Genome-Wide Array-Based Comparative Genomic Hybridization Reveals Multiple Amplification Targets and Novel Homozygous Deletions in Pancreatic Carcinoma Cell Lines

Abstract

Pancreatic carcinomas display highly complex chromosomal abnormalities, including many structural and numerical aberrations. There is ample evidence indicating that some of these abnormalities, such as recurrent amplifications and homozygous deletions, contribute to tumorigenesis by altering expression levels of critical oncogenes and tumor suppressor genes. To increase the understanding of gene copy number changes in pancreatic carcinomas and to identify key amplification/deletion targets, we applied genome-wide array-based comparative genomic hybridization to 31 pancreatic carcinoma cell lines. Two different microarrays were used, one containing 3,565 fluorescence in situ hybridization-verified bacterial artificial chromosome clones and one containing 25,468 cDNA clones representing 17,494 UniGene clusters. Overall, the analyses revealed a high genomic complexity, with several copy number changes detected in each case. Specifically, 60 amplicons at 32 different locations were identified, most frequently located within 8q (8 cases), 12p (7 cases), 7q (5 cases), 18q (5 cases), 19q (5 cases), 6p (4 cases), and 8p (4 cases). Amplifications of 8q and 12p were mainly clustered at 8q23-24 and 12p11-12, respectively, whereas amplifications on other chromosome arms were more dispersed. Furthermore, our analyses identified several novel homozygously deleted segments located to 9p24, 9p21, 9q32, 10p12, 10q22, 12q24, and 18q23. The individual complexity and aberration patterns varied substantially among cases, i.e., some cell lines were characterized mainly by high-level amplifications, whereas others showed primarily whole-arm imbalances and homozygous deletions. The described amplification and deletion targets are likely to contain genes important in pancreatic tumorigenesis.

Microarray analyses reveal strong influence of DNA copy number alterations on the transcriptional patterns in pancreatic cancer: implications for the interpretation of genomic amplifications

“…For cDNA clones lacking gene symbol annotation, clone identities are shown with the corresponding UniGene cluster number. The 29 pancreatic carcinoma cell lines analysed comprised LPC1p, LPC2p, LPC3p, LPC4p, LPC5m, LPC6p, LPC7m, LPC8p, LPC10m, LPC11p, LPC12m, LPC13p, LPC14p, and LPC15p, which have been previously described (Jonson et al, 1999;Heidenblad et al, 2004), and the established cell lines AsPC-1, BxPC-3, Capan-2, CFPAC-1, DANG, Hs700T, Hs766T, HupT3, HupT4, PANC-1, PaTu8902, PaTu8988S, PaTu8988T, SU.86.86, and SW1990 obtained from cell repositories. Expression profiling was performed using cDNA microarrays, containing 25 648 different cDNA clones (17,494 UniGene clusters) obtained from the Swegene DNA microarray resource center at Lund University (http://swegene.onk.lu.se).…”

“…The figure shows 98/71 genes/ESTs differentially expressed, as determined by a Bonferroni adjusted t-test (Po0.05) using the TIGR MeV suite of software (Saeed et al, 2003), in cases with 8q24/12p11-12 amplifications as compared with their unaffected counterparts. The cases were selected based on previous conventional and array-based CGH data (Mahlama¨ki et al, 1997;Heidenblad et al, 2004). Expression levels are presented on a log2-based pseudocolor scale (color code on top), where red and green indicate over-and underexpression, respectively, and gray denotes missing values.…”

“…The DNA copy number variations, which range from homozygous deletions to high-level amplifications, are believed to constitute key genetic alterations in the development and progression of this tumor type. To investigate the genome-wide transcriptional effects of gene copy number alterations, in particular of genomic amplifications, we performed expression profiling analyses of 29 pancreatic carcinoma cell lines using cDNA microarrays, and compared the results with matching genomic profiling data (Heidenblad et al, 2004). The analysis of the combined genomic and expression data sets allowed us to (i) determine the genome-wide association between gene copy number and expression levels in pancreatic cancer, (ii) identify 67 recurrently overexpressed genes in seven precisely mapped commonly amplified regions, and (iii) analyse the genome-wide mRNA effects two of the most frequently amplified regions in pancreatic cancer, 8q23-24 and 12p11-12.…”

“…We previously determined the exact genomic boundaries of 60 amplicons in the investigated cell lines (Heidenblad et al, 2004). In total, 491 (36%) of the UniGene clusters harboring clones in these segments were found to show more than twofold increased expression (Supplementary Table 1).…”

A multiplex PCR predictor for aCGH success of FFPE samples

“…In the past, we have noticed that this was not solved by repeating aCGH experiments, even when DNA was isolated from new sections from the same tissue blocks (Van Beers et al, 2005). Nevertheless, it is possible to obtain high-quality data using archival DNA samples in array CGH experiments ( Figure 1) (Gray et al, 1994;Ried et al, 1995;Albertson and Pinkel, 2003;Heidenblad et al, 2004;Loo et al, 2004;Devries et al, 2005), even from 20-year-old tissue blocks, provided that robust procedures, high-quality reagents and 'good' sample DNA quality are being used. A 'good sample quality' definition and an assay to determine this FFPE DNA sample quality would therefore be of great value.…”