(Background and Aim) The genetic instabilities in colorectal cancer can be divided into two distinct types, with over 85 % of tumors showing chromosomal aberrations, and 10 % of tumors demonstrating microsatellite instability. Though comparative genomic hybridization (CGH) was initially used to detect chromosomal copy number changes, it was difficult to identify key molecule for carcinogenesis because of low resolution. In this study, we sought to identify the key molecule in colorectal cancer pathogenesis by using high-density single nucleotide polymorphism (SNP) array.
(Materials and Methods) We analyzed 35 colorectal cancer cell lines and 20 primary colorectal cancer for areas of DNA copy number changes and loss of heterozygosity (LOH) through genome-wide analysis of 116, 204 SNPs distributed across all human chromosomes with Affymetrix GeneChip Human Mapping 100 K Set. Copy Number Analyzer for GeneChip (CNAG), our original analysis tool that automates the detection of copy number and regions of LOH, was used for data analysis. The results from the array analysis were verified in additional 22 primary colorectal cancers by genomic quantitative real-time PCR, FISH and LOH analysis.
(Results)Over all, in both cell lines and primary tumors, gain of 8q, 13q, 20p and 20q, loss of 8p and 18q and LOH of 5q, 14q, 17p and 18q were detected with high frequency. Moreover, allelic homozygosity was detected at 5q, 14q, 17p and 18q frequently in both cell lines and primary tumors. In detail, narrow (less than 1 Mbp) 23 and 32 regions of high grade amplification (n ≥ 5, in cell line or n ≥ 4, in primary tumor) were found in at least two cell lines and at least one primary tumor, respectively. The size of minimal common amplified region was 42 Kbp. Eleven regions were newly detected as candidates for amplification. Among them, copy number gain of CCAMP1 (lab name, Chr.7p21.1), CCAMP3 (Chr.7p15.2), CCAMP9 (Chr.17q11.2) were identified in 9/22, 11/22, 8/22 of additional primary tumors, respectively. Twenty-four and 6 regions of homozygous deletion were detected in at least one cell line and in at least one primary tumor, respectively. The size of minimal deletion was 11 Kbp. Seventeen regions were newly detected as candidates for homozygous deletion. Among them, LOH of CCDEL1 (Chr.4q22.1) and CCDEL2 (Chr.6q26) were detected in 9/22 and 10/22 of additional primary tumors, respectively.
(Conclusion) Affymetrix GeneChip Human Mapping 100 K Set and CNAG revealed that allelic homozygosity may be one of the mechanisms of colorectal carciniogenesis and by using these methods, we could detect minimal region of copy number alteration, which had never been able to detect by other methods because of its limitation of resolution. The novel amplified and deleted regions discovered in this study may be significant and useful for early diagnosis and novel therapy based on molecular targeting in colorectal cancer.
98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA