ABSTRACT
Intratumoral heterogeneous MYCN amplification (hetMNA) is an unusual event in neuroblastoma with unascertained biological and clinical implications. Diagnosis is based on the detection of MYCN amplification surrounded by non-amplified tumor cells by fluorescence in situ hybridization (FISH). To better define the genetic features of hetMNA tumors, we studied the Spanish cohort of neuroblastic tumors by FISH and single nucleotide polymorphism arrays. We compared hetMNA tumors with homogeneous MNA (homMNA) and nonMNA tumors with 11q deletion (nonMNA w11q-). Of 1091 primary tumors, 28 were hetMNA by FISH. Intratumoral heterogeneity of 1p, 2p, 11q and 17q was closely associated with hetMNA tumors when analyzing different pieces for each case. For chromosome 2, 16 cases showed 2p intact, 4 focal gain at 2p24.3 and 8 MNA. The lengths of the smallest regions of overlap (SROs) for 2p gains and 1p deletions were between the SRO lengths observed in homMNA and nonMNA w11q- tumors. Co-occurrence of 11q- and +17q was frequently found with the largest SROs for both aberrations. The evidence for and frequency of different genetic subpopulations representing a hallmark of the hetMNA subgroup of NB indicates, on one hand, the presence of a considerable genetic instability with different SRO of either gains and losses compared with those of the other NB groups and highlights and, on the other hand, the need for multiple sampling from distant and macroscopically and microscopically distinct tumor areas. Narrowing down the different SRO for both deletions and gains in NB groups would be crucial to pinpointing the candidate gene(s) and the critical gene dosage with prognostic and therapeutic significance. This complexity of segmental chromosomal aberration patterns reinforces the necessity for a larger cohort study using FISH and pangenomic techniques to develop a suitable therapeutic strategy for these patients.
Subject(s)
Gene Dosage/genetics , Neuroblastoma/genetics , Nuclear Proteins/genetics , Oncogene Proteins/genetics , Adolescent , Adult , Aged , Aged, 80 and over , Child , Child, Preschool , Chromosome Deletion , Chromosomes, Human, Pair 1/genetics , Chromosomes, Human, Pair 11/genetics , Chromosomes, Human, Pair 17/genetics , Chromosomes, Human, Pair 2/genetics , Cohort Studies , Humans , In Situ Hybridization, Fluorescence , Middle Aged , N-Myc Proto-Oncogene Protein , Neuroblastoma/classification , Polymorphism, Single Nucleotide/genetics , Young AdultABSTRACT
BACKGROUND: The prognostic impact of segmental chromosome alterations (SCAs) in children older than 1 year, diagnosed with localised unresectable neuroblastoma (NB) without MYCN amplification enrolled in the European Unresectable Neuroblastoma (EUNB) protocol is still to be clarified, while, for other group of patients, the presence of SCAs is associated with poor prognosis. METHODS: To understand the role of SCAs we performed multilocus/pangenomic analysis of 98 tumour samples from patients enrolled in the EUNB protocol. RESULTS: Age at diagnosis was categorised into two groups using 18 months as the age cutoff. Significant difference in the presence of SCAs was seen in tumours of patients between 12 and 18 months and over 18 months of age at diagnosis, respectively (P=0.04). A significant correlation (P=0.03) was observed between number of SCAs per tumour and age. Event-free (EFS) and overall survival (OS) were calculated in both age groups, according to both the presence and number of SCAs. In older patients, a poorer survival was associated with the presence of SCAs (EFS=46% vs 75%, P=0.023; OS=66.8% vs 100%, P=0.003). Moreover, OS of older patients inversely correlated with number of SCAs (P=0.002). Finally, SCAs provided additional prognostic information beyond histoprognosis, as their presence was associated with poorer OS in patients over 18 months with unfavourable International Neuroblastoma Pathology Classification (INPC) histopathology (P=0.018). CONCLUSIONS: The presence of SCAs is a negative prognostic marker that impairs outcome of patients over the age of 18 months with localised unresectable NB without MYCN amplification, especially when more than one SCA is present. Moreover, in older patients with unfavourable INPC tumour histoprognosis, the presence of SCAs significantly affects OS.
Subject(s)
Neuroblastoma/genetics , Peripheral Nervous System Neoplasms/genetics , Chromosome Aberrations , Comparative Genomic Hybridization , Disease-Free Survival , Gene Amplification , Humans , Infant , Kaplan-Meier Estimate , N-Myc Proto-Oncogene Protein , Neuroblastoma/diagnosis , Neuroblastoma/mortality , Nuclear Proteins/genetics , Oncogene Proteins/genetics , Peripheral Nervous System Neoplasms/diagnosis , Peripheral Nervous System Neoplasms/mortality , PrognosisABSTRACT
Neuroblastoma tumor cells show complex combinations of genetic aberrations, and to date many different methods have been used for their detection. To apply genome-wide techniques, such as Multiplex Ligation-dependent Probe Amplification (MLPA), in routine diagnosis their validation is appropriate and necessary. DNA copy number alterations in 129 cases of neuroblastic tumors were detected using MPLA, and the results validated by Fluorescence In Situ Hybridization (FISH) (MYCN gene, 1p36, 11q and 17q). Kappa index values showed very good concordance between the two techniques in detecting homogeneous MYCN amplification (1); 11q deletion (0.908) and 17q gain (0.922). The validation results showed that MLPA is a highly efficient technique for diagnosis based on the genetic aberrations in relevant regions in neuroblastoma, showing a high concordance with FISH.
