Different Pattern of Chromosomal AlÃ-eleLoss in Multiple Hepatocellular Carcinomas as Evidence of Their Multifocal Origin1


One of the most problematic aspects of surgery for hepatocellular carcinoma (HCC) is the frequent development of multiple tumors. Deter mination of the origin of multiple tumors, i.e., multifocal or metastatic, is important for predicting the clinical course of the disease after surgery. In order to clarify the origin of multiple tumors of HCC genetically, we examined patterns of loss of heterozygosity (LOH) on chromosome 16 for ON A isolated from 43 HCCs resected from 19 patients by analysis of restriction fragment length polymorphism. The cases were classified macroand microscopically into 3 groups: multifocal origin; metastatic origin; and undetermined. Classification based on morphological features was shown to be well correlated with patterns of LOH in multiple tumors of HCC. Different patterns of LOH on chromosome 16 were detected in 8 of 11 patients with tumors of morphologically multifocal origin, whereas they were detected in none of 5 patients with tumors of morphologically metastatic origin. Among five patients with tumors of morphologically undetermined origin, a difference of LOH pattern among the tumors was detected in two, whereas in the other three, the pattern was identical between the tumors. A different pattern of LOH among HCCs arising in situ showed that they were composed of different clones, strongly sug gesting their independent clonal origin and multifocal development. These results show that not only appropriate morphological observation but also examination of the LOH pattern on a particular chromosome is useful in diagnosis of multifocal HCC. INTRODUCTION HCC4 is one of the most frequent malignancies in Africa and Asia (1). In Japan, about 20,000 persons die of HCC every year (2). Recent progress in diagnostic imaging techniques, e.g., ultrasonography, computed tomography, magnetic resonance imaging, and selective angiography (3) and also surgical meth ods (4) has made surgical therapy for HCCs feasible at a relatively early stage in an increasing number of patients. How ever, the frequent multiple occurrence and high recurrence rate of HCCs in patients with chronic viral hepatitis and/or liver cirrhosis are major obstacles which hinder the curability of the disease (5, 6). It has usually been difficult to determine whether multiple and/or recurrent HCCs occur multifocally or originate from a single tumor through metastasis. The cell origin of multiple HCCs has been classified conventionally as multifocal or met astatic based mainly on macroand microscopic observations. However, it seems necessary to confirm the propriety of mor phological criteria by carrying out analyses at the molecular level. If it were possible to clarify more definitely the origin of such multiple tumors, evaluation of clinical stage and postop erative outcome would be made much more accurate. Received 10/14/91; accepted 12/30/91. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1Supported in part by a (¡rant-in-Aid for the Comprehensive 10-Year Strategy for Cancer Control from the Ministry of Health and Welfare of Japan. 2Awardee of a Research Resident Fellowship from the Foundation for Pro motion of Cancer Research. 3To whom requests for reprints should be addressed. 4The abbreviations used are: HCC, hepatocellular carcinoma; LOH, loss of heterozygosity; RFLP, restriction fragment length polymorphism; Several previous studies using DNA hybridization techniques have examined the origin of multiple HCCs by analysis of the pattern of hepatitis B virus DNA integration (7-9). However, this method was applicable for only a portion of cases, because the majority of HCCs in Japan are not associated with hepatitis B virus infection (10). We have shown that LOH on chromo some 16 occurs frequently (52%) in primary tumors of human HCC (11,12) and that the region showing the LOH is variable among tumors (12). Accordingly, we considered that examina tion of LOH on chromosome 16 might help to reveal the clonality and origin of multiple HCCs. The purpose of the present study was to determine whether examination of the LOH pattern in multiple HCCs would be useful for differen tiating HCCs arising multifocally from those occurring through metastasis. First, we classified multiple HCCs into three groups, i.e., multifocal origin, metastatic origin, and undetermined, according to the criteria based on morphological findings. Then, we examined the association of the LOH pattern on chromo some 16 and morphological classification of origin of multiple HCCs. MATERIALS AND METHODS Patients. The materials used were tissue samples from 43 tumors and nontumorous areas of liver, for which adequate amount were available, from 19 patients who had received surgical treatment for multiple HCCs, occurring synchronously or metachronously, at the National Cancer Center Hospital, Tokyo, Japan, between January 1988 and December 1989. Among these patients, 17 were Japanese, one was Korean, and one was Chinese. In 13 cases, multiple (2 or 3) HCCs were resected on the same day; in 4 cases, one tumor had been resected initially, and then another liver tumor or extrahepatic metastatic tumor had been removed later; in 2 cases, 2 tumors were resected initially, and another tumor was resected later. In all cases, the histolÃ3gica! findings for the noncancerous liver were chronic active hepatitis, prccirrhosis, or liver cirrhosis. Morphological Criteria for Multifocal HCC. The cases of multiple HCC were classified into 3 groups: multifocal; metastatic; and undeter mined. Multiple HCCs in the liver were diagnosed as metastatic in origin from a single tumor when they were components of portal vein tumor thrombi, tumors apparently growing in contiguity with such thrombi, or when they were distributed as multiple small satellite nodules surrounding a larger main tumor (13). Other multiple HCCs in the liver were considered to be of potentially multifocal origin and were further divided into a group in which a multifocal origin was highly likely or a group in which multifocal or metastatic origin could not be determined, using the following morpho logical criteria, (a) Multiple early HCCs, or concurrent early HCC and advanced HCC, were considered to be of multifocal origin, since no metastasis of early HCC has yet been detected (13, 14). Early HCC denotes a small tumor (usually 2.0 cm or less in diameter) composed solely of very well differentiated HCC, or HCC of Edmondson-Steiner grade I (13-15) grossly conserving the preexisting structure of cirrhosis or chronic hepatitis within the tumor (13, 14). (b) When a peripheral area of very well differentiated HCC, or Grade I, was observed around an area of HCC showing poorer differentiation in all multiple HCCs or in a smaller one, the tumors were considered to be primary since the presence of such an area indicates the emergence of more aggressive HCC within early HCC (16). (c) Presence of a significant difference in 1504 on April 20, 2017. © 1992 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from MULTIFOCAL HEPATOCELLULARCARCINOMA Table 1 Clinical and pathological profiles of multiple HCC cases Case Size (cm) HistolÃ3gica! grade" Peripheral Grade I area Case Size (cm) HistolÃ3gica! grade Peripheral Grade I area

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@inproceedings{Tsuda2006DifferentPO, title={Different Pattern of Chromosomal AlÃ-eleLoss in Multiple Hepatocellular Carcinomas as Evidence of Their Multifocal Origin1}, author={Hitoshi Tsuda and Tatsuya Oda and Michiie Sakamoto and Setsuo HirohashP}, year={2006} }