| Co-Investigator(Kenkyū-buntansha) |
SUZUKI Kenji NIIGATA UNIVERSITY, SCHOOL OF MEDICINE, ASSISTANT, 医学部, 助手 (00303123)
OHGOSHI Shogo NIIGATA UNIVERSITY, University Medical Hospital, ASSISTANT, 医学部・附属病院, 助手 (70231199)
ODANI Shohji NIIGATA UNIVERSITY, FACULTY OF SCIENCE, PROFESSOR, 理学部, 教授 (60018702)
鈴木 康史 新潟大学, 医学部, 助手 (50270939)
小方 則夫 新潟大学, 医学部・附属病院, 助手 (70204063)
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| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 1999: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1998: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1997: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Research Abstract |
Hepatocellular carcinoma (HCC) is one of the most prevalent human cancers, and the prevention of intra and extrahepatic metastasis of HCC is still difficult even with the development of several therapeutic modalities. We have investigated serological features indicating a malignant potential of HCC. Although, in general, the serum concentration of alpha-fetoprotein (AFP) has been used for the diagnosis of HCC, we have shown that the relative amount of the Lens culinaris agglutinin (LCA)-reactive species of AFP is significantly greater in HCC than in non-neoplastic liver diseases. In addition, we have elucidated that the higher proportion of (LCA)-reactive species of AFP is significantly associated with the tumor invasiveness. The molecular basis of the LCA-reactive species of AFP is the fucosylation at the innermost N-acetylglucosamine residue of the biantennary sugar chain of AFP. Furthermore, it was reported that N-linked β1-6 branched oligosaccharides might contribute directly to th
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e malignant phenotype including metastatic potential of several types of cancers including HCC. It is supposed that these phenotypic changes must be provided by GDP-L-Fuc : N-acethyl-β-D-glucosaminide : β1-6 fucosyltransferase (αFT), which catalyzes the addition of fucose from GDP-fucose through an αl-6 linkage to the reducing end of N-acetylglucosamine residue of N-linked oligosaccharides of glycoproteins, and N-acetylglucosaminyltransferase V (GnT V), which catalyzes β1-6 branching, respectively.
Therefore, in this research term, at first we determined weather the amount of message of αFT and GnT V in each liver tissue were really associated with each enzyme activity in the tissues and the serum. Consequently, quantitative reverse transcription coupled polymerase chain reaction revealed that the quantity of each mRNA in HCC was significantly associated with each enzyme activity in HCC and the corresponding serum. From the point of these observations, next we tried to investigate effects of antisense RNAs against αFT and GnT V. In order to evaluate the effects in vivo, we employed the high-level gene expression system based on tetracycline-responsive element (TRE). Although this system has a great advantage of capability for strict regulation of gene expression in vivo according to the presence or absence of tetracycline, it is relatively difficult to establish stable cell lines. Because it requires transfection of two critical components, the tetracycline-controlled transactivator (TA) and TRE fused to the gene of interest. Furthermore, none of genes for the selection is integrated into the plasmid consisting of TRE and the gene of interest. Finally, three components have to be transformed including a selectable marker. We have finished to clone αFT and GnT V genes in an appropriate construct, and create a stable cell line of TA. In a near future, we will be able to develop double-stable cell lines, and elucidate a biological significance of fucosylation and glucosaminylation at a molecular level on cancer invasiveness. Less
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