Neuroblastoma is a malignancy of the developing sympathetic nervous system that often presents with widespread metastatic disease, resulting in survival rates of less than 50%1. To determine the spectrum of somatic mutation in high-risk neuroblastoma, we studied 240 cases using a combination of whole exome, genome and transcriptome sequencing as part of the Therapeutically Applicable Research to Generate Effective Treatments (TARGET) initiative. Here we report a low median exonic mutation frequency of 0.60 per megabase (0.48 non-silent), and remarkably few recurrently mutated genes in these tumors. Genes with significant somatic mutation frequencies included ALK (9.2% of cases), PTPN11 (2.9%), ATRX (2.5%, an additional 7.1% had focal deletions), MYCN (1.7%, a recurrent p.Pro44Leu alteration), and NRAS (0.83%). Rare, potentially pathogenic germline variants were significantly enriched in ALK, CHEK2, PINK1, and BARD1. The relative paucity of recurrent somatic mutations in neuroblastoma challenges current therapeutic strategies reliant upon frequently altered oncogenic drivers.
The majority of neuroblastoma patients have tumors that initially respond to chemotherapy, but a large proportion of patients will experience therapy-resistant relapses. The molecular basis of this aggressive phenotype is unknown. Whole genome sequencing of 23 paired diagnostic and relapsed neuroblastomas showed clonal evolution from the diagnostic tumor with a median of 29 somatic mutations unique to the relapse sample. Eighteen of the 23 relapse tumors (78%) showed mutations predicted to activate the RAS-MAPK signaling pathway. Seven events were detected only in the relapse tumor while the others showed clonal enrichment. In neuroblastoma cell lines we also detected a high frequency of activating mutations in the RAS-MAPK pathway (11/18, 61%) and these lesions predicted for sensitivity to MEK inhibition in vitro and in vivo. Our findings provide the rationale for genetic characterization of relapse neuroblastoma and show that RAS-MAPK pathway mutations may function as a biomarker for new therapeutic approaches to refractory disease.
BackgroundGene expression profiling is being widely applied in cancer research to identify biomarkers for clinical endpoint prediction. Since RNA-seq provides a powerful tool for transcriptome-based applications beyond the limitations of microarrays, we sought to systematically evaluate the performance of RNA-seq-based and microarray-based classifiers in this MAQC-III/SEQC study for clinical endpoint prediction using neuroblastoma as a model.ResultsWe generate gene expression profiles from 498 primary neuroblastomas using both RNA-seq and 44 k microarrays. Characterization of the neuroblastoma transcriptome by RNA-seq reveals that more than 48,000 genes and 200,000 transcripts are being expressed in this malignancy. We also find that RNA-seq provides much more detailed information on specific transcript expression patterns in clinico-genetic neuroblastoma subgroups than microarrays. To systematically compare the power of RNA-seq and microarray-based models in predicting clinical endpoints, we divide the cohort randomly into training and validation sets and develop 360 predictive models on six clinical endpoints of varying predictability. Evaluation of factors potentially affecting model performances reveals that prediction accuracies are most strongly influenced by the nature of the clinical endpoint, whereas technological platforms (RNA-seq vs. microarrays), RNA-seq data analysis pipelines, and feature levels (gene vs. transcript vs. exon-junction level) do not significantly affect performances of the models.ConclusionsWe demonstrate that RNA-seq outperforms microarrays in determining the transcriptomic characteristics of cancer, while RNA-seq and microarray-based models perform similarly in clinical endpoint prediction. Our findings may be valuable to guide future studies on the development of gene expression-based predictive models and their implementation in clinical practice.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-015-0694-1) contains supplementary material, which is available to authorized users.
Tumor infiltration with Vα24-invariant NKT cells (NKTs) associates with favorable outcome in neuroblastoma and other cancers. Although NKTs can be directly cytotoxic against CD1d + cells, the majority of human tumors are CD1d -. Therefore, the role of NKTs in cancer remains largely unknown. Here, we demonstrate that CD68 + tumor-associated monocytes/macrophages (TAMs) represented the majority of CD1d-expressing cells in primary human neuroblastomas. TAMs stimulated neuroblastoma growth in human cell lines and their xenografts in NOD/SCID mice via IL-6 production. Indeed, TAMs produced IL-6 in primary tumors and in the BM of patients with metastatic neuroblastoma. Gene expression analysis using TaqMan low-density arrays of 129 primary human neuroblastomas without MYCN amplification revealed that high-level expression of TAM-specific genes (CD14, CD16, IL6, IL6R, and TGFB1) was associated with poor 5-year event-free survival. While NKTs were not cytotoxic against neuroblastoma cells, they effectively killed monocytes pulsed with tumor cell lysate. The killing of monocytes was CD1d restricted because it was inhibited by a CD1d-specific mAb. Cotransfer of human monocytes and NKTs to tumor-bearing NOD/SCID mice decreased monocyte number at the tumor site and suppressed tumor growth compared with mice transferred with monocytes alone. Thus, killing of TAMs reveals what we believe to be a novel mechanism of NKT antitumor activity that relates to the disease outcome.
Background-Neuroblastoma is a malignancy of the developing sympathetic nervous system that most commonly affects young children and is often lethal. The etiology of this embryonal cancer is not known.
These data suggest that interactions between tumor and inflammatory cells may contribute to the clinical metastatic neuroblastoma phenotype, improve prognostication, and reveal novel therapeutic targets.
These data indicate a unique role of exosomic miR-21 and miR-155 in the cross-talk between NBL cells and human monocytes in the resistance to chemotherapy, through a novel exosomic miR-21/TLR8-NF-кB/exosomic miR-155/TERF1 signaling pathway.
We conducted a SNP-based genome-wide association study (GWAS) focused on the high-risk subset of neuroblastoma1. As our previous unbiased GWAS showed strong association of common 6p22 SNP alleles with aggressive neuroblastoma2, we now restricted our analysis to 397 high-risk cases compared to 2,043 controls. We detected new significant association of six SNPs at 2q35 within the BARD1 gene locus (Pallelic = 2.35×10−9 − 2.25×10−8). Each SNP association was confirmed in a second series of 189 high-risk cases and 1,178 controls (Pallelic = 7.90×10−7 − 2.77×10−4). The two most significant SNPs (rs6435862, rs3768716) were also tested in two additional independent high-risk neuroblastoma case series, yielding combined allelic odds-ratios of 1.68 each (P = 8.65×10−18 and 2.74×10−16, respectively). Significant association was also found with known BARD1 nsSNPs. These data show that common variation in BARD1 contributes to the etiology of the aggressive and most clinically relevant subset of human neuroblastoma.
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