Detection and mapping of amplified DNA sequences in breast cancer by comparative genomic hybridization.

Abstract

Comparative genomic hybridization was applied to 5 breast cancer cell lHes and 33 primary tumors to discover and map regions of the genome with increased DNAsequence copy-number. Two-thirds of primary tumors and almost all cell lines showed increased DNA-sequence copynumber affecting a total of 26 chromosomal subregions. Most of these loci were distinct from those of currently known amplified genes in breast cancer, with sequences originating from 17q22-q24 and 20q13 showing the highest frequency of amplification. The results indicate that these chromosomal regions may contain previously unknown genes whose increased expression contributes to breast cancer progression.Chromosomal regions with increased copy-number often spanned tens of Mb, suggesting involvement of more than one gene in each region.Increased expression of specific genes plays an important role in the pathogenesis of solid tumors (1-3). Gene amplification, characterized by distinct cytogenetic structures, such as homogeneously stained regions, double-minute chromosomes (1-7), is commonly found in tumor cells and is considered an important mechanism by which tumor cells gain increased levels of expression of critical genes. Increased copy-numbers also occur as a result of extensive chromosomal rearrangements, such as duplications, isochromosomes, extra marker chromosomes, and acentric chromosomal fragments that may affect the gene dosage of numerous genes simultaneously. In breast cancer, cytogenetic evidence of increased DNA-sequence copy-number is common (4-7). For example, homogeneously stained regions have been found in 60% of primary breast carcinomas (7). Although genetic analysis has found amplification of oncogenes, such as ERBB2 (17q12), MYC (8q24), PRADII CYCLIN D (11q13), FLG (8p12), BEK (10q24), and IGFR-1/FES (15q24-q25) (8-12), in most cases these do not explain the presence of large homogeneously stained regions (13). Thus, amplification of currently unknown genes may often occur in breast cancer.We have recently developed a method, comparative genomic hybridization (CGH), for surveying entire genomes for DNA-sequence copy-number variation (14, 15). In CGH, the relative intensities of tumor DNA (detected using green fluorescence) and normal reference DNA (detected with red fluorescence) after hybridization to normal metaphase chromosomes is used to reveal and map regions of increased DNA-sequence copy number (14-16). These loci are visualized as chromosomal region(s) with predominantly green fluorescence ( Fig. 1) and quantified by digital image analysis as an increased green-to-red fluorescence intensity ratio (Fig. 2). As no specific probes or previous knowledge of aberrations is required, CGH is especially suitable for identification and mapping of previously unknown DNA copy-number changes that may highlight locations of important genes. In the present study, we have used CGH to identify and map increases in DNA-sequence copy number in 15 breast cancer cell lines and 33 uncultured primary breast tumors. MATERIALS AND ME...

The role of the MKK6/p38 MAPK pathway in Wip1-dependent regulation of ErbB2-driven mammary gland tumorigenesis

“…Thus, it would be of interest to know whether a simultaneous It remains to be determined whether there is an oncogene within the 17q23 breast cancer amplicon whose overexpression could drive tumorigenesis in transgenic mice. The 17q23 amplification occurs fairly frequently (B18%) in primary human breast cancers (Kallioniemi et al, 1994;Sinclair et al, 2003) and there are three potential oncogenes, RPS6KB1, TBX-2 and WIP1, which have been identified within this genomic locus (Sinclair et al, 2003). However, overexpression of either WIP1, Tbx2 or both in MMTVtransgenic mice is not sufficient for tumor formation (present study and data not shown).…”

“…This site has been found to be a hot spot for gene amplifications in multiple types of human cancer. The initial observation of regional gain at chromosome 17q22-23 using a comparative genomic hybridization (CGH) technique detected amplification in B18% of primary breast tumors (Kallioniemi et al, 1994), giving its name to the locus as the 17q23 breast cancer amplicon. Three potential oncogenes, RPS6KB1, TBX-2 and WIP1 have been found within this genomic locus at chromosome 17q22-23 (Sinclair et al, 2003).…”

Up-regulation of CIR1/CROC1 expression upon cell immortalization and in tumor-derived human cell lines

“…Inhibition of differentiation would be a prerequisite for the maintenance of proliferative capacity of immortalized cells; (iii) CROC1/UEV-1 maps to chromosome 20q13.2 (Sancho et al, 1998), a region which is moderately ampli®ed in many types of cancer (e.g. Kallioniemi et al, 1994;Muleris et al, 1994;Tanner et al, , 1995Brinkman et al, 1996) and highly ampli®ed in others, e.g. breast cancer, where it correlates with aggressiveness and poor prognosis (Tanner et al, 1995).…”

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“…The relatively low CGH concordances and correlations involving 19q alterations resulted, in part, from using microsatellite markers and FISH probes that map to 19q13.3, a region in human gliomas that often contains deletions below the resolution of CGH. Furthermore, CGH has known artifacts at both 1p and 19q (Kallioniemi et al, 1994;Kim et al, 1995;Mohapatra et al, 1995).…”