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Comparative Genomic Hybridization Analysis of 38 Breast Cancer Cell Lines: A Basis for Interpreting Complementary DNA Microarray Data¹

Cancer Genetics Branch, National Human Genome Research Institute, NIH, Bethesda, Maryland 20892-4470 [F. F., Y. C., R. V., O. M., Y. J., G. C. G., A. K., O-P. K.]; Laboratory of Cancer Genetics, Institute of Medical technology, University of Tampere and Tampere University Hospital, FIN-33521 Tampere, Finland [E. H. M.]; and Department of Radiation Oncology, Division of Radiation and Cancer Biology, University of Michigan Medical School, Ann Arbor, Michigan 48109-0948 [S. P. E.]

	ABSTRACT

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Breast cancer cell lines provide a useful starting point for the discovery and functional analysis of genes involved in breast cancer. Here, we studied 38 established breast cancer cell lines by comparative genomic hybridization (CGH) to determine recurrent genetic alterations and the extent to which these cell lines resemble uncultured tumors. The following chromosomal gains were observed: 8q (75%), 1q (61%), 20q (55%), 7p (44%), 3q (39%), 5p (39%), 7q (39%), 17q (33%), 1p (30%), and 20p (30%), and the most common losses were: 8p (58%), 18q (58%), 1p (42%), Xp (42%), Xq (42%), 4p (36%), 11q (36%), 18p (33%), 10q (30%), and 19p (28%). Furthermore, 35 recurrent high-level amplification sites were identified, most often involving 8q23 (37%), 20q13 (29%), 3q25-q26 (24%), 17q22-q23 (16%), 17q23-q24 (16%), 1p13 (11%), 1q32 (11%), 5p13 (11%), 5p14 (11%), 11q13 (11%), 17q12-q21 (11%), and 7q21-q22 (11%). A comparison of DNA copy number changes found in the cell lines with those reported in 17 published studies (698 tumors) of uncultured tumors revealed a substantial degree of overlap. CGH copy number profiles may facilitate identification of important new genes located at the hotspots of such chromosomal alterations. This was illustrated by analyzing expression levels of 1236 genes using cDNA microarrays in four of the cell lines. Several highly overexpressed genes (such as RCH1 at 17q23, TOPO II at 17q21-q22, as well as CAS and MYBL2 at 20q13) were involved in these recurrent DNA amplifications.

In conclusion, DNA copy number profiles were generated by CGH for most of the publicly available breast cancer cell lines and were made available on a web site (http://www.nhgri.nih.gov/DIR/CGB/CR2000). This should facilitate the correlative analysis of gene expression and copy number as illustrated here by the finding by cDNA microarrays of several overexpressed genes that were amplified.

	INTRODUCTION

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Several hundred studies on genetic alterations in cancer have been published using CGH³ (1) . There are at least 19 published CGH studies on breast cancer, reporting genetic alterations in a total of 792 cases. These studies have also identified associations of these genetic alterations with tumor grade (2) , histological subtype (3, 4, 5) , metastasis (6) , genetic predisposition (7) , as well as patient survival (8) . A number of recurrent genetic alterations have been reported, such as gains of 1q, 8q, and 20q, as well as losses of 1p, 16q, and 18q. Furthermore, specific high-level amplifications have been observed at chromosomal sites that do not coincide with the locations of the classical breast cancer oncogenes, such as 1q32, 8q23, 17q23-q25, and 20q11-q13. Several genes have already been implicated to be involved in these amplification sites, including the ZNF217, NABC1, BTAK/STK1/Aurora II, AIB1, and CAS-1 genes at 20q13 (9, 10, 11, 12) , as well as the ribosomal protein S6 kinase at 17q23 (13 , 14) . Despite this progress, there are numerous sites in the genome that are often amplified in breast cancer where the genes involved have not yet been identified.

Most of the published studies of breast cancer by CGH have analyzed unique clinical series of breast carcinomas. Therefore, any genetic alterations discovered by CGH can only be followed in the same tissue material, which usually is not accessible to other investigators. Whereas CGH can be easily performed from even small, formalin-fixed, archival tumor tissues, cloning genes involved in genetic rearrangements requires the availability of large quantities of fresh tissue material. Here, we performed a CGH study of 38 breast cancer cell lines, which comprise the majority of the publicly available breast cancer cell lines. Eleven of these represent novel cell lines recently characterized by us (15) . The CGH profiles from all 38 cell lines are available on a web site.⁴ Furthermore, a cDNA microarray analysis of 1236 genes was performed in four cases to study the relationship of gene expression and genomic copy number.

	MATERIALS AND METHODS

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CGH.
The following breast carcinoma cell lines were studied: MCF7, MDA-436, MDA-361, MDA-468, MDA-157, MDA-175 VI, MDA-453, MDA-134, MDA-231, MDA-435, MDA-415, HS578T, SKBR3, BT474, BT549, BT20, BT483, ZR-75–30, UACC-812, and HBL-100 from American Type Culture Collection (Manassas, VA); ZR-75–1-wt strain from Dr. Jeff Moscow (National Cancer Institute, Bethesda, MD); SUM-149, SUM-1315, SUM-159, SUM-185, SUM-190, SUM-206, SUM-225, and SUM-229, SUM-44, SUM-52, and SUM-102 from Dr. Steve Ethier’s laboratory (University of Michigan); MX-1, KPL-1, CAL-51, COLO-824, EFM-19, and EFM-192A from German Collection of Microorganisms and Cell Cultures (Braunschweig, Germany); and MPE-600 from Vysis Inc. (Downers Grove, IL). CGH was carried out essentially as described previously (15) . Normal male versus female hybridizations were used as negative controls and for ensuring the linearity of the hybridization (16) . Chromosomal regions where CGH ratios were >1.2 were considered as gained, and those regions where the ratio was <0.8 were considered as lost. High-level amplifications were defined as small regions with a ratio >1.4. To define recurrent, independent regions of amplification, a high-level copy number increase had to be present in a localized chromosomal region in two or more cases.

FISH.
FISH with probes to Cyclin-D1, ERBB2, and MYC (Vysis Inc.) was done on 37 of 38 cell lines and with bacterial artificial chromosome probes specific to CAS, RCH1, TOPO II, and MYBL2 on four cell lines. Dual color FISH analysis was done on interphase nuclei as described before (15) . The slides were hybridized with SpectrumOrange-labeled probes for the genes of interest with the corresponding SpectrumGreen-labeled centromeric probe for chromosomes 8, 11, 17, and 20 as reference probes. Because the cell lines are genetically rather homogeneous, 20 nonoverlapping nuclei with intact morphology based on 4',6-diamidino-2-phenylindole counterstaining were scored to determine the mean copy number of the gene-specific probes relative to the chromosome-specific reference probe.

Fluorescent cDNA Microarrays.
The cDNA microarray used for our experiments consisted of 1236 cDNA clones selected from the Unigene set (17) and printed on glass slides as described earlier (18 , 19) . Four cell lines with distinct, high-level amplifications were used in the cDNA microarray hybridizations. In all experiments, cancer cell line RNA was compared with normal, mammary gland RNA (Clontech, Palo Alto, CA) as a reference. Total RNA was extracted from breast cancer cell lines using the RNeasy kit (Qiagen, Valencia, CA). The labeling, hybridization, and washing procedures were done as described previously (20) . Briefly, 60–100 µg of total RNA extracted from tumor cell lines, and 100 µg of total RNA obtained from control sample were reverse-transcribed by oligo dT-primed polymerization using Superscript II reverse transcriptase (LTI Inc.) and Cy5-dUTP and Cy3-dUTP as fluorescent nucleotides (Amersham). After the labeling, the Cy5- and Cy3-labeled cDNAs were combined with 8 µg of poly(dA) (Pharmacia, Bridgewater, NJ), 4 µg of Escherichia coli tRNA (Sigma Chemical Co., St. Louis, MO), and 10 µg of Cot-1 DNA (Life Technologies, Inc., Rockville, MD) in 0.15% SDS, and 3x SSC. The hybridization mixture was incubated for 2 min at 98°C and then for 10 s at 4°C, and it was applied to a cDNA microarray slide. The hybridization was carried out for 16 h at 65°C. The slides were washed in 0.5x SSC and 0.01% SDS for 2 min each at room temperature.

Fluorescence intensities of the 1236 probe targets on the slide were measured using a custom-designed laser confocal microscope containing a scanning stage, appropriate excitation and emission filters, and two photomultiplier tube detectors for two fluorescent emission channels (20) . Intensity data were integrated over 225-µm² pixels and recorded at 16 bits. The Cy3 and Cy5 images were scanned independently through two separated channels, the pixel intensities of which were integrated over a 15-µm² area and recorded at 16 bits. The color images were formed by assigning tumor intensity values into the red channel and control intensity into the green channel.

The data obtained were analyzed using the ArraySuite program, developed at the National Human Genome Research Institute on the IPLab Spectrum platform (21) . To determine the actual target region based on the information from both red and green pixel values, a segmentation method was used. After background subtraction, fluorescent intensity of a particular target on the slide was calculated by averaging the intensity of every pixel inside the detected target region, and the difference of expression levels of the target was determined by taking the ratios of the R:G. The ratios were normalized for differential efficiencies of labeling, hybridization, and detection based on 88 preselected internal control genes that are usually stable for most experiments (R:G ratio close to 1.0; Ref. 20 ). A 99% confidence interval was used throughout the experiments to test the significance of differentially expressed genes. A gene is determined up- or down-regulated when the tumor:normal ratios of expression are not within the 99% confidence interval. The statistical significance of the data for each experiment is determined within that experiment and not across experiments. For this analysis, we used a filter that included all genes exhibiting an intensity at least 1000 (on a scale of 0–65535 fluorescent units) for either the red (Cy5) or green (Cy3) channels.

	RESULTS

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CGH Studies on Chromosomal Gains, Losses, and Amplifications.
On average, 19 genetic changes were found in breast cancer cell lines (range from 14 to 60), including 9 losses (range from 4 to 25) and 10 gains (range from 3 to 42)/cell line. An example of CGH ratio profiles is shown in Fig. 1 , and a summary of the different regions of gains and losses is shown in an ideogram format in Fig. 2 . The most common gains were at: 8q (75%), 1q (61%), 20q (55%), 7p (44%), 3q (39%), 5p (39%), 7q (39%), 17q (33%), 1p (30%), and 20p (30%). In addition, partial or complete losses of chromosomal regions were observed at: 8p (58%), 18q (58%), 1p (42%), Xp (42%), Xq (42%), 4p (36%), 11q (36%), 18p (33%), 10q (30%), and 19p (28%).

View larger version (28K): [in this window] [in a new window] [Download PPT slide]   Fig. 1. CGH analysis of BT483 breast cancer cell line. The green:red fluorescence ratio profiles are shown along with ± 1 SD. For each profile: black vertical line (middle), a ratio of 1.0; red line (left), a ratio of 0.8; and green line (right), a ratio of 1.2. The regions of gain (right) and loss (left) are marked as green and red bars, respectively, next to the chromosome ideograms.

View larger version (48K): [in this window] [in a new window] [Download PPT slide]   Fig. 2. A summary ideogram of gains (right) and losses (left) of chromosomal regions seen by CGH in 38 breast cancer cell lines. Chromosomal regions where CGH ratios were >1.2 were considered as gained, and those regions where the ratio was <0.8 were considered as lost.

View larger version (68K): [in this window] [in a new window] [Download PPT slide]   Fig. 3. An overview of the most common gains and losses reported in our study of 38 breast cancer cell lines as well as 17 published CGH studies of 698 breast tumors. The number of tumors analyzed in each study is indicated in the parenthesis. Green squares, the most common gains in a particular study; red squares, the most common losses. Squares with both colors, a gain and a loss for the same chromosome arm. Approximately 10 of the most common changes were indicated for each study. Despite the similarities of genetic changes observed, many of these studies focused at a specific subtype of breast cancer. Study 1 focused on grade I tumors, studies 2 and 6 focused on grade III tumors, study 3 focused on high-grade ductal carcinoma in situ, study 4 focused on node-positive tumors, studies 5, 7, and 15 focused on node-negative tumors, study 8 focused on hypodiploid cancers, study 12 focused on infiltrating lobular breast cancer, and study 14 focused on metastatic tumors.

View larger version (24K): [in this window] [in a new window] [Download PPT slide]   Fig. 4. A summary ideogram of regions of high-level amplification by CGH in the 38 breast cancer cell lines. A high-level amplification was defined as small regions (one-to-three chromosomal bands wide) with a highly elevated ratio (ratio >1.4).

View larger version (65K): [in this window] [in a new window] [Download PPT slide]   Fig. 5. Fluorescent cDNA microarray analysis of four breast cancer cell lines using an array of 1236 cDNA clones. The picture shows the original cDNA microarray image for each cell line and a histogram of the R:G ratios for each cell line, including the percentage of down- and up-regulated genes. Right, a FISH image illustrating the amplification of an overexpressed gene in the interphase nuclei of the corresponding cell lines.

	DISCUSSION

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DNA amplification is a prominent mechanism of oncogene up-regulation in breast cancer, as best illustrated by the association of ERBB2 amplification and overexpression with poor prognosis in breast cancer (31) , a finding that led to the development and implementation of anti-ERBB2 therapy. As illustrated by this and other CGH studies, DNA amplification often takes places at multiple regions of the genome. Identification of all genes involved in these amplicons would be challenging with traditional techniques. An alternative to positional cloning of amplification target genes is provided by large-scale cDNA microarray analysis of expression levels of numerous genes at once. In our cDNA microarray survey of 1236 genes, we identified several highly overexpressed genes, which mapped to the regions of amplification defined by CGH. These included TOPO II at 17q22, RCH1 at 17q23, and MYBL2 and CAS at 20q13. FISH verified the amplifications of all of these highly overexpressed genes in at least one of the four cell lines analyzed. Similarly, we recently identified the ribosomal S6 kinase gene (at 17q23) highly up-regulated in our study of the MCF7 cell line by a cDNA microarray containing 5000 cDNA clones (13) . The S6 kinase gene was significantly overexpressed and also highly amplified in all of the cell lines with 17q23 amplification.

Finally, although the majority of chromosomal changes identified in the breast cancer cell lines were also present in uncultured tumors, it will be critical to validate the presence of newly identified gene amplifications in vivo in uncultured primary breast cancers. Using our recently described high-throughput tissue microarray technology, a series of 372 primary breast cancer specimens were screened for the amplification of the MYBL2 gene, which was identified in this study to be highly overexpressed in breast cancer cell lines. Up to 7% of primary breast cancers studied showed MYBL2 amplification (32) . In a similar fashion, we found the S6 kinase gene to be highly amplified in 9% of 668 primary breast tumors. In addition, we were able to show a significant association between amplification of the gene and poor prognosis (13) .

The results shown in this study illustrate the powerful approach of defining putative amplification target genes by combining CGH information with results from cDNA microarray analysis, followed by quickly surveying large numbers of uncultured tumors with the tissue microarray technology to study the clinical significance of such newly discovered gene amplifications.

	ACKNOWLEDGMENTS

 
We thank Nasser Z. Parsa, Cheryl Ammerman, and Michele Dziubinski for excellent technical assistance and Darryl Leja for help with the technical illustrations. We also thank Vysis, Inc. for the bacterial artificial chromosome probes.

	FOOTNOTES

 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Supported in part by Grant DAMD17-94-J4382 (to S. P. E.), Department of Defense, breast cancer program, and by Academy of Finland (to E. H. M.), Sigrid Juselius Foundation and Tampere University Hospital.

² To whom requests for reprints should be addressed, at Cancer Genetics Branch, National Human Genome Research Institute, NIH, Building 49, Room 4A24, 49 Convent Drive, MSC 4470, Bethesda, MD 20892-4470. Phone: (301) 435-2896; Fax: (301) 402-7957; E-mail: okalli{at}nhgri.nih.gov

³ The abbreviations used are: CGH, comparative genomic hybridization; FISH, fluorescent in situ hybridization; R:G, red intensity:green intensity.

⁴ Internet address: http://www.nhgri.nih.gov/DIR/CGB/CR2000.

Received 10/27/99. Accepted 6/ 6/00.

	REFERENCES

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 

1. Knuutila S., Björkqvist A. M., Autio K., Tarkkanen M., Wolf M., Monni O., Szymanska J., Larramendy M. L., Tapper J., Pere H., El-Rifai W., Hemmer S., Wasenius V-M., Vidgren V., Zhu Y. DNA copy number amplifications in human neoplasms: review of comparative genomic hybridization studies. Am. J. Pathol., 152: 1107-1123, 1998.[Abstract]
2. Roylance R., Gorman P., Harris W., Liebmann R., Barnes D., Hanby A., Sheer D. Comparative genomic hybridization of breast tumors stratified by histological grade reveals new insights into the biological progression of breast cancer. Cancer Res., 59: 1433-1436, 1999.[Abstract/Free Full Text]
3. Schwendel A., Richard F., Langreck H., Kaufmann O., Lage H., Winzer K. J., Petersen I., Dietel M. Chromosome alterations in breast carcinomas: frequent involvement of DNA losses including chromosomes 4q and 21q. Br. J. Cancer, 78: 806-811, 1998.[Medline]
4. James L. A., Mitchell E. L., Menasce L., Varley J. M. Comparative genomic hybridisation of ductal carcinoma in situ of the breast: identification of regions of DNA amplification and deletion in common with invasive breast carcinoma. Oncogene, 14: 1059-1065, 1997.[Medline]
5. Nishizaki T., Chew K., Chu L., Isola J., Kallioniemi A., Weidner N., Waldman F. M. Genetic alterations in lobular breast cancer by comparative genomic hybridization. Int. J. Cancer, 74: 513-517, 1997.[Medline]
6. Kuukasjärvi T., Karhu R., Tanner M., Kähkönen M., Schaffer A., Nupponen N., Pennanen S., Kallioniemi A., Kallioniemi O. P., Isola J. Genetic heterogeneity and clonal evolution underlying development of asynchronous metastasis in human breast cancer. Cancer Res., 57: 1597-1604, 1997.[Abstract/Free Full Text]
7. Tirkkonen M., Kainu T., Loman N., Johannsson O. T., Olsson H., Barkardottir R. B., Kallioniemi O. P., Borg A. Somatic genetic alterations in BRCA2-associated and sporadic male breast cancer. Genes Chromosomes Cancer, 24: 56-61, 1999.[Medline]
8. Isola J. J., Kallioniemi O. P., Chu L. W., Fuqua S. A., Hilsenbeck S. G., Osborne C. K., Waldman F. M. Genetic aberrations detected by comparative genomic hybridization predict outcome in node-negative breast cancer. Am. J. Pathol., 147: 905-911, 1995.[Abstract]
9. Collins C., Rommens J. M., Kowbel D., Godfrey T., Tanner M., Hwang S. I., Polikoff D., Nonet G., Cochran J., Myambo K., Jay K. E., Froula J., Cloutier T., Kuo W. L., Yaswen P., Dairkee S., Giovanola J., Hutchinson G. B., Isola J., Kallioniemi O. P., Palazzolo M., Martin C., Ericsson C., Pinkel D., Albertson D., Li W-B., Gray J. W. Positional cloning of ZNF217 and NABC1: genes amplified at 20q13. 2 and overexpressed in breast carcinoma. Proc. Natl. Acad. Sci. USA, 95: 8703-8708, 1998.[Abstract/Free Full Text]
10. Anzick S. L., Kononen J., Walker R. L., Azorsa D. O., Tanner M. M., Guan X. Y., Sauter G., Kallioniemi O. P., Trent J. M., Meltzer P. S. AIB1, a steroid receptor coactivator amplified in breast and ovarian cancer. Science (Washington DC), 277: 965-968, 1997.[Abstract/Free Full Text]
11. Sen S., Zhou H., White R. A. A putative serine/threonine kinase encoding gene BTAK on chromosome 20q13 is amplified and overexpressed in human breast cancer cell lines. Oncogene, 14: 2195-2200, 1997.[Medline]
12. Brinkmann U., Gallo M., Polymeropoulos M. H., Pastan I. The human CAS (cellular apoptosis susceptibility) gene mapping on chromosome 20q13 is amplified in BT474 breast cancer cells and part of aberrant chromosomes in breast and colon cancer cell lines. Genome Res., 6: 187-194, 1996.[Abstract/Free Full Text]
13. Bärlund, M., Forozan, F., Kononen, J., Bubendorf, L., Chen, Y., Bittner, M. L., Torhorst, J., Haas, P., Bucher, C., Sauter, G., Kallioniemi, O-P., and Kallioniemi, A. Detecting activation of ribosomal protein S6 kinase complementary DNA and tissue microarray analysis. J. Natl. Cancer Inst., in press, 2000.
14. Couch F. J., Wang X. Y., Wu G. J., Qian J., Jenkins R. B., James C. D. Localization of PS6K to chromosomal region 17q23 and determination of its amplification in breast cancer. Cancer Res., 59: 1408-1411, 1999.[Abstract/Free Full Text]
15. Forozan F., Veldman R., Ammerman C. A., Parsa N. Z., Kallioniemi A., Kallioniemi O-P., Ethier S. P. Molecular cytogenetic analysis of 11 new breast cancer cell lines. Br. J. Cancer, 81: 1328-1334, 1999.[Medline]
16. Karhu R., Kähkönen M., Kuukasjärvi T., Pennanen S., Tirkkonen M., Kallioniemi O. Quality control of CGH: impact of metaphase chromosomes and the dynamic range of hybridization. Cytometry, 28: 198-205, 1997.[Medline]
17. Schuler, G. D., Boguski, M. S., Stewart, E. A., Stein, L. D., Gyapay, G., Rice, K., White, R. E., Rodriguez-Tomé, P., Aggarwal, A., Bajorek, E., Bentolila, S., Birren, B. B., Butler, A., Castle, A. B., Chiannilkulchai, N., Chu, A., Clee, C., Cowles, S., Day, P. J. R., Dibling, T., East, C., Drouot, N., Dunham, I., Duprat, S., Edwards, C., Fan, J-B., Fang, N., Fizames, C., Garrett, C., Green, L., Hadley, D., Harris, M., Harrison, P., Brady, S., Hicks, A., Holloway, E., Hui, L., Hussain, S., LouisDit-Sully, C., Ma, J., MacGilvery, A., Mader, C., Maratukulam, A., Matise, T. C., McKusick, K. B., Morissette, J., Mungall, A., Muselet, D., Nusbaum, H. C., Page, D. C., Peck, A., Perkins, S., Piercy, M., Qin, F., Quackenbush, J., Ranby, S., Reif, T., Rozen, S., Sanders, C., She, X., Silva, J., Slonim, D. K., Soderlund, C., Sun, W-L., Tabar, P., Thangarajah, T., Vega-Czarny, N., Vollrath, D., Voyticky, S., Wilmer, T., Wu, X., Adams, M. D., Auffray, C., Walter, N. A. R., Brandon, R., Dehejia, A., Goodfellow, P. N., Houlgatte, R., Hudson, J. R., Jr., Ide, S. E., Iorio, K. R., Lee, W. Y., Seki, N., Nagase, T., Ishikawa, K., Nomura, N., Phillips, C., Polymeropoulos, M. H., Sandusky, M., Schmitt, K., Berry, R., Swanson, K., Torres, R., Venter, J. C., Sikela, J. M., Beckmann, J. S., Weissenbach, J., Myers, R. M., Cox, D. R., James, M. R., Bentley, D., Deloukas, P., Lander, E. S., and Hudson, T. J. A Gene Map of the Human Genome. Science (Washington DC), 274: 540–546, 1996.
18. Duggan D. J., Bittner M., Chen Y., Meltzer P., Trent J. M. Expression profiling using cDNA microarrays. Nat. Genet., 21: 10-14, 1999.[Medline]
19. Ermolaeva O., Rastogi M., Pruitt K. D., Schuler G. D., Bittner M. L., Chen Y., Simon R., Meltzer P., Trent J. M., Boguski M. S. Data management and analysis for gene expression arrays. Nat. Genet., 20: 19-23, 1998.[Medline]
20. Khan J., Simon R., Bittner M., Chen Y., Leighton S. B., Pohida T., Smith P. D., Jiang Y., Gooden G. C., Trent J. M., Meltzer P. S. Gene expression profiling of alveolar rhabdomyosarcoma with cDNA microarrays. Cancer Res., 58: 5009-5013, 1998.[Abstract/Free Full Text]
21. Chen Y., Dougherty E. R., Bittner M. L. Ratio-based decisions and the quantitative analysis of cDNA microarray images. J. Biomed. Optics, 2: 364-374, 1997.
22. Adeyinka A., Mertens F., Idvall I., Bondeson L., Ingvar C., Mitelman F., Pandis N. Different patterns of chromosomal imbalances in metastasising and non-metastasising primary breast carcinomas. Int. J. Cancer, 84: 370-375, 1999.[Medline]
23. Buerger H., Otterbach F., Simon R., Poremba C., Diallo R., Decker T., Riethdorf L., Brinkschmidt C., Dockhorn-Dworniczak B., Boecker W. Comparative genomic hybridization of ductal carcinoma in situ of the breast-evidence of multiple genetic pathways. J. Pathol., 187: 396-402, 1999.[Medline]
24. Moore E., Magee H., Coyne J., Gorey T., Dervan P. A. Widespread chromosomal abnormalities in high-grade ductal carcinoma in situ of the breast. Comparative genomic hybridization study of pure high-grade DCIS. J. Pathol., 187: 403-409, 1999.[Medline]
25. Hermsen M. A., Baak J. P., Meijer G. A., Weiss J. M., Walboomers J. W., Snijders P. J., van Diest P. J. Genetic analysis of 53 lymph node-negative breast carcinomas by CGH and relation to clinical, pathological, morphometric, and DNA cytometric prognostic factors. J. Pathol., 186: 356-362, 1998.[Medline]
26. Tanner M. M., Karhu R. A., Nupponen N. N., Borg A., Baldetorp B., Pejovic T., Ferno M., Killander D., Isola J. J. Genetic aberrations in hypodiploid breast cancer: frequent loss of chromosome 4 and amplification of cyclin D1 oncogene. Am. J. Pathol., 153: 191-199, 1998.[Abstract/Free Full Text]
27. Tirkkonen M., Tanner M., Karhu R., Kallioniemi A., Isola J., Kallioniemi O. P. Molecular cytogenetics of primary breast cancer by CGH. Genes Chromosomes Cancer, 21: 177-184, 1998.[Medline]
28. Courjal F., Theillet C. Comparative genomic hybridization analysis of breast tumors with predetermined profiles of DNA amplification. Cancer Res., 57: 4368-4377, 1997.[Abstract/Free Full Text]
29. Kallioniemi A., Kallioniemi O. P., Piper J., Tanner M., Stokke T., Chen L., Smith H. S., Pinkel D., Gray J. W., Waldman F. M. Detection and mapping of amplified DNA sequences in breast cancer by comparative genomic hybridization. Proc. Natl. Acad. Sci. USA, 91: 2156-2160, 1994.[Abstract/Free Full Text]
30. Ried T., Just K. E., Holtgreve-Grez H., du Manoir S., Speicher M. R., Schrock E., Latham C., Blegen H., Zetterberg A., Cremer T., Auer G. Comparative genomic hybridization of formalin-fixed, paraffin-embedded breast tumors reveals different patterns of chromosomal gains and losses in fibroadenomas and diploid and aneuploid carcinomas. Cancer Res., 55: 5415-5423, 1995.[Abstract/Free Full Text]
31. Ross J. S., Fletcher J. A. The HER-2/neu oncogene: prognostic factor, predictive factor and target for therapy. Semin. Cancer Biol., 9: 125-138, 1999.[Medline]
32. Kononen J., Bubendorf L., Kallioniemi A., Bärlund M., Schraml P., Leighton S., Torhorst J., Mihatsch M. J., Sauter G., Kallioniemi O. P. Tissue microarrays for high-throughput molecular profiling of tumor specimens. Nat. Med., 4: 844-847, 1998.[Medline]

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				B. Xing, C. M. T. Greenwood, and S. B. Bull A hierarchical clustering method for estimating copy number variation Biostat., July 1, 2007; 8(3): 632 - 653. [Abstract] [Full Text] [PDF]

				Z. Q. Yang, K. L. Streicher, M. E. Ray, J. Abrams, and S. P. Ethier Multiple Interacting Oncogenes on the 8p11-p12 Amplicon in Human Breast Cancer Cancer Res., December 15, 2006; 66(24): 11632 - 11643. [Abstract] [Full Text] [PDF]

				M. Dolled-Filhart, L. Ryden, M. Cregger, K. Jirstrom, M. Harigopal, R. L. Camp, and D. L. Rimm Classification of breast cancer using genetic algorithms and tissue microarrays. Clin. Cancer Res., November 1, 2006; 12(21): 6459 - 6468. [Abstract] [Full Text] [PDF]

				C. Chen, A. K. Seth, and A. E. Aplin Genetic and Expression Aberrations of E3 Ubiquitin Ligases in Human Breast Cancer Mol. Cancer Res., October 1, 2006; 4(10): 695 - 707. [Abstract] [Full Text] [PDF]

				J. Russo, S. V. Fernandez, P. A. Russo, R. Fernbaugh, F. S. Sheriff, H. M. Lareef, J. Garber, and I. H. Russo 17-Beta-estradiol induces transformation and tumorigenesis in human breast epithelial cells FASEB J, August 1, 2006; 20(10): 1622 - 1634. [Abstract] [Full Text] [PDF]

				C. Ginestier, N. Cervera, P. Finetti, S. Esteyries, B. Esterni, J. Adelaide, L. Xerri, P. Viens, J. Jacquemier, E. Charafe-Jauffret, et al. Prognosis and Gene Expression Profiling of 20q13-Amplified Breast Cancers Clin. Cancer Res., August 1, 2006; 12(15): 4533 - 4544. [Abstract] [Full Text] [PDF]

				D. A. Engler, G. Mohapatra, D. N. Louis, and R. A. Betensky A pseudolikelihood approach for simultaneous analysis of array comparative genomic hybridizations Biostat., July 1, 2006; 7(3): 399 - 421. [Abstract] [Full Text] [PDF]

				N. Maqani, A. Belkhiri, C. Moskaluk, S. Knuutila, A. A. Dar, and W. El-Rifai Molecular Dissection of 17q12 Amplicon in Upper Gastrointestinal Adenocarcinomas Mol. Cancer Res., July 1, 2006; 4(7): 449 - 455. [Abstract] [Full Text] [PDF]

				M. J. Ellis Functional Analysis of the Breast Cancer Genome J. Clin. Oncol., April 10, 2006; 24(11): 1649 - 1650. [Full Text] [PDF]

				A A Ghazani, N C R Arneson, K Warren, and S J Done Limited tissue fixation times and whole genomic amplification do not impact array CGH profiles. J. Clin. Pathol., March 1, 2006; 59(3): 311 - 315. [Abstract] [Full Text] [PDF]

				P K Lovelock, S Healey, W Au, E Y M Sum, A Tesoriero, E M Wong, S Hinson, R Brinkworth, A Bekessy, O Diez, et al. Genetic, functional, and histopathological evaluation of two C-terminal BRCA1 missense variants J. Med. Genet., January 1, 2006; 43(1): 74 - 83. [Abstract] [Full Text] [PDF]

				L. Melchor, S. Alvarez, E. Honrado, J. Palacios, A. Barroso, O. Diez, A. Osorio, and J. Benitez The Accumulation of Specific Amplifications Characterizes Two Different Genomic Pathways of Evolution of Familial Breast Tumors Clin. Cancer Res., December 15, 2005; 11(24): 8577 - 8584. [Abstract] [Full Text] [PDF]

				V. Gelsi-Boyer, B. Orsetti, N. Cervera, P. Finetti, F. Sircoulomb, C. Rouge, L. Lasorsa, A. Letessier, C. Ginestier, F. Monville, et al. Comprehensive Profiling of 8p11-12 Amplification in Breast Cancer Mol. Cancer Res., December 1, 2005; 3(12): 655 - 667. [Abstract] [Full Text] [PDF]

				L. D. Miller, J. Smeds, J. George, V. B. Vega, L. Vergara, A. Ploner, Y. Pawitan, P. Hall, S. Klaar, E. T. Liu, et al. From The Cover: An expression signature for p53 status in human breast cancer predicts mutation status, transcriptional effects, and patient survival PNAS, September 20, 2005; 102(38): 13550 - 13555. [Abstract] [Full Text] [PDF]

				F. Reyal, N. Stransky, I. Bernard-Pierrot, A. Vincent-Salomon, Y. de Rycke, P. Elvin, A. Cassidy, A. Graham, C. Spraggon, Y. Desille, et al. Visualizing Chromosomes as Transcriptome Correlation Maps: Evidence of Chromosomal Domains Containing Co-expressed Genes--A Study of 130 Invasive Ductal Breast Carcinomas Cancer Res., February 15, 2005; 65(4): 1376 - 1383. [Abstract] [Full Text] [PDF]

				J. Zhou, T. Suzuki, A. Kovacic, R. Saito, Y. Miki, T. Ishida, T. Moriya, E. R. Simpson, H. Sasano, and C. D. Clyne Interactions between Prostaglandin E2, Liver Receptor Homologue-1, and Aromatase in Breast Cancer Cancer Res., January 15, 2005; 65(2): 657 - 663. [Abstract] [Full Text] [PDF]

				M. Zollo, A. Andre, A. Cossu, M. C. Sini, A. D'Angelo, N. Marino, M. Budroni, F. Tanda, G. Arrigoni, and G. Palmieri Overexpression of h-prune in Breast Cancer is Correlated with Advanced Disease Status Clin. Cancer Res., January 1, 2005; 11(1): 199 - 205. [Abstract] [Full Text] [PDF]

				G. Wang, E. Maher, C. Brennan, L. Chin, C. Leo, M. Kaur, P. Zhu, M. Rook, J. L. Wolfe, and G. M. Makrigiorgos DNA amplification method tolerant to sample degradation Genome Res., November 1, 2004; 14(11): 2357 - 2366. [Abstract] [Full Text] [PDF]

				A. B. Moffa, S. L. Tannheimer, and S. P. Ethier Transforming Potential of Alternatively Spliced Variants of Fibroblast Growth Factor Receptor 2 in Human Mammary Epithelial Cells Mol. Cancer Res., November 1, 2004; 2(11): 643 - 652. [Abstract] [Full Text] [PDF]

				B. Orsetti, M. Nugoli, N. Cervera, L. Lasorsa, P. Chuchana, L. Ursule, C. Nguyen, R. Redon, S. du Manoir, C. Rodriguez, et al. Genomic and Expression Profiling of Chromosome 17 in Breast Cancer Reveals Complex Patterns of Alterations and Novel Candidate Genes Cancer Res., September 15, 2004; 64(18): 6453 - 6460. [Abstract] [Full Text] [PDF]

				M Lacroix, R-A Toillon, and G Leclercq Stable 'portrait' of breast tumors during progression: data from biology, pathology and genetics Endocr. Relat. Cancer, September 1, 2004; 11(3): 497 - 522. [Abstract] [Full Text] [PDF]

				A. Somasiri, J. S. Nielsen, N. Makretsov, M. L. McCoy, L. Prentice, C. B. Gilks, S. K. Chia, K. A. Gelmon, D. B. Kershaw, D. G. Huntsman, et al. Overexpression of the Anti-Adhesin Podocalyxin Is an Independent Predictor of Breast Cancer Progression Cancer Res., August 1, 2004; 64(15): 5068 - 5073. [Abstract] [Full Text] [PDF]

				M. Zheng, R. Simon, M. Mirlacher, R. Maurer, T. Gasser, T. Forster, P. A. Diener, M. J. Mihatsch, G. Sauter, and P. Schraml TRIO Amplification and Abundant mRNA Expression Is Associated with Invasive Tumor Growth and Rapid Tumor Cell Proliferation in Urinary Bladder Cancer Am. J. Pathol., July 1, 2004; 165(1): 63 - 69. [Abstract] [Full Text] [PDF]

				X.-Y. Guan, J. M-W. Fung, N.-F. Ma, S.-H. Lau, L.-S. Tai, D. Xie, Y. Zhang, L. Hu, Q.-L. Wu, Y. Fang, et al. Oncogenic Role of eIF-5A2 in the Development of Ovarian Cancer Cancer Res., June 15, 2004; 64(12): 4197 - 4200. [Abstract] [Full Text] [PDF]

				G. Wang, C. Brennan, M. Rook, J. L. Wolfe, C. Leo, L. Chin, H. Pan, W.-H. Liu, B. Price, and G. M. Makrigiorgos Balanced-PCR amplification allows unbiased identification of genomic copy changes in minute cell and tissue samples Nucleic Acids Res., May 21, 2004; 32(9): e76 - e76. [Abstract] [Full Text] [PDF]

				Y.-M. Jeng, S.-Y. Peng, C.-Y. Lin, and H.-C. Hsu Overexpression and Amplification of Aurora-A in Hepatocellular Carcinoma Clin. Cancer Res., March 15, 2004; 10(6): 2065 - 2071. [Abstract] [Full Text] [PDF]

				M. E. Ray, Z. Q. Yang, D. Albertson, C. G. Kleer, J. G. Washburn, J. A. Macoska, and S. P. Ethier Genomic and Expression Analysis of the 8p11-12 Amplicon in Human Breast Cancer Cell Lines Cancer Res., January 1, 2004; 64(1): 40 - 47. [Abstract] [Full Text] [PDF]

				R. I. Skotheim, A. Kallioniemi, B. Bjerkhagen, F. Mertens, H. R. Brekke, O. Monni, S. Mousses, N. Mandahl, G. Soeter, J. M. Nesland, et al. Topoisomerase-II{alpha} Is Upregulated in Malignant Peripheral Nerve Sheath Tumors and Associated With Clinical Outcome J. Clin. Oncol., December 15, 2003; 21(24): 4586 - 4591. [Abstract] [Full Text] [PDF]

				P. Kauraniemi, T. Kuukasjarvi, G. Sauter, and A. Kallioniemi Amplification of a 280-Kilobase Core Region at the ERBB2 Locus Leads to Activation of Two Hypothetical Proteins in Breast Cancer Am. J. Pathol., November 1, 2003; 163(5): 1979 - 1984. [Abstract] [Full Text] [PDF]

				F Al-Mulla, M Al-Maghrebi, and G Varadharaj Expressive genomic hybridisation: gene expression profiling at the cytogenetic level Mol. Pathol., August 1, 2003; 56(4): 210 - 217. [Abstract] [Full Text] [PDF]

				N. M. Luscombe, T. E. Royce, P. Bertone, N. Echols, C. E. Horak, J. T. Chang, M. Snyder, and M. Gerstein ExpressYourself: a modular platform for processing and visualizing microarray data Nucleic Acids Res., July 1, 2003; 31(13): 3477 - 3482. [Abstract] [Full Text] [PDF]

				C. Montagna, M.-S. Lyu, K. Hunter, L. Lukes, W. Lowther, T. Reppert, B. Hissong, Z. Weaver, and T. Ried The Septin 9 (MSF) Gene Is Amplified and Overexpressed in Mouse Mammary Gland Adenocarcinomas and Human Breast Cancer Cell Lines Cancer Res., May 1, 2003; 63(9): 2179 - 2187. [Abstract] [Full Text] [PDF]

				M. A. Dressman, A. Baras, R. Malinowski, L. B. Alvis, I. Kwon, T. M. Walz, and M. H. Polymeropoulos Gene Expression Profiling Detects Gene Amplification and Differentiates Tumor Types in Breast Cancer Cancer Res., May 1, 2003; 63(9): 2194 - 2199. [Abstract] [Full Text] [PDF]

				N. S. Williams, R. B. Gaynor, S. Scoggin, U. Verma, T. Gokaslan, C. Simmang, J. Fleming, D. Tavana, E. Frenkel, and C. Becerra Identification and Validation of Genes Involved in the Pathogenesis of Colorectal Cancer Using cDNA Microarrays and RNA Interference Clin. Cancer Res., March 1, 2003; 9(3): 931 - 946. [Abstract] [Full Text] [PDF]

				A K Banerjee, P H Rabbitts, and J George Lung cancer * 3: Fluorescence bronchoscopy: clinical dilemmas and research opportunities Thorax, March 1, 2003; 58(3): 266 - 271. [Abstract] [Full Text] [PDF]

				J. Gollub, C. A. Ball, G. Binkley, J. Demeter, D. B. Finkelstein, J. M. Hebert, T. Hernandez-Boussard, H. Jin, M. Kaloper, J. C. Matese, et al. The Stanford Microarray Database: data access and quality assessment tools Nucleic Acids Res., January 1, 2003; 31(1): 94 - 96. [Abstract] [Full Text] [PDF]

				A. Bar-Shira, J. H. Pinthus, U. Rozovsky, M. Goldstein, W. R. Sellers, Y. Yaron, Z. Eshhar, and A. Orr-Urtreger Multiple Genes in Human 20q13 Chromosomal Region Are Involved in an Advanced Prostate Cancer Xenograft Cancer Res., December 1, 2002; 62(23): 6803 - 6807. [Abstract] [Full Text] [PDF]

				L. F. A. Wessels, T. van Welsem, A. A. M. Hart, L. J. van't Veer, M. J. T. Reinders, and P. M. Nederlof Molecular Classification of Breast Carcinomas by Comparative Genomic Hybridization: a Specific Somatic Genetic Profile for BRCA1 Tumors Cancer Res., December 1, 2002; 62(23): 7110 - 7117. [Abstract] [Full Text] [PDF]

				J. Climent, J. A. Martinez-Climent, D. Blesa, M. J. Garcia-Barchino, R. Saez, D. Sanchez-Izquierdo, P. Azagra, A. Lluch, and J. Garcia-Conde Genomic Loss of 18p Predicts an Adverse Clinical Outcome in Patients with High-Risk Breast Cancer Clin. Cancer Res., December 1, 2002; 8(12): 3863 - 3869. [Abstract] [Full Text] [PDF]

				J. R. Pollack, T. Sorlie, C. M. Perou, C. A. Rees, S. S. Jeffrey, P. E. Lonning, R. Tibshirani, D. Botstein, A.-L. Borresen-Dale, and P. O. Brown Microarray analysis reveals a major direct role of DNA copy number alteration in the transcriptional program of human breast tumors PNAS, October 1, 2002; 99(20): 12963 - 12968. [Abstract] [Full Text] [PDF]

				S. Mohr, G. D. Leikauf, G. Keith, and B. H. Rihn Microarrays as Cancer Keys: An Array of Possibilities J. Clin. Oncol., July 15, 2002; 20(14): 3165 - 3175. [Abstract] [Full Text] [PDF]

				J. J. Oh, A. R. West, M. C. Fishbein, and D. J. Slamon A Candidate Tumor Suppressor Gene, H37, from the Human Lung Cancer Tumor Suppressor Locus 3p21.3 Cancer Res., June 1, 2002; 62(11): 3207 - 3213. [Abstract] [Full Text] [PDF]

				A. Dellas, J. Torhorst, E. Schultheiss, M. J. Mihatsch, and H. Moch DNA Sequence Losses on Chromosomes 11p and 18q Are Associated with Clinical Outcome in Lymph Node-negative Ductal Breast Cancer Clin. Cancer Res., May 1, 2002; 8(5): 1210 - 1216. [Abstract] [Full Text] [PDF]

				R. I. Skotheim, O. Monni, S. Mousses, S. D. Fossa, O.-P. Kallioniemi, R. A. Lothe, and A. Kallioniemi New Insights into Testicular Germ Cell Tumorigenesis from Gene Expression Profiling Cancer Res., April 1, 2002; 62(8): 2359 - 2364. [Abstract] [Full Text] [PDF]

				P. Platzer, M. B. Upender, K. Wilson, J. Willis, J. Lutterbaugh, A. Nosrati, J. K. V. Willson, D. Mack, T. Ried, and S. Markowitz Silence of Chromosomal Amplifications in Colon Cancer Cancer Res., February 1, 2002; 62(4): 1134 - 1138. [Abstract] [Full Text] [PDF]

				T. Ueno, A. Tangoku, S. Yoshino, T. Abe, H. Toshimitsu, T. Furuya, S. Kawauchi, A. Oga, M. Oka, and K. Sasaki Gain of 5p15 Detected by Comparative Genomic Hybridization as an Independent Marker of Poor Prognosis in Patients with Esophageal Squamous Cell Carcinoma Clin. Cancer Res., February 1, 2002; 8(2): 526 - 533. [Abstract] [Full Text] [PDF]

				P. Kauraniemi, M. Barlund, O. Monni, and A. Kallioniemi New Amplified and Highly Expressed Genes Discovered in the ERBB2 Amplicon in Breast Cancer by cDNA Microarrays Cancer Res., November 1, 2001; 61(22): 8235 - 8240. [Abstract] [Full Text] [PDF]

				M. Tsuneizumi, M. Emi, H. Nagai, H. Harada, G. Sakamoto, F. Kasumi, S. Inoue, T. Kazui, and Y. Nakamura Overrepresentation of the EBAG9 Gene at 8q23 Associated with Early-Stage Breast Cancers Clin. Cancer Res., November 1, 2001; 7(11): 3526 - 3532. [Abstract] [Full Text] [PDF]

				A. Maitra, I. I. Wistuba, C. Washington, A. K. Virmani, R. Ashfaq, S. Milchgrub, A. F. Gazdar, and J. D. Minna High-Resolution Chromosome 3p Allelotyping of Breast Carcinomas and Precursor Lesions Demonstrates Frequent Loss of Heterozygosity and a Discontinuous Pattern of Allele Loss Am. J. Pathol., July 1, 2001; 159(1): 119 - 130. [Abstract] [Full Text] [PDF]

				M. Bärlund, O. Monni, J. Kononen, R. Cornelison, J. Torhorst, G. Sauter, O.-P. Kallioniemi, and A. Kallioniemi Multiple Genes at 17q23 Undergo Amplification and Overexpression in Breast Cancer Cancer Res., October 1, 2000; 60(19): 5340 - 5344. [Abstract] [Full Text]

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