テロメラーゼ機能・テロメア維持機構の解明とその臨床応用

• Project/Area Number: 08265104
• Research Category: Grant-in-Aid for Scientific Research on Priority Areas (A)
• Allocation Type: Single-year Grants
• Research Institution: Hiroshima University
• Principal Investigator: 井出 利憲 広島大学, 医学部, 教授 (60012746)
• Co-Investigator(Kenkyū-buntansha): 清宮 啓之 癌研究所, 化学療法センター, 研究員 (50280623) ; 檜山 佳子 (檜山 桂子) 広島大学, 医学部, 助手 (60253069) ; 柴沼 質子 昭和大学, 薬学部, 助教授 (60245876) ; 清水 素行 東京歯科大学, 歯学部, 助教授 (80130756) ; 石川 冬木 東京工業大学, 生命理工学部, 教授 (30184493)
• Project Period (FY): 1999
• Project Status: Completed (Fiscal Year 1999)
• Budget Amount: ¥90,500,000 (Direct Cost: ¥90,500,000); Fiscal Year 1999: ¥7,500,000 (Direct Cost: ¥7,500,000); Fiscal Year 1998: ¥25,000,000 (Direct Cost: ¥25,000,000); Fiscal Year 1997: ¥29,000,000 (Direct Cost: ¥29,000,000); Fiscal Year 1996: ¥29,000,000 (Direct Cost: ¥29,000,000)
• Keywords: テロメア維持機構 / 染色体外テロメア断片DNA / テロメラーゼ / 不死化 / 癌診断 / テロメア / hTERT / 癌診断法 / 制癌剤 / TRAP / HPA法 / テロメラーゼ遺伝子 / テロメラーゼ複合体 / 診断マーカー / 癌早期診断 / 細胞不死化

Research Abstract

テロメラーゼ非依存的テロメア維持機構は、テロメア維持機構の理解のために重要であるばかりでなく、ヒト癌の一部に見られることや、テロメラーゼ活性を持つ癌細胞をテロメラーゼ阻害剤で処理した時に耐性細胞として出現するなど、制癌の面でも重要である。本機構を有する細胞は、異常に長いテロメア末端(23kbp以上)を持つことが特長とされてきたが、実は2-4kbpと短く、これが染色体外テロメア断片DNA(ECTR-DNA)と大きな複合体を形成していることがわかった。細胞内では、このような複合体形成によって、DNAポリメラーゼによるテロメア延長が起きるものと推定された。ヒト線維芽細胞は、テロメラーゼ遺伝子導入だけでは不死化できず、不死化にはSV40のT抗原遺伝子の働きも必要であることがわかったが、必要とされる機能はT抗原機能の全部ではなく、T抗原機能のごく一部かあるいはt抗原機能で代行されものと推定するまでに限局された。これに対して、ヒト血管内皮細胞ではテロメラーゼ遺伝子導入だけで不死化する可能性が有ることがわかった。テロメア・テロメラーゼの高感度・定量的・簡便な測定法を改良し、肝組織において前癌組織の半数に、慢性肝疾患を含む非癌部より明らかに高い活性を認め、患者に対する対処への指針の一つになると考えられた。肝組織のテロメアは、年齢とともに、年1回程度の細胞分裂に相当する短縮を示した。慢性肝炎、肝硬変では年齢によらずテロメアは短く、中には分裂限界に近いサイズを示すケースも見られることなどが、初めて示された。このように、本研究の目標としたすべてについて大きな進展があり、テロメア・テロメラーゼと細胞の分裂寿命・不死化、癌の発生・進展に関する基礎的研究と、診断・治療に結び付ける応用的研究の両面にわたって、大きく寄与することができた。

Research Products

• [Publications] Tahara,H., et al: "Immuno-histochemical detection of human telomerase catalytic component, hTERT, in human colorectal tumor and non-tumor tissue sections"Oncogene. 18. 1561-1567 (1999)
• [Publications] Kitamoto,M. and Ide,T: "Telomerase activity in precancerous hepatic nodules"Cancer. 85. 245-246 (1999)
• [Publications] Sugimoto,M, et al: "A new aspect of Epstein Barr virus-transformed human B-lymphoblastoid cell lines : senescence and immortalization"Mech.Aging.Dev.. 107. 51-60 (1999)
• [Publications] Nakamura,A et al: "Four novel mutations of the Fanconi anemia group A gene(FAA) in Japanese patients"J.Hum.Genet. 44. 48-51 (1999)
• [Publications] Kakuo,S. et al: "Human is a unique species among primates in terms of telomere length"Biochem.Biophys.Res.Commun. 263. 308-314 (1999)
• [Publications] Yasui,W. et al: "Immunohistochemical detection of human telomerase reverse transcriptase in normal mucosa and precancerous lesions of the stomach"Jpn.J.Cancer Res.. 90. 589-595 (1999)
• [Publications] Harada,K. et al: "Telomerase activity in central nervous system malignant limphoma"Cancer. 86. 1050-1055 (1999)
• [Publications] Nakamura,Y. et al: "Simple, rapid, sensitive and quantitative detection of telomere repeats in cell lysate by hybridization protection assay(HPA)"Clin.Chem.. 45. 1718-1724 (1999)
• [Publications] Sugimoto,M. et al: "Wrong use of "immortalization" for B-lympho-blastoid cell lines transformed by Epstein-Barr virus"J.Virol. 73. 9690-9691 (1999)
• [Publications] Ishii,Y. et al: "Telomerase activity in hybrids between telomerase-negative and telomerase-positive immortal human cells is repressed in the defferent complementation groups but not in the same complementation group of immortality"Mech.Aging Dev.. 110. 175-193 (1999)
• [Publications] Nakamura,Y. et al: "Quantitative re-evaluation of telomerase activity in cancerous and noncancerous gastrointestinal tissues"Mol.Carcinog.. 26. 312-320 (1999)
• [Publications] Okui,M. et al: "APG-2 protein, a member of the heat shock protein 110 family, is expressed high in various tissues during the rat development, but exceptionally low in adult heart"Neurochem.Internatl.. (in press). (2000)
• [Publications] Okui,M. et al: "MNB/DYRK gene cloned from the Down syndrome critical region is expressed high in rat brain and heart during development"Genomics. (in press). (2000)
• [Publications] Takahashi,S. et al: "Expression of telomerase component genes in hepatocellular carcinomas"Eur.J.Cancer. (in press). (2000)
• [Publications] Takaishi,H. et al: "Precancerous hepatic nodules had significant levels of telomerase activity by sensitive quantitation using hybridization protection assay"Cancer. (in press). (2000)
• [Publications] Aikata,H. et al: "Telomere reduction in human liver tissues with age and chronic inflammation"Exp.Cell.Res.. (in press). (2000)
• [Publications] Nakayama,J.: "Telomerase activation by the catalytic coMponent gene TRT in human primary hbroblasts and hepatocellular carcinomas." Nature Genetics. 181. 65-68 (1998)
• [Publications] Yamada,M.: "Y'-Help 1,a DNA helrcase encoded by the yeast subtelomere Y'element in induced in survivors defective for telomerase" J.Biol.Chem.273. 203-206 (1998)
• [Publications] Tanaka,H.: "Evidence for a putative telomerase repressor gene in the 3p14,2-p21 region." Genes Chromosomes Cancer. 23. 123-133 (1998)
• [Publications] Hiyama,K.: "Telomerase activity as a novel marker of lung cancer and immune-associated lung diseases." Int.J.Mol.Med. 1. 545-549 (1998)
• [Publications] Tahara,H: "Immo-histochemical detecter of human telomerase catalytic component,hTERT a human colorectal tumor and non-tumor tissuesection." Oncogene. 18 (in press). (1999)
• [Publications] Nishiya,N.: "Hic-5 a paxillin homologue,lnets to the protein tyrosine phosphatase PEST (PTD-PEST) trough its LIM3/domainan." J.Biol.Chem. (in press). (1999)
• [Publications] Katoh, M.: "Telomerase-negative immortal cells carry a repressor-function for telomerase activity." Mol.Carcinog.(in press).
• [Publications] Nakayama, J.: "Telomerase activation by the catalytic component gene TRT in human primary fibroblasts and hepatocellular carcinomas." Nature Genetics. 18・1. 65-68 (1998)
• [Publications] Murakami, J.: "Telomerase activity in ovarian tumors." Cancer. 80. 1085-1092 (1997)
• [Publications] Nakayama, J.: "TLP1:A gene encoding a protein component of mammalian telomerase is a novel member of WD-repeats family." Cell. 88. 875-884 (1997)
• [Publications] Shibanuma, M.: "Induction of senescence-like phenotypes by the forced expression of hic-5 encoding a novel LIM motif protein in immortalized human fibroblasts." Mol.Cell.Biol.17. 1224-1235 (1997)
• [Publications] Hiyama, E.: "Telomerase activity is detected in pancreatic cancer but not in benign tumors." Cancer Research. 57. 326-331 (1997)
• [Publications] Yasumoto, S., et al.: "Telomerase activity in normal human epthelial cells." Oncogene. 13・4. 433-439 (1996)
• [Publications] Tatematsu, K., et al.: "A novel quantitative "Stretch PCR assay" that detects a dramatic increase in telomerase activity during the progression of myeloid leukemias." Oncogene. 13・11. 2265-2274 (1996)
• [Publications] Hiyama, E., et al.: "Telomerase activity is detected in pancreatic cancer but not in benign tumors." Cancer Research. 57・2. 326-331 (1997)
• [Publications] Shibanuma, M., et al.: "Introduction of senescence-like phenotypes by the forced expression of hic-5 encoding a novel LIM motif protein in immortalized human fibroblasts." Mol. Cell. Biol.17・3. 1224-1235 (1997)
• [Publications] Nakashio, R., et al.: "Significance of telomerase activity in the diagnosis of small differentiated hepatocellular carcinomas." Int. J. Cancer. (in press). (1997)
• [Publications] Shimizu, M., et al.: "Genetic regulation of telomerase in a multiple pathways model." Human Cell. (in press). (1997)
• [Publications] 井出利憲,他: "DNAの複製・修復と発癌" 羊土社, 172 (1996)