Heterogeneity of the MYCN Oncogene in Neuroblastoma

Abstract

Purpose: MYCN amplification is an important therapy-stratifying marker in neuroblastoma.Fluorescence in situ hybridization with signal detection on the single-cell level allows a critical judgement of MYCN intratumoral heterogeneity. Experimental Design: The MYCN status was investigated by fluorescence in situ hybridization at diagnosis and relapse. Heterogeneity was defined as the simultaneous presence of amplified cells (z5 cells per slide) and nonamplified cells within one tumor or sequential change of the amplification status during the course of the disease. Likewise, heterogeneity can be detected between primary tumor and metastasis. Results: From 1,341 patients analyzed, 1,071 showed no amplification, 250 showed homogeneous amplification, and 20 patients showed MYCN heterogeneity. Of the patients with heterogeneity, 12 of 20 had clusters of MYCN amplifications, 3 of 20 had amplified single cells, 3 of 20 showed MYCN amplifications in the bone marrow but not in the primary tumor, and 2 of 20 acquired MYCN amplification during the course of the disease. All stage 4 patients were treated according to high-risk protocols; 7 of 8 later progressed. Four patients with localized disease were treated according to high-risk protocol because of MYCN-amplified clusters; 1of 4 later progressed. One patient treated with mild chemotherapy experienced progression. Seven patients with localized/4S disease underwent no chemotherapy: 4 of 5 patients with MYCN heterogeneity at diagnosis remained disease-free, and 1of 5 experienced local progression. Two patients had normal MYCN status at diagnosis but acquired MYCN amplification during the course of the disease. Conclusion: MYCN heterogeneity is rare. Our results suggest that small amounts of MYCNamplified cells are not correlated to adverse outcomes. More patients with heterogeneity are warranted to clarify the role of MYCN heterogeneity for risk classification.

Revised Risk Estimation and Treatment Stratification of Low- and Intermediate-Risk Neuroblastoma Patients by Integrating Clinical and Molecular Prognostic Markers

“…Of note, this time frame matches the turnaround time that is currently required for the detection of genetic alterations, that is, the determination of the genomic status of MYCN and chromosome 1p. Second, tumor heterogeneity is present in a small fraction of neuroblastoma tumor (36), raising the possibility that the genomic profile might not adequately reflect the underlying tumor behavior. To prevent misclassifications due to tumor heterogeneity, it is intended to perform expression profiles from RNA of at least two separate parts of the tumor specimens of each patient.…”

Genomic Copy Number Profiling Using Circulating Free Tumor DNA Highlights Heterogeneity in Neuroblastoma

“…Indeed, spatial heterogeneity with differences in the genomic profile between different sites or even different clones within a neuroblastoma, or between a primary neuroblastoma and its metastatic sites, have been described, concerning MYCN, SCA, or mutations including ALK (15,16). Temporal heterogeneity has been shown to be of importance in neuroblastoma progression, with an accumulation of additional SCA, and clonal evolution with new mutations, such as ALK or other MAPK mutations emerging at time of progression (17)(18)(19).…”

2p24 Gain Region Harboring MYCN Gene Compared with MYCN Amplified and Nonamplified Neuroblastoma

“…Furthermore, because the copy number can be determined at the single-cell level, FISH has the capability to demonstrate intercellular heterogeneity in MYCN amplification within a given tumor cell population. 28,29 The International Neuroblastoma Risk Group Biology Committee defined 2p gain as equal number of MYCN and 2p signals, exceeding 2q signals in mostly one or two copies. A balanced ratio between the MYCN specific signals and the signal numbers of the reference probe on the chromosome 2q arm defines a MYCN nonamplified tumor (MYCN normal status).…”