Proteomic analysis of tumor-derived nanovesicles and its application for innovative diagnostic method.

这是一项具有挑战性的研究，旨在探索肿瘤细胞衍生的纳米囊泡（外泌体）上的生物标志物，并研究其作为癌症早期检测和分子表征的创新诊断标志物以及癌症治疗决策的潜力。在这里，我们对外来体的蛋白质进行了集中的蛋白质组分析，这些外来体最近被揭示为从包括肿瘤细胞在内的各种细胞分泌的膜囊泡。首先，通过超离心法从细胞培养上清液中收集肿瘤细胞来源的外泌体。最终收集的产物通过电子显微镜的形态学、通过动态光散射的粒度和通过蛋白质印迹的四跨膜蛋白（其是外泌体的标记物）的表达来确认为纯化的外泌体。然后，从纯化的外泌体中提取蛋白，并通过质谱分析表达的蛋白。这些实验表明，在肿瘤来源的外来体中可检测到各种肿瘤相关的膜蛋白。未来，我们将通过从患者外周血中分离肿瘤源性外泌体并分析其肿瘤相关蛋白来测试基于外泌体的体外诊断的可行性。

This is a challenging study to explore biomarkers on tumor cell-derived nanovesicles (exosomes) and to investigate their potential as innovative diagnostic markers for early detection and molecular characterization of cancer, and for decision making in cancer treatment. Here, we performed a focused proteome analysis for the proteins of exosomes, which are recently revealed to be membrane vesicles secreted from various cells including tumor cells. At first, tumor cell-derived exosomes were collected by ultracentrifugation method from cell culture supernatant. The finally collected products were confirmed to be purified exosomes by morphology of electron microscopy, particle size by dynamic light scattering and expression of tetraspanins, which were markers of exosomes, by western blotting. Then, proteins were extracted from the purified exosomes and analyzed the expressed proteins by mass spectrometry. These experiments revealed that various tumor-related membrane proteins were detectable in tumor-derived exosomes. In the future, we will test the feasibility of exoxome-based in vitro diagnosis by isolating the tumor-derived exoxomes from the peripheral blood of patients and profiling tumor-related proteins of them.