High definition cytogenetics and oligonucleotide aCGH analyses of cisplatin-resistant ovarian cancer cells
Array comparative genomic hybridization (aCGH) is a key platform to assess cancer genomic profiles. Many structural genomic aberrations cannot be detected by aCGH alone. We have applied molecular cytogenetic analyses including spectral karyotyping, multicolor banding, and fluorescence in situ hybrid...
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| Published in | Genes chromosomes & cancer Vol. 47; no. 5; pp. 427 - 436 |
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| Main Authors | , , , , , , , , |
| Format | Journal Article |
| Language | English |
| Published |
Hoboken
Wiley Subscription Services, Inc., A Wiley Company
01.05.2008
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| Subjects | |
| Online Access | Get full text |
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| Summary: | Array comparative genomic hybridization (aCGH) is a key platform to assess cancer genomic profiles. Many structural genomic aberrations cannot be detected by aCGH alone. We have applied molecular cytogenetic analyses including spectral karyotyping, multicolor banding, and fluorescence in situ hybridization with aCGH to comprehensively investigate the genomic aberrations associated with cisplatin resistance in A2780 ovarian cancer cells. A2780 is a well‐established model of chemotherapeutic resistance with distinct karyotypic abnormalities in the parental and cisplatin‐resistant cells. Cytogenetic analysis revealed that two unbalanced translocations, der(8)t(1;8) and der(X)t(X;1), and loss of chromosome 13 were present only in the resistant line. Our aCGH analyses detected imbalances affecting an additional 10.59% of the genome in the cisplatin‐resistant cells compared with the parental. DNA copy number changes included deletions at 1p10–p22.1, 8p23.3, and Xq13.1‐pter, and a duplication of 8q11.22‐q23. Cryptic genomic aberrations associated with concurrent localized changes of specific gene expression included a homozygous deletion of 0.38 Mb at 1p21.3 adjacent to SNX7, and an insertional transposition of 0.85 Mb from 13q12.12 into chromosome 22. This latter rearrangement led to an overexpression of four contiguous genes that flanked one of the breakpoint regions in chromosome 13. Furthermore, 17 genes showed differential expression correlating with genomic gain or loss between the resistant and parent lines, validated by a second expression array platform. These results highlight the integration of comprehensive profiling to determine relationships of genomic aberrations and genes associated with an in vitro drug resistance model in ovarian cancer. This article contains Supplementary Material available at http://www.interscience.wiley.com/jpages/1045‐2257/suppmat. © 2008 Wiley‐Liss, Inc. |
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| Bibliography: | istex:9EDF8A3CCF32A29791A034D0AD2C82F9D156E236 Supplementary Figure 1. A. A2780par aCGH profile for chromosome 6. (ii) A2780par SKY (inverted DAPI in the left and pseudo color on right) and (iii) XCyte 1mBAND in for the derivative chromosome der(6)t(1;6)(q24;q22) contains the 1q22 region of chromosome 1. B. A2780cis aCGH profile for chromosome 6. (ii) A2780cis SKY (inverted DAPI in the left and pseudo color on right) and (iii) XCyte 1 mBAND for the same derivative chromosome der(6)t(1;6)(q21.2;q22) as in A2780par which also contains the 1q22-pter region of chromosome 1.Supplementary Figure 2. A. (i) A2780cis SKY (inverted DAPI on left and pseudo color on right) and XCyte X mBAND of the normal chromosome X and (ii) the derivative chromosome X, der(X)t(X;1)(q13.1;q11), which is associated with loss of Xq12-q23 in A2780cis . Two copies of the Xpter-Xq12 region are represented by the purple line. B. (i) A2780cis aCGH view of chromosome X. The blue box is the zoomed in view of the Xq12 breakpoint that was localized to the androgen receptor locus (inset). However, there was no significant enrichment of differentially expressed genes in the A2780cis cells, panel B (ii). All aCGH and gene expression data plotted represent the average of dye flip experiments (materials and methods). All ADM1 settings include a threshold of 10 and a filter of 2 probes with a minimum log2ratio fold change of 0.2. High scoring intervals in A2780cis cells that pass these filters are shaded in pink and their genomic locations denoted by solid red vertical lines in the aCGH plots. C. (i) FISH validation using BAC clone RP11-51D15 which maps to the Xq26.3 region, showing two signals in A2780par cell lines on chromosome X, and (ii) one signal in the A2780cis cell line. The inverted DAPI and real color images are shown.Supplementary Table 1. High Resolution Drug-Specific Copy Number Aberrations as Detected by ADM1 The table lists the bands as determined by the start and end base pair positions exported from aCGH analytics along with the number of probes present in that region. The blue arrow denotes drug-specific aberrant regions that were detected by SKY, m-BAND, and aCGH. The ADM1 score of each interval, which represents the deviation from the average log2ratios from its expected value, is indicated along with the log2ratios for that region. Where log2ratio>0.5 represents a one or greater copy gain and ratios of <-0.5 represents a one or greater copy loss. All CGH data represent the average of dye-flip experiments for each of the cell lines.Supplementary Table 2. Relative Percentages of Genomic Imbalance in A2780cis in Comparison to A2780par Sum of the total genomic imbalances based on the regions detected by the ADM1 algorithm for each cell line was tabulated. Cisplatin specific genomic imbalances were calculated by subtracting A2780cis total genomic imbalance from A2780par and divided by the total human genome of 3000 Mb.Supplementary Table 3. Results of Differential Expression Enrichment Analysis for A2780cis Specific Regions on Chromosomes 1, 8, and 13 Genomic regions on chromosomes 1 and 13 are enriched in genes that are down-regulated in A2780cis compared to the A2780par line; genomic region on chromosome 8 is enriched and up-regulated in the ovarian drug resistant cell line. Genes with larger than 2-fold expression change were considered differentially expressed. Enrichment p-values are calculated based on the hyper-geometric distribution.Supplementary Table 4. Coordinated Genomic Change and Gene Expression Between A2780 Resistant and Parent Cell Lines using 2 Separate Expression PlatformsThese 17 genes have coordinated down or up regulation with the areas of genomic change. The values are represented as differences in the log normalized expression values using both the Affymertix and Agilent platforms. There was complete concordance for all 17 genes whereby expression differences between the resistant and parent cell lines was dependent of genomic loss or gain. Ontario Cancer Research Network Ontario Cancer Biomarker Network ark:/67375/WNG-ZZDCX30K-9 ArticleID:GCC20547 ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
| ISSN: | 1045-2257 1098-2264 |
| DOI: | 10.1002/gcc.20547 |