Biallelic deletion and loss of expression of genes at FRA2G region in tumor derived cell lines.

Common fragile sites (CFSs) are regions of chromosome instability, that extend for hundreds or thousands of kilobases, at which gaps or breaks are observed when cells are exposed to appropriate culture conditions and particularly to replication stress (Sutherland et al., 1998). CFSs are expressed, with different frequency, on chromosomes of all analysed individuals and therefore have to be considered a constitutive feature of normal chromosomes. Many evidences suggest that CFSs are causally related to cancer. In fact these sites are localised in chromosome bands recurrently involved in rearrangements in cancer cells (Richards, 2001), are sites of preferential viral integration (e.g. Wilke et al., 1996), and may also be involved in amplification events as the breakage points in breakage-fusion-bridge cycles (Coquelle et al., 1997; Ciullo et al., 2002; Hellman et al., 2002). Recently, authors showed that miRNA genes are frequently located at fragile sites as well as in cancer-associated genomic regions (Colin et al., 2004). Moreover more conclusive evidences supporting the relation between CFSs and tumourigenesis derive from cloning and characterization of some CFSs. Ten CFSs have been molecularly characterised, FRA2G (Limongi et al., 2003), FRA3B (Wilke et al., 1996),FRA6E (Denison et al., 2003), FRA6F (Morelli et al., 2002), FRA7G (Huang et al., 1998), FRA7H (Mishmar et al., 1998), FRA7I (Ciullo et al., 2002), FRA9E (Callahan et al., 2003), FRA16D (Mangelsdorf et al., 2000; Paige et al., 2000; Ried et al., 2000), FRAXB (Artl et al., 2002). Many of them have been associated with cancer-specific rearrangements, frequently deletions, so as with gene inactivation involving known or putative tumor-suppressor (TS) genes mapping in the fragile regions. FRA2G, an aphidicolin- and DAPI-induced common fragile site located at 2q31 region, was characterized by us using a FISH-based approach to identify a BAC contig of over 1 Mb that spans the fragile region (Limongi et al., 2003). Recurrent deletions, that may suggest the presence of TS genes, are observed at this region in some neoplasms, mostly leukemias and lymphomas (Mitelman et al., 2004). According to the human genome sequence public data base (http://www.ncbi.nih.gov/mapview/), to date twelve genes have been localized within the FRA2G region. In this work, nine of these genes have been investigated for biallelic deletions and loss of expression (LOE) in a panel of nineteen cancer cell lines, eight of which derived from leukemias and lymphomas.