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，Hanash教授及其同事利用蛋白质组学分析首次报道肺腺癌血浆中经常存在针对Annexin-I和II的自身抗体（37-40%），此后许多研究人员在癌症患者中发现了自身抗体。我们还应用基于蛋白质组学的新型分析来鉴定用于诊断肺腺癌的候选生物标志物。识别针对癌症/肿瘤相关抗原的自身抗体比基因分析方法更容易和简单，这种技术，即所谓的自身抗体组学，有望应用于血浆中癌症诊断生物标志物的筛选。 [方法]将培养的人肺癌细胞株（A549）在溶解缓冲液中处理，将可溶性组分应用到样品溶液中，用二维分离器分离可溶性蛋白质。电泳后转移到PVDF膜上，封闭PVDF膜后，将血浆（500倍稀释）与膜上分离的蛋白质反应，用ECLplus试剂盒检测阳性斑点。凝胶上切下与 ECL 阳性点相对应的阳性点，并通过飞行时间质谱仪 (UltraFlex) 或离子阱型质谱仪 ( [结果与结论]我们在肺腺癌患者血浆中发现了抗α-enlace、伴侣蛋白、peroxiredoxin-6自身抗体等8种生物标志物。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