Molecular cloning of c-met 6.0 Kb abnormal transcript

c-met 6.0 Kb异常转录本的分子克隆

• 批准号: 05670175
• 负责人: YOKOZAKI Hiroshi
• 金额: $ 1.41万
• 依托单位: HIROSHIMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05670175/
• 关键词: Gastric cancer; Oncogene; C-met; Abnormal transcript; Molecular cloning

The expression of 6.0 Kb c-met abnormal transcript was proven to be closely correlated with progression and metastasis of gastric cancer. The aim of this grant aided research termed between 1993 and 1995 was to clone this c-met abnormal transcript and clarify the nature of c-met transcripts in neoplastic and non-neoplastic gastric tissue. The results and estimations of the research are summarized as following :1. The attempt for the molecular cloning of 6.0 Kb c-metabnormal transcript was conducted with (1) cDNA library from gastric cancer cell line KATO-III, (2) high molecular weight pAP3 neo cDNA library from MKN-28 stomach cancer cell line and (3) genomic DNA library from normal human peripheral white blood cell using pmetH probe. Although several positive clones were obtained from each library system, the full length 6.0 Kb abnormal transcript could not be available. The cause of the cloning failure was considered to be due to the large size as well as the rarity of the target transcript. Some highly sensitive like lonk RT-PCR cloning may improve the cloning efficiency.2. Fragments of c-met cDNA were analyzed by RT-PCR method. Gastric cancer tissue as well as corresponding normal gastric mucosa was found to express a splice variant lacking nt 2265 to nt 2319 of c-metcDNA preferentially (Yokozaki, H., et al., Oncology Reports, 3,1996 (in press) ).3. Loss of heterozygosity (LOH) on the long arm of the chromosome 7 including c-met locus in human gastric carcinoma tissues was analyzed extensively. Cases with LOH at D7S95 showed significantly worse prognosis than those without the LOH.This finding suggested that D7S95 locus on 7q may contain a candidate suppressor gene for the progression of gastric carcinoma.

6.0Kb c-met异常转录本的表达与胃癌的进展和转移密切相关。这项在1993年至1995年期间资助的研究的目的是克隆这种c-met异常转录本，并阐明肿瘤性和非肿瘤性胃组织中c-met转录本的性质。本研究的主要结果与估计如下：1.用pmetH探针分别从人胃癌细胞株KATO-Ⅲ、人胃癌细胞株MKN-28和正常人外周血白色细胞基因组DNA文库中克隆了6.0Kb的c-metabormal转录本。虽然从每个文库系统中获得了几个阳性克隆，但不能获得全长6.0Kb的异常转录本。克隆失败的原因被认为是由于目标转录本的大尺寸以及稀有性。一些高灵敏度的克隆如lonk RT-PCR克隆可提高克隆效率.用RT-PCR方法分析c-met cDNA片段。发现胃癌组织以及相应的正常胃粘膜优先表达缺乏c-metcDNA的nt 2265至nt 2319的剪接变体（Yokozaki，H.，例如，肿瘤学报告，1996年3月（印刷中））。3.本研究对胃癌组织中包括c-met基因在内的7号染色体长臂上的杂合性缺失（洛）进行了广泛的分析。D 7 S95位点洛缺失者的预后明显差于无洛缺失者，提示D 7 S95位点可能是胃癌发生发展的一个候选抑制基因。

项目成果

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Kuniyasu, H.: "A berrant expression of c-met mRNA in human gastric carcinomas" International Journal of Cancer. 55. 72-75 (1993)

Kuniyasu, H.：“c-met mRNA 在人类胃癌中的异常表达”国际癌症杂志。

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