大腸癌に対する新規腫瘍抗原遺伝子

结直肠癌新肿瘤抗原基因

• 批准号: 15790727
• 负责人: 落合 大樹
• 金额: $ 1.98万
• 依托单位: Keio University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Young Scientists (B)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-15790727/
• 关键词: SEREX; 腫瘍マーカー; 大腸癌

【方法】癌抗原を同定する方法として,SEREX (serological identification of antigens by recombinant expression cloning)法をもちいて新規のCT antigenを同定し,その大腸癌における発現を検討した.【方法】glioma細胞株U87MGおよびT98GよりmRNAを精製し,逆転写により合成したcDNAをλ Zap Expressファージベクターに組み込んでcDNA libraryを作製した.ファージより誘導したrecombinant蛋白はニトロセルロース膜に転写後,glioma患者血清と反応させスクリーニングした.陽性クローンを単離後,ファージに組み込まれたcDNAインサートをPCRで増幅,塩基配列を決定しDNAデータベースを検索した.また,単離された抗原の遺伝子発現を正常組織,各種癌細胞株および大腸癌臨床検体をもちいたRT-PCR法により解析した.【結果】52個の陽性クローンを単離,遺伝子データーベースで解析し,新規癌抗原KU-GB-1を同定した.KU-GB-1 mRNAは正常組織では精巣のみに発現していた.KU-GB-1 mRNAは癌細胞株(glioma,肺癌,食道癌,膵癌,膀胱癌,前立腺癌,胃癌,大腸癌)に広く発現を認めた.Northern Blot解析でも同様の結果が得られた.大腸癌臨床検体を用いたRT-PCR法では癌部にKU-GB-1 mRNAの強発現を認めた.【結語】新規CT antigen KU-GB-1は大腸癌に広く発現を認めた.今後更に遺伝子および蛋白レベルの発現解析を進め,新たな大腸癌診断法への応用を検討していく.

[Methods] SEREX (serological identification of antigens by recombinant expression cloning) was used to identify the new CT antigen and detect the occurrence of colorectal cancer. Methods Glioma cell line U87MG and T98G mRNA was purified and cDNA library was prepared. The recombinant protein was induced by the enzyme and the serum of glioma patients was detected by ELISA. The DNA sequence was determined by PCR amplification. In addition, RT-PCR was used to analyze the expression of antigen in normal tissues, various cancer cell lines and clinical specimens of colorectal cancer. The results showed that 52 positive genes were isolated and analyzed, and the new cancer antigen KU-GB-1 was identified. KU-GB-1 mRNA was detected in normal tissues. KU-GB-1 mRNA was detected in cancer cell lines (glioma, lung cancer, esophageal cancer, bladder cancer, prostate cancer, gastric cancer, colorectal cancer). Northern blot analysis showed the same results. Expression of KU-GB-1 mRNA in Colorectal Cancer by RT-PCR [Conclusion] New CT antigen KU-GB-1 anti-colorectal cancer detection and detection. In the future, the analysis of gene expression and protein expression will be further developed, and the application of new diagnostic methods for colorectal cancer will be discussed.