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In order to clarify early molecular events involved in liver carcinogenesis, we analyzed 53 liver-cirrhosis nodules (LCNs) from five patients and 13 micro-hepatocellular carcinoma (HCC) nodules from one patient and looked for alterations of microsatellites in genomic DNA after carefully preparing the tissue samples by laser-capture microdissection (LCM). Allelotyping was done with 20 markers corresponding to anonymous microsatellites and 13 corresponding to tumor suppressor genes (TSGs) that had shown significant alterations in HCCs. We detected both loss of heterozygosity (LOH) and microsatellite shifts (MS). Overall, 24 of 53 (47%) of LCNs showed LOH with any of the informative markers used in the study, reflecting that proportion of LCNs with clonal growth. The fractional allelic loss (FAL) index, an indication of total genomic complexity, was not significantly different between LCN and micro-HCC nodules, but their profiles of alteration were different. These profiles were classified into three groups: (1) LCN profile—allelic loss at chromosomal arms 1q and 14q, TBP and BRCA1; (2) HCC profile—LOH at 4q, 6q, 7q, 17p, NF1, IGFIIr and p53 in micro-HCC nodules; these changes in early lesions were identical to those seen in mature HCCs; (3) Common profile—LOH at NF1 and 6q, including IGFIIr, common to both LCN and HCC. No LCN showed LOH at p53 and Rb, loci that are generally altered in HCCs. However, 12 intra-tumoral nodules examined had lost p53 in all informative cases, although the loss of Rb was a late event. These results suggest that early genomic profiles confined to LCNs, and additional profiles that can be observed when liver tissue undergoes malignant transformation, support a model of multi-step development of HCC. [Copyright &y& Elsevier]

Insulin-like growth factor binding protein-3 (IGFBP-3) is postulated to be a mediator of growth suppression signals. Reduced expression of the IGFBP-3 was observed in nine out of 12 human hepatocellular carcinomas (HCC) (75%). Promoter hypermethylation of the IGFBP-3 was detected in four out of 12 HCCs (33%) although mutations were not identified. The expression of IGFBP-3 was restored by the demethylating agent 5-aza-2′-deoxycytidine in HCC cell line with promoter hypermethylation (HepG2). As IGFBP-3 functions like a tumor suppressor gene, it may be used as a therapeutic target for HCC. [Copyright &y& Elsevier]