Neuroblastoma - Molecular Biology

Overview

Neuroblastoma tumour cells are characterised by a wide diversity of somatic genetic mutations. Some common genetic features include:
  • Amplification of the MYCN gene is one of the most established genetic prognostic factors. Amplified tumours are mostly (though not exclusively) found in children aged over 1 year at diagnosis with advanced stage disease . Other genes, such as DDX1 are often co-amplified with MYCN.
  • Deletion of material from the chromosome 1p36 region is also associated with adverse prognosis. This is thought to be a candidate region for a suppressor gene which has yet to be identified.
  • Gain of 17q material is the most frequent genetic abnormality in neuroblastoma. Unbalanced 17q gain is an adverse prognostic factor and is strongly associated with adverse clinical features, 1p deletion, and MYCN amplification.
  • Expression of TRKA in contrast is a favourable genetic feature. This is associated with low stage and age under 1 yrear at diagnosis. TRKA is frequently supressed in MYCN amplified Tumours. Other members of the TRK neurotrophine receptor gene family, TRKB and TRKC, are also implicated in neuroblastoma.
See also: Familial Neuroblastoma and Genetic Susceptibility.

See also: Neuroblastoma - clinical resources (21)

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

Tag cloud generated 10 March, 2017 using data from PubMed, MeSH and CancerIndex

Mutated Genes and Abnormal Protein Expression (102)

How to use this data tableClicking on the Gene or Topic will take you to a separate more detailed page. Sort this list by clicking on a column heading e.g. 'Gene' or 'Topic'.

GeneLocationAliasesNotesTopicPapers
MYCN 2p24.3 NMYC, ODED, MODED, N-myc, bHLHe37 Amplification
Prognostic
Overexpression
-MYCN amplification in Neuroblastoma
-ABCC1 (MRP1) Overexpression in Neuroblastoma
1205
CASP8 2q33-q34 CAP4, MACH, MCH5, FLICE, ALPS2B, Casp-8 Methylation
Prognostic
Epigenetics
-CASP8 Inactivation in Neuroblastoma
118
NTRK2 9q22.1 TRKB, trk-B, GP145-TrkB Prognostic
-NTRK2 expression in Neuroblastoma
109
ALK 2p23 CD246, NBLST3 -ALK and Neuroblastoma
-ALK mutations in Familial Neuroblastoma
92
PHOX2B 4p12 PMX2B, NBLST2, NBPhox -PHOX2B germline mutations in familial neuroblastoma
-PHOX2B and Neuroblastoma
-PHOX2B and Monitoring of Residual Disease
56
CD44 11p13 IN, LHR, MC56, MDU2, MDU3, MIC4, Pgp1, CDW44, CSPG8, HCELL, HUTCH-I, ECMR-III -CD44 and Neuroblastoma
63
BIRC5 17q25 API4, EPR-1 Overexpression
-Survivin Expression in Neuroblastoma
60
NTRK3 15q25 TRKC, gp145(trkC) Prognostic
-NTRK3 expression in Neuroblastoma
41
TP53 17p13.1 P53, BCC7, LFS1, TRP53 -P53 and Neuroblastoma
32
ID2 2p25 GIG8, ID2A, ID2H, bHLHb26 -ID2 Expression in Neuroblastoma
30
BDNF 11p14.1 ANON2, BULN2 -BDNF and Neuroblastoma
26
DDX1 2p24 DBP-RB, UKVH5d Amplification
-DDX1 Amplification in Neuroblastoma
22
NME1 17q21.3 NB, AWD, NBS, GAAD, NDKA, NM23, NDPKA, NDPK-A, NM23-H1 -NME1 and Neuroblastoma
18
NRAS 1p13.2 NS6, CMNS, NCMS, ALPS4, N-ras, NRAS1 -NRAS and Neuroblastoma
17
NTRK1 1q21-q22 MTC, TRK, TRK1, TRKA, Trk-A, p140-TrkA Prognostic
-NTRK1 expression in Neuroblastoma
17
REST 4q12 XBR, NRSF -REST and Neuroblastoma
17
PTER 10p12 HPHRP, RPR-1 -PTER and Neuroblastoma
15
NGFR 17q21-q22 CD271, p75NTR, TNFRSF16, p75(NTR), Gp80-LNGFR -NGFR and Neuroblastoma
13
VEGFA 6p12 VPF, VEGF, MVCD1 -VEGFA Expression in Neuroblastoma
13
FGF2 4q26 BFGF, FGFB, FGF-2, HBGF-2 -FGF2 and Neuroblastoma
13
ASCL1 12q23.2 ASH1, HASH1, MASH1, bHLHa46 -ASCL1 and Neuroblastoma
13
PARK2 6q25.2-q27 PDJ, PRKN, AR-JP, LPRS2 -PARK2 and Neuroblastoma
11
MAX 14q23 bHLHd4 -MAX and Neuroblastoma
11
KIF1B 1p36.2 KLP, CMT2, CMT2A, CMT2A1, HMSNII, NBLST1 -KIF1B and Neuroblastoma
11
CHD5 1p36.31 CHD-5 -CHD5 and Neuroblastoma
11
BARD1 2q35 -BARD1 polymorphisms in Neuroblastoma
10
IGF1R 15q26.3 IGFR, CD221, IGFIR, JTK13 -IGF1R Expression in Neuroblastoma
10
EFNB2 13q33 HTKL, EPLG5, Htk-L, LERK5 -EFNB2 expression in Neuroblastoma
10
ABCC1 16p13.1 MRP, ABCC, GS-X, MRP1, ABC29 Overexpression
-ABCC1 (MRP1) Overexpression in Neuroblastoma
9
VIP 6q25 PHM27 -VIP and Neuroblastoma
9
LMO1 11p15.4 TTG1, RBTN1, RHOM1 -LMO1 and Neuroblastoma
8
VEGFC 4q34.3 VRP, Flt4-L, LMPH1D -VEGFC Expression in Neuroblastoma
8
TOP1 20q12-q13.1 TOPI -TOP1 and Neuroblastoma
8
MAGEA1 Xq28 CT1.1, MAGE1 -MAGEA1 and Neuroblastoma
8
MAGEA3 Xq28 HIP8, HYPD, CT1.3, MAGE3, MAGEA6 -MAGEA3 and Neuroblastoma
8
TGFA 2p13 TFGA -TGFA Expression in Neuroblastoma
8
EPHB6 7q33-q35 HEP -EPHB6 and Neuroblastoma
8
BCHE 3q26.1-q26.2 E1, CHE1, CHE2 -BCHE and Neuroblastoma
7
TNFRSF10D 8p21 DCR2, CD264, TRUNDD, TRAILR4, TRAIL-R4 -TNFRSF10D and Neuroblastoma
7
ATRX Xq21.1 JMS, SHS, XH2, XNP, ATR2, SFM1, MRX52, RAD54, MRXHF1, RAD54L, ZNF-HX -ATRX and Neuroblastoma
7
NME2 17q21.3 PUF, NDKB, NDPKB, NM23B, NDPK-B, NM23-H2 -NME2 and Neuroblastoma
7
CAMTA1 1p36.31-p36.23 CANPMR -CAMTA1 and Neuroblastoma
7
EFNB3 17p13.1 EFL6, EPLG8, LERK8 -EFNB3 Expression in Neuroblastoma
6
BIN1 2q14 AMPH2, AMPHL, SH3P9 -Reduced BIN1 expression in MYCN amplified Neuroblastoma
6
CHAT 10q11.2 CMS6, CMS1A, CMS1A2, CHOACTASE -CHAT and Neuroblastoma
6
DLK1 14q32 DLK, FA1, ZOG, pG2, DLK-1, PREF1, Delta1, Pref-1 -DLK1 and Neuroblastoma
6
TP73 1p36.3 P73 -TP73 and Neuroblastoma
6
CDK5 7q36 LIS7, PSSALRE -CDK5 and Neuroblastoma
5
CHGA 14q32 CGA -CHGA and Neuroblastoma
5
NBL1 1p36.13 NB, DAN, NO3, DAND1, D1S1733E -NBL1 and Neuroblastoma
5
SOD1 21q22.11 ALS, SOD, ALS1, IPOA, hSod1, HEL-S-44, homodimer -SOD1 and Neuroblastoma
4
PIK3CD 1p36.2 APDS, PI3K, IMD14, p110D, P110DELTA -PIK3CD and Neuroblastoma
4
HOXC6 12q13.3 CP25, HOX3, HOX3C, HHO.C8 -HOXC6 and Neuroblastoma
4
VEGFB 11q13.1 VRF, VEGFL -VEGFB Expression in Neuroblastoma
4
CNTF 11q12.1 HCNTF -CNTF and Neuroblastoma
4
LIN28B 6q21 CSDD2 GWS
Prognostic
-LIN28B and Neuroblastoma
4
ZMYND10 3p21.3 BLU, FLU, CILD22 -ZMYND10 and Neuroblastoma
4
MIR107 10q23.31 MIRN107, miR-107 -MicroRNA mir-107 and Neuroblastoma
4
RBL2 16q12.2 Rb2, P130 -RBL2 and Neuroblastoma Differentiation
4
EPHB2 1p36.1-p35 DRT, EK5, ERK, CAPB, Hek5, PCBC, EPHT3, Tyro5 -EPHB2 Expression in Neuroblastoma
3
ODC1 2p25 ODC -ODC1 and Neuroblastoma
3
LMO4 1p22.3 -LMO4 and Neuroblastoma
3
TNFRSF25 1p36.2 DR3, TR3, DDR3, LARD, APO-3, TRAMP, WSL-1, WSL-LR, TNFRSF12 -TNFRSF25 and Neuroblastoma
3
CDK9 9q34.1 TAK, C-2k, CTK1, CDC2L4, PITALRE -CDK9 and Neuroblastoma
3
PDGFA 7p22 PDGF1, PDGF-A -PDGFA and Neuroblastoma
3
ENO1 1p36.2 NNE, PPH, MPB1, ENO1L1 -ENO1 and Neuroblastoma
3
TNFRSF10C 8p22-p21 LIT, DCR1, TRID, CD263, TRAILR3, TRAIL-R3, DCR1-TNFR -TNFRSF10C and Neuroblastoma
3
MYCL 1p34.2 LMYC, L-Myc, MYCL1, bHLHe38 -MYCL and Neuroblastoma
3
CD81 11p15.5 S5.7, CVID6, TAPA1, TSPAN28 -CD81 and Neuroblastoma
3
RASSF5 1q32.1 RAPL, Maxp1, NORE1, NORE1A, NORE1B, RASSF3 Methylation
-RASSF5 methylation in neuroblastoma
3
HACE1 6q16.3 GWAS
Prognostic
-HACE1 and Neuroblastoma
3
P2RX7 12q24 P2X7 -P2RX7 and Neuroblastoma
3
EXTL1 1p36.1 EXTL -EXTL1 and Neuroblastoma
3
SSTR2 17q24 -SSTR2 and Neuroblastoma
3
CDK7 5q12.1 CAK1, HCAK, MO15, STK1, CDKN7, p39MO15 -CDK7 and Neuroblastoma
3
MIRLET7E 19q13.41 LET7E, let-7e, MIRNLET7E, hsa-let-7e -MicroRNA let-7e and Neuroblastoma
2
SCFV 14 -SCFV and Neuroblastoma
2
LGI1 10q24 EPT, ETL1, ADLTE, ADPAEF, ADPEAF, IB1099, EPITEMPIN -LGI1 and Neuroblastoma
2
GAS7 17p13.1 MLL/GAS7 -GAS7 and Neuroblastoma
2
RASSF7 11p15.5 HRC1, HRAS1, C11orf13 Methylation
-RASSF7 methylation in neuroblastoma
2
MCF2 Xq27 DBL, ARHGEF21 -MCF2 and Neuroblastoma
2
TERC 3q26 TR, hTR, TRC3, DKCA1, PFBMFT2, SCARNA19 -TERC Expression in Neuroblastoma
2
MAPKAPK2 1q32 MK2, MK-2, MAPKAP-K2 -MAPKAPK2 and Neuroblastoma
2
ANGPT2 8p23.1 ANG2, AGPT2 -ANGPT2 Expression in Neuroblastoma
2
NEFL 8p21 NFL, NF-L, NF68, CMT1F, CMT2E, PPP1R110 -NEFL and Neuroblastoma
2
POU2F2 19q13.2 OCT2, OTF2, Oct-2 -POU2F2 and Neuroblastoma
2
GAGE1 Xp11.23 CT4.1, GAGE-1 -GAGE1 and Neuroblastoma
2
CD276 15q23-q24 B7H3, B7-H3, B7RP-2, 4Ig-B7-H3 -CD276 and Neuroblastoma
2
SRGAP3 3p25.3 WRP, MEGAP, SRGAP2, ARHGAP14 -SRGAP3 and Neuroblastoma
1
CCKBR 11p15.4 GASR, CCK-B, CCK2R -CCKBR and Neuroblastoma
1
SEPT7 7p14.2 CDC3, CDC10, SEPT7A, NBLA02942 -SEPT7 Expression in Neuroblastoma
1
IL24 1q32 C49A, FISP, MDA7, MOB5, ST16, IL10B -IL24 and Neuroblastoma
1
HOXD11 2q31.1 HOX4, HOX4F -HOXD11 and Neuroblastoma
1
PNN 14q21.1 DRS, DRSP, SDK3, memA -PNN and Neuroblastoma
1
RASSF6 4q13.3 Methylation
-RASSF6 methylation in neuroblastoma
1
LRRN2 1q32.1 GAC1, LRRN5, LRANK1, FIGLER7 -LRRN2 and Neuroblastoma
1
SSTR1 14q13 SS1R, SS1-R, SRIF-2, SS-1-R -SSTR1 Expression in Neuroblastoma
1
CDK12 17q12 CRK7, CRKR, CRKRS -CDK12 and Neuroblastoma
1
E2F3 6p22 E2F-3 -E2F3 and Neuroblastoma
1
EPAS1 2p21-p16 HLF, MOP2, ECYT4, HIF2A, PASD2, bHLHe73 -EPAS1 and Neuroblastoma
1
PAFAH1B2 11q23.3 HEL-S-303 -PAFAH1B2 and Neuroblastoma
1
MYBL2 20q13.1 BMYB, B-MYB Prognostic
-MYBL2 and Neuroblastoma

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications

Zheng J, Zhang R, Zhu J, et al.
Lack of Associations between XPC Gene Polymorphisms and Neuroblastoma Susceptibility in a Chinese Population.
Biomed Res Int. 2016; 2016:2932049 [PubMed] Free Access to Full Article Related Publications
Neuroblastoma is one of the most malignant solid tumors in infants and young children. No more than 40% of neuroblastoma patients can survive for longer than five years after it has been diagnosed. XPC protein is a pivotal factor that recognizes DNA damage and starts up the nucleotide excision repair (NER) in mammalian cells. This makes up the first group to defend against the cancer. Previous studies have identified that XPC gene polymorphisms were associated with various types of cancer. However, the associations between XPC gene polymorphisms and neuroblastoma risk have not yet been studied. We investigated the associations between three XPC gene polymorphisms (rs2228001 A>C, rs2228000 C>T, and rs2229090 G>C) and neuroblastoma risk with 256 neuroblastoma patients and 531 healthy controls in a Chinese Han population. Odds ratios and 95% confidence intervals were used to access the association between these three polymorphisms and neuroblastoma risk. No significant association was detected between these three polymorphisms and neuroblastoma risk in the overall analysis as well as in the stratification analysis. These results suggest that none of these three polymorphisms may be associated with the risk of neuroblastoma in the Chinese Han population.

Kawashima M, Kojima M, Ueda Y, et al.
Telomere biology including TERT rearrangements in neuroblastoma: a useful indicator for surgical treatments.
J Pediatr Surg. 2016; 51(12):2080-2085 [PubMed] Related Publications
PURPOSE: Our telomere biology study of neuroblastomas (NBLs) has revealed that unfavorable NBLs acquired telomere stabilization by telomerase activation or ALT (alternative lengthening of telomeres). Recently, genomic rearrangements in a region proximal to the telomerase reverse transcriptase (TERT) gene have been discovered in NBLs. In this study, TERT rearrangements were examined in NBLs along with their relationship to other aspects of telomere biology.
METHODS: In 121 NBLs, including 67 cases detected by mass-screening whose telomere length, telomerase activity, ALT with ATRX/DAXX alterations, and MYCN amplification were already known, TERT rearrangements were examined using GeneChip SNP arrays.
RESULTS: The 11 ATRX/DAXX mutated ALT cases and 29 cases with high telomerase activity showed poor prognosis. MYCN amplification and TERT rearrangements were independently detected in 16 and 13 cases, respectively, and these alterations were significantly correlated with high telomerase activity. In 81 infant cases, MYCN amplification, TERT rearrangements and ATRX mutations were detected in 3, 4, and 3 cases, respectively. Among them, 6 cases showed progression or recurrences.
CONCLUSIONS: Telomere stabilization in NBLs is acquired by telomerase activation through MYCN amplification, TERT rearrangements or by ALT. Since these tumors usually show progression and recurrence, complete resection should be considered, even in infant cases.
LEVEL OF EVIDENCE: Prognosis study, level III.

Li Z, Xu Z, Xie Q, et al.
miR-1303 promotes the proliferation of neuroblastoma cell SH-SY5Y by targeting GSK3β and SFRP1.
Biomed Pharmacother. 2016; 83:508-513 [PubMed] Related Publications
Neuroblastoma (NB) is one of the most common solid tumors in children, many microRNAs regulate progression and development of NB. Here, we found miR-1303 was upregulated in NB cells and tissues, miR-1303 overexpression promoted the proliferation of SH-SY5Y NB cell investigated by MTT assay, colony formation assay and anchorage-independent growth ability assay, while miR-1303 knockdown reduced this effect. mechanism analysis suggested glycogen synthase kinase 3 beta (GSK3β) and secreted frizzled-related protein 1 (SFRP1) were the target of miR-1303, luciferase assay revealed miR-1303 directly bound to the 3'UTR of GSK3β and SFRP1. miR-1303 increased expression of MYC and CyclinD1, and decreased the expression of p21 and p27, and further demonstrated miR-1303 promotes NB proliferation. Moreover, there was a negative correlation between miR-1303 expression and GSK3β and SFRP1 expression in NB tissues, confirming GSK3β and SFRP1 were the targets of miR-1303 in NB tissues. Collectively, our findings suggested miR-1303 promotes NB proliferation by targeting GSK3β and SFRP1, and might be a target for NB therapy.

Salet MC, Vogels R, Brons P, et al.
Maturation toward neuronal tissue in a Ewing sarcoma of bone after chemotherapy.
Diagn Pathol. 2016; 11(1):74 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Ewing sarcoma is the second most common bone tumor, occurring mainly in children and young adults. It shows a typical primitive, small round cell morphology and a characteristic fusion oncogene involving EWSR1 and members of the ETS family in most of the cases. Neuronal maturation after chemotherapy is a rare phenomenon and we herein describe such an exceptional case.
CASE PRESENTATION: An 8-year old boy was diagnosed with a Ewing sarcoma in the left femur. On biopsy the morphology was typical and there was an EWSR1-FLI1 gene fusion. He underwent neo-adjuvant chemotherapy and resection of the tumor. On microscopic evaluation, part of the tumor showed ganglioneuroblastoma-like differentiation with expression of neuronal markers. The continued presence of EWSR1 rearrangement in both the blue round cell component and the ganglioneuroblastoma-like component was shown by FISH analysis.
CONCLUSIONS: In conclusion, this case describes the possibility of a Ewing sarcoma to differentiate into a ganglioneuroblastoma-like lesion after neo-adjuvant chemotherapy treatment; the prognostic value of this phenomenon remains questionable.

Chen Y, Tsai YH, Tseng BJ, et al.
Suppression of miR-19b enhanced the cytotoxic effects of mTOR inhibitors in human neuroblastoma cells.
J Pediatr Surg. 2016; 51(11):1818-1825 [PubMed] Related Publications
BACKGROUND: Mammalian target of rapamycin (mTOR) inhibitors exert significant antitumor effects on several cancer cell types. In this study, we investigated the effects of mTOR inhibitors, in particular the regulation of the microRNA, in neuroblastoma cells.
METHODS: AZD8055 (a new mTOR inhibitor)- or rapamycin-induced cytotoxic effects on neuroblastoma cells were studied. Western blotting was used to investigate the expression of various proteins in the mTOR pathway. MicroRNA precursors and antagomirs were transfected into cells to manipulate the expression of target microRNA.
RESULTS: AZD8055 exerted stronger cytotoxic effects than rapamycin in neuroblastoma cells (p<0.03). In addition, AZD8055 suppressed the mTOR pathway and increased the expression of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in the neuroblastoma cells. AZD8055 significantly decreased miR-19b expression (p<0.005); in contrast, rapamycin increased miR-19b expression (p<0.05). Transfection of miR-19b antagomir into the neuroblastoma cells mimicked the effects of AZD8055 treatment, whereas miR-19b overexpression reversed the effects of AZD8055. Combination of miR-19b knockdown and rapamycin treatment significantly improved the sensitivity of neuroblastoma cells to rapamycin (p<0.02).
CONCLUSION: Suppression of miR-19b may enhance the cytotoxic effects of mTOR inhibitors in neuroblastoma cells.

Wachowiak R, Mayer S, Kaifi J, et al.
Prognostic Impact of Activated Leucocyte Cell Adhesion Molecule (ALCAM/CD166) in Infantile Neuroblastoma.
Anticancer Res. 2016; 36(8):3991-5 [PubMed] Related Publications
BACKGROUND/AIM: Activated leukocyte cell adhesion molecule (ALCAM/CD166) as a member of the 'immunoglobulin superfamily' is known to be involved in cancer cell proliferation and migration. The aim of this study was to investigate the expression of ALCAM in neuroblastoma tissues.
MATERIALS AND METHODS: ALCAM expression was analyzed in primary neuroblastoma specimens by immunohistochemistry on microarray sections. Histopathological and clinical data were correlated with ALCAM expression and survival analysis was performed.
RESULTS: Sixty-six children were included in the study. Strong expression of ALCAM was detected in 52 (79%) of the samples. Weak expression was significantly correlated with the International Neuroblastoma Staging System (INSS) stage (p=0.024) and positive n-MYC amplification (p=0.019). Recurrence-free survival (RFS) and overall survival (OS) were significantly shorter if ALCAM was expressed weakly (p=0.032 and p=0.001).
CONCLUSION: Weak ALCAM expression was significantly correlated with established markers for poor prognosis, as well as shorter RFS and OS. ALCAM might be considered as a prognostic marker for infantile neuroblastoma.

Powers JT, Tsanov KM, Pearson DS, et al.
Multiple mechanisms disrupt the let-7 microRNA family in neuroblastoma.
Nature. 2016; 535(7611):246-51 [PubMed] Free Access to Full Article Related Publications
Poor prognosis in neuroblastoma is associated with genetic amplification of MYCN. MYCN is itself a target of let-7, a tumour suppressor family of microRNAs implicated in numerous cancers. LIN28B, an inhibitor of let-7 biogenesis, is overexpressed in neuroblastoma and has been reported to regulate MYCN. Here we show, however, that LIN28B is dispensable in MYCN-amplified neuroblastoma cell lines, despite de-repression of let-7. We further demonstrate that MYCN messenger RNA levels in amplified disease are exceptionally high and sufficient to sponge let-7, which reconciles the dispensability of LIN28B. We found that genetic loss of let-7 is common in neuroblastoma, inversely associated with MYCN amplification, and independently associated with poor outcomes, providing a rationale for chromosomal loss patterns in neuroblastoma. We propose that let-7 disruption by LIN28B, MYCN sponging, or genetic loss is a unifying mechanism of neuroblastoma development with broad implications for cancer pathogenesis.

Sheikh A, Takatori A, Hossain MS, et al.
Unfavorable neuroblastoma prognostic factor NLRR2 inhibits cell differentiation by transcriptional induction through JNK pathway.
Cancer Sci. 2016; 107(9):1223-32 [PubMed] Free Access to Full Article Related Publications
The novel human gene family encoding neuronal leucine rich repeat (NLRR) proteins were identified as prognostic markers from our previous screening of primary neuroblastoma (NB) cDNA libraries. Of the NLRR gene family members, NLRR1 and NLRR3 are associated with the regulation of cellular proliferation and differentiation, respectively. However, the functional regulation and clinical significance of NLRR2 in NB remain unclear. Here, we evaluated the differential expression of NLRR2, where high expressions of NLRR2 were significantly associated with a poor prognosis of NB (P = 0.0009), in 78 NBs. Enforced expression of NLRR2 in NB cells enhanced cellular proliferation and induced resistance to retinoic acid (RA)-mediated cell growth inhibition. In contrast, knockdown of NLRR2 exhibited growth inhibition effects and enhanced RA-induced cell differentiation in NB cells. After RA treatment, NLRR2 expression was increased and correlated with the upregulation of c-Jun, a member of the activator protein-1 (AP-1) family in NB cells. Moreover, the expressions of NLRR2 and c-Jun were suppressed by treatment with a JNK inhibitor, which ameliorated the promoter activity of the NLRR2 gene while knockdown of c-Jun reduced NLRR2 expression. We then searched AP-1 binding consensus in the NLRR2 promoter region and confirmed c-Jun recruitment at a consensus. Conclusively, NLRR2 must be an inducible gene regulated by the JNK pathway to enhance cell survival and inhibit NB cell differentiation. Therefore, NLRR2 should have an important role in NB aggressiveness and be a potential therapeutic target for the treatment of RA resistant and aggressive NB.

Speleman F, Park JR, Henderson TO
Neuroblastoma: A Tough Nut to Crack.
Am Soc Clin Oncol Educ Book. 2016; 35:e548-57 [PubMed] Related Publications
Neuroblastoma, an embryonal tumor arising from neural crest-derived progenitor cells, is the most common solid tumor in childhood, with more than 700 cases diagnosed per year in the United States. In the past several decades, significant advances have been made in the treatment of neuroblastoma. Treatment advances reflect improved understanding of the biology of neuroblastoma. Although amplification of MYCN was discovered in the early 1980s, our understanding of neuroblastoma oncogenesis has advanced in the last decade as a result of high-throughput genomic analysis, exome and whole-genome sequencing, genome-wide association studies, and synthetic lethal drug screens. Our refined understanding of neuroblastoma biology and genetics is reflected in improved prognostic stratification and appropriate tailoring of therapy in recent clinical trials. Moreover, for high-risk neuroblastoma, a disease that was uniformly fatal 3 decades ago, recent clinical trials incorporating autologous hematopoietic transplant and immunotherapy utilizing anti-GD2 antibody plus cytokines have shown improved event-free and overall survival. These advances have resulted in a growing population of long-term survivors of neuroblastoma. Examination of the late effects and second malignant neoplasms (SMNs) in both older generations of survivors and more recently treated survivors will inform both design of future trials and surveillance guidelines for long-term follow-up. As a consequence of advances in understanding of the biology of neuroblastoma, successful clinical trials, and refined understanding of the late effects and SMNs of survivors, the promise of precision medicine is becoming a reality for patients with neuroblastoma.

Walsh KM, Whitehead TP, de Smith AJ, et al.
Common genetic variants associated with telomere length confer risk for neuroblastoma and other childhood cancers.
Carcinogenesis. 2016; 37(6):576-82 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
Aberrant telomere lengthening is an important feature of cancer cells in adults and children. In addition to somatic mutations, germline polymorphisms in telomere maintenance genes impact telomere length. Whether these telomere-associated polymorphisms affect risk of childhood malignancies remains largely unexplored. We collected genome-wide data from three groups with pediatric malignancies [neuroblastoma (N = 1516), acute lymphoblastic leukemia (ALL) (N = 958) and osteosarcoma (N = 660)] and three control populations (N = 6892). Using case-control comparisons, we analyzed eight single nucleotide polymorphisms (SNPs) in genes definitively associated with interindividual variation in leukocyte telomere length (LTL) in prior genome-wide association studies: ACYP2, TERC, NAF1, TERT, OBFC1, CTC1, ZNF208 and RTEL1 Six of these SNPs were associated (P < 0.05) with neuroblastoma risk, one with leukemia risk and one with osteosarcoma risk. The allele associated with longer LTL increased cancer risk for all these significantly associated SNPs. Using a weighted linear combination of the eight LTL-associated SNPs, we observed that neuroblastoma patients were predisposed to longer LTL than controls, with each standard deviation increase in genotypically estimated LTL associated with a 1.15-fold increased odds of neuroblastoma (95%CI = 1.09-1.22; P = 7.9×10(-7)). This effect was more pronounced in adolescent-onset neuroblastoma patients (OR = 1.46; 95%CI = 1.03-2.08). A one standard deviation increase in genotypically estimated LTL was more weakly associated with osteosarcoma risk (OR = 1.10; 95%CI = 1.01-1.19; P = 0.017) and leukemia risk (OR = 1.07; 95%CI = 1.00-1.14; P = 0.044), specifically for leukemia patients who relapsed (OR = 1.19; 95%CI = 1.01-1.40; P = 0.043). These results indicate that genetic predisposition to longer LTL is a newly identified risk factor for neuroblastoma and potentially for other cancers of childhood.

Wang L, Ding Y, Wei L, et al.
Recurrent Olfactory Neuroblastoma Treated With Cetuximab and Sunitinib: A Case Report.
Medicine (Baltimore). 2016; 95(18):e3536 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
Olfactory neuroblastoma (ONB) is a rare cancer originating in the olfactory epithelium of the nasal vault. The recurrence rate of ONB is high, as the standard treatment of surgery followed by radiotherapy and/or chemotherapy is usually unsuccessful. The use of targeted therapy based on individual genomic variations after cancer relapse has not been reported. Here, we present the case of a 44-year-old man who was diagnosed with recurrent ONB and treated with a regimen developed using whole exome sequencing. Potential targets were first identified and then matched to appropriate drugs. Gene mutations in the genes encoding EGFR, FGFR2, KDR, and RET were discovered in the patient's tumor tissue by whole exome sequencing and the patient was treated with a combination of the targeted drugs cetuximab and sunitinib. Five days after treatment, enhancement magnetic resonance imaging showed a 65% reduction in tumor size, and the Visual analog scale headache scores went down to 2/10 from 10/10. Repeat imaging at 1 month showed a complete response.This study represents the first demonstration of an effective personalized treatment of ONB by targeted drugs, and sheds light on how precision medicine can be used to treat recurrent ONB that fails to respond to routine tumor resection, radiotherapy, and/or chemotherapy.

Kumar MD, Dravid A, Kumar A, Sen D
Gene therapy as a potential tool for treating neuroblastoma-a focused review.
Cancer Gene Ther. 2016; 23(5):115-24 [PubMed] Related Publications
Neuroblastoma, a solid tumor caused by rapid division of undifferentiated neuroblasts, is the most common childhood malignancy affecting children aged <5 years. Several approaches and strategies developed and tested to cure neuroblastoma have met with limited success due to different reasons. Many oncogenes are deregulated during the onset and development of neuroblastoma and thus offer an opportunity to circumvent this disease if the expression of these genes is restored to normalcy. Gene therapy is a powerful tool with the potential to inhibit the deleterious effects of oncogenes by inserting corrected/normal genes into the genome. Both viral and non-viral vector-based gene therapies have been developed and adopted to deliver the target genes into neuroblastoma cells. These attempts have given hope to bringing in a new regime of treatment against neuroblastoma. A few gene-therapy-based treatment strategies have been tested in limited clinical trials yielding some positive results. This mini review is an attempt to provide an overview of the available options of gene therapy to treat neuroblastoma.

Tashnizi AH, Jaberipour M, Razmkhah M, et al.
Tumour suppressive effects of WEE1 gene silencing in neuroblastomas.
J Cancer Res Ther. 2016 Jan-Mar; 12(1):221-7 [PubMed] Related Publications
AIM OF STUDY: WEE1, a member of serine/threonine protein kinase family is the master inhibitor of cyclin-dependent kinase 1 in cell cycle. Over-expression of WEE1 in glioblastomas (GBMs) and some other cancers has been shown. Here, we investigated the expression of WEE1 in 13 brain samples from GBM patients and two GBM cell lines. Further to that, we asked whether if knocking down WEE1 expression in the cell lines change tumor cells' reaction.
MATERIALS AND METHODS: All brain tumor samples were collected after confirmed pathological diagnosis. Western blotting was used to screen the expression of WEE1 and a panel of tumor markers. As a model of WEE1 gene silencing with small hairpin RNA (shRNA) technology in GBMs, A172, and U373GM cell lines were transfected with four WEE1 specific shRNAs. The growth characteristics of the cells and the expression of a panel of downstream genes were investigated after gene suppression.
RESULTS: All GBMs and both cell lines over-expressed WEE1. Transduction of the cell lines with different shRNAs suppressed WEE1 expression with different extent and pooling of four shRNAs together resulted in additive effect. Suppression of WEE1 not only repressed cellular growth but also changed the profile of gene expression of the cells. Quantitative real-time polymerase chain reaction showed also reduced expression of genes such as hypoxia-inducible factor-1, B-cell lymphoma-2, vascular endothelial growth factor, and p53 with crucial roles in tumor survival and invasiveness.
CONCLUSION: These results highlight the key role of WEE1 suppression to combat GBMs. Moreover, it showed beneficial possibilities of WEE1 suppression with different anticancer approaches for neurological malignancies.

Dyberg C, Papachristou P, Haug BH, et al.
Planar cell polarity gene expression correlates with tumor cell viability and prognostic outcome in neuroblastoma.
BMC Cancer. 2016; 16:259 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
BACKGROUND: The non-canonical Wnt/Planar cell polarity (PCP) signaling pathway is a major player in cell migration during embryonal development and has recently been implicated in tumorigenesis.
METHODS: Transfections with cDNA plasmids or siRNA were used to increase and suppress Prickle1 and Vangl2 expression in neuroblastoma cells and in non-tumorigenic cells. Cell viability was measured by trypan blue exclusion and protein expression was determined with western blotting. Transcriptional activity was studied with luciferase reporter assay and mRNA expression with real-time RT-PCR. Immunofluorescence stainings were used to study the effects of Vangl2 overexpression in non-tumorigenic embryonic cells. Statistical significance was tested with t-test or one-way ANOVA.
RESULTS: Here we show that high expression of the PCP core genes Prickle1 and Vangl2 is associated with low-risk neuroblastoma, suppression of neuroblastoma cell growth and decreased Wnt/β-catenin signaling. Inhibition of Rho-associated kinases (ROCKs) that are important in mediating non-canonical Wnt signaling resulted in increased expression of Prickle1 and inhibition of β-catenin activity in neuroblastoma cells. In contrast, overexpression of Vangl2 in MYC immortalized neural stem cells induced accumulation of active β-catenin and decreased the neural differentiation marker Tuj1. Similarly, genetically modified mice with forced overexpression of Vangl2 in nestin-positive cells showed decreased Tuj1 differentiation marker during embryonal development.
CONCLUSIONS: Our experimental data demonstrate that high expression of Prickle1 and Vangl2 reduce the growth of neuroblastoma cells and indicate different roles of PCP proteins in tumorigenic cells compared to normal cells. These results suggest that the activity of the non-canonical Wnt/PCP signaling pathway is important for neuroblastoma development and that manipulation of the Wnt/PCP pathway provides a possible therapy for neuroblastoma.

Braekeveldt N, Wigerup C, Tadeo I, et al.
Neuroblastoma patient-derived orthotopic xenografts reflect the microenvironmental hallmarks of aggressive patient tumours.
Cancer Lett. 2016; 375(2):384-9 [PubMed] Related Publications
Treatment of high-risk childhood neuroblastoma is a clinical challenge which has been hampered by a lack of reliable neuroblastoma mouse models for preclinical drug testing. We have previously established invasive and metastasising patient-derived orthotopic xenografts (PDXs) from high-risk neuroblastomas that retained the genotypes and phenotypes of patient tumours. Given the important role of the tumour microenvironment in tumour progression, metastasis, and treatment responses, here we analysed the tumour microenvironment of five neuroblastoma PDXs in detail. The PDXs resembled their parent tumours and retained important stromal hallmarks of aggressive lesions including rich blood and lymphatic vascularisation, pericyte coverage, high numbers of cancer-associated fibroblasts, tumour-associated macrophages, and extracellular matrix components. Patient-derived tumour endothelial cells occasionally formed blood vessels in PDXs; however, tumour stroma was, overall, of murine origin. Lymphoid cells and lymphatic endothelial cells were found in athymic nude mice but not in NSG mice; thus, the choice of mouse strain dictates tumour microenvironmental components. The murine tumour microenvironment of orthotopic neuroblastoma PDXs reflects important hallmarks of aggressive and metastatic clinical neuroblastomas. Neuroblastoma PDXs are clinically relevant models for preclinical drug testing.

Cavalli M, Pan G, Nord H, et al.
Allele-specific transcription factor binding to common and rare variants associated with disease and gene expression.
Hum Genet. 2016; 135(5):485-97 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
Genome-wide association studies (GWAS) have identified a large number of disease-associated SNPs, but in few cases the functional variant and the gene it controls have been identified. To systematically identify candidate regulatory variants, we sequenced ENCODE cell lines and used public ChIP-seq data to look for transcription factors binding preferentially to one allele. We found 9962 candidate regulatory SNPs, of which 16 % were rare and showed evidence of larger functional effect than common ones. Functionally rare variants may explain divergent GWAS results between populations and are candidates for a partial explanation of the missing heritability. The majority of allele-specific variants (96 %) were specific to a cell type. Furthermore, by examining GWAS loci we found >400 allele-specific candidate SNPs, 141 of which were highly relevant in our cell types. Functionally validated SNPs support identification of an SNP in SYNGR1 which may expose to the risk of rheumatoid arthritis and primary biliary cirrhosis, as well as an SNP in the last intron of COG6 exposing to the risk of psoriasis. We propose that by repeating the ChIP-seq experiments of 20 selected transcription factors in three to ten people, the most common polymorphisms can be interrogated for allele-specific binding. Our strategy may help to remove the current bottleneck in functional annotation of the genome.

Taran K, Wysocka A, Sitkiewicz A, et al.
Evaluation of potential prognostic value of Bmi-1 gene product and selected markers of proliferation (Ki-67) and apoptosis (p53) in the neuroblastoma group of tumors.
Postepy Hig Med Dosw (Online). 2016; 70:110-6 [PubMed] Related Publications
INTRODUCTION: Cancer in children is a very important issue in pediatrics. The least satisfactory treatment outcome occurs among patients with clinically advanced neuroblastomas. Despite much research, the biology of this tumor still remains unclear, and new prognostic factors are sought. The Bmi-1 gene product is a currently highly investigated protein which belongs to the Polycomb group (PcG) and has been identified as a regulator of primary neural crest cells. It is believed that Bmi‑1 and N-myc act together and are both involved in the pathogenesis of neuroblastoma. The aim of the study was to assess the potential prognostic value of Bmi-1 protein and its relations with mechanisms of proliferation and apoptosis in the neuroblastoma group of tumors.
MATERIAL/METHODS: 29 formalin-fixed and paraffin-embedded neuroblastoma tissue sections were examined using mouse monoclonal antibodies anti-Bmi-1, anti-p53 and anti-Ki-67 according to the manufacturer's instructions.
RESULTS: There were found statistically significant correlations between Bmi-1 expression and tumor histology and age of patients.
CONCLUSIONS: Bmi-1 seems to be a promising marker in the neuroblastoma group of tumors whose expression correlates with widely accepted prognostic parameters. The pattern of BMI-1 expression may indicate that the examined protein is also involved in maturation processes in tumor tissue.

Zhang R, Zou Y, Zhu J, et al.
The Association between GWAS-identified BARD1 Gene SNPs and Neuroblastoma Susceptibility in a Southern Chinese Population.
Int J Med Sci. 2016; 13(2):133-8 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
A previous genome-wide association study (GWAS) has found that some common variations in the BARD1 gene were associated with neuroblastoma susceptibility especially for high-risk subjects, and the associations have been validated in Caucasians and African-Americans. However, the associations between BARD1 gene polymorphisms and neuroblastoma susceptibility have not been studied among Asians, not to mention Chinese subjects. In the present study, we investigated the association of three BARD1 polymorphisms (rs7585356 G>A, rs6435862 T>G and rs3768716 A>G) with neuroblastoma susceptibility in 201 neuroblastoma patients and 531 controls using TaqMan methodology. Overall, none of these polymorphisms was significantly associated with neuroblastoma susceptibility. However, stratified analysis showed a more profound association between neuroblastoma risk and rs6435862 TG/GG variant genotypes among older children (adjusted OR=1.55, 95% CI=1.04-2.31), and children with adrenal gland-originated disease (adjusted OR=2.94, 95% CI=1.40-6.18), or with ISSN clinical stages III+IV disease (adjusted OR=1.75, 95% CI=1.09-2.84). Similar results were observed for the variant genotypes of rs3768716 A>G polymorphism among these three subgroups. Our results suggest that the BARD1 rs6435862 T>G and rs3768716 A>G polymorphisms may contribute to increased susceptibility to neuroblastoma, especially for the subjects at age ≥12 months, with adrenal gland-originated or with late clinical stage neuroblastoma. These findings need further validation by prospective studies with larger sample size with subjects enrolled from multicenter, involving different ethnicities.

Woodfield SE, Zhang L, Scorsone KA, et al.
Binimetinib inhibits MEK and is effective against neuroblastoma tumor cells with low NF1 expression.
BMC Cancer. 2016; 16:172 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
BACKGROUND: Novel therapies are needed for children with high-risk and relapsed neuroblastoma. We hypothesized that MAPK/ERK kinase (MEK) inhibition with the novel MEK1/2 inhibitor binimetinib would be effective in neuroblastoma preclinical models.
METHODS: Levels of total and phosphorylated MEK and extracellular signal-regulated kinase (ERK) were examined in primary neuroblastoma tumor samples and in neuroblastoma cell lines by Western blot. A panel of established neuroblastoma tumor cell lines was treated with increasing concentrations of binimetinib, and their viability was determined using MTT assays. Western blot analyses were performed to examine changes in total and phosphorylated MEK and ERK and to measure apoptosis in neuroblastoma tumor cells after binimetinib treatment. NF1 protein levels in neuroblastoma cell lines were determined using Western blot assays. Gene expression of NF1 and MEK1 was examined in relationship to neuroblastoma patient outcomes.
RESULTS: Both primary neuroblastoma tumor samples and cell lines showed detectable levels of total and phosphorylated MEK and ERK. IC50 values for cells sensitive to binimetinib ranged from 8 nM to 1.16 μM, while resistant cells did not demonstrate any significant reduction in cell viability with doses exceeding 15 μM. Sensitive cells showed higher endogenous expression of phosphorylated MEK and ERK. Gene expression of NF1, but not MEK1, correlated with patient outcomes in neuroblastoma, and NF1 protein expression also correlated with responses to binimetinib.
CONCLUSIONS: Neuroblastoma tumor cells show a range of sensitivities to the novel MEK inhibitor binimetinib. In response to binimetinib, sensitive cells demonstrated complete loss of phosphorylated ERK, while resistant cells demonstrated either incomplete loss of ERK phosphorylation or minimal effects on MEK phosphorylation, suggesting alternative mechanisms of resistance. NF1 protein expression correlated with responses to binimetinib, supporting the use of NF1 as a biomarker to identify patients that may respond to MEK inhibition. MEK inhibition therefore represents a potential new therapeutic strategy for neuroblastoma.

Ramraj SK, Aravindan S, Somasundaram DB, et al.
Serum-circulating miRNAs predict neuroblastoma progression in mouse model of high-risk metastatic disease.
Oncotarget. 2016; 7(14):18605-19 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
BACKGROUND: Circulating miRNAs have momentous clinical relevance as prognostic biomarkers and in the progression of solid tumors. Recognizing novel candidates of neuroblastoma-specific circulating miRNAs would allow us to identify potential prognostic biomarkers that could predict the switch from favorable to high-risk metastatic neuroblastoma (HR-NB).
RESULTS: Utilizing mouse models of favorable and HR-NB and whole miRnome profiling, we identified high serum levels of 34 and low levels of 46 miRNAs in animals with HR-NB. Preferential sequence homology exclusion of mouse miRNAs identified 25 (11 increased; 14 decreased) human-specific prognostic marker candidates, of which, 21 were unique to HR-NB. miRNA QPCR validated miRnome profile. Target analysis defined the candidate miRNAs' signal transduction flow-through and demonstrated their converged roles in tumor progression. miRNA silencing studies verified the function of select miRNAs on the translation of at least 14 target proteins. Expressions of critical targets that correlate tumor progression in tissue of multifarious organs identify the orchestration of HR-NB. Significant (>10 fold) increase in serum levels of miR-381, miR-548h, and miR-580 identify them as potential prognostic markers for neuroblastoma progression.
CONCLUSION: For the first time, we identified serum-circulating miRNAs that predict the switch from favorable to HR-NB and, further imply that these miRNAs could play a functional role in tumor progression.

Bogen D, Brunner C, Walder D, et al.
The genetic tumor background is an important determinant for heterogeneous MYCN-amplified neuroblastoma.
Int J Cancer. 2016; 139(1):153-63 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
Amplification of MYCN is the signature genetic aberration of 20-25% of neuroblastoma and a stratifying marker associated with aggressive tumor behavior. The detection of heterogeneous MYCN amplification (hetMNA) poses a diagnostic dilemma due to the uncertainty of its relevance to tumor behavior. Here, we aimed to shed light on the genomic background which permits hetMNA in neuroblastoma and tied the occurrence to other stratifying markers and disease outcome. We performed SNP analysis using Affymetrix Cytoscan HD arrays on 63 samples including constitutional DNA, tumor, bone marrow and relapse samples of 26 patients with confirmed hetMNA by MYCN-FISH. Tumors of patients ≤18m were mostly aneuploid with numeric chromosomal aberrations (NCAs), presented a prominent MNA subclone and carried none or a few segmental chromosomal aberrations (SCAs). In older patients, tumors were mostly di- or tetraploid, contained a lower number of MNA cells and displayed a multitude of SCAs including concomitant 11q deletions. These patients often suffered disease progression, tumor dissemination and relapse. Restricted to aneuploid tumors, we detected chromosomes with uniparental di- or trisomy (UPD/UPT) in almost every sample. UPD11 was exclusive to tumors of younger patients whereas older patients featured UPD14. In this study, the MNA subclone appears to be constraint by the tumor environment and thus less relevant for tumor behavior in aggressive tumors with a high genomic instability and many segmental aberrations. A more benign tumor background and lower tumor stage may favor an outgrowth of the MNA clone but tumors generally responded better to treatment.

Pinto N, Mayfield JR, Raca G, et al.
Segmental Chromosomal Aberrations in Localized Neuroblastoma Can be Detected in Formalin-Fixed Paraffin-Embedded Tissue Samples and Are Associated With Recurrence.
Pediatr Blood Cancer. 2016; 63(6):1019-23 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
BACKGROUND: Array comparative genomic hybridization (CGH) analyses of frozen tumors have shown strong associations between the pattern of chromosomal aberrations and outcome in patients with advanced-stage neuroblastoma. New platforms for analyzing chromosomal aberrations using formalin-fixed paraffin-embedded (FFPE) tissue have recently been developed. We sought to determine whether chromosomal microarray analysis (CMA) using FFPE tumors is feasible and if segmental chromosomal aberrations were prognostic of recurrence in localized neuroblastoma.
METHODS: Patients with MYCN nonamplified International Neuroblastoma Staging System stage 1 and 2 disease who recurred were identified. CMA was performed with diagnostic FFPE samples using OncoScan™ FFPE Express 2.0. The prognostic significance of chromosomal pattern was validated in 105 patients with available CGH results.
RESULTS: In 26 evaluable patients, 11 recurred locally, nine had metastatic relapse, and six remained progression free >3 years from diagnosis. No chromosomal aberrations were identified in four tumors. Numerical chromosomal aberrations (NCAs) without segmental chromosomal aberration (SCA) were identified in 11 patients: six progressed locally, two had metastatic progression and 3 remained progression-free. Eleven patients had SCAs: four progressed locally, six developed metastatic progression and one remained progression-free. Five or more SCAs were only detected in tumors from patients who developed metastases (P = 0.0004). In the validation cohort, SCAs were associated with inferior event-free survival (EFS) compared to NCA (5-year EFS 68% ± 8.3% vs. 91% ± 3.6%, respectively; P = 0.0083).
CONCLUSIONS: It is feasible to evaluate chromosomal aberrations using FFPE neuroblastoma tissue. SCA is associated with inferior EFS in localized neuroblastoma patients, and multiple SCAs may be predictive of metastatic relapse.

Gu L, Chu P, Lingeman R, et al.
The Mechanism by Which MYCN Amplification Confers an Enhanced Sensitivity to a PCNA-Derived Cell Permeable Peptide in Neuroblastoma Cells.
EBioMedicine. 2015; 2(12):1923-31 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
Dysregulated expression of MYC family genes is a hallmark of many malignancies. Unfortunately, these proteins are not amenable to blockade by small molecules or protein-based therapeutic agents. Therefore, we must find alternative approaches to target MYC-driven cancers. Amplification of MYCN, a MYC family member, predicts high-risk neuroblastoma (NB) disease. We have shown that R9-caPep blocks the interaction of PCNA with its binding partners and selectively kills human NB cells, especially those with MYCN amplification, and we now show the mechanism. We found elevated levels of DNA replication stress in MYCN-amplified NB cells. R9-caPep exacerbated DNA replication stress in MYCN-amplified NB cells and NB cells with an augmented level of MYC by interfering with DNA replication fork extension, leading to Chk1 dependence and susceptibility to Chk1 inhibition. We describe how these effects may be exploited for treating NB.

Naftali O, Maman S, Meshel T, et al.
PHOX2B is a suppressor of neuroblastoma metastasis.
Oncotarget. 2016; 7(9):10627-37 [PubMed] Article available free on PMC after 01/06/2017 Related Publications
Paired like homeobox 2B (PHOX2B) is a minimal residual disease (MRD) marker of neuroblastoma. The presence of MRD, also referred to as micro-metastases, is a powerful marker of poor prognosis in neuroblastoma. Lung metastasis is considered a terminal event in neuroblastoma. Lung micro-metastatic neuroblastoma (MicroNB) cells show high expression levels of PHOX2B and possess a less malignant and metastatic phenotype than lung macro metastatic neuroblastoma (MacroNB) cells, which hardly express PHOX2B. In vitro assays showed that PHOX2B knockdown in MicroNB cells did not affect cell viability; however it decreased the migratory capacity of the MicroNB-shPHOX2B cells. An orthotopic inoculation of MicroNB-shPHOX2B cells into the adrenal gland of nude mice resulted in significantly larger primary tumors and a heavier micro-metastatic load in the lungs and bone-marrow, than when control cells were inoculated. PHOX2B expression was found to be regulated by methylation. The PHOX2B promoter in MacroNB cells is significantly more methylated than in MicroNB cells. Demethylation assays using 5-azacytidine demonstrated that methylation can indeed inhibit PHOX2B transcription in MacroNB cells. These pre-clinical data strongly suggest that PHOX2B functions as a suppressor of neuroblastoma progression.

Whittle SB, Reyes S, Du M, et al.
A Polymorphism in the FGFR4 Gene Is Associated With Risk of Neuroblastoma and Altered Receptor Degradation.
J Pediatr Hematol Oncol. 2016; 38(2):131-8 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Outcomes for children with high-risk neuroblastoma are poor, and improved understanding of the mechanisms underlying neuroblastoma pathogenesis, recurrence, and treatment resistance will lead to improved outcomes. Aberrant growth factor receptor expression and receptor tyrosine kinase signaling are associated with the pathogenesis of many malignancies. A germline polymorphism in the FGFR4 gene is associated with increased receptor expression and activity and with decreased survival, treatment resistance, and aggressive disease for many malignancies. We therefore investigated the role of this FGFR4 polymorphism in neuroblastoma pathogenesis.
MATERIALS AND METHODS: Germline DNA from neuroblastoma patients and matched controls was assessed for the FGFR4 Gly/Arg388 polymorphism by RT-PCR. Allele frequencies were assessed for association with neuroblastoma patient outcomes and prognostic features. Degradation rates of the FGFR4 Arg388 and Gly388 receptors and rates of receptor internalization into the late endosomal compartment were measured.
RESULTS: Frequency of the FGFR4 AA genotype and the prevalence of the A allele were significantly higher in patients with neuroblastoma than in matched controls. The Arg388 receptor demonstrated slower degradation than the Gly388 receptor in neuroblastoma cells and reduced internalization into multivesicular bodies.
CONCLUSIONS: The FGFR4 Arg388 polymorphism is associated with an increased prevalence of neuroblastoma in children, and this association may be linked to differences in FGFR4 degradation rates. Our study provides the first evidence of a role for FGFR4 in neuroblastoma, suggesting that FGFR4 genotype and the pathways regulating FGFR4 trafficking and degradation may be relevant for neuroblastoma pathogenesis.

Decock A, Ongenaert M, Van Criekinge W, et al.
DNA methylation profiling of primary neuroblastoma tumors using methyl-CpG-binding domain sequencing.
Sci Data. 2016; 3:160004 [PubMed] Free Access to Full Article Related Publications
Comprehensive genome-wide DNA methylation studies in neuroblastoma (NB), a childhood tumor that originates from precursor cells of the sympathetic nervous system, are scarce. Recently, we profiled the DNA methylome of 102 well-annotated primary NB tumors by methyl-CpG-binding domain (MBD) sequencing, in order to identify prognostic biomarker candidates. In this data descriptor, we give details on how this data set was generated and which bioinformatics analyses were applied during data processing. Through a series of technical validations, we illustrate that the data are of high quality and that the sequenced fragments represent methylated genomic regions. Furthermore, genes previously described to be methylated in NB are confirmed. As such, these MBD sequencing data are a valuable resource to further study the association of NB risk factors with the NB methylome, and offer the opportunity to integrate methylome data with other -omic data sets on the same tumor samples such as gene copy number and gene expression, also publically available.

Soriano A, París-Coderch L, Jubierre L, et al.
MicroRNA-497 impairs the growth of chemoresistant neuroblastoma cells by targeting cell cycle, survival and vascular permeability genes.
Oncotarget. 2016; 7(8):9271-87 [PubMed] Free Access to Full Article Related Publications
Despite multimodal therapies, a high percentage of high-risk neuroblastoma (NB) become refractory to current treatments, most of which interfere with cell cycle and DNA synthesis or function, activating the DNA damage response (DDR). In cancer, this process is frequently altered by deregulated expression or function of several genes which contribute to multidrug resistance (MDR). MicroRNAs are outstanding candidates for therapy since a single microRNA can modulate the expression of multiple genes of the same or different pathways, thus hindering the development of resistance mechanisms by the tumor. We found several genes implicated in the MDR to be overexpressed in high-risk NB which could be targeted by microRNAs simultaneously. Our functional screening identified several of those microRNAs that reduced proliferation of chemoresistant NB cell lines, the best of which was miR-497. Low expression of miR-497 correlated with poor patient outcome. The overexpression of miR-497 reduced the proliferation of multiple chemoresistant NB cell lines and induced apoptosis in MYCN-amplified cell lines. Moreover, the conditional expression of miR-497 in NB xenografts reduced tumor growth and inhibited vascular permeabilization. MiR-497 targets multiple genes related to the DDR, cell cycle, survival and angiogenesis, which renders this molecule a promising candidate for NB therapy.

Kiessling MK, Curioni-Fontecedro A, Samaras P, et al.
Targeting the mTOR Complex by Everolimus in NRAS Mutant Neuroblastoma.
PLoS One. 2016; 11(1):e0147682 [PubMed] Free Access to Full Article Related Publications
High-risk neuroblastoma remains lethal in about 50% of patients despite multimodal treatment. Recent attempts to identify molecular targets for specific therapies have shown that Neuroblastoma RAS (NRAS) is significantly mutated in a small number of patients. However, few inhibitors for the potential treatment for NRAS mutant neuroblastoma have been investigated so far. In this in-vitro study, we show that MEK inhibitors AZD6244, MEK162 and PD0325901 block cell growth in NRAS mutant neuroblastoma cell lines but not in NRAS wild-type cell lines. Several studies show that mutant NRAS leads to PI3K pathway activation and combined inhibitors of PI3K/mTOR effectively block cell growth. However, we observed the combination of MEK inhibitors with PI3K or AKT inhibitors did not show synergestic effects on cell growth. Thus, we tested single mTOR inhibitors Everolimus and AZD8055. Interestingly, Everolimus and AZD8055 alone were sufficient to block cell growth in NRAS mutant cell lines but not in wild-type cell lines. We found that Everolimus alone induced apoptosis in NRAS mutant neuroblastoma. Furthermore, the combination of mTOR and MEK inhibitors resulted in synergistic growth inhibition. Taken together, our results show that NRAS mutant neuroblastoma can be targeted by clinically available Everolimus alone or in combination with MEK inhibitors which could impact future clinical studies.

Hu L, Uzhameckis D, Hedborg F, Blomberg J
Dynamic and selective HERV RNA expression in neuroblastoma cells subjected to variation in oxygen tension and demethylation.
APMIS. 2016 Jan-Feb; 124(1-2):140-9 [PubMed] Related Publications
We studied HERV expression in cell lines after hypoxia, mitogenic stimulation, and demethylation, to better understand if hypoxia may play a role in ERV activation also within the nervous system, as represented by neuroblastoma cell lines. The level of RNA of four human ERV groups (HERVs) (HERVE, I/T, H, and W), and three housekeeping genes, of different cell lines including A549, COS-1, Namalwa, RD-L and Vero-E6, as well as human neuroblastoma cell lines SH-SY5Y, SK-N-DZ, and SK-N-AS were studied using reverse transcription and real-time quantitative PCR (QPCR). During the course of recovery from hypoxia a pronounced and selective activation of RNA expression of HERVW-like sequences, but not of HERVE, I/T, H, and three housekeeping genes, was found in the neuroblastoma cell lines, most pronounced in SK-N-DZ. In the SK-N-DZ cell line, we also tested the expression of HERVs after chemical treatments. HERVW-like sequences were selectively upregulated by 5-azacytidine, a demethylating agent. Some HERVW loci seem especially responsive to hypoxia and demethylation. HERV expression in neuroblastoma cells is selectively and profoundly influenced by some physiological and chemical stimuli.

H'Mida Ben Brahim D, Trabelsi S, Chabchoub I, et al.
Assessment of MYCN amplification status in Tunisian neuroblastoma: CISH and MLPA combining approach.
Tunis Med. 2015 Aug-Sep; 93(8-9):527-31 [PubMed] Related Publications
BACKGROUND: Neuroblastoma (NB) shows a complex combination of genetic aberrations. Some of them represent poor genetic prognosis factors that require specific and intensive chemotherapy. MYCN amplification consists of the major bad outcome prognostic factor, it is indeed frequently observed in aggressive neuroblastomas. To date different methods are used for MYCN status detection.
OBJECTIVES: The primary aim of our study was to provide a critical assessment of MYCN status using 2 molecular techniques CISH and MLPA. We also focused on the correlation between neuroblastoma genetic markers and patient's clinical course among 15 Tunisian patients.
METHODS: we developed a descriptive study that includes 15 pediatric Tunisian patients referred to our laboratory from 2004 to 2011. We reported the analysis of fresh and FFPE NB tumors tissues.
RESULTS: No significant correlation was found between COG grade and patients overall survival. Assessment of NMYC gene copy number by kappa statistic test revealed high concordance between CISH and MLPA tests (kappa coefficient = 0.02).
CONCLUSION: Despite misdiagnosing of MYCN status fewer than 5 copies, MLPA remains an effective molecular technique that enables a large panel of genomic aberrations screening. Thus combining CISH and MLPA is an effective molecular approach adopted in our laboratory. Our results allow pediatric oncologists to set up the first Neuroblastoma therapeutic strategy based on molecular markers in Tunisia.

Recurrent Chromosome Abnormalities

Selected list of common recurrent structural abnormalities

This is a highly selective list aiming to capture structural abnormalies which are frequesnt and/or significant in relation to diagnosis, prognosis, and/or characterising specific cancers. For a much more extensive list see the Mitelman Database of Chromosome Aberrations and Gene Fusions in Cancer.

del(1p36) in Neuroblastoma

Enomoto H, Ozaki T, Takahashi E, et al.
Identification of human DAN gene, mapping to the putative neuroblastoma tumor suppressor locus.
Oncogene. 1994; 9(10):2785-91 [PubMed] Related Publications
The expression of DAN gene (previously designated as N03 gene) is significantly reduced in a variety of transformed rat fibroblasts, including v-src- (SR-3Y1), SV40- and v-mos-transformed 3Y1 cells, compared with that in parental 3Y1 cells. Recently, DAN gene has been shown to possess a tumor suppressive activity when it is overexpressed in SR-3Y1 cells (Ozaki & Sakiyama, 1994). To assess the involvement of DAN gene with human neoplasms, we have isolated human DAN counterpart from a normal lung cDNA library by using rat DAN cDNA as a probe, and determined its chromosomal location. Human DAN gene mapped to chromosome 1p36.11-p36.13, which is well known to show highly significant linkage with the genesis and/or progression of human neuroblastoma. Southern blot analysis on tumor DNA from 26 patients with neuroblastoma has detected three patients showing genomic rearrangement or deletion within or closely linked to the DAN gene locus. Collectively, we propose that human DAN gene is a possible candidate for a tumor suppressor gene of human neuroblastoma.

White PS, Maris JM, Beltinger C, et al.
A region of consistent deletion in neuroblastoma maps within human chromosome 1p36.2-36.3.
Proc Natl Acad Sci U S A. 1995; 92(12):5520-4 [PubMed] Free Access to Full Article Related Publications
Deletion of the short arm of human chromosome 1 is the most common cytogenetic abnormality observed in neuroblastoma. To characterize the region of consistent deletion, we performed loss of heterozygosity (LOH) studies on 122 neuroblastoma tumor samples with 30 distal chromosome 1p polymorphisms. LOH was detected in 32 of the 122 tumors (26%). A single region of LOH, marked distally by D1Z2 and proximally by D1S228, was detected in all tumors demonstrating loss. Also, cells from a patient with a constitutional deletion of 1p36, and from a neuroblastoma cell line with a small 1p36 deletion, were analyzed by fluorescence in situ hybridization. Cells from both sources had interstitial deletions of 1p36.2-36.3 which overlapped the consensus region of LOH defined by the tumors. Interstitial deletion in the constitutional case was confirmed by allelic loss studies using the panel of polymorphic markers. Four proposed candidate genes--DAN, ID3 (heir-1), CDC2L1 (p58), and TNFR2--were shown to lie outside of the consensus region of allelic loss, as defined by the above deletions. These results more precisely define the location of a neuroblastoma suppressor gene within 1p36.2-36.3, eliminating 33 centimorgans of proximal 1p36 from consideration. Furthermore, a consensus region of loss, which excludes the four leading candidate genes, was found in all tumors with 1p36 LOH.

Spieker N, Beitsma M, van Sluis P, et al.
An integrated 5-Mb physical, genetic, and radiation hybrid map of a 1p36.1 region implicated in neuroblastoma pathogenesis.
Genes Chromosomes Cancer. 2000; 27(2):143-52 [PubMed] Related Publications
Common genetic aberrations of neuroblastoma are deletions of the short arm of chromosome 1 (1p36) and MYCN amplification. Our deletion analysis of 25 tumor cell lines and 171 tumors strongly suggests that 1p harbors several tumor suppressor loci. Distinct loci are involved in MYCN single-copy versus MYCN-amplified neuroblastoma. Deletions in MYCN single-copy tumors have a shortest region of overlap (SRO) of 20 cM at 1p36.3. MYCN-amplified tumors have large deletions with an SRO of about 60 cM, from 1p36.1 to the telomere. This SRO is defined by D1S7 (1p36.1), which was the most distal locus retained. Therefore, a suppressor gene associated with MYCN-amplified tumors probably maps within a few megabases distal of D1S7. In order to map this locus, we further refined this SRO. We mapped the breakpoint of the MYCN-amplified neuroblastoma with the smallest 1p deletion between 56.6 and 57.2 cM from 1pter. Pulsed-field gel electrophoresis and radiation hybrid mapping were used to construct a 5-Mb physical map of this region. The map includes the region from 82.73 till 92.89 cR from 1pter. About half of it was isolated in P1 and PAC clones. The region harbors the genes FGR, SLC9A1, HMG17, EXTL1, AML2, RH, OP18, four ESTs, and a newly identified gene with a transcript size of approximately 7 Kb. Several of the mapped genes have a putative role in cell growth, differentiation, and morphogenesis. Genes Chromosomes Cancer 27:143-152, 2000.

Maris JM, Weiss MJ, Guo C, et al.
Loss of heterozygosity at 1p36 independently predicts for disease progression but not decreased overall survival probability in neuroblastoma patients: a Children's Cancer Group study.
J Clin Oncol. 2000; 18(9):1888-99 [PubMed] Related Publications
PURPOSE: To determine the independent prognostic significance of 1p36 loss of heterozygosity (LOH) in a representative group of neuroblastoma patients.
PATIENTS AND METHODS: Diagnostic tumor specimens from 238 patients registered onto the most recent Children's Cancer Group phase III clinical trials were assayed for LOH with 13 microsatellite polymorphic markers spanning chromosome band 1p36. Allelic status at 1p36 was correlated with other prognostic variables and disease outcome.
RESULTS: LOH at 1p36 was detected in 83 (35%) of 238 neuroblastomas. There was a correlation of 1p36 LOH with age at diagnosis greater than 1 year (P = .026), metastatic disease (P<.001), elevated serum ferritin level (P<.001), unfavorable histopathology (P<.001), and MYCN oncogene amplification (P<.001). LOH at 1p36 was associated with decreased event-free survival (EFS) and overall survival (OS) probabilities (P<.0001). For the 180 cases with single-copy MYCN, 1p36 LOH status was highly correlated with decreased EFS (P = .0002) but not OS (P = .1212). Entering 1p36 LOH into a multivariate regression model suggested a trend toward an independent association with decreased EFS (P = .0558) but not with decreased OS (P = .3687). Furthermore, allelic status at 1p36 was the only prognostic variable that was significantly associated with decreased EFS in low-risk neuroblastoma patients (P = .0148).
CONCLUSION: LOH at 1p36 is independently associated with decreased EFS, but not OS, in neuroblastoma patients. Determination of 1p36 allelic status may be useful for predicting which neuroblastoma patients with otherwise favorable clinical and biologic features are more likely to have disease progression.

del(9p) in Neuroblastoma

Takita J, Hayashi Y, Kohno T, et al.
Deletion map of chromosome 9 and p16 (CDKN2A) gene alterations in neuroblastoma.
Cancer Res. 1997; 57(5):907-12 [PubMed] Related Publications
We reported previously that loss of heterozygosity (LOH) on chromosomes 2q, 9p and 18q frequently occurs in neuroblastoma and that patients with 9p LOH in the tumors showed statistically significant association with an advanced stage of the disease and poor prognosis. To determine the role of chromosome 9 loss in neuroblastoma, we performed deletion mapping of chromosome 9 in 80 cases of neuroblastoma using 11 polymorphic microsatellite markers and a restriction fragment length porymorphism marker. LOH at one or more loci on chromosome 9 was detected in 33 of 80 cases (41%). Chromosome 9p was lost in 24 of 80 cases (32%), whereas chromosome 9q was lost in 18 of 80 cases (23%). There were two commonly deleted regions mapped to 9p21 between the D9S171 marker and the IFNB1 marker and 9q34-qter distal to the D9S176 marker. In addition, patients with LOH at 9p21 but not at 9q34-qter in the tumors showed statistically significant association with poor prognosis (P = 0.023). Because the commonly deleted regions at 9p21 includes the p16 (CDKN2A) gene, the status of the p16 gene was further examined in 80 fresh tumors and 19 cell lines of neuroblastoma. A missense mutation was detected at codon 52 in a fresh tumor. The p16 gene was not expressed in 13 of 19 cell lines (72%), and 5 of the 13 cell lines displayed methylation of the CpG island surrounding the first exon of the p16 gene. These results suggest that the p16 gene is a candidate tumor suppressor gene for neuroblastoma, and its inactivation may contribute to the progression of neuroblastoma.

Marshall B, Isidro G, Martins AG, Boavida MG
Loss of heterozygosity at chromosome 9p21 in primary neuroblastomas: evidence for two deleted regions.
Cancer Genet Cytogenet. 1997; 96(2):134-9 [PubMed] Related Publications
The genes responsible for the development of neuroblastoma following in vivo deletion or mutation are largely unknown. We have performed loss of heterozygosity studies on a series of 24 Portuguese primary neuroblastomas using 6 polymorphic markers located at chromosome 9p21 spanning the p16/MTS1/CDKN2, p15/MTS2/CDKN2B, and the interferon alpha and beta genes. Loss of heterozygosity was observed in 4 of the 24 tumors (17%), a somewhat lower percentage than a previous study that identified patients by a mass screening program. A correlation was also observed between 9p21 LOH and 1p36 LOH in our group of tumors. Two distinct regions of 9p21 deletion were observed: one located in the region adjacent to the markers D9S162 and D9S1747 and a second located centromerically of the p16 gene near the D9S171 marker. The latter region is exclusive of the p16 gene. This result suggests the presence of at least one other tumor suppressor gene at 9p21, apart from the p16 and p15 genes, which may be of importance to the development of neuroblastoma.

Gain of Chromosome 17q in Neuroblastoma

Gain of extra 17q material is the most frequent genetic abnormality in neuroblastoma. Unbalanced (partial) gain is associated with 1p deletion and MYCN amplification; in some cases 1p deletion can be caused by t(1;17) translocation.

Brinkschmidt C, Poremba C, Christiansen H, et al.
Comparative genomic hybridization and telomerase activity analysis identify two biologically different groups of 4s neuroblastomas.
Br J Cancer. 1998; 77(12):2223-9 [PubMed] Free Access to Full Article Related Publications
Chromosomal aberrations of 20 stage 4s neuroblastomas were analysed by comparative genomic hybridization (CGH). In a subset of 13/20 tumours, telomerase activity was evaluated by the telomeric repeat amplification protocol (TRAP). The CGH data were compared with the CGH results of ten stage 1 and 2 (stage 1/2) and 22 stage 3 and 4 (stage 3/4) neuroblastomas. A total of 17/20 stage 4s neuroblastomas did not progress clinically, whereas tumour progression with lethal outcome occurred in 3/20 cases. The CGH data of clinically non-progressing stage 4s tumours revealed a high rate of whole-chromosome aberrations (73.4%) with an overrepresentation of mainly chromosomes 2, 6, 7, 12, 13, 17, 18 and an underrepresentation of mainly chromosomes 3, 4, 11, 14. MYCN amplification or 1p deletion was observed in only 1/27 or 2/17 clinically non-progressing stage 4s tumours respectively, whereas all three progressive stage 4s neuroblastomas showed MYCN amplification, 1p deletion and, in 2/3 cases, distal 17q gains. Except for one case, telomerase activity was not observed in non-progressing stage 4s neuroblastomas. In contrast, 4s tumours with lethal outcome revealed elevated telomerase activity levels. Our data suggest that stage 4s neuroblastomas belong to two biologically different groups, one of which displays the genetic features of localized stage 1/2 tumours, whereas the other mimics advanced stage 3/4 neuroblastomas.

Plantaz D, Mohapatra G, Matthay KK, et al.
Gain of chromosome 17 is the most frequent abnormality detected in neuroblastoma by comparative genomic hybridization.
Am J Pathol. 1997; 150(1):81-9 [PubMed] Free Access to Full Article Related Publications
Neuroblastoma behavior is variable and outcome partially depends on genetic factors. However, tumors that lack high-risk factors such as MYCN amplification or 1p deletion may progress, possibly due to other genetic aberrations. Comparative genomic hybridization summarizes DNA copy number abnormalities in a tumor by mapping them to their positions on normal metaphase chromosomes. We analyzed 29 tumors from nearly equal proportions of children with stage I, II, III, IV, and IV-S disease by comparative genomic hybridization. We found two classes of copy number abnormalities: whole chromosome and partial chromosome. Whole chromosome losses were frequent at 11, 14, and X. The most frequent partial chromosome losses were on 1p and 11q. Gains were most frequent on chromosome 17 (72% of cases). The two patterns of gain for this chromosome were whole 17 gain and 17q gain, with 17q21-qter as a minimal common region of gain. Other common gains were on chromosomes 7, 6, and 18. High level amplifications were detected at 2p23-25 (MYCN region), at 4q33-35, and at 6p11-22. Chromosome 17q gains were associated with 1p and/or 11q deletions and advanced stage. The high frequency of chromosome 17 gain and its association with bad prognostic factors suggest an important role for this chromosome in the development of neuroblastoma.

Caron H
Allelic loss of chromosome 1 and additional chromosome 17 material are both unfavourable prognostic markers in neuroblastoma.
Med Pediatr Oncol. 1995; 24(4):215-21 [PubMed] Related Publications
In neuroblastoma, N-myc amplification and loss of heterozygosity for the short arm of chromosome 1 (LOH 1p) are common genetic abnormalities. We have recently shown that the presence of additional material of the long arm of chromosome 17 (add.17q) also occurs relatively frequently. In the present study, we analyzed a series of 55 tumors for LOH 1p, N-myc amplification and add.17q, using Southern blot analysis with polymorphic DNA probes of pairs of tumor and constitutional DNA. We determined the correlation of these parameters with clinical variables, such as age, stage, serum lactate dehydrogenase (LDH) and ferritin and also with outcome. LOH 1p occurred in 20 out of 55 cases (36%) and was found more often in stage III/IV tumors and in the older age group, although both correlations were not statistically significant. N-myc amplification was only demonstrated in 12 tumors with concomitant LOH 1p and was not present in the 35 cases without LOH 1p. Add.17q was found in 20/53 (38%) informative cases. LOH 1p was shown to be the most significant predictor of a poor outcome (P < 0.00001), independent of age and stage. LOH 1p is also of prognostic value in those cases without N-myc amplification, indicating a stronger prognostic value for LOH 1p. Add.17q was also associated with an unfavourable prognosis, although this was less significantly then with LOH 1p (P = 0.00004).

Bown N, Cotterill S, Lastowska M, et al.
Gain of chromosome arm 17q and adverse outcome in patients with neuroblastoma.
N Engl J Med. 1999; 340(25):1954-61 [PubMed] Related Publications
BACKGROUND: Gain of genetic material from chromosome arm 17q (gain of segment 17q21-qter) is the most frequent cytogenetic abnormality of neuroblastoma cells. This gain has been associated with advanced disease, patients who are > or =1 year old, deletion of chromosome arm 1p, and amplification of the N-myc oncogene, all of which predict an adverse outcome. We investigated these associations and evaluated the prognostic importance of the status of chromosome 17.
METHODS: We compiled molecular cytogenetic analyses of chromosome 17 in primary neuroblastomas in 313 patients at six European centers. Clinical and survival information were collected, along with data on 1p, N-myc, and ploidy.
RESULTS: Unbalanced gain of segment 17q21-qter was found in 53.7 percent of the tumors, whereas the chromosome was normal in 46.3 percent. The gain of 17q was characteristic of advanced tumors and of tumors in children > or =1 year of age and was strongly associated with the deletion of 1p and amplification of N-myc. No tumor showed amplification of N-myc in the absence of either deletion of 1p or gain of 17q. Gain of 17q was a significant predictive factor for adverse outcome in univariate analysis. Among the patients with this abnormality, overall survival at five years was 30.6 percent (95 percent confidence interval, 21 to 40 percent), as compared with 86.0 percent (95 percent confidence interval, 78 to 91 percent) among those with normal 17q status. in multivariate analysis, gain of 17q was the most powerful prognostic factor, followed by the presence of stage 4 disease and deletion of 1p (hazard ratios, 3.4, 2.3, and 1.9, respectively).
CONCLUSIONS: Gain of chromosome segment 17q21-qter is an important prognostic factor in children with neuroblastoma.

Abel F, Ejeskär K, Kogner P, Martinsson T
Gain of chromosome arm 17q is associated with unfavourable prognosis in neuroblastoma, but does not involve mutations in the somatostatin receptor 2(SSTR2) gene at 17q24.
Br J Cancer. 1999; 81(8):1402-9 [PubMed] Free Access to Full Article Related Publications
Deletion of chromosome arm 1p and amplification of the MYCN oncogene are well-recognized genetic alterations in neuroblastoma cells. Recently, another alteration has been reported; gain of the distal part of chromosome arm 17q. In this study 48 neuroblastoma tumours were successfully analysed for 17q status in relation to known genetic alterations. Chromosome 17 status was detected by fluorescence in situ hybridization (FISH). Thirty-one of the 48 neuroblastomas (65%) showed 17q gain, and this was significantly associated with poor prognosis. As previously reported, 17q gain was significantly associated with metastatic stage 4 neuroblastoma and more frequently detected than both deletion of chromosome arm 1p and MYCN amplification in tumours of all stages. 17q gain also showed a strong correlation to survival probability (P = 0.0009). However, the most significant correlation between 17q gain and survival probability was observed in children with low-stage tumours (stage 1, 2, 3 and 4S), with a survival probability of 100% at 5 years from diagnosis for children with tumours showing no 17q gain compared to 52.5% for those showing 17q gain (P = 0.0021). This suggests that 17q gain as a prognostic factor plays a more crucial role in low-stage tumours. Expression of the somatostatin receptor 2 (SSTR2), localized in chromosome region 17q24, has in previous studies been shown to be positively related to survival in neuroblastoma. A point mutation in the SSTR2 gene has earlier been reported in a human small-cell lung cancer. In this study, mutation screening of the SSTR2 gene in 43 neuroblastoma tumours was carried out with polymerase chain reaction-based single-stranded conformation polymorphism/heteroduplex (SSCP/HD) and DNA sequencing, and none of the tumours showed any aberrations in the SSTR2 gene. These data suggest that mutations in the SSTR2 gene are uncommon in neuroblastoma tumours and do not correlate with either the 17q gain often seen or the reason some tumours do not express SSTR2 receptors. Overall, this study indicates that gain of chromosome arm 17q is the most frequently occurring genetic alteration, and that it is associated with established prognostic factors.

Godfried MB, Veenstra M, v Sluis P, et al.
The N-myc and c-myc downstream pathways include the chromosome 17q genes nm23-H1 and nm23-H2.
Oncogene. 2002; 21(13):2097-101 [PubMed] Related Publications
Gain of chromosome 17q material is the most frequent genetic abnormality in neuroblastomas. The common region of gain is at least 375 cR large, which has precluded the identification of genes with a role in neuroblastoma pathogenesis. Neuroblastoma also frequently show amplification of the N-myc oncogene, which correlates closely with 17q gain. Both events are strong predictors of unfavorable prognosis. To identify genes that are part of the N-myc downstream pathway, we constructed SAGE libraries of an N-myc transfected and a control cell line. This identified the chromosome 17q genes nm23-H1 and nm23-H2 as being 6-10 times induced in the N-myc expressing cells. Northern and Western blot analysis confirmed this up-regulation. Time-course experiment shows that both genes are induced within 4 h after N-myc is switched on. Furthermore, we demonstrate also that c-myc can up-regulate nm23-H1 and nm23-H2 expression. Neuroblastoma tumor and cell line panels reveal a striking correlation between N-myc amplification and mRNA and protein expression of both nm23 genes. We show that the nm23 genes are located at the edge of the common region of chromosome 17q gain previously described in neuroblastoma cell lines. Our findings suggest that nm23-H1 and nm23-H2 expression is increased by 17q gain in neuroblastoma and can be further up-regulated by myc overexpression. These observations suggest a major role for nm23-H1 and nm23-H2 in tumorigenesis of unfavorable neuroblastomas.

14q Deletions in Neuroblastoma

Theobald M, Christiansen H, Schmidt A, et al.
Sublocalization of putative tumor suppressor gene loci on chromosome arm 14q in neuroblastoma.
Genes Chromosomes Cancer. 1999; 26(1):40-6 [PubMed] Related Publications
RFLP and microsatellite analysis with 23 polymorphic markers spanning the entire long arm of chromosome 14 in 108 neuroblastomas showed allelic loss in 19 out of 107 (18%) informative tumors, placing 14q among the most frequently affected chromosomal regions in neuroblastoma. One minimal deletion region could be sublocalized in 17 of 19 cases between markers D14S1 and D14S16, and a second one between markers D14S17 and D14S23 in band 14q32. Furthermore, breakpoints in bands 14q23 and 14q12 were detected. These results suggest the presence of at least two putative tumor suppressor gene loci on chromosome 14. Survival analyses revealed no prognostic impact of allelic loss of 14q in neuroblastoma. Genes Chromosomes Cancer 26:40-46, 1999.

Hoshi M, Otagiri N, Shiwaku HO, et al.
Detailed deletion mapping of chromosome band 14q32 in human neuroblastoma defines a 1.1-Mb region of common allelic loss.
Br J Cancer. 2000; 82(11):1801-7 [PubMed] Free Access to Full Article Related Publications
Neuroblastoma (NB) is a well-known malignant disease in infants, but its molecular mechanisms have not yet been fully elucidated. To investigate the genetic contribution of abnormalities on the long arm of chromosome 14 (14q) in NB, we analysed loss of heterozygosity (LOH) in 54 primary NB samples using 12 microsatellite markers on 14q32. Seventeen (31%) of 54 tumours showed LOH at one or more of the markers analysed, and the smallest common region of allelic loss was identified between D14S62 and D14S987. This region was estimated to be 1-cM long from the linkage map. Fluorescence in situ hybridization also confirmed the loss. There was no statistical correlation between LOH and any clinicopathologic features, including age, stage, amplification of MYCN and ploidy. We further constructed a contig spanning the lost region using bacterial artificial chromosome and estimated this region to be approximately 1.1-Mb by pulsed-field gel electrophoresis. Our results will contribute to cloning and characterizing the putative tumour-associated gene(s) in 14q32 in NB.

Thompson PM, Seifried BA, Kyemba SK, et al.
Loss of heterozygosity for chromosome 14q in neuroblastoma.
Med Pediatr Oncol. 2001; 36(1):28-31 [PubMed] Related Publications
BACKGROUND: Neuroblastoma is a genetically heterogeneous disease, with subsets of tumors demonstrating rearrangements of several genomic regions. Preliminary studies from several groups have identified loss of heterozygosity (LOH) for the long arm of chromosome 14 (14q) in 20-25% of primary neuroblastomas.
PROCEDURE: To determine precisely the frequency and extent of 14q deletions, we performed LOH analysis for a large series of primary neuroblastomas using a panel of 11 highly polymorphic markers.
RESULTS: LOH was detected in 83 of 372 tumors (22%). Although the majority of tumors with allelic loss demonstrated allelic loss for all informative markers, 13 cases showed LOH for only a portion of 14q. A single consensus region of deletion, which was shared by all tumors with 14q LOH, was defined within 14q23-q32 between D14S588 and the 14q telomere. Allelic loss for 14q was strongly correlated with the presence of 11q LOH (P < 0.001 ) and inversely correlated with MYCN amplification (P= 0.04).
CONCLUSIONS: LOH for 14q was evident in all clinical risk groups, indicating that this abnormality may be a universal feature of neuroblastoma tumor development. These findings suggest that a tumor suppressor gene involved in the initiation or progression of neuroblastoma is located within distal 14q.

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