Comparative Genomic Hybridization Analysis Of Breast Cancer

OBJECTIVE: Breast cancer is one of the most severe malignancies of women. The clinical and molecular epidemiological data indicated that the genetic events played an important role in the carcinogenesis of breast cancer. In this series, we applied comparative genomic hybridization (CGH) to analyze 34 primary invasive ductal breast carcinomas (IDC) to acquire the genetic alterations in Chinese women. METHODS: Tumor DNA and normal reference DNA were isolated by Proteinase K digestion and phenol/chloroform/isoamylalcohol extraction, then tumor DNA was labeled by nick translation with Biotin-16-dUTP, and reference placental DNA, with digoxigenin-11-dUTP. The final size of the labeled fragments was 600～2000bp. Equal amounts of tumor and reference DNA probes were mixed and precipitated with 20μg unlabeled Cot-1 DNA. The probes were dissolved in 5μl of hybridization buffer. The hybridization mixture was applied on slides and hybridized for 60～72 hours at 37℃ in a moist chamber. After hybridization, the tumor DNA probes were detected with avidin-FITC, biotinylated goat-anti-avidin, and a second layer of avidin-FITC. The reference DNA probes were detected with mouse anti-digoxigenin, rabbit anti-mouse antibody-TRITC, and sheep anti-rabbit antibody-TRITC. The chromosomes were counterstained with 4,6-diamidino-2-phenylindole (DAPI). Regions of gain or loss of DNA sequences were observed as changes in the ratio of the intensities of the two fluorochromes along the chromosome target. 0.8 and 1.2 were chosen as thresholds for the identification of DNA copy number decreases (below 0.8) and increases (above 1.2). RESULT: All of the 34 primary invasive ductal carcinomas showed DNA-sequence copy number changes involving one or more regions of the genome. Recurrent genetic alterations include gains of chromosome 1q (59%), 16p (50%), 17q (44%), 8q (38%), 11q (32%), 20q (32%), 1p (24%), 20p (24%), 19q (21%) and 19p(18%), and losses of chromosome 6q (15%), 8p (12%), 18 (12%), 4q (9%), X (9%) and 17p (9%). High-level amplifications were on chromosomes 1q32, 8p, 11q13, 17q, and 20q. The incidence of chromosome DNA losses was lower than previously reported. CONCLUSION: There were nonrandom chromosome DNA gains and losses in breast cancer. The aberrant chromosome sites probably contain candidate oncogenes or anti-oncogenes associated with breast cancer.