**3.3 Fluorescence in situ hybridization (FISH) analysis of pathological archives with BACs**

It is now known that there are extensive somatic changes, including multiple point mutations (Wood, Parsons et al. 2007; Velculescu 2008), copy number alterations (Weir, Woo et al. 2007; Kubo, Kuroda et al. 2009), and further complex rearrangements (Campbell, Stephens et al. 2008) tumors. But when and where these genetic changes occur during human cancer development remains unclear. Human archival tissue blocks contain

by using overlapping clones (Ishkanian, Malloff et al. 2004; Lockwood, Chari et al. 2006). These platforms provide sufficiently strong signals to detect single-copy change, and are able to accurately define the boundaries of genomic aberrations, which can possibly be utilized in archival formalin-fixed, paraffin-embedded (FFPE) tissue (Johnson, Hamoudi et

High amounts of high-quality BAC DNA are needed to obtain good array performance (Ylstra, van den Ijssel et al. 2006). BACs DNA yield is generally low when isolated from *E. coli* (Pinkel and Albertson 2005). Because of the low yields of DNA from isolated BAC clones, DNA amplification is required to generate sufficient quantities of adequately pure BAC DNA for the assay. Therefore a tiling path array is costly and highly labor intensive. In addition, as BAC probes are representative of the human genome, they also contain repetitive sequences, which can result in nonspecific hybridization (Tan,

BAC technique has increasing been applied in detecting structural changes in chromosomes, such as copy number aberrations (CANs) and rearrangement. Mosse et al (Mosse, Greshock et al. 2005) generated 4135 BAC clones spanning the human genome at about 1.0 Mb resolution as targets for array-based comparative genomic hybridization (aCGH) experiments (Greshock, Naylor et al. 2004). They measured the relevance of neuroblastoma DNA copy number changes (CNAs) in forty-two human neuroblastoma cell lines. They found that all cell lines exhibited CNAs ranging from 2% to 41% of the genome. Chromosome 17 showed the highest frequency of CANs. The most frequent region of gain with high-level amplification localized to 2p24.22-2p24.3 detected in 81% of cell lines (Mosse, Greshock et al. 2005). Potential oncogenes such as MYCN, NAG and DDX1 were located in these regions. The less frequent region of gain localized to 17q23.2, 17q23.3-17q24.1, 17q24.1-17q24.2, 17q25.2-17q25.3 was detected in about 70% of the cell lines. Potential oncogene BIRC5 was localized in these regions. Although gain of 17q material was common, this low level gain of chromosomal material was rather complicated (Mosse, Greshock et al. 2005). The most frequent hemizygous deletion localized to a 4.0 Mb region at 1p36.23-36.32, was detected in 60% of the cell lines. Potential tumor suppressor genes TP73, CHD5, RPL22 and HKR3 were also localized. A 10.4 Mb region at 11q23.3-11q25 was detected in 36% of the cell lines, and the potential tumor suppressor gene CHEK1 was found there as well (Mosse, Greshock et al. 2005). Overall, the array CGH could be reliable in examining DNA copy number aberrations including single copy gain or loss. Compared to the data with standard techniques, data from array CGH correlates well with known aberrations detected by standard techniques. Therefore, array

**3.2 Measurement of neuroblastoma DNA copy number aberrations (CNAs)** 

CGH can be applied to identify novel regions of genomic imbalance.

**3.3 Fluorescence in situ hybridization (FISH) analysis of pathological archives with** 

It is now known that there are extensive somatic changes, including multiple point mutations (Wood, Parsons et al. 2007; Velculescu 2008), copy number alterations (Weir, Woo et al. 2007; Kubo, Kuroda et al. 2009), and further complex rearrangements (Campbell, Stephens et al. 2008) tumors. But when and where these genetic changes occur during human cancer development remains unclear. Human archival tissue blocks contain

al. 2006; Little, Vuononvirta et al. 2006).

Lambros et al. 2007).

**BACs** 

specimens of human tumors in various stages of development, which are precious in the post-human-genome-sequencing era. Based on their findings and other's work, Sugimura et al (Sugimura, Mori et al. 2010) stated that the intensive application BAC clones as probes for FISH that have exact 'addresses' in the whole genome will become a useful diagnostic tools for pathologists. Thousands of BAC clones are commercially available, and any of them can be used as FISH probes. Sugimura et al tested 100 BAC probes containing different kinase loci in a gastric, colorectal, and lung cancer detection sets (20 cases for each organ) by using tissue microarray (TMA)-FISH technology (Sugimura, Mori et al. 2010). Sugimura et al found that unexpected kinase loci were amplified in a significant proportion of human common solid tumors (Sugimura, Mori et al. 2010). Combinatory chemistry has generated many drugs by targeting kinase genes or their products. Thus, amplification of specific regions on certain kinase genes are amenable to pharmacological intervention which could result in the target specific therapy. Therefore it is reasonable to believe that the FISH-BACs diagnostic system combined with particular kinase probes may provide the practical basis of individual cancer therapy.
