Cholangiocarcinoma (CCA) includes a heterogeneous group of biliary cancers with poor prognosis. Several conditions, such as primary sclerosing cholangitis (PSC), are risk factors. Noninvasive differential diagnosis between intrahepatic CCA and hepatocellular carcinoma (HCC) is sometimes difficult. Accurate noninvasive biomarkers for PSC, CCA, and HCC are not available. In the search for novel biomarkers, serum extracellular vesicles (EV) were isolated from CCA (n = 43), PSC (n = 30), or HCC (n = 29) patients and healthy individuals (control, n = 32); and their protein content was characterized. By using nanoparticle tracking analysis, serum EV concentration was found to be higher in HCC than in all the other groups. Round morphology (by transmission electron microscopy), size (∼180 nm diameter by nanoparticle tracking analysis), and markers (clusters of differentiation 9, 63, and 81 by immunoblot) indicated that most serum EV were exosomes. Proteome profiles (by mass spectrometry) revealed multiple differentially expressed proteins among groups. Several of these proteins showed high diagnostic values with maximum area under the receiver operating characteristic curve of 0.878 for CCA versus control, 0.905 for CCA stage I‐II versus control, 0.789 for PSC versus control, 0.806 for noncirhottic PSC versus control, 0.796 for CCA versus PSC, 0.956 for CCA stage I‐II versus PSC, 0.904 for HCC versus control, and 0.894 for intrahepatic CCA versus HCC. Proteomic analysis of EV derived from CCA human cells in vitro revealed higher abundance of oncogenic proteins compared to EV released by normal human cholangiocytes. Orthotopic implant of CCA human cells in the liver of immunodeficient mice resulted in the release to serum of EV containing some similar human oncogenic proteins. Conclusion: Proteomic signatures found in serum EV of CCA, PSC, and HCC patients show potential usefulness as diagnostic tools. (Hepatology 2017;66:1125‐1143).

中文翻译：

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血清细胞外囊泡含有原发性硬化性胆管炎和胆管癌的蛋白质生物标志物

胆管癌 (CCA) 包括一组异质性预后较差的胆道癌。一些疾病，例如原发性硬化性胆管炎 (PSC)，是危险因素。肝内 CCA 和肝细胞癌 (HCC) 之间的无创鉴别诊断有时很困难。没有准确的 PSC、CCA 和 HCC 无创生物标志物。在寻找新的生物标志物时，从 CCA（n = 43）、PSC（n = 30）或 HCC（n = 29）患者和健康个体（对照，n = 32）中分离出血清细胞外囊泡 (EV)；并对它们的蛋白质含量进行了表征。通过使用纳米粒子追踪分析，发现 HCC 中的血清 EV 浓度高于所有其他组。圆形形态（通过透射电子显微镜）、尺寸（通过纳米粒子追踪分析发现直径约为 180 nm）、和标记物（免疫印迹的分化簇 9、63 和 81）表明大多数血清 EV 是外泌体。蛋白质组谱（通过质谱法）揭示了组间多种差异表达的蛋白质。这些蛋白质中的一些显示出高诊断值，CCA 与对照的受试者操作特征曲线下的最大面积为 0.878，CCA I-II 期与对照为 0.905，PSC 与对照为 0.789，非热性 PSC 与对照为 0.806，CCA 为 0.796与 PSC 相比，CCA I-II 期与 PSC 相比为 0.956，HCC 与对照组为 0.904，肝内 CCA 与 HCC 为 0.894。体外来自 CCA 人类细胞的 EV 的蛋白质组学分析显示，与正常人胆管细胞释放的 EV 相比，致癌蛋白的丰度更高。在免疫缺陷小鼠的肝脏中原位植入 CCA 人类细胞导致 EV 释放到血清中，其中包含一些类似的人类致癌蛋白。结论：在 CCA、PSC 和 HCC 患者的血清 EV 中发现的蛋白质组学特征显示出作为诊断工具的潜在有用性。（肝病学 2017；66：1125-1143）。