These tumor-specific types of copy number profiles determined the copy number phenotypes of tumors. Thus, tumors that differ in their histology, in the genes they contain that are inactivated, in their response to therapy or in other clinicopathological parameters, can be readily distinguished on the basis of the types of genetic instability displayed and on the selection of the genes that are altered. Analysis of the numbers and types of aberrations in the aCGH copy number profiles [23] from mismatch repair (MMR)-proficient and MMR-deficient colorectal cancer cell lines, confirmed the cytogenetic observations; however MMR-deficient cell lines with alterations in MSH2 showed fewer aberrations than those with alterations in MLH1. The aCGH analysis of microsatellite-stable, near-diploid bowel cancers suggested that this group is heterogeneous, one type having few (<6) chromosomal-scale changes and the other having more (>10) changes and resembling micro- satellite-stable, non diploid cancers [22]. A comparison of changes in copy number in breast tumors based on histological subtype and estrogen receptor (ER) status showed that ER-positive infiltrating ductal carcinoma had a higher frequency of gain in 16p13 and loss in 16q21 than ER-negative infiltrating ductal carcinoma, while ER-positive infiltrating lobular carcinoma differed from ER-positive infiltrating ductal carcinoma in the frequency of gain in 1q and loss in 11q and in having high-level amplification in 1q32, 8p23, 11q13 and 11q14 [19]. In a study of liposarcoma, copy number profiles had greater power to discriminate between dedifferentiated and pleomorphic subtypes than expression profiling [20]. Specific loss at 8p23.2 was associated with an advanced stage of prostate cancer, and gain at 11q13.1 was predictive for postoperative recurrence independent of stage and grade [18]. Comparison with an independent set of metastatic samples from prostate cancers revealed approximately 40 candidate markers associated with metastatic potential-copy number aberrations at these loci may define “metastatic” genotypes. A study of bladder tumors failed to find any significant relationship between changes in copy number and tumor stage and grade [17]. However, analysis of changes in copy number at particular loci showed that certain aberrations occurred together (e.g., gains of ERBB2 and CCNE1), whereas alterations in the copy number of loci for genes that function in the same pathway, such as gains of CCND1 and E2F3,were found to be ‘complementary’ as they did not occur in the same tumors [17]. This study suggests that copy number profiles may be useful for understanding deregulation of cellular control pathways in solid tumors. The general applicability of these observations should be confirmed in future as more aCGH studies of other tumor types are published.

      Array Comparative Genomic Hybridization for High-Resolution Analysis and Fine-Mapping of the Regions of Change. Several studies have taken advantage of the higher resolution afforded by aCGH to map more precisely the boundaries and amplification maxima of amplified regions [24-30]. Once these are known, candidate oncogenes that map within the region or at the amplification maximum, and tumor suppressor genes within the regions of loss will readily be identified from the genome sequence database. Investigation of expression levels of these candidate genes in tumors and cell lines can then be used to determine which are most likely to contribute to the disease phenotype and to be the ‘driver gene(s)’ for amplification [7,10,16]. The capability of aCGH to provide high-resolution mapping of variation in copy number has been demonstrated in the case of breast tumors, where the fine structure of amplicons has been resolved and potential candidate genes identified. Genomic profiles indicated severe rearrangements of chromosome 17 in breast cancers. This chromosome was subdivided into 13 consensus segments: four regions showing mainly losses, six regions showing mainly gains, and three regions showing either gains or losses [25]. High-resolution analysis of DNA copy number of 22q in ovarian cancers displayed three distinct gene copy number profiles: one with heterozygous terminal deletion of various sizes, the second with the coexistence of heterozygous deletion and different patterns of low-copy-number-gain of the proximal half of 22q, and the third with continuous deletion encompassing the entire 22q [27]. In Barrett’s adenocarcinoma, changes in DNA sequence copy number for a panel of about 50 genes were identified, most of which have not previously been described: these included gains of SNRPN, GNLY, NME1, DDX15, LAMA3, and losses of PDGFB, AKT3, RASSFI, FHIT, CDKN2A, and SAS [29]. Analysis of ovarian cancer cell lines by aCGH detected multiple novel changes. For example, amplification on 11q22 near the progesterone receptor gene and a cluster of matrix metal­loproteinase loci, and potential oncogenes (gene for cyclin E, PIK3C2G) or tumor suppressor genes (CDKN2C, gene for SMAD4-interacting protein, RASSF2) were mapped to other detected regions [26]. Using aCGH, consistent sub­megabase deletions (550 kb region in 11q13 and 300 kb region in 13q12) were identified in low-grade oligoden­driomas [30]. Novel regions of chromosomal amplification at 6p21, 5p13, and 12q14 were discovered in gastric cancer [28].

      Array Comparative Genomic Hybridization and Gene-Expression Profiling Data. The parallel analysis of cytogenetic and transcriptional profiling data has revealed that changes in DNA copy number can have noticeable effects on gene expression. The dependence of gene expression on DNA copy number has been observed in aneu­ploid tumors and tumor derived cell lines, and in tissues obtained from patients with inherited trisomy disorders [31-33]. Regional expression changes have been shown to associate with cytogenetic abnormalities as determined by aCGH and traditional CGH in hepatocellular carcinoma and in clear cell renal cell carcinoma [32]. Parallel micro­array measurements of mRNA levels in breast tumors revealed the degree to which variation in gene copy number contributes to variation in gene expression of tumor cells. Thus, 62% of highly amplified genes show moderately or highly elevated expression; DNA copy number influences gene expression across a wide range of DNA copy number alterations (deletions, low-, mid- and high-level amplification); on average, a 2-fold change in DNA copy number is associated with a corresponding 1.5-fold change in mRNA levels; and overall, at least 12% of all variation in gene expression among breast tumors is directly attributable to underlying variation in gene copy number [33]. A good correspondence between copy number alterations and changes in gene expression has also been found by genome-wide aCGH and global expression profiling in other investigations [31].