High-Resolution Analysis of Chromosomal Breakpoints and Genomic Instability Identifies PTPRD as a Candidate Tumor Suppressor Gene in Neuroblastoma

Abstract

Although neuroblastoma is characterized by numerous recurrent, large-scale chromosomal imbalances, the genes targeted by such imbalances have remained elusive. We have applied whole-genome oligonucleotide array comparative genomic hybridization (median probe spacing 6 kb) to 56 neuroblastoma tumors and cell lines to identify genes involved with disease pathogenesis. This set of tumors was selected for having either 11q loss or MYCN amplification, abnormalities that define the two most common genetic subtypes of metastatic neuroblastoma. Our analyses have permitted us to map large-scale chromosomal imbalances and high-level amplifications at exon-level resolution and to identify novel microdeletions and duplications. Chromosomal breakpoints (n = 467) generating imbalances >2 Mb were mapped to intervals ranging between 6 and 50 kb in size, providing substantial information on each abnormality. For example, breakpoints leading to large-scale hemizygous loss of chromosome 11q were highly clustered and preferentially associated with segmental duplications. High-level amplifications of MYCN were extremely complex, often resulting in a series of discontinuous regions of amplification. Imbalances (n = 540) <2 Mb long were also detected. Although the majority (78%) of these imbalances mapped to segmentally duplicated regions and primarily reflect constitutional copy number polymorphisms, many subtle imbalances were detected that are likely somatically acquired alterations and include genes involved with tumorigenesis, apoptosis, or neural cell differentiation. The most frequent microdeletion involved the PTPRD locus, indicating a possible tumor suppressor function for this gene.

Genes Proximal and Distal to MYCN Are Highly Expressed in Human Neuroblastoma as Visualized by Comparative Expressed Sequence Hybridization

“…4,5,14,53 A number of highly coamplified regions that were discontinuous regarding the MYCN locus and located at varying distances from MYCN were described, demonstrating the unique constitution of the MYCN amplicon. 4,5,14,53 Interestingly, only one group of researchers found high-level amplification of two genes (SNTG2 and TPO) situated at 936 kb and 1.4 Mb, respectively, from 2pter, occurring in one of 10 MNA samples.…”

MicroRNA-34a functions as a potential tumor suppressor by inducing apoptosis in neuroblastoma cells

“…The region of 1p loss is quite large, containing many potential candidate genes. Furthermore, smaller homozygous deletions that might further pinpoint the genes of interest are exceedingly rare Stallings et al, 2006) and do not define a single shortest region of overlap (Schleiermacher et al, 1994). Expression microarray studies of primary tumors have also led some investigators to suggest that multiple genes on chromosome 1p contribute to NB pathogenesis (Janoueix-Lerosey et al, 2004;Wang et al, 2006), a concept supported by the fact that in vitro functional studies have shown that more than one gene from chromosome 1p can have anti-tumorigenic effects when ectopically expressed in NB cell lines (Ejeskar et al, 2005;Valentijn et al, 2005).…”

“…For example, two miRNAs (miR-15 and miR-16) mapping to chromosomal region 13q14 are frequently deleted and downregulated in chronic lymphocytic leukemia and thus act as tumor suppressor genes (Calin et al, 2002). MiRNAs acting in a dominant oncogenic manner are illustrated by miR-21 on chromosome 17q, which, when overexpressed in some forms of cancer have anti Figure 1 Analysis of 1p loss in 13 primary tumors and the SK-N-AS cell line using oligonucleotide array CGH, as described previously (Stallings et al, 2006). The vertical axis of each plot represents the fluorescence ratio of tumor to normal DNA on a log 2 scale, whereas the horizontal axis represents genomic position on chromosome 1p in base pairs.…”

“…An in silico search for miRNAs encoded within the 1p region identified miR-34a (1p36.23) and miR-30e (1p34.2) as potential candidates. Using array comparative genomic hybridization (CGH) data on a published set of tumors, we determined that the region encoding miR-34a is located in the minimal region of 1p loss (Figure 1) whereas the region encoding miR-30e is lost in 79% of tumors (Stallings et al, 2006). Using realtime PCR analysis, the expression of miR-34a was generally reduced in NB primary tumors as compared to normal adrenal gland (Figure 2), and tumors with 1p loss have a 30% reduction in miR-34a expression relative to tumors with an intact 1p locus.…”

High-content analysis of cancer genome DNA alterations

“…to study DNA alterations on a genome-wide scale, including array CGH with very high genome resolution [4], new methylation detection systems [5], and last but not least, single-molecule sequencing technologies [6] (Box 1). In this review we focus on the recent biological discoveries that have been made with these new tools along with the large-scale efforts using older tools.…”