Identifications of diagnostic biomarkers specific binding to soluble proteins of adenocarcinoma A 549 cell lines by an autoantibodiomics

通过自身抗体组学鉴定与腺癌 A 549 细胞系可溶性蛋白特异性结合的诊断生物标志物

基本信息

批准号：
17590501
负责人：
NAKANISHI Toyofumi
金额：
$ 2.3万
依托单位：
Osaka Medical College
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

项目摘要

At 2001, Prof. Hanash and his colleagues first reported that the autoantibodies against Annexin-I ＆ II frequently (37-40%) were existed in plasma of lung adenocarcinoma using proteomics-based analysis. Thereafter many researchers have been identified autoantibody in cancer patients' plasma against targeted proteins derived from cancer cells using the same technique. We also applied the novel proteomics-based analysis to identify the candidate biomarkers for the diagnosis of lung adenocarcinoma. The advantages of the identification of autoantibodies against cancer/tumor related antigens were more easy and simple than the gene analytical method. This technique, so-called autoantibodiomics, was expected to apply the screening of cancer diagnostic biomarkers in plasma. In this project we identified candidate new lung cancer diagnosis markers.[Method] We treated a cultured human lung cancer cell strain (A549) in solubilization buffer solution and soluble fractions were applied a sample solution. The soluble proteins were separated with a two-dimensional electrophoresis and then transferred on a PVDF membrane. After blocking the PVDF membrane, plasma (500 times dilution) as a primary antibody was reacted with separated proteins on membrane, detected a positive spots using ECLplus reagent Kit. On the other hand, silver-staining gel was cut-off a positive spots corresponding to the ECL positive spots and analyzed them by a time-of-flight mass spectrometer (UltraFlex) or an ion trap type mass spectrometer ( LCQDECA) and a database searched a provided result and identified target proteins.[Results and conclusion] We found eight kinds of biomarkers, anti α-enlace, chaperonin, peroxiredoxin-6 autoantibodies, in patients' plasma of lung adenocarcinoma. We concluded that these autoantibodies were candidate biomarker to diagnose lung adenocarcinoma.

2001年，汉纳什教授及其同事首先报道说，使用基于蛋白质组学的分析，在肺腺癌的血浆中经常存在针对膜联蛋白I＆II的自身抗体（37-40％）。此后，许多研究人员使用相同的技术鉴定出癌症患者血浆中的自身抗体对癌细胞的靶向蛋白质。我们还应用了基于蛋白质组学的新分析来识别候选生物标志物，以诊断肺腺癌。鉴定针对癌症/肿瘤相关抗原的自身抗体的优势比基因分析方法更容易和简单。该技术是所谓的自身抗体，预计将在血浆中应用癌症诊断生物标志物。在这个项目中，我们确定了候选新的肺癌诊断标记。[方法]我们在溶液缓冲液溶液中处理了培养的人肺癌细胞菌株（A549），并应用了固体级分。用二维电泳分离固体蛋白质，然后在PVDF膜上转移。阻断PVDF膜后，将血浆（500次稀释）作为原代抗体在膜上与分离的蛋白质反应，并使用ECLPLUS试剂盒检测到阳性斑点。另一方面，白色染色凝胶被切除一个与ECL阳性斑点相对应的阳性斑点，并通过飞行时间质谱仪（Ultraflex）或离子陷阱型质谱仪（LCQDECA）进行了分析，数据库搜索了提供的结果并确定了八个靶标蛋白。肺腺癌患者血浆中过氧蛋白6自身抗体。我们得出的结论是，这些自身抗体是诊断肺腺癌的候选生物标志物。

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Proteomic-based approach identifying autoantibody against peroxiredoxin VI as a novel serum marker in esophageal squamous cell carcinoma.

基于蛋白质组学的方法鉴定抗过氧化还原蛋白 VI 的自身抗体作为食管鳞状细胞癌的新型血清标志物。

DOI：
发表时间：
2006
期刊：
Clin.Cancer Res. 12(21)
影响因子：
0
作者：
Fujita Y;Nakanishi T;Hiramatsu M;Mabuchi H;Miyamoto Y;Miyamoto A;Shimizu A;Tanigawa N.
通讯作者：
Tanigawa N.

Detection and characterization of variant and modified structures of proteins in blood and tissues by mass spectrometry

通过质谱法检测和表征血液和组织中蛋白质的变异和修饰结构