Assessment of genetic changes in hepatocellular carcinoma by comparative genomic hybridization analysis: relationship to disease stage, tumor size, and cirrhosis

Abstract

Hepatocellular carcinoma (HCC) is a common and highly malignant tumor that is prevalent in Southeast Asia. Although the etiological factors associated are now well recognized, the interactions between individual factors and the molecular mechanisms by which they lead to cancer remain unclear. Cytogenetic analysis on HCC has been limited because of poor hepatocyte growth in vitro. The recently developed technique of comparative genomic hybridization (CGH), however, permits screening of the entire genome without the need of cell culture. CGH was applied to the study of genomic aberrations in 67 surgically resected samples of HCC, 3 of adenomatous hyperplasia (AH), and 12 of nontumorous cirrhotic liver surrounding the tumors. All samples were from patients of a racially and etiologically homogeneous population in Southern China, where chronic hepatitis B virus infection is the main etiological factor. CGH analysis of the HCC samples revealed frequent copy number gain of 1q (48/67 cases, 72%), 8q (32/67 cases, 48%), 17q (20/67 cases, 30%), and 20q (25/67 cases, 37%) and common losses on 4q (29/67 cases, 43%), 8p (25/67 cases, 37%), 13q (25/67 cases, 37%), and 16q (20/67 cases, 30%). Our finding of a high incidence of 1q gain strongly suggested this aberration was associated with the development of HCC. Genomic abnormalities were detected in 1 of the 3 AH specimens but absent in all 12 cirrhotic tissues surrounding the tumor. Clinical staging classified 3/67 HCC cases as T1, 53 cases as T2, and 11 cases as T3. No significant difference in the pattern of genomic imbalances was detected between stages T2 and T3. A significant copy number loss of 4q11-q23 was, however, identified in those tumors larger than 3 cm in diameter. Of particular interest was the identification of 8q copy number gain in all 12 cases of HCC that arose in a noncirrhotic liver, compared with only 20/55 cases in HCC arising in a cirrhotic liver. We suggest that 8q over-representation is likely associated with a growth advantage and proliferative stimulation that have encouraged malignant changes in the noncirrhotic human liver.

Figure 1.

CGH aberrations detected in case H2. A CGH image of hybridized chromosomes is shown with the corresponding fluorescence ratio profile plotted alongside the chromosome ideogram. Green regions represent gains, whereas red highlights the losses. The mean ratio profile of 12 analyzed chromosomes or more (n ≤ 12, pink line) is depicted with the 95% confidence interval (gold lines). Red and green lines represent thresholds for chromosomal losses (0.75) and gains (1.25). An amplicon on 1q21-22 and high-level gain of 8q can be readily identified by the intense green signals. The corresponding ratio profiles exceeded 1.5 (see Materials and Methods). The CGH profile of this specimen suggests high-level copy gains of 1q21-22 and 8q, low-level gains of 1q, 9q32-qter, 10p, and 16pter-p11.2, and losses of 2q32-qter, 4q28-qter, 7q32-qter, 8p, 10q, 11q, 12p, 13q12-q31, 14q, and 16q.

Figure 2.

Summary of gains and losses of DNA sequences identified by CGH in 47 HCCs >3 cm in diameter. Gains are shown on the right side of the chromosome ideogram and losses on the left. High-level gains are shown as thick lines. Each vertical line represents the affected chromosomal region seen in a single tumor specimen.