cells. This technique is easily applied to spheres-obtained from cell lines cultured under appropriate conditions-that are enriched in stem cells. The possibility of constructing TMA using cell lines (grown in adherence and as spheres) and samples of the corresponding tumors or normal tissues may allow the direct comparison of original tumors with in vitro-expanded cell lines.

Evaluation of two commercial and three home-made fixatives for the substitution of formalin: a formaldehyde-free laboratory is possible.

BACKGROUND: Formaldehyde (HCHO) is a gas (available as a 37% concentrated solution, stabilized with methanol). The 10% dilution (approximately 4% formaldehyde) has been used as a fixative since the end of the 19th century. Alternative fixatives are also commercially available or may be prepared in-house in laboratories. Statements by the IARC, along with other USA agencies (CalEPA, RoC/NTP) on the carcinogenicity of formaldehyde for humans renders its substitution in Pathology Departments necessary since the annual use of formalin may exceed 3,500 liters for a medium-large laboratory. To achieve a "formalin-free laboratory" we tested straightforward-to-make fixatives along with registered reagents offered as formalin substitutes. METHODS: More than two hundreds specimens were fixed in parallel with in-laboratory made fixatives PAGA (Polyethylenglycol, ethyl Alcohol, Glycerol, Acetic acid), two zinc-based fixatives (ZBF, Z7), and commercially-available alternatives (RCL2 and CellBlock). Tissue micro arrays were used for morphological and immunohistochemical comparison. Extraction of RNA was carried out to evaluate preservation of nucleic acids. RESULTS: Differences compared to formalin fixation were evident in alcohol-based fixatives, mainly restricted to higher stain affinity and considerable tissue shrinkage. Conversely, nuclear detail was superior with these alcohol-based formulas compared to formalin or glyoxale-based recipes. RNA extraction was superior for Z7, PAGA and RCL2 with regard to concentration but relatively comparable regarding quality. CONCLUSIONS: Abolition of the human carcinogen formaldehyde from pathology laboratories is possible even in contexts whereby commercial alternatives to formalin are unavailable or are too expensive for routine use, and aspiration devices are lacking or not adequately serviced. The use of known formulations, possibly with simple and not-noxious ("alimentary grade") constituents, comparable with registered proprietary products, may expand the search for the ideal fixative combining satisfactory morphology with improved preservation of nucleic acids and proteins as well as being easy and safe to dispose of.

Circulating Extracellular Vesicles Contain Liver-Derived RNA Species as Indicators of Severe Cholestasis-Induced Early Liver Fibrosis in Mice.

Aims: Biliary diseases represent around 10% of all chronic liver diseases and affect both adults and children. Currently available biochemical tests detect cholestasis but not early liver fibrosis. Circulating extracellular vesicles (EVs) provide a noninvasive, real-time molecular snapshot of the injured organ. We thus aimed at searching for a panel of EV-based biomarkers for cholestasis-induced early liver fibrosis using mouse models. Results: Progressive and detectable histological evidence of collagen deposition and liver fibrosis was observed from day 8 after bile duct ligation (BDL) in mice. Whole transcriptome and small RNA sequencing analyses of circulating EVs revealed differentially enriched RNA species after BDL versus sham controls. Unsupervised hierarchical clustering identified a signature that allowed for discrimination between BDL and controls. In particular, 151 microRNAs (miRNAs) enriched in BDL-derived EVs were identified, of which 66 were conserved in humans. The liver was an important source of circulating EVs in BDL animals as evidenced by the enrichment of several hepatic mRNAs, such as Albumin and Haptoglobin. Interestingly, among experimentally validated miRNAs, miR192-5p, miR194-5p, miR22-3p, and miR29a-3p showed similar enrichment patterns also in EVs derived from 3,5-diethoxycarboncyl-1,4-dihydrocollidine-treated (drug-induced severe cholestasis) but not in mice with mild phenotype or non-cholestatic liver fibrosis. Innovation: A panel of mRNAs and miRNAs contained in circulating EVs, when combined, indicates hepatic damage and fibrosis in mice and represents promising biomarkers for human severe cholestasis-induced liver fibrosis. Conclusion: Analysis of EV-based miRNAs, in combination with hepatic injury RNA markers, can detect early cholestatic liver injury and fibrosis in mice. Antioxid. Redox Signal. 36, 480-504.

Phosphoinositide Conversion Inactivates R-RAS and Drives Metastases in Breast Cancer.

Breast cancer is the most prevalent cancer and a major cause of death in women worldwide. Although early diagnosis and therapeutic intervention significantly improve patient survival rate, metastasis still accounts for most deaths. Here it is reported that, in a cohort of more than 2000 patients with breast cancer, overexpression of PI3KC2α occurs in 52% of cases and correlates with high tumor grade as well as increased probability of distant metastatic events, irrespective of the subtype. Mechanistically, it is demonstrated that PI3KC2α synthetizes a pool of PI(3,4)P2 at focal adhesions that lowers their stability and directs breast cancer cell migration, invasion, and metastasis. PI(3,4)P2 locally produced by PI3KC2α at focal adhesions recruits the Ras GTPase activating protein 3 (RASA3), which inactivates R-RAS, leading to increased focal adhesion turnover, migration, and invasion both in vitro and in vivo. Proof-of-concept is eventually provided that inhibiting PI3KC2α or lowering RASA3 activity at focal adhesions significantly reduces the metastatic burden in PI3KC2α-overexpressing breast cancer, thereby suggesting a novel strategy for anti-breast cancer therapy.

STAT3 induces breast cancer growth via ANGPTL4, MMP13 and STC1 secretion by cancer associated fibroblasts.

In the tumor microenvironment, Cancer Associated Fibroblasts (CAFs) become activated by cancer cells and increase their secretory activity to produce soluble factors that contribute to tumor cells proliferation, invasion and dissemination to distant organs. The pro-tumorigenic transcription factor STAT3 and its canonical inducer, the pro-inflammatory cytokine IL-6, act conjunctly in a positive feedback loop that maintains high levels of IL-6 secretion and STAT3 activation in both tumor and stromal cells. Here, we demonstrate that STAT3 is essential for the pro-tumorigenic functions of murine breast cancer CAFs both in vitro and in vivo, and identify a STAT3 signature significantly enriched for genes encoding for secreted proteins. Among these, ANGPTL4, MMP13 and STC-1 were functionally validated as STAT3-dependent mediators of CAF pro-tumorigenic functions by different approaches. Both in vitro and in vivo CAFs activities were moreover impaired by MMP13 inhibition, supporting the feasibility of a therapeutic approach based on inhibiting STAT3-induced CAF-secreted proteins. The clinical potential of such an approach is supported by the observation that an equivalent CAF-STAT3 signature in humans is expressed at high levels in breast cancer stromal cells and characterizes patients with a shorter disease specific survival, including those with basal-like disease.

Analysis of different medulloblastoma histotypes by two-dimensional gel and MALDI-TOF.

OBJECTIVE: The purpose of this study is to detect different protein profiles in medulloblastoma (MDB) that may be clinically relevant and to check the correspondence of histological classification of MDB with proteomic profiles. MATERIALS AND METHODS: Surgical specimens, snap frozen at the time of neurosurgery, entered the proteomic study. Eight samples from patients (age range, 4 months-26 years) with different MDB histotypes (five classic, one desmoplastic/nodular, one with extensive nodularity, and one anaplastic) were analyzed by two-dimensional gel electrophoresis. One sample for each histotype was further characterized by matrix-assisted laser desorption/ionization time of flight mass spectrometry analysis. RESULTS: Eighty-six unique proteins were identified and compared to histology, with the determination of proteins expressed by single histotypes and of a smaller number of proteins shared by two or three histotypes. The sharp difference of protein expression was found to be in agreement with WHO histological classification, with the identification of type-specific proteins with limited overlapping between histotypes. CONCLUSION: Proteomic analysis confirmed and strengthened the difference between histotypes as biologically relevant. Cluster analysis enhanced the distance of extensive nodularity MDB from other histotypes. Possible innovative approaches to therapy may rely upon a proteomic-based classification of MDB tightly correlated to histology. The utility of snap freezing tumoral samples must be stressed and should become a mandatory task for pathologists.

Medulloblastoma in adults: an analysis of clinico-pathological, molecular and treatment factors.

BACKGROUND: Medulloblastoma is a highly malignant, embryonal tumor, which is rare in adults, and shows distinct clinical, histopathological, molecular and treatment response features. METHODS: We retrospectively investigated 44 adults (age 17-48 years) with an histological diagnosis of medulloblastoma, and in 23 immunohistochemistry was used to identif y the molecular subgroups. We analyzed demographic, diagnostic, therapeutic and cognitive data, and correlated with PFS (progression-free-survival) and OS (overall survival). RESULTS: We observed a male prevalence and a median age of 31 years. Symptoms at onset were related to infratentorial location, while myeloradicular and/or cranial nerve involvement was rare. Histological examination showed the classic variant in 75% of patients, the desmoplastic/nodular in 23% and the anaplastic in one. As for molecular diagnosis, 17 patients were SHH and 6 non-WNT/non-SHH (5 group 4 and 1 group 3), while no WNT subgroup was found. The SHH subgroup had a prevalence of high-risk patients and leptomeningeal involvement. Patients underwent grosstotal or subtotal/partial resection, and craniospinal irradiation, followed in 20 cases by adjuvant chemotherapy. Median OS and PFS were 16.9 and 12 years, respectively. Metastatic disease at presentation and subtotal/partial resection were associated with worse prognosis, while the addition of chemotherapy did not yield a significant advantage over radiotherapy alone. Cognitive impairment in long-term survivors was limited and late relapses occurred in 15% of patients. CONCLUSIONS: Future studies with adequate sample size and long-term follow-up should prospectively investigate the role of surgery and adjuvant therapies across the different molecular subgroups to see whether a personalized approach is feasible.

A complex karyotype including a t(2;11) in a paediatric ependymoma: case report and review of the literature.

Ependymomas are glial tumours representing approximately 5-10% of all intracranial tumours and are the third most common primary brain tumour in childhood. Only a few karyotypic studies on paediatric ependymomas have been published and no specific chromosomal aberration has been specifically related to this type of cancer. We performed cytogenetic analysis of an ependymoma in an 11-year-old boy. Our patient showed a complex karyotype, characterized by a near-tetraploidy and a sole structural unbalanced aberration: der(2)t(2;11)(q11.2;q13.1), which has not been described before. We here discuss such cytogenetic findings, comparing our data with those reported in the literature.

Accuracy of core-needle biopsy after contrast-enhanced ultrasound in soft-tissue tumours.

OBJECTIVE: Percutaneous biopsies are gaining acceptance in the diagnosis of soft-tissue tumours. Sampling in the most representative area is not easy in sarcomas of huge dimension. We hypothesised that ultrasound (US) contrast medium could identify the representative area for focus core-needle biopsy (CNB) METHODS: This is a retrospective cohort series of 115 soft-tissue masses treated from January 2007 to November 2008. Accuracy of US-guided CNB after contrast-enhanced US (CEUS) was determined by comparing the histology of the biopsy with the definitive diagnosis in 105 surgically excised samples (42 benign, 63 malignant) and with the expected outcome in the remaining ten malignant cases not surgically treated. A myxoid component was present in 21 sarcomas (34.4%). RESULTS: Of samples, 94.8% were adequate for diagnosis with 97.1% sensitivity and 92.5% specificity. Sensitivity and specificity in specific histopathological subgroupings were 100%, and in grading definition they were 100% and 96.8%. DISCUSSION: US-guided CNB is safe and effective. US contrast medium depicts tumour vascular supply and identifies the representative area(s) for sampling. Sensitivity and specificity are also high in subgrouping and grading, including myxoid types. Discussion about biopsy is part of the essential multidisciplinary strategy for these tumours.

Medulloblastoma variants: age-dependent occurrence and relation to Gorlin syndrome--a new clinical perspective.

PURPOSE: We aimed to test the hypothesis that medulloblastoma (MB) variants show a different age distribution and clinical behavior reflecting their specific biology, and that MB occurring at very young age is associated with cancer predisposition syndromes such as Gorlin syndrome (GS). EXPERIMENTAL DESIGN: We investigated the frequency, age distribution, location, response to treatment, outcome, and association with familial cancer predisposition syndromes in a series of 82 cases of MB in patients ages <14 years diagnosed at the Giannina Gaslini Children's Hospital, Genoa, between 1987 and 2004. RESULTS: Desmoplastic MB and MB with extensive nodularity (MBEN), were present in 22 of 82 cases (27%) and were more frequent in children ages

Liver-Specific siRNA-Mediated Stat3 or C3 Knockdown Improves the Outcome of Experimental Autoimmune Myocarditis.

Myocarditis can lead to autoimmune disease, dilated cardiomyopathy, and heart failure, which is modeled in the mouse by cardiac myosin immunization (experimental autoimmune myocarditis [EAM]). Signal transducer and activator of transcription 3 (STAT3) systemic inhibition exerts both preventive and therapeutic effects in EAM, and STAT3 constitutive activation elicits immune-mediated myocarditis dependent on complement C3 and correlating with activation of the STAT3-interleukin 6 (IL-6) axis in the liver. Thus, liver-specific STAT3 inhibition may represent a therapeutic option, allowing to bypass the heart toxicity, predicted by systemic STAT3 inhibition. We therefore decided to explore the effectiveness of silencing liver Stat3 and C3 in preventing EAM onset and/or the recovery of cardiac functions. We first show that complement C3 and C5 genetic depletion significantly prevents the onset of spontaneous myocarditis, supporting the complement cascade as a viable target. In order to interfere with complement production and STAT3 activity specifically in the liver, we took advantage of liver-specific Stat3 or C3 small interfering (si)RNA nanoparticles, demonstrating that both siRNAs can significantly prevent myocarditis onset and improve the recovery of heart functions in EAM. Our data demonstrate that liver-specific Stat3/C3 siRNAs may represent a therapeutic option for autoimmune myocarditis and suggest that complement levels and activation might be predictive of progression to dilated cardiomyopathy.

Immunohistochemical and proteomic profile of melanotic medulloblastoma.

We present the case of a 6-year-old male affected by an infratentorial tumor. Histological diagnosis was melanotic medulloblastoma. Immunohistochemistry showed in the melanin rich areas positive cells for HMB45. We performed a proteomic study to compare protein profiles in melanotic versus non-melanotic areas. Protein profiles of different areas of the tumor displayed similarity, with the exception of seven proteins. In accordance with the hypothesis that melanotic medulloblastomas produce oculo-cutaneous melanin, proteomic analysis showed melanocytic-associated antigens and epidermal autoantigen 450K in the pigmented nodule; both these proteins have a significant role as markers of melanotic elements.

The retinoblastoma family member pRb2/p130 is an independent predictor of survival in human soft tissue sarcomas.

PURPOSE: pRb2/p130, a member of the Retinoblastoma gene family, has been shown to be a powerful prognostic factor in several malignancies. We sought to evaluate pRb2/p130 protein expression and its clinical effect in patients affected with soft tissue sarcomas (STS). EXPERIMENTAL DESIGN: Expression of pRb2/p130 was evaluated by immunohistochemistry on formalin-fixed, paraffin-embedded sections in 41 STSs. Results obtained were correlated with clinicopathologic variables and disease-free and overall survival (OS) in univariate and multivariate analysis. RESULTS: Expression of pRb2/p130 was diminished in 25 (61%) tumors, whereas the remaining ones (39%) were classified as high expressors. No correlation between pRb2/p130 expression and clinicopathologic variables was observed. However, a direct relationship between pRb2/p130 expression and clinical outcome of the patients was found in the subgroup of nonmetastatic tumors (n = 31). In univariate analysis, reduced pRb2/p130 expression was a negative prognostic factor and correlated with shorter disease-free survival (P = 0.021) and OS (P = 0.017) survival. In multivariate analysis, reduced pRb2/p130 expression was confirmed to be an independent predictor of shorter OS when considered together with tumor stage and grading (risk ratio, 7.893; confidence interval, 1.618-38.509; P = 0.011). CONCLUSIONS: This study shows for the first time the potential prognostic value of pRb2/130 expression evaluated on formalin-fixed, paraffin-embedded sections in STSs patients. pRb2/p130 immunoreactivity can be used to predict OS in patients with nonmetastatic STSs and, therefore, may represent a new prognostic marker.

Ultrastructural evidence of ependymal differentiation in a genetically proven atypical teratoid/rhabdoid tumor.

INTRODUCTION: We describe a case of genetically proven atypical teratoid/rhabdoid tumor (ATRT), showing ultrastructural evidence of ependymal differentiation. Such differentiation has not been reported so far in ATRT. CONCLUSION: This finding supports the concept that ATRTs as the majority of central nervous system embryonal tumors may derive from an immature and pluripotent neuroectodermal cell capable of differentiating along multiple lineages.

Proteomic identification of heat shock protein 27 as a differentiation and prognostic marker in neuroblastoma but not in Ewing's sarcoma.

Neuroblastoma (NB) and Ewing's sarcoma (ES) cell lines were analysed by two-dimensional gel electrophoresis (2-DE) searching for new diagnostic/prognostic markers. Protein expression profiles displayed a high degree of similarity with the exception of marked heat shock protein (HSP) 27 and less marked HSP60 and HSP70 family up-modulations in NB cells. HSP27, which showed peculiar variability in different NB cell preparations, responded to all trans-retinoic acid treatment in NB cells but not in ES cells at gene and protein expression levels. Immunohistochemistry studies showed different behaviours of HSP27 and HSP70 expression in NB and ES biopsies. HSP27 was less expressed, whereas HSP70 was more expressed in the immature areas of NB. HSP27 expression showed positive and statistically significant correlation with favourable prognosis, and HSP27 expression also negatively correlated with increasing aggressiveness of histological type. In ES, both chaperones were expressed without characteristic patterns. Our results suggest that HSP27, after further clinical validations, could be used as a marker of neuronal differentiation in vivo for the assessment of the biological behaviour of NB and for the risk stratification of patients.

A Virus-Like-Particle immunotherapy targeting Epitope-Specific anti-xCT expressed on cancer stem cell inhibits the progression of metastatic cancer in vivo.

Aggressive forms of breast cancer, such as Her2+ and triple negative breast cancer (TNBC), are enriched in breast cancer stem cells (BCSC) and have limited therapeutic options. BCSC represent a key cellular reservoir for relapse, metastatic progression and therapeutic resistance. Their ability to resist common cytotoxic therapies relies on different mechanisms, including improved detoxification. The cystine-glutamate antiporter protein xCT (SLC7A11) regulates cystine intake, conversion to cysteine and subsequent glutathione synthesis, protecting cells against oxidative and chemical insults. Our previous work showed that xCT is highly expressed in tumorspheres derived from breast cancer cell lines and downregulation of xCT altered BCSC function in vitro and inhibited pulmonary metastases in vivo. We further strengthened these observations by developing a virus-like-particle (VLP; AX09-0M6) immunotherapy targeting the xCT protein. AX09-0M6 elicited a strong antibody response against xCT including high levels of IgG2a antibody. IgG isolated from AX09-0M6 treated mice bound to tumorspheres, inhibited xCT function as assessed by reactive oxygen species generation and decreased BCSC growth and self-renewal. To assess if AX09-0M6 impacts BCSC in vivo seeding, Her2+ TUBO-derived tumorspheres were injected into the tail vein of AX09-0M6 or control treated female BALB/c mice. AX09-0M6 significantly inhibited formation of pulmonary nodules. To evaluate its ability to impact metastases, AX09-0M6 was administered to mice with established subcutaneous 4T1 tumors. AX09-0M6 administration significantly hampered tumor growth and development of pulmonary metastases. These data show that a VLP-based immunization approach inhibits xCT activity, impacts BCSC biology and significantly reduces metastatic progression in preclinical models.

The RNA-binding protein ESRP1 promotes human colorectal cancer progression.

Epithelial splicing regulatory protein 1 (ESRP1) is an epithelial cell-specific RNA binding protein that controls several key cellular processes, like alternative splicing and translation. Previous studies have demonstrated a tumor suppressor role for this protein. Recently, however, a pro-metastatic function of ESRP1 has been reported. We thus aimed at clarifying the role of ESRP1 in Colorectal Cancer (CRC) by performing loss- and gain-of-function studies, and evaluating tumorigenesis and malignancy with in vitro and in vivo approaches. We found that ESRP1 plays a role in anchorage-independent growth of CRC cells. ESRP1-overexpressing cells grown in suspension showed enhanced fibroblast growth factor receptor (FGFR1/2) signalling, Akt activation, and Snail upregulation. Moreover, ESRP1 promoted the ability of CRC cells to generate macrometastases in mice livers. High ESRP1 expression may thus stimulate growth of cancer epithelial cells and promote colorectal cancer progression. Our findings provide mechanistic insights into a previously unreported, pro-oncogenic role for ESRP1 in CRC, and suggest that fine-tuning the level of this RNA-binding protein could be relevant in modulating tumor growth in a subset of CRC patients.

Crucial Role of FLVCR1a in the Maintenance of Intestinal Heme Homeostasis.

AIMS: The maintenance of heme homeostasis, mucosa cell renewal, and redox environment in the intestine is essential to permit digestion, absorption, cell proliferation, cell apoptosis, and immune response and to avoid the development of gut disorders. The feline leukemia virus, subgroup C, receptor 1a (FLVCR1a) is a heme exporter expressed in almost all cell types, including intestinal cells. This work investigates the role of FLVCR1a in the intestine, taking advantage of an intestine-specific conditional Flvcr1a-knockout mouse and of FLVCR1a-depleted Caco2 cells. RESULTS: The data show that FLVCR1a does not participate in the absorption of dietary heme, whereas it is involved in the export of de novo synthesized heme from intestinal cells. The loss of Flvcr1a is associated with a decrease of intestinal cell proliferation and with alterations in the peculiar homeostasis of proliferating cells, including the maintenance of their redox status. The involvement of FLVCR1a in these processes renders this exporter crucial for the survival of mice in a model of ulcerative colitis. INNOVATION: These findings shed light on the role of heme export in the dietary heme absorption process and unravel a new role for heme export in the control of mucosal renewal and in proliferating cell redox status and metabolic activity, demonstrating a crucial role for FLVCR1a in maintaining intestinal homeostasis in both physiologic and pathologic situations. CONCLUSION: By exporting the excess of de novo synthesized heme from intestinal cells, FLVCR1a participates in the control of intestinal mucosa homeostasis.

Proteomic analysis of extracellular vesicles from medullospheres reveals a role for iron in the cancer progression of medulloblastoma.

BACKGROUND: Medulloblastoma (MB) is the most common malignant childhood brain tumor with the propensity to disseminate at an early stage, and is associated with high morbidity. New treatment strategies are needed to improve cure rates and to reduce life-long cognitive and functional deficits associated with current therapies. Extracellular Vesicles (EVs) are important players in cell-to-cell communication in health and diseases. A clearer understanding of cell-to-cell communication in tumors can be achieved by studying EV secretion in medullospheres. This can reveal subtle modifications induced by the passage from adherent to non-adherent growth, as spheres may account for the adaptation of tumor cells to the mutated environment. METHODS: Formation of medullospheres from MB cell lines stabilized in adherent conditions was obtained through culture conditioning based on low attachment flasks and specialized medium. EVs collected by ultracentrifugation, in adherent conditions and as spheres, were subjected to electron microscopy, NanoSight measurements and proteomics. RESULTS: Interestingly, iron carrier proteins were only found in EVs shed by CSC-enriched tumor cell population of spheres. We used iron chelators when culturing MB cell lines as spheres. Iron chelators induced a decrease in number/size of spheres and in stem cell populations able to initiate in vitro spheres formation. CONCLUSIONS: This work suggests a not yet identified role of iron metabolism in MB progression and invasion and opens the possibility to use chelators as adjuvants in anti-tumoral chemotherapy.