ヒトTNF遺伝子を導入したヒト線維芽細胞の抗腫瘍効果

• Project/Area Number: 09877045
• Research Category: Grant-in-Aid for Exploratory Research
• Allocation Type: Single-year Grants
• Research Field: Experimental pathology
• Research Institution: Okayama University
• Principal Investigator: 難波 正義 岡山大学, 医学部, 教授 (80069004)
• Co-Investigator(Kenkyū-buntansha): 井上 裕介 岡山大学, 医学部, 助手 (90304302) ; 辻 俊也 岡山大学, 医学部, 助手 (10304304) ; 近藤 格 岡山大学, 医学部, 助手 (30284061) ; 三原 浩一郎 岡山大学, 医学部, 助手 (50229788)
• Project Period (FY): 1997 – 1998
• Project Status: Completed (Fiscal Year 1998)
• Budget Amount: ¥1,700,000 (Direct Cost: ¥1,700,000); Fiscal Year 1998: ¥600,000 (Direct Cost: ¥600,000); Fiscal Year 1997: ¥1,100,000 (Direct Cost: ¥1,100,000)
• Keywords: TNF / 癌治療 / TNF産生ヒト細胞 / カプセル化 / TNF-α / Human cells / Transfection / Cancer treatment / Radicals / Antioxydant / In Vitro

Research Abstract

コバルト60ガンマー線照射で不死化した脱瘍性をもたないヒト線維芽細胞にTNF-αを導入し,TNF-αを産生する細胞のクローンを取得した.この細胞を培養した培地(Conditioned medium:CM)を,ヒト由来の肺癌(A549),卵巣癌(PA-1),肝癌(HLE,HLF,JHH-7),試験管内で癌化したヒト線維芽細胞(KMST-6/RAS)の培養培地に終濃度10%添加すると,これらの細胞の増殖を45-92%抑制した.この細胞増殖抑制作用はCMの濃度依存性であった.また,この細胞増殖抑制作用は細胞の産生ずるTNF-αであることを,抗TNF-α抗体で証明した.一方,ヒト由来の正常線維芽細胞(KMS-6),子宮癌(HeLa),肝癌(PLC/PLF/5,JI-II-I-l,HuH-6,HepG2)などの細胞の増殖は抑制しなかった.したがって,TNF-α産生細胞は約50%のヒト腫瘍の増殖を抑えることが分かった.
このTNF-α産生細胞を,臨床的に応用できるかどうかを検討するために,この細胞をスフエロイド培養し,そのスフエロイドをアガロース,カルボキシメチール-レルローズでカプセル化した.このカプセル化TNF細胞は培地に1カ月間にわたってTNFを産生した.

Research Products

• [Publications] Fushimi,K.: "Establishment of a human fibroblast cell line producing tumor necrosis factor α (KMST6/TNF) and growth inhibitory effects of its conditioned medium on malignant cells in culture." In Vitro Cell Dev Biol. 34. 463-467 (1998)
• [Publications] Nakamura,K.: "Inhibitory effects of antioxidants on neonatal rat cardiac myocyte hypertrophy induced by tumor necrosis factor-α and angiotensin II." Circulation. 98. 794-799 (1998)
• [Publications] Tsuji T.: "Cyclin E overexpression responsible for growth of human hepatic tumors with p21^<WAF1/CIP1/SDI1>" Biochem.Biophys.Res.Commun.242 (2). 317-321 (1998)
• [Publications] Miyazaki,M.: "Transforming growth factor-β1 stimulates or inhibits cell growth via down-or up-regulation of p21/Waf1." Biochem.Biophys.Res.Commun.246. 873-880 (1998)
• [Publications] Tagashira,H.: "Reduced metastatic potential and c-myc overexpression of colon adenocarcinoma cells (Colon 26 line) transfected with nm23-R2/rat nucleoside diphosphate kinase α isoform." Int.J.Mol.Med,. 2. 25-68 (1998)
• [Publications] Kondo,T.: "Characteristics of intracellular transferrin produced by human fibroblasts : Its posttranscriptional regulation and association with tubulin." Exp.Cell Res.242. 38-44 (1998)
• [Publications] Namba,M., et al.: "Transformation of normal human fibroblasts into immortalized cells with the mutant p53 gene and X-rays." Int.J.Cancer. 70. 135-140 (1997)
• [Publications] Namba,M., et al.: "The telomere lengthening mechanism in telomerase-negative immortal human cells does not involve the telomerase RNA subunit." Hum.Mol.Genet.6. 921-926 (1997)
• [Publications] Namba,M., et al.: "Screening the p53 status of human cell lines using a yeast functional assay." Mol.Carcinog.19. 243-253 (1997)
• [Publications] Namba,M., et al.: "The p53 gene status and other cellular characteristics of human cell lines maintained in our laboratory." Tiss.Cult.Res.Commun.16. 173-178 (1997)
• [Publications] Namba,M., et al.: "Immortalization of mutant p53-transfected human fibroblasts by treatment with either 4-nitroquinoline 1-oxide or X-rays." In Vitro Cell.Dev.Biol.33・7. 628-632 (1997)
• [Publications] Namba,M., et al.: "Establishment of a human fibroblast cell line producing tumor necrosis factor α (KMST-6/TNF) and growth inhibitory effects of its conditioned medium on malignant cells in culture." In Vitro 1998. (in press).