To identify candidate genes that could be used as diagnostic and therapeutic targets for hepatocellular carcinoma (HCC), we searched for the genes that are overexpressed in HCC by combining representational difference analysis and microarray. Genes such as glypican-3 (GPC3), insulin-like growth factor 2, long-chain fatty-acid-coenzyme A ligase 4, farnesyl diphosphate synthase were frequently identified in our screening. Northern blot analysis with these four genes confirmed their overexpression in HCC. Among them we found that GPC3 transcript is upregulated in six out of seven cases of HCC. Immunoblot and immunohistochemical staining using polyclonal anti-GPC3 antibodies further confirmed that GPC3 protein is indeed increased in HCC tumor samples. We also found that GPC3 is secreted into culture media from cell lines derived from HCC. We conclude that GPC3 is a good molecular marker for HCC. (Cancer Sci 2003; 94: 259-262) lypican-3 (GPC3) is a member of the glypican family of heparan-sulfate proteoglycans, which are linked to the cell surface through a glycosylphosphatidylinositol anchor.1) GPC3 loss-of-function mutation in human causes type 1 Simpson-Golabi-Behmel syndrome (SGBS1), an X-linked condition characterized by pre-and postnatal overgrowth.2) GPC3 knockout mice indeed exhibited several phenotypic features of SGBS1. [3][4][5] These findings together with cell line-specific promotion of apoptosis by OCI-5/GPC3 6) suggest that GPC3 plays a negative role in cell proliferation and an apoptosis-inducing role in specific tissues.Consistent with the above idea, GPC3 expression is frequently silenced by promoter methylation in ovarian cancer cell lines, 7) rat mesothelioma cell lines and human primary tumors, 8) and breast cancer cell lines. 9) In addition, ectopic expression of GPC3 inhibited growth in some of the above cell lines, suggesting a tumor-suppressive role of GPC3. In contrast, GPC3 is known to be overexpressed in hepatocellular carcinoma, 10,11) neuroblastoma and Wilms' tumor cells.12) The role of GPC3 in these tumors is not known. It is also not known whether GPC3 protein is indeed increased in these tumors.Hepatocellular carcinoma (HCC) is one of the most common tumors worldwide and is one of the leading causes of death among cancer patients in Korea. Identification of genes that are overexpressed in HCC not only helps our understanding of tumorigenesis, but also helps to develop diagnostic and therapeutic targets. In this study, we combined representational difference analysis (RDA) 13) and microarray 14) to identify genes that are frequently overexpressed in HCC tumor samples. Since GPC3 was the most frequently obtained gene in our screening, we further evaluated it as a tumor marker for HCC. Materials and MethodsTumor samples and cell lines. HCC tumor tissues and corresponding normal liver tissues were obtained from patients (Table 1) undergoing surgery in Kyungpook National University Hospital (Daegu, Korea) with the approval of the human research review committee and the patients' consent. C...

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Urolithiasis and calcium oxalate crystal deposition diseases are still significant medical problems. In the course of nephrocalcin cDNA cloning, we have identified FKBP-12 as an inhibitory molecule of calcium oxalate crystal growth. lambdagt 11 cDNA libraries were constructed from renal carcinoma tissues and screened for nephrocalcin cDNA clones using anti-nephrocalcin antibody as a probe. Clones expressing recombinant proteins, which appeared to be antigenically cross-reactive to nephrocalcin, were isolated and their DNA sequences and inhibitory activities on the calcium oxalate crystal growth were determined. One of the clone lambda gt 11 #31-1 had a partial fragment (80 bp) of FKBP-12 cDNA as an insert. Therefore, a full-length FKBP-12 cDNA was PCR-cloned from the lambda gt 11 renal carcinoma cDNA library and was subcloned into an expression vector. The resultant recombinant FKBP-12 exhibited an inhibitory activity on the calcium oxalate crystal growth (Kd=10(-7) M). Physiological effect of the extracellular FKBP-12 was investigated in terms of macrophage activation and proinflammatory cytokine gene induction. Extracellular FKBP-12 failed to activate macrophages even at high concentrations. FKBP-12 seems an anti-stone molecule for the oxalate crystal deposition disease and recurrent stone diseases.

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