High-resolution analysis of genomic copy number alterations in bladder cancer by microarray-based comparative genomic hybridization

Abstract

We have screened 22 bladder tumour-derived cell lines and one normal urothelium-derived cell line for genome-wide copy number changes using array comparative genomic hybridization (CGH). Comparison of array CGH with existing multiplex-fluorescence in situ hybridization (M-FISH) results revealed excellent concordance. Regions of gain and loss were defined more accurately by array CGH, and several small regions of deletion were detected that were not identified by M-FISH. Numerous genetic changes were identified, many of which were compatible with previous results from conventional CGH and loss of heterozygosity analyses on bladder tumours. The most frequent changes involved complete or partial loss of 4q (83%) and gain of 20q (78%). Other frequent losses were of 18q (65%), 8p (65%), 2q (61%), 6q (61%), 3p (56%), 13q (56%), 4p (52%), 6p (52%), 10p (52%), 10q (52%) and 5p (43%). We have refined the localization of a region of deletion at 8p21.2-p21.3 to an interval of approximately 1 Mb. Five homozygous deletions of tumour suppressor genes were confirmed, and several potentially novel homozygous deletions were identified. In all, 15 high-level amplifications were detected, with a previously reported amplification at 6p22.3 being the most frequent. Real-time PCR analysis revealed a novel candidate gene with consistent overexpression in all cell lines with the 6p22.3 amplicon.

Role of E2F3 expression in modulating cellular proliferation rate in human bladder and prostate cancer cells

“…This is consistent with the observation that the presence of the 6p22 amplicon correlates with tumour cell proliferation rate (Tomovska et al, 2001) and with the observed correlation between expression of E2F3 and the Ki67 proliferation marker (Oeggerli et al, 2004). In contrast, we failed to find evidence to support the view that CDKAL1 represents the bladder cancer oncogene within the 6p22 amplicon (Hurst et al, 2004): knocking down expression of this gene did not alter the BrdU incorporation. The possibility that CDKAL1, which is consistently overexpressed within the 6p22 amplicon, makes some other contribution to tumour development in bladder cancer cells cannot be excluded.…”

“…We also demonstrated that in all these three cell lines, amplification of the E2F3 gene was accompanied by E2F3 mRNA overexpression and high-level expression of the E2F3 proteins. In Northern blot analyses, we first confirmed the observation of Hurst et al (2004) that the amplification of the CDKAL1 gene in these cell lines is also accompanied by mRNA overexpression (Figure 1a). The bladder cancer cell line 5637 that exhibits amplification and overexpression of both E2F3 (Figure 2) and CDKAL1 was then chosen for small interfering RNA (siRNA)-mediated knockdown studies.…”

“…Oeggerli et al (2004) reported similar results, and on the basis of parallel analyses of E2F3 protein and the proliferation marker Ki67 suggested that E2F3 overexpression may provide a growth advantage to tumour cells by activating cellular proliferation in a subset of bladder cancers. In comparison, Hurst et al (2004) examined amplification of expression levels of the same genes within the 6p22 amplicon, but concluded that the CDKAL1 gene immediately adjacent to E2F3, rather than E2F3 itself, showed the clearest amplicon-related expression in tumour cell lines. In their study, CDKAL1 was therefore proposed as a putative bladder cancer oncogene.…”

Genetic analysis of multifocal superficial urothelial cancers by array-based comparative genomic hybridisation

“…The most common alterations of the chromosome arms were 9q loss (79%), 9p loss (75%), 11p loss (67%), 20q gain (50%), 17p loss (46%), and 11q loss (38%). Alterations of these regions were also reported in two previous array-CGH studies of bladder cancer (Veltman et al, 2003;Hurst et al, 2004). Homozygous deletions were observed at the locus where clones (9p21) containing the MTAP gene and CDKN2B (p16) gene were mapped in eight tumours of two patients, whereas high-level amplifications were not detected.…”

High-resolution analysis of chromosome rearrangements on 8p in breast, colon and pancreatic cancer reveals a complex pattern of loss, gain and translocation

“…Davidson et al, 2000;AbdelRahman et al, 2001;Karan et al, 2003;Hurst et al, 2004;Hwang et al, 2004), and more recently array CGH has begun to provide more detail of these losses (e.g. Paris et al, 2003;Douglas et al, 2004;Hurst et al, 2004;Nakao et al, 2004;Garcia et al, 2005). Distal 8p also frequently shows loss of heterozygosity (LOH), which would often reflect loss through unbalanced translocation (for references see Adams et al, 2005).…”