Novel Recurrent Genetic Imbalances in Human Hepatocellular Carcinoma Cell Lines Identified by Comparative Genomic Hybridization

Abstract

To search for recurrent and specific genomic alterations in human hepatocellular carcinoma (HCC), we examined 18 cell lines by comparative genomic hybridization (CGH), a molecular cytogenetic approach that allows positional identification of gains and losses of DNA sequences of the entire tumor genome. We report here a distinct pattern of multiple recurrent DNA copy-number gains and losses that include alterations frequently seen in other neoplasias as well as changes potentially specific for HCC. The most frequent gains were localized on 1p34. Human hepatocellular carcinoma (HCC) is one of the most common cancers worldwide and accounts for 90% of all liver carcinomas. It is also one of the most common visceral malignant tumors and is known to have an unfavorable prognosis. 1,2 Different factors, such as infection with hepatitis B virus (HBV), hepatitis C virus, dietary aflatoxin, alcohol consumption, and exposure to chemical carcinogens have been implicated in the etiology of HCC. 1,2 Both chemical carcinogens and HBV cause DNA damage in liver cells manifested at the chromosome level as deletions, duplications, and translocations. 3 The central role of sequential accumulation of genetic alterations during development of cancer is now firmly established. 4,5 In fact, nonrandom and recurrent chromosome abnormalities have been identified in all human cancers that have been examined in sufficient numbers to allow statistical evaluation. 6 However, in HCC the cytogenetic studies are sporadic and cumulative data, based on analysis of eight adenocarcinomas of the liver, comprise only two recurrent breakpoints on the short arm of chromosome 1. 6 Recently, we reported unbalanced translocations of chromosomes 1 and 3 with the breakpoints nonrandomly involving regions 1p36 and 3p14, respectively, in two HCC cell lines. 7 On the other hand, allelic losses detected in HCC involve multiple regions of the genome, none of which has been assigned primary importance (also reviewed in Fourel 29 and Grisham 30 ).Comparative genomic hybridization (CGH) is a highly specific molecular cytogenetic approach that allows positional identification of gains and losses of DNA sequences in the entire tumor genome. It is based on competitive fluorescence in situ hybridization (FISH) of differentially labeled total genomic DNA from appropriate control tissue and tumor DNA to normal chromosome spreads. 31 The ratio of the fluorescence intensities generated by the two different fluorochromes incorporated into test and control DNA is used to differentiate chromosome regions with a normal genomic content, from those showing over-or underrepresentation of sequences. The spatial resolution of CGH mapping is approximately 10 megabasepair (Mbp) for lowcopy-number gains and losses and close to 2 Mbp for high-copy-number amplifications. [32][33][34][35] This analytic approach has already been applied over a wide range of tumor specimens, cell lines, and archival materials to define chromosomal imbalances. CGH studies of solid tumors, such as prostate c...

Y chromosome loss and other genomic alterations in hepatocellular carcinoma cell lines analyzed by CGH and CGH array

“…HCC cell lines are useful material for study of hepatocarcinogenesis as an in vitro model. The frequent gains of 1q, 7, 8q, 17q, and 20q and the frequent losses of 1p, 4q, 8p, 13q, 16q, 17p, and Y were observed in HCC cell lines [11,15,16]. These results suggest the existence of oncogenes and/or tumor suppressor genes on these chromosomal regions that play an important role in hepatocarcinogenesis.…”

“…In the present study, we investigated 5 Korean HCC cell lines and found that gains of genomic material were common on chromosome 7 and chromosome regions 1q and 8q and losses on 4q, 16q, and the Y chromosome (Tables 1 and 2). Previously reported chromosomal studies on primary HCC tissues and HCC cell lines have suggested that the frequent structural abnormality at 1q is involved in HCC [15,16,26,27]. Increases in copy numbers of the LAMC2, TGFB2, and AKT3 genes at 1q have been reported in 19 HCC tissues by CGH array [28].…”

Detection of Chromosomal Aberrations in Well-Differentiated Hepatocellular Carcinoma by Bright-Field In Situ Hybridization

“…The new technique described in this study employs results obtained by CGH (4,13,14). Even in well-differentiated HCC, numerous recurrent aberrations have been found (2).…”

“…Because of the time and effort required for conventional cytogenetics, this technique is usually not appropriate. The recently developed comparative genomic hybridization (CGH) has revealed promising results (2,4,11,13,14), but it is also of limited value because of the duration and amount of tissue required. By contrast, fluorescence in situ hybridization (FISH) yields results that are evaluable by simple counting of fluorescence signals in conventional biopsies (12).…”

Increased expression of glycosyl‐phosphatidylinositol anchor attachment protein 1 (GPAA1) is associated with gene amplification in hepatocellular carcinoma

“…All these markers are located at chromosomal regions showing little DNA copy number variations in HCC samples. [21][22][23][24] A 21-bp oligomer complementary to the microsatellite CA repeat (5 0 FAM-ACA CAC ACA CAC ACA CAC ACA-TAMRA) was used as the probe for the reference pool. All primer pairs were tested for PCR efficiency individually and demonstrated >90% efficiency using the 10.5 CA repeat probe.…”