SELDI-TOF-MS联合蛋白质组学实验数据库对结直肠癌血清标志物的鉴定

Combined use of surface-enhanced laser desorption/ionization time-of-flight mass spectrometry and empirical proteomic ontology knowledge base for identification of serum biomarkers for colorectal cancer

目的:探讨表面增强激光解析离子化飞行时间质谱仪(SELDI-TOF-MS)联合蛋白质组学实验数据库方法研究结直肠癌患者血清标志物的可行性.方法:应用SELDI-TOF-MS对169例结直肠癌患者与83例正常人血清进行了蛋白质谱检测,通过Biomarker WizardTM与Biomarker PatternsTM软件分析发现差异蛋白,建立结直肠癌诊断模型;用蛋白质组学实验数据库检索鉴定出有意义的差异蛋白,并用ELISA法对结直肠癌患者及正常人血清样本中差异蛋白表达进行验证.结果:发现34种差异蛋白,以质荷比(m/z)为2873,3163,6121,7778的4种差异蛋白组成的分类树诊断模型,在学习模式下其诊断结直肠癌的灵敏度为91.57%,特异度为90.53%;在测试模式下分别为82.25%,80.72%.其中m/z为7778的蛋白峰强度在结直肠癌患者血清中明显高于正常人(P<0.01),数据库检索结果提示该蛋白为血小板因子4(PF4).ELISA法测定结直肠癌患者血清PF4水平显著高于正常人(P<0.01),其灵敏度及特异度分别为61.1%,76.9%.结论:SELDI-TOF-MS联合蛋白质组学实验数据库的方法简便可行,可为蛋白芯片研究提供新思路;PF4(m/z=7778)在结直肠癌患者血清中明显升高,是结直肠癌患者新的血清学标志物.

AIM：To determine the feasibility of combined use of surface-enhanced laser desorption/ionization time-of-flight mass spectrometry （SELDI-TOF-MS） and empirical proteomic ontology knowledge base （EPO-KB） for identification of serum biomarkers for colorectal cancer. METHODS：Serum samples from 169 colorectal cancer patients and 83 age-and gender-matched healthy individuals were analyzed by SELDI-TOF-MS using immobilized metal affinity capture 30 （IMAC30） proteinchip arrays. The resulting SELDI-TOF-MS spectral data were analyzed using the Biomarker WizardTM and Biomarker PatternsTM software to find differential proteins and develop a classification tree for colorectal cancer. The differential proteins were identified by searching the EPO-KB and verified by enzyme-linked immunosorbent assay （ELISA）. RESULTS：A total of 34 mass peaks were identified. Four peaks at mass：charge ratios （m/z） of 2873,3163,6121 and 7778 were used to construct the classification tree. The classification tree could effectively distinguish colorectal cancer samples from control samples,achieving a sensitivity and specificity of 91.57% and 90.53% in training mode,and 82.25% and 80.72% in test mode,respectively. Database search revealed that the peak at a m/z of 7778 corresponded to platelet factor 4 （PF4）. ELISA detection of PF4 showed that serum PF4 level was significantly higher in colorectal cancer patients than in healthy controls （P〈0.01）,with a sensitivity and specificity of 61.1% and 76.9%,respectively. CONCLUSION：Combined use of SELDI-TOF-MS and EPO-KB is feasible for identification of serum biomarkers for colorectal cancer. PF4 （m/ z at 7778） is significantly upregulated in colorectal cancer patients,providing a new biomarker for colorectal cancer.