Tsangaris George Th, Papathanasiou Chrissa, Adamopoulos Panagiotis G, Scorilas Andreas, Vorgias Constantinos E, Prodromou Neofytos, Stathopoulou Foteini Tzortzatou, Stravopodis Dimitrios J, Anagnostopoulos Athanasios K
Proteomics Research Unit, Biomedical Research Foundation of the Academy of Athens, Athens, Greece.
Hematology/Oncology Unit, First Department of Pediatrics, University of Athens, Aghia Sophia Children's Hospital, Athens, Greece.
Cancer Genomics Proteomics. 2017 Mar-Apr;14(2):127-136. doi: 10.21873/cgp.20025.
BACKGROUND/AIM: Proteomics based on high-resolution mass spectrometry (MS) is the tool of choice for the analysis of protein presence, modifications and interactions, with increasing emphasis on the examination of tumor tissues. Application of MS-based proteomics offers a detailed picture of tumor tissue characteristics, facilitating the appreciation of different tumor entities, whilst providing reliable and fast results for therapeutic marker targeting and prognostic factor assessment. Through use of the high analytical resolution of nano-high-pressure liquid chromatography (nanoHPLC) and the high resolution of an Orbitrap Elite mass spectrometer, the present study aimed to provide knowledge on the proteome of the generally unknown entity of pediatric ependymal tumors.
Ten resected specimens of childhood ependymoma were analyzed through a one-dimensional (1D) nanoLC-MS/MS approach. Method optimization steps were undertaken for both the sample preparation/protein extraction procedure and LC parameters, aiming to achieve the highest possible identification rates.
Following method optimization, each nanoLC-MS/MS run resulted in identification of more than 5,000 proteins and more than 25,000 peptides for every analyzed sample, thus detailing the greater part of the ependymoma proteome. Identified proteins were found to spread throughout all known tumor categories regarding their molecular function and subcellular localization.
Through the proposed nanoLC-MS/MS method herein we report, for the firs time, the ependymoma proteome database. A large number of similarities regarding proteome content are revealed compared to other two pediatric brain tumor entities; astrocytomas and medulloblastomas. Furthermore, through our approach, the majority of currently proposed markers for ependymoma (e.g. nucleolin, nestin, Ki67 and laminin subunit A2) as well as all major key players of the phosphoinositide 3-kinase pathway (seemingly implicated in ependymoma), were definitely detected.
背景/目的：基于高分辨率质谱（MS）的蛋白质组学是分析蛋白质存在、修饰和相互作用的首选工具，目前越来越重视对肿瘤组织的检测。基于MS的蛋白质组学应用能够详细描绘肿瘤组织特征，有助于鉴别不同的肿瘤实体，同时为治疗标志物靶向和预后因素评估提供可靠且快速的结果。通过使用纳米高压液相色谱（nanoHPLC）的高分析分辨率和Orbitrap Elite质谱仪的高分辨率，本研究旨在提供关于一般未知的小儿室管膜瘤实体蛋白质组的知识。
采用一维（1D）nanoLC-MS/MS方法分析10例儿童室管膜瘤切除标本。对样品制备/蛋白质提取程序和液相色谱参数进行方法优化步骤，旨在实现尽可能高的鉴定率。
经过方法优化，每次nanoLC-MS/MS运行可为每个分析样品鉴定出5000多种蛋白质和25000多种肽段，从而详细描述了室管膜瘤蛋白质组的大部分内容。已鉴定的蛋白质在分子功能和亚细胞定位方面分布于所有已知的肿瘤类别中。
通过本文报道的nanoLC-MS/MS方法，我们首次报告了室管膜瘤蛋白质组数据库。与其他两种小儿脑肿瘤实体——星形细胞瘤和髓母细胞瘤相比，揭示了蛋白质组含量方面的大量相似性。此外，通过我们的方法，明确检测到了目前大多数提出的室管膜瘤标志物（如核仁素、巢蛋白、Ki67和层粘连蛋白亚基A2）以及磷脂酰肌醇3-激酶途径的所有主要关键分子（似乎与室管膜瘤有关）。
