miR-210-3p enriched extracellular vesicles from hypoxic neuroblastoma cells stimulate migration and invasion of target cells.

BACKGROUND: Tumor hypoxia stimulates release of extracellular vesicles (EVs) that facilitate short- and long-range intercellular communication and metastatization. Albeit hypoxia and EVs release are known features of Neuroblastoma (NB), a metastasis-prone childhood malignancy of the sympathetic nervous system, whether hypoxic EVs can facilitate NB dissemination is unclear. METHODS: Here we isolated and characterized EVs from normoxic and hypoxic NB cell culture supernatants and performed microRNA (miRNA) cargo analysis to identify key mediators of EVs biological effects. We then validated if EVs promote pro-metastatic features both in vitro and in an in vivo zebrafish model. RESULTS: EVs from NB cells cultured at different oxygen tensions did not differ for type and abundance of surface markers nor for biophysical properties. However, EVs derived from hypoxic NB cells (hEVs) were more potent than their normoxic counterpart in inducing NB cells migration and colony formation. miR-210-3p was the most abundant miRNA in the cargo of hEVs; mechanistically, overexpression of miR-210-3p in normoxic EVs conferred them pro-metastatic features, whereas miR-210-3p silencing suppressed the metastatic ability of hypoxic EVs both in vitro and in vivo. CONCLUSION: Our data identify a role for hypoxic EVs and their miR-210-3p cargo enrichment in the cellular and microenvironmental changes favoring NB dissemination.

Verteporfin induces apoptosis and reduces the stem cell-like properties in Neuroblastoma tumour-initiating cells through inhibition of the YAP/TAZ pathway.

Neuroblastoma is an embryonal malignancy of early childhood arising from the embryonic sympatho-adrenal lineage of the neural crest. About half of all cases are currently classified as high-risk of disease recurrence, with an overall survival rate of less than 40% at 5 years despite intensive therapy. Recent studies on matched primary tumours and at the relapse revealed downregulation of genes transcriptionally silenced by YAP as significant association with neuroblastoma relapse. Here, we evaluated the pharmacological targeting of YAP/TAZ with the YAP/TAZ-TEAD inhibitor Verteporfin (VP) in Tumour Initiating Cells (TICs) derived from High-Risk Neuroblastoma patients. VP treatment suppresses YAP/TAZ expression, induces apoptosis and causes the re-organization of the cytoskeleton reducing cells migration and clonogenic ability. Moreover, VP reduces the percentage of side population cells and ABC transporters involved in drug resistance, and the percentage of stem cell subpopulations CD133+ and CD44+ of TICs. Finally, we demonstrated that VP sensitizes TICs to the standard drugs used for neuroblastoma therapy etoposide and cis-platin opening the way to use VP as drug repositioning candidate for recurrent neuroblastoma.

Chromosome instability in neuroblastoma.

Neuroblastoma is a neural crest-derived tumor that accounts for 7-10% of all malignancies in children and ~15% of all childhood cancer-associated mortalities. Approximately 50% of patients are characterized as high-risk (HR) and have an overall survival of <40% at 5 years from diagnosis. HR patients with unfavorable prognosis exhibit several structural copy number variations (CNVs), whereas localized tumors belonging to patients in the low- and intermediate-risk classes, have favorable outcomes and display several numerical CNVs. Taken together these results are indicative of chromosome instability (CIN) in neuroblastoma tumor cells. The present review discusses multiple aspects of CIN including methods of measuring CIN, CIN targeting as a therapeutic strategy in cancer and the effects of CIN in neuroblastoma development and aggressiveness with particular emphasis on the CIN gene signature associated with HR neuroblastoma patients.

Resveratrol treatment reduces the appearance of tubular aggregates and improves the resistance to fatigue in aging mice skeletal muscles.

Resveratrol (RES) is a polyphenolic compound found in grapes, peanuts, and in some berries. RES has been reported to exhibit antioxidant, anti-inflammatory, anti-proliferative properties, and to target mitochondrial-related pathways in mammalian cells and animal models. Therefore, RES is currently advised as supplement in the diet of elderly individuals. Although it is hypothesized that some of RES beneficial actions likely arise from its action on the skeletal muscle, the investigation of RES effects on this tissue remains still elusive. This study reports the effects of a 0,04% RES-supplemented diet for six months, on the skeletal muscle properties of C57/BL6 aging mice. The analysis of the morphology, protein expression, and functional-mechanical properties of selected skeletal muscles in treated compared to control mice, revealed that treated animals presented less tubular aggregates and a better resistance to fatigue in an ex-vivo contraction test, suggesting RES as a good candidate to reduce age-related alterations in muscle.

SQSTM1 gene analysis and gene-environment interaction in Paget's disease of bone.

Even though SQSTM1 gene mutations have been identified in a consistent number of patients, the etiology of Paget's disease of bone (PDB) remains in part unknown. In this study we analyzed SQSTM1 mutations in 533 of 608 consecutive PDB patients from several regions, including the high-prevalence area of Campania (also characterized by increased severity of PDB, higher number of familial cases, and peculiar phenotypic characteristics as giant cell tumor). Eleven different mutations (Y383X, P387L, P392L, E396X, M401V, M404V, G411S, D423X, G425E, G425R, and A427D) were observed in 34 of 92 (37%) and 43 of 441 (10%) of familial and sporadic PDB patients, respectively. All five patients with giant cell tumor complicating familial PDB were negative for SQSTM1 mutations. An increased heterogeneity and a different distribution of mutations were observed in southern Italy (showing 9 of the 11 mutations) than in central and northern Italy. Genotype-phenotype analysis showed only a modest reduction in age at diagnosis in patients with truncating versus missense mutations, whereas the number of affected skeletal sites did not differ significantly. Patients from Campania had the highest prevalence of animal contacts (i.e., working or living on a farm or pet ownership) without any difference between patients with or without mutation. However, when familial cases from Campania were considered, animal contacts were observed in 90% of families without mutations. Interestingly, a progressive age-related decrease in the prevalence of animal contacts, as well as a parallel increase in the prevalence of SQSTM1 mutations, was observed in most regions except in the subgroup of patients from Campania. Moreover, patients reporting animal contacts showed an increased number of affected sites (2.54 +/- 2.0 versus 2.19 +/- 1.9, p < .05) over patients without animal contacts. This difference also was evidenced in the subgroup of patients with SQSTM1 mutations (3.84 +/- 2.5 versus 2.76 +/- 2.2, p < .05). Overall, these data suggest that animal-related factors may be important in the etiology of PDB and may interact with SQSTM1 mutations in influencing disease severity.

Vitamin D receptor gene polymorphisms predict acquired resistance to clodronate treatment in patients with Paget's disease of bone.

Bisphosphonates are first-choice drugs for treatment of Paget's disease of bone (PDB); nevertheless, acquired resistance to bisphosphonate therapy has been described in PDB patients. The 1,25(OH)(2)D(3)/vitamin D receptor (VDR) system influences the effectiveness of antiresorptive treatments in metabolic bone disorders. This study evaluated the relationship between acquired resistance to clodronate treatment and BsmI, TaqI, and FokI VDR polymorphisms in Caucasian patients with polyostotic PDB (n = 84). We also evaluated the influence of mutations in exons 7 and 8 of the sequestosome 1 (SQSTM1) gene on the occurrence of this phenomenon. All patients were treated from diagnosis for several cycles with intravenous clodronate infusion (1500 mg/cycle). Acquired resistance to clodronate treatment was defined as the failure of total alkaline phosphatase serum levels to be suppressed to at least 50% of the patient's previous highest levels during a subsequent treatment course with the same compound, which produced a >50% response after the first exposure. During an observation period of 10.6 +/- 2.7 years, 31 PDB patients (36.9%) showed acquired resistance to clodronate. It was observed that the bb and TT VDR genotypes as well as a lower persistence of the biochemical response to the first treatment course were significantly and independently associated with the risk of developing resistance to clodronate treatment. SQSTM1 gene mutations, considered altogether, did not influence the occurrence of this phenomenon. Our results indicate that 3'VDR allelic variants and duration of biochemical response to the first treatment course are independent predictors of acquired resistance to clodronate treatment in patients with polyostotic PDB.