Identification of novel CSF biomarkers for neurodegeneration and their validation by a high-throughput multiplexed targeted proteomic assay.
Identification of novel CSF biomarkers for neurodegeneration and their validation by a high-throughput multiplexed targeted proteomic assay.
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DOI：
10.1186/s13024-015-0059-y
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发表时间：
2015-12-01
影响因子：
15.1
通讯作者：
Mills K
中科院分区：
文献类型：
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作者：
Heywood WE;Galimberti D;Bliss E;Sirka E;Paterson RW;Magdalinou NK;Carecchio M;Reid E;Heslegrave A;Fenoglio C;Scarpini E;Schott JM;Fox NC;Hardy J;Bhatia K;Heales S;Sebire NJ;Zetterberg H;Mills K
目前许多神经退行性疾病还没有有效的治疗方法。用于识别和分层这些疾病的可靠生物标志物对于未来新疗法的开发非常重要。路易体痴呆 (LBD) 被认为是一种诊断不足的痴呆症，需要标记物来区分 LBD 和其他形式的痴呆症，例如阿尔茨海默病 (AD)。这项工作描述了对非神经退行性对照和 LBD 患者的脑脊液 (CSF) 进行无标记蛋白质组学分析。利用这项技术，我们鉴定出了几种潜在的 LBD 新型标记物。然后将它们与之前发表的研究中的其他生物标志物结合起来，创建 10 分钟多重靶向和转化 MRM-LC-MS/MS 检测。该测试用于在更大的样本组中验证我们的新检测方法，其中包括对照组以及阿尔茨海默病和帕金森病 (PD) 的其他神经退行性疾病。通过蛋白质组分析，LBD CSF 中 38 种蛋白质的表达显着改变（p<0.05）。靶向 MRM-LC-MS/MS 检测显示 4 种蛋白可特异性识别 LBD 中的 AD：外核苷酸焦磷酸酶/磷酸二酯酶 2 (p < 0.0001)、溶酶体相关膜蛋白 1 (p < 0.0001)、食欲素原 (p < 0.0017) 和转甲状腺素蛋白(p < 0.0001)。与对照组相比，AD 和 LBD 中 19 种蛋白质显着升高，其中 4 种蛋白质是新的（苹果酸脱氢酶 1、血清淀粉样蛋白 A4、GM2 激活蛋白和前塞塞辛）。蛋白质 DJ1 仅在 PD 组中显着升高，而在 LBD 或 AD 样本中均没有显着升高。通过 ELISA 测定，观察 AD 疾病组中运甲状腺素蛋白、GM2 激活蛋白和 IGF2 与阿尔茨海默病相关淀粉样蛋白 β-42 水平的相关性（r2 ≥ 0.39，p ≤ 0.012）。在所有组中，胱抑素 C、泛素和骨桥蛋白与磷酸化 tau 水平均表现出很强的显着线性关系（r2 ≥ 0.4，p ≤ 0.03），而苹果酸脱氢酶和载脂蛋白 E 仅在 AD 和 LBD 疾病组中与磷酸化 tau 和总 tau 水平呈线性关系。利用蛋白质组学，我们鉴定了几种潜在的新型神经退行性标记物，并随后使用快速多重质谱测试对其进行了验证。这个有针对性的蛋白质组平台可以测量神经退行性变的常见标志物，这些标志物与现有的诊断标志物以及一些有可能显示 AD 与 LBD 之间变化的标志物相关。本文的在线版本 (doi:10.1186/s13024-015-0059-y) 包含补充材料，可供授权用户使用。
Currently there are no effective treatments for many neurodegenerative diseases. Reliable biomarkers for identifying and stratifying these diseases will be important in the development of future novel therapies. Lewy Body Dementia (LBD) is considered an under diagnosed form of dementia for which markers are needed to discriminate LBD from other forms of dementia such as Alzheimer’s Disease (AD). This work describes a Label-Free proteomic profiling analysis of cerebral spinal fluid (CSF) from non-neurodegenerative controls and patients with LBD. Using this technology we identified several potential novel markers for LBD. These were then combined with other biomarkers from previously published studies, to create a 10 min multiplexed targeted and translational MRM-LC-MS/MS assay. This test was used to validate our new assay in a larger cohort of samples including controls and the other neurodegenerative conditions of Alzheimer’s and Parkinson’s disease (PD). Thirty eight proteins showed significantly (p < 0.05) altered expression in LBD CSF by proteomic profiling. The targeted MRM-LC-MS/MS assay revealed 4 proteins that were specific for the identification of AD from LBD: ectonucleotide pyrophosphatase/phosphodiesterase 2 (p < 0.0001), lysosome-associated membrane protein 1 (p < 0.0001), pro-orexin (p < 0.0017) and transthyretin (p < 0.0001). Nineteen proteins were elevated significantly in both AD and LBD versus the control group of which 4 proteins are novel (malate dehydrogenase 1, serum amyloid A4, GM2−activator protein, and prosaposin). Protein-DJ1 was only elevated significantly in the PD group and not in either LBD or AD samples. Correlations with Alzheimer-associated amyloid β-42 levels, determined by ELISA, were observed for transthyretin, GM2 activator protein and IGF2 in the AD disease group (r2 ≥ 0.39, p ≤ 0.012). Cystatin C, ubiquitin and osteopontin showed a strong significant linear relationship (r2 ≥ 0.4, p ≤ 0.03) with phosphorylated–tau levels in all groups, whilst malate dehydrogenase and apolipoprotein E demonstrated a linear relationship with phosphorylated-tau and total-tau levels in only AD and LBD disease groups. Using proteomics we have identified several potential and novel markers of neurodegeneration and subsequently validated them using a rapid, multiplexed mass spectral test. This targeted proteomic platform can measure common markers of neurodegeneration that correlate with existing diagnostic makers as well as some that have potential to show changes between AD from LBD. The online version of this article (doi:10.1186/s13024-015-0059-y) contains supplementary material, which is available to authorized users.