Neuroblastic tumors include the neuroblastomas, ganglioneuroblastomas, and ganglioneuromas. Clinical behavior of these developmental malignancies varies from regression to aggressive growth with ...metastatic dissemination. Several clinical, histological, genetic, and biological features are associated with this diversity of clinical presentations. The calcium-sensing receptor (CaSR) is a G-protein coupled receptor with a key role in calcium homeostasis. We have previously reported that it is expressed in benign, differentiated neuroblastic tumors, but silenced by genetic and epigenetic events in unfavorable neuroblastomas. We have now analyzed three functionally relevant polymorphisms clustered at the signal transduction region of the CaSR (rs1801725, rs1042636 and rs1801726) to assess if genetic variants producing a less active receptor are associated with more aggressive disease course.
Polymorphisms were analyzed in DNA samples from 65 patients using specific Taqman Genotyping Assays.
Mildly inactivating variant rs1801725 was associated with clinical stage 4 (P = 0.002) and the histological subgroup of undifferentiated neuroblastomas (P = 0.046). Patients harboring this polymorphism had significantly lower overall (P = 0.022) and event-free survival (P = 0.01) rates than those who were homozygous for the most common allele among Caucasians. However, this single locus genotype was not independently associated with outcome in multivariate analyses. Conversely, the tri-locus haplotype TAC was independently associated with an increased risk of death in the entire cohort (Hazard Ratio = 2.45; 95% Confidence Interval 1.14-5.29; P = 0.022) and also in patients diagnosed with neuroblastomas (Hazard Ratio = 2.74; 95% Confidence Interval 1.20-6.25; P = 0.016).
The TAC haplotype includes the moderately inactivating variant rs1801725 and absence of the gain-of-function rs1042636 polymorphism. Thus, its association with metastatic disease and poor outcome would add to our previous data and further support that inactivation of the CaSR gene is a mechanism associated with neuroblastoma malignant behavior.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
To define the DNA methylation landscape of neuroblastoma and its clinicopathological impact.
Microarray DNA methylation data were analyzed and associated with functional/regulatory genome annotation ...data, transcriptional profiles and clinicobiological parameters.
DNA methylation changes in neuroblastoma affect not only promoters but also intragenic and intergenic regions at cytosine-phosphate-guanine (CpG) and non-CpG sites, and target functional chromatin domains of development and cancer-related genes such as
. Tumors with diverse clinical risk showed differences affecting CpG and, remarkably, non-CpG sites. Non-CpG methylation observed essentially in clinically favorable cases was associated with the differentiation status of neuroblastoma and expression of key genes such as
.
This epigenetic fingerprint of neuroblastoma provides new insights into the pathogenesis and clinical behavior of this pediatric tumor.
Ewing sarcoma (EwS) is an aggressive tumor that affects adolescents and young adults. EwS is defined by a chromosomal translocation, EWSR1-FLI1 being the most common, that causes genome reprogramming ...through remodeling of enhancers. Here, we describe an unexpected function of RING1B, which is highly expressed in EwS. While retaining its repressive activity at Polycomb developmental regulated genes, RING1B colocalizes with EWSR1-FLI1 at active enhancers. We demonstrate that RING1B is necessary for the expression of key EWSR1-FLI1 targets by facilitating oncogene recruitment to their enhancers. Knockdown of RING1B impairs growth of tumor xenografts and expression of genes regulated by EWSR1-FLI1 bound enhancers. Pharmacological inhibition of AURKB with AZD1152 increases H2Aub levels causing down-regulation of RING1B/EWSR1-FLI1 common targets. Our findings demonstrate that RING1B is a critical modulator of EWSR1-FLI1-induced chromatin remodeling, and its inhibition is a potential therapeutic strategy for the treatment of these tumors.
Molecular classification of medulloblastoma is critical for the treatment of this brain tumor. Array-based DNA methylation profiling has emerged as a powerful approach for brain tumor classification. ...However, this technology is currently not widely available. We present a machine-learning decision support system (DSS) that enables the classification of the principal molecular groups—WNT, SHH, and non-WNT/non-SHH—directly from quantitative PCR (qPCR) data. We propose a framework where the developed DSS appears as a user-friendly web-application—EpiGe-App—that enables automated interpretation of qPCR methylation data and subsequent molecular group prediction. The basis of our classification strategy is a previously validated six-cytosine signature with subgroup-specific methylation profiles. This reduced set of markers enabled us to develop a methyl-genotyping assay capable of determining the methylation status of cytosines using qPCR instruments. This study provides a comprehensive approach for rapid classification of clinically relevant medulloblastoma groups, using readily accessible equipment and an easy-to-use web-application.
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•A PCR-based machine-learning strategy for medulloblastoma subgroup classification•MB subgroup-specific DNA methylation profiles analyzed using PCR methyl-genotyping•Automated website analysis of PCR methylation data predicts MB molecular subgroups•EpiGe, a comprehensive approach for rapid classification of MB principal subgroups
Health technology; Cancer; Machine learning
Fundoscopy is the standard method for diagnosis and follow‐up of intraocular retinoblastoma, but it is sometimes insufficient to discern whether tumors are inactivated following treatments. In this ...work, we hypothesized that the amount of conserved nuclear DNA sequences in the cell‐free DNA (cfDNA) fraction of the aqueous humor (AH) might complement fundoscopy for retinoblastoma follow‐up. To address our hypothesis, we developed highly sensitive droplet digital polymerase chain reaction (ddPCR) methods to quantify highly conserved DNA sequences of nucleus‐encoded genes (GAPDH and B4GALNT1) and of a mitochondrial gene, MT‐ATP6. We obtained AH samples during intravitreal treatments. We analyzed 42 AH samples from 25 patients with intraocular retinoblastoma and 11 AH from controls (non‐cancer patients). According to clinical criteria, we grouped patients as having progression‐free or progressive retinoblastoma. cfDNA concentration in the AH was similar in both retinoblastoma groups. Copy counts for nucleus‐derived sequences of GAPDH and B4GALNT1 were significantly higher in the AH from patients with progressive disease, compared to the AH from progression‐free patients and control non‐cancer patients. The presence of mitochondrial DNA in the AH explained that both retinoblastoma groups had similar cfDNA concentration in AH. The optimal cut‐off point for discriminating between progressive and progression‐free retinoblastomas was 108 GAPDH copies per reaction. Among patients having serial AH samples analyzed during their intravitreal chemotherapy, GAPDH copies were high and decreased below the cut‐off point in those patients responding to chemotherapy. In contrast, one non‐responder patient remained with values above the cut‐off during follow‐up, until enucleation. We conclude that the measurement of conserved nuclear gene sequences in AH allows follow‐up of intraocular retinoblastoma during intravitreal treatment. The method is applicable to all patients and could be relevant for those in which fundoscopy evaluation is inconclusive.
The germline TP53 p.R337H mutation is reported as the most common germline TP53 variant. It exists at a remarkably high frequency in the population of southeast Brazil as founder mutation in two ...distinct haplotypes with the most frequent co-segregating with the p.E134∗ variant of the XAF1 tumor suppressor and an increased cancer risk. Founder mutations demonstrate linkage disequilibrium with neighboring genetic polymorphic markers that can be used to identify the founder variant in different geographic regions and diverse populations. We report here a shared haplotype among Brazilian, Portuguese, and Spanish families and the existence of three additional distinct TP53 p.R337H alleles. Mitochondrial DNA sequencing and Y-STR profiling of Brazilian carriers of the founder TP53 p.R337H allele reveal an excess of Native American haplogroups in maternal lineages and exclusively European haplogroups in paternal lineages, consistent with communities established through male European settlers with extensive intermarriage with Indigenous women. The identification of founder and independent TP53 p.R337H alleles underlines the importance for considering the haplotype as a functional unit and the additive effects of constitutive polymorphisms and associated variants in modifier genes that can influence the cancer phenotype.
p.R337H is the most common germline TP53 variant that exists as a founder and as independent alleles. In this context, the haplotype as a functional unit needs to be considered due to the additive effects of constitutive polymorphisms and associated variants in modifier genes that can influence cancer susceptibility and phenotype.
The goals of this work were to identify factors favoring patient‐derived xenograft (PDX) engraftment and study the association between PDX engraftment and prognosis in pediatric patients with Ewing ...sarcoma, osteosarcoma, and rhabdomyosarcoma. We used immunodeficient mice to establish 30 subcutaneous PDX from patient tumor biopsies, with a successful engraftment rate of 44%. Age greater than 12 years and relapsed disease were patient factors associated with higher engraftment rate. Tumor type and biopsy location did not associate with engraftment. PDX models retained histology markers and most chromosomal aberrations of patient samples during successive passages in mice. Model treatment with irinotecan resulted in significant activity in 20 of the PDXs and replicated the response of rhabdomyosarcoma patients. Successive generations of PDXs responded similarly to irinotecan, demonstrating functional stability of these models. Importantly, out of 68 tumor samples from 51 patients with a median follow‐up of 21.2 months, PDX engraftment from newly diagnosed patients was a prognostic factor significantly associated with poor outcome (p = 0.040). This association was not significant for relapsed patients. In the subgroup of patients with newly diagnosed Ewing sarcoma classified as standard risk, we found higher risk of relapse or refractory disease associated with those samples that produced stable PDX models (p = 0.0357). Overall, our study shows that PDX engraftment predicts worse outcome in newly diagnosed pediatric sarcoma patients.
Neuroblastic tumors (NBT) derive from neural crest stem cells (NCSC). Histologically, NBT are composed by neuroblasts and Schwannian cells. In culture, neuroblastic (N-), substrate-adherent (S-) and ...intermediate phenotype (I-) cell subtypes arise spontaneously.
Here, neuroblastoma (NB) cell line subtypes were characterized according to embryonic peripheral nervous system development markers (GAP43, Phox2b, Sox10, c-kit, GD2, NF68, vimentin, S100beta, calcyclin and ABCG2), morphological features, gene expression and differentiation potential. I-type cells were investigated as a bipotential (neuronal and glial) differentiation stage.
Positive immunostaining of NCSC (GAP43, c-kit, NF68, vimentin and Phox2b) and undifferentiated cell (ABCG2) markers was observed in all NB subtypes. N- and I-type cells displayed cytoplasmic membrane GD2 staining, while nuclear calcyclin was restricted to S-type. N- and I-type cells showed similar phenotype and immunoreactivity pattern. Differential gene expression was associated with each cell subtype. N- and I-type cells displayed similar differentiation capacity towards neuronal and glial lineage fates. S-type cells, upon induction, did not show a neuronal-like phenotype, despite gene expression changes.
Results suggest that N- and I-type NB cell subtypes represent an immature bilineage stage, able to progress towards neuronal and glial fates upon induction of differentiation. S-type cells appear irreversibly committed to a glial lineage fate.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Highlights ► We describe a neuroblastic subpopulation coexpressing neuronal and glial markers. ► The GD2+ /S100A6+ subpopulation is more common in low risk neuroblastoma. ► GD2+ /S100A6+ neuroblasts ...populate the perivascular niche. ► GD2+ /S100A6+ neuroblasts represent tumor glial precursor cells.
Neuroblastoma (NB) is one of the most frequently occurring extracranial solid tumors of childhood (Maris et al., 2007 1; Brodeur, 2003 2). Probability of cure varies according to patient's age, ...extent of disease and tumor biology (Maris et al., 2007 1; Brodeur, 2003 2; Cohn et al., 2009 3). However, the etiology of this developmental tumor is unknown. Recent evidence has shown that pediatric solid tumors, including NB, harbor a paucity of recurrent genetic mutations, with a significant proportion of recurrent events converging on epigenetic mechanisms (Cheung et al., 2012 4; Molenaar et al., 2012 5; Pugh et al., 2013 6; Sausen et al., 2013 7.
We have analyzed the DNA methylome of neuroblastoma using high-density microarrays (Infinium Human Methylation 450k BeadChip) to define the epigenetic landscape of this pediatric tumor and its potential clinicopathological impact. Here, we provide the detail of methods and quality control parameters of the microarray data used for the study. Methylation data has been deposited at NCBI Gene Expression Omnibus data repository, accession number GSE54719; superseries record GSE54721.
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