Analysis of chromosome breakpoints in neuroblastoma at sub‐kilobase resolution using fine‐tiling oligonucleotide array CGH

Abstract

Understanding the genes and genetic pathways targeted by recurrent chromosomal imbalances in malignancy, along with the molecular mechanisms that generate the imbalances, are important problems in cancer biology. In this report, we demonstrate that oligonucleotide array CGH (oaCGH) analysis can routinely map chromosomal imbalance breakpoints at exon-level resolution, including imbalances that are single copy number genomic alterations. Different tiling-path array designs were used in this study: a whole-genome array with a 6-kb median probe spacing and fine-tiling arrays for selected genomic regions with either 50- or 140-bp median probe spacing. In both array formats, oligonucleotide probes were of isothermal design and were tiled through genic and inter-genic regions. Whole-genome oaCGH analysis of two neuroblastoma cell lines and three primary tumors led to the identification of 58 chromosomal breakpoints that generated 45 large-scale partial chromosomal imbalances (> 2 Mb). An unexpectedly high proportion (34%) of these breakpoint intervals mapped to regions containing segmental duplications. In addition, 88 smaller-sized regions (< 2 Mb) of imbalance were detected, the majority of which mapped to segmentally duplicated regions and may reflect constitutional copy number polymorphisms. The chromosomal breakpoints for 12 recurrent abnormalities exhibited in neuroblastoma tumors and cell lines, including MYCN amplicon boundaries, loss of 3p, loss of 11q, and gain of 17q, could be mapped to intervals ranging from 50 bp to 10 kb in size using high-density fine-tiling oligonucleotide microarrays. Fine-tiling oaCGH analysis provides an unprecedented level of resolution, allowing detailed mapping of recurrent unbalanced chromosomal abnormalities. Supplementary material for this article can be found on the Genes, Chromosomes, and Cancer website at http://www.interscience.wiley.com/jpages/1045-2257/suppmat/index.html.

Characterization of amplicons in neuroblastoma: High‐resolution mapping using DNA microarrays, relationship with outcome, and identification of overexpressed genes

“…As expected, the MYCN oncogene is the most frequent gene targeted by amplification in NB. The extensive variation in the MYCN amplicon size and complexity that we have noticed is consistent with previous reports (Selzer et al, 2005;Stallings et al, 2006;Lastowska et al, 2007;Mosse et al, 2007). Amplification of the MYC and MYCL1 genes belonging to the same family of transcription factors as MYCN were rarely observed in our series.…”

Characterization of amplicons in neuroblastoma: High‐resolution mapping using DNA microarrays, relationship with outcome, and identification of overexpressed genes

“…As expected, the MYCN oncogene is the most frequent gene targeted by amplification in NB. The extensive variation in the MYCN amplicon size and complexity that we have noticed is consistent with previous reports (Selzer et al, 2005;Stallings et al, 2006;Lastowska et al, 2007;Mosse et al, 2007). Amplification of the MYC and MYCL1 genes belonging to the same family of transcription factors as MYCN were rarely observed in our series.…”

Molecular cloning and analysis of breakpoints on ring chromosome 17 in a patient with autism

“…Our results clearly support the notion that CGH on high resolution DNA arrays is an extremely potent approach to breakpoint cloning in chromosomal rearrangements with gains or losses of genetic material (Selzer et al, 2005). Similarly, painting of these arrays with labelled derivative chromosomes can shed light on balanced translocations (Gribble et al, 2007).…”

Identification of candidate genes involved in neuroblastoma progression by combining genomic and expression microarrays with survival data

“…It confirms the coamplification and overexpression of previously defined amplified genes (Chen et al, 2004;Seltzer et al, 2005) and the poor correlation between MYCN expression and copy number (Nisen et al, 1988;Slavc et al, 1990). While the extensive variation in amplicon size and complexity we observe is consistent with previous reports (Figure 1; Seltzer et al, 2005;Stallings et al, 2006), the present analysis defines for the first time over 30 genes as being co-amplified at both the DNA and expression level, several of which have been implicated in other neoplasias (Table 3). This complexity suggests that the relationship between gene expression/copy number within the MYCN amplicon and disease outcome, which has produced conflicting results using lower resolution methodologies (Weber et al, 2004;De Preter et al, 2005), may warrant renewed investigation.…”

Characterization of amplicons in neuroblastoma: High‐resolution mapping using DNA microarrays, relationship with outcome, and identification of overexpressed genes

“…As expected, the MYCN oncogene is the most frequent gene targeted by amplification in NB. The extensive variation in the MYCN amplicon size and complexity that we have noticed is consistent with previous reports (Selzer et al, 2005;Stallings et al, 2006;Lastowska et al, 2007;Mosse et al, 2007). Amplification of the MYC and MYCL1 genes belonging to the same family of transcription factors as MYCN were rarely observed in our series.…”

Molecular cloning and analysis of breakpoints on ring chromosome 17 in a patient with autism

“…Our results clearly support the notion that CGH on high resolution DNA arrays is an extremely potent approach to breakpoint cloning in chromosomal rearrangements with gains or losses of genetic material (Selzer et al, 2005). Similarly, painting of these arrays with labelled derivative chromosomes can shed light on balanced translocations (Gribble et al, 2007).…”

Identification of candidate genes involved in neuroblastoma progression by combining genomic and expression microarrays with survival data

“…It confirms the coamplification and overexpression of previously defined amplified genes (Chen et al, 2004;Seltzer et al, 2005) and the poor correlation between MYCN expression and copy number (Nisen et al, 1988;Slavc et al, 1990). While the extensive variation in amplicon size and complexity we observe is consistent with previous reports (Figure 1; Seltzer et al, 2005;Stallings et al, 2006), the present analysis defines for the first time over 30 genes as being co-amplified at both the DNA and expression level, several of which have been implicated in other neoplasias (Table 3). This complexity suggests that the relationship between gene expression/copy number within the MYCN amplicon and disease outcome, which has produced conflicting results using lower resolution methodologies (Weber et al, 2004;De Preter et al, 2005), may warrant renewed investigation.…”

Characterization of amplicons in neuroblastoma: High‐resolution mapping using DNA microarrays, relationship with outcome, and identification of overexpressed genes

“…As expected, the MYCN oncogene is the most frequent gene targeted by amplification in NB. The extensive variation in the MYCN amplicon size and complexity that we have noticed is consistent with previous reports (Selzer et al, 2005;Stallings et al, 2006;Lastowska et al, 2007;Mosse et al, 2007). Amplification of the MYC and MYCL1 genes belonging to the same family of transcription factors as MYCN were rarely observed in our series.…”

Molecular cloning and analysis of breakpoints on ring chromosome 17 in a patient with autism

“…Our results clearly support the notion that CGH on high resolution DNA arrays is an extremely potent approach to breakpoint cloning in chromosomal rearrangements with gains or losses of genetic material (Selzer et al, 2005). Similarly, painting of these arrays with labelled derivative chromosomes can shed light on balanced translocations (Gribble et al, 2007).…”

Identification of candidate genes involved in neuroblastoma progression by combining genomic and expression microarrays with survival data

“…It confirms the coamplification and overexpression of previously defined amplified genes (Chen et al, 2004;Seltzer et al, 2005) and the poor correlation between MYCN expression and copy number (Nisen et al, 1988;Slavc et al, 1990). While the extensive variation in amplicon size and complexity we observe is consistent with previous reports (Figure 1; Seltzer et al, 2005;Stallings et al, 2006), the present analysis defines for the first time over 30 genes as being co-amplified at both the DNA and expression level, several of which have been implicated in other neoplasias (Table 3). This complexity suggests that the relationship between gene expression/copy number within the MYCN amplicon and disease outcome, which has produced conflicting results using lower resolution methodologies (Weber et al, 2004;De Preter et al, 2005), may warrant renewed investigation.…”