Jung et al. BMC Cancer 2012, 12:382 http://www.biomedcentral.com/1471-2407/12/382 adjusting for age, stage, ER, PR, and HER2. SAS version 9.1 (SAS Institute Inc., NC) was used and P-values less than 0.05 were considered significant in all statistical analyses. Results General characteristics of copy number alterations in EBC Genome-wide CNAs of the 48 EBCs (discovery set) were examined individually, as described in the Methods (Figure 1A). The median number of CNAs per each sample was 10 (range 1–21). Frequency plots of CNAs in the discovery set showed that alterations were not randomly distributed, but were clustered in several hot regions across the whole chromosomes (Figures 1A and B). The overall CNA frequency profiles were similar for stage I and II groups, and among the molecular subtypes (Additional file 1: Figure S1). Of all 4,396 CNAs, copy number gains on 16p13.3, 16p12.3, and 17q25, and copy number losses on 16q21, 17p12, 17p13, and 20q11.1 were significantly more frequent in the stage II groups based on unadjusted P values (P < 0.05), but none after multiple comparison correction (Additional file 1: Table S2). On average, 376 Mb (13% of the whole genome) per EBC tissue specimen showed chromosomal alterations. Recurrently altered chromosomal regions in EBC Except for a few entire chromosomal arm changes, the majority of copy number alterations were regional Page 4 of 11 changes, and some of them were observed repeatedly in the discovery set. Chromosomal segments that covered overlapping CNAs appeared in at least 30% of the sam- ples with P < 0.05 and were defined as RARs (RAR-G for gains and RAR-L for losses, respectively). A total of 23 RARs (18 RAR-Gs and 5 RAR-Ls) were defined in the discovery set (Table 2). Figure 2 illustrates the RAR on 17q12 as an example. Of the 23 RARs, 15 RARs detected commonly in both stage I and II groups can be considered earlier events in breast tumorigenesis. Two RARs (RAR-G14 and –G15), which appeared in less than 10% of the stage I samples but in over 40% of the stage II samples, appear to be relatively later events (Table 2). There were six RARs observed in over 50% of the discovery samples: RAR-G2 (1q21.2-q21.3), RAR-G7 (8q24.13), RAR-G8 (8q24.13-21), RAR-G9 (8q24.3), RAR-G10 (8q24.3), and RAR-L1 (8p23.1-p22). All six highly common RARs were earlier events, as described above. Their high prevalence and early occurrence sug- gest that potential driver cancer genes may be included within the segments. Some cancer-related genes are located in these highly common, early appearing RARs: MCL1, CTSK, ARNT, S100A10, PTK2, PTP4A3, and PSCA in the RAR-Gs; and DLC1, PINX1, and GATA4 in the RAR-Ls. Many known or putative cancer-related genes are also located in other RARs. For example, the ERBB2, GRB2, and MMP families, as well as the TPM3, PYGO2, CKS1B, MUC1, CCT3, PRCC, UBE2C, and Figure 1 Genome-wide profiles and frequency plot of chromosomal alterations in 48 breast cancer samples. (A) The genomic alteration profiles of 48 early breast cancers are presented in individual lanes. A total of 24,107 probes are mapped according to the UCSC genome browser (Human NCBI36/hg18) and ordered by chromosomal position from 1pter to Yqter (X-axis). Tumor vs. reference intensity ratios are plotted in different color scales reflecting the extent of genomic gains (red) and losses (green), as indicated on the reference color bar (log2 scale). (B) Frequencies of copy number gains and losses in the 48 breast cancers. The green bars denote the copy number gains and the red bars denote the copy number losses. Boundaries of individual chromosomes are indicated by vertical bars.