纳米磁珠联合质谱技术筛选大肠癌患者血清特异性蛋白标志物

目的：通过纳米磁珠联合质谱技术筛选并鉴定大肠癌患者特异性蛋白标志物及在大肠癌早期诊断中的意义。方法采用弱阳离子纳米磁珠联合MALDI-TOF-MS质谱技术分析102例大肠癌患者和124例健康对照者血清蛋白表达谱，并建立蛋白分类模型，采用LCQ液质联用质谱（LC-MS）技术鉴定已筛选出的特异性蛋白，并用该模型在双盲模式下从使用未建立模型的其它样本中选取59例大肠癌血清标本（Dukes A、B、C期各33例、18例和8例）和正常体检者90例进行检测以验证其诊断的准确性。结果大肠癌组与健康对照组间有8个蛋白表达存在明显差异（P〈0.05），其中大肠癌患者中有3个蛋白质高表达。差异蛋白质峰[质荷比（m/z）2963.17、4095.13、5906.83和38146.5]为最佳的大肠癌患者血清学诊断模型，敏感性为82.3%，特异性为90.3%，阳性预测值为90.3%，ROC曲线下面积为0.922。在双盲模式下的诊断准确率为89.83%，排除率为94.44%。其差异蛋白峰和大肠癌的分期无明显相关。LC-MS成功鉴定到m/z 2963.17（myosin-7B）、m/z 5906.83（Myoferlin）。结论纳米磁珠联合质谱技术对大肠癌患者血清学诊断具有较高灵敏度和特异性，并通过LCQ液质联用质谱成功鉴定到的myosin-7B和Myoferlin可能有助于大肠癌的早期诊断。

Objective To screen and identify early specific protein markers of colorectal cancer （CRC）by MALDI-TOF-MS technology. Methods Weak cationic nano magnetic beads combined with MALDI-TOF-MS were used to profile differentially expressed proteins between 102 cases of CRC patients and 124 healthy controls in the serum protein,then constructing protein classification as diagnosis model and identifying specific proteins by LCQ liquid chromatography-mass spectrometry. Selected serum specimens of 59 patients with CRC and 90 normal healthy persons from samples in unestablished model were used to verify the accuracy of diagnosis. Results We found that eight difference protein peaks between CRC patients and healthy group（P〈0.05） and three proteins showed high expression in the CRC patients. A diagnostic model with four proteomic features （m/z 2963.17,4095.13,5906.83 and 38146.5） of CRC patients was generated using Biomarker Patterns Software 5.0 with best performance, the sensitivity was 82.3%,specificity was 90.3%,the positive predictive value was 90.3%,and area under the ROC curve was 0.922. The accuracy rate of diagnosis in double blind mode was89.83%,removal rate was 94.44%. The difference between the protein peaks and the staging of CRC hadno correlation. The identification results showed that m/z 2963.17 （MYH7B,myosin-7B） and m/z 5906.83 （MYOF Isoform 1 of Myoferlin） by LCQ mass spectrometry coupled with liquid. Conclusion Myosin-7B and Myoferlin screened by nano magnetic beads combined with MALDI-TOF-MS technology may contribute to early diagnosis of CRC.