Institute for Clinical Chemistry, Medical Faculty Mannheim of the University of Heidelberg, Heidelberg, Germany.
Proteomics Clin Appl. 2012 Jan;6(1-2):60-78. doi: 10.1002/prca.201100058. Epub 2011 Dec 27.
Clinical proteomic profiling by mass spectrometry (MS) aims at uncovering specific alterations within mass profiles of clinical specimens that are of diagnostic value for the detection and classification of various diseases including cancer. However, despite substantial progress in the field, the clinical proteomic profiling approaches have not matured into routine diagnostic applications so far. Their limitations are mainly related to high-abundance proteins and their complex processing by a multitude of endogenous proteases thus making rigorous standardization difficult. MS is biased towards the detection of low-molecular-weight peptides. Specifically, in serum specimens, the particular fragments of proteolytically degraded proteins are amenable to MS analysis. Proteases are known to be involved in tumour progression and tumour-specific proteases are released into the blood stream presumably as a result of invasive progression and metastasis. Thus, the determination of protease activity in clinical specimens from patients with malignant disease can offer diagnostic and also therapeutic options. The identification of specific substrates for tumour proteases in complex biological samples is challenging, but proteomic screens for proteases/substrate interactions are currently experiencing impressive progress. Such proteomic screens include peptide-based libraries, differential isotope labelling in combination with MS, quantitative degradomic analysis of proteolytically generated neo-N-termini, monitoring the degradation of exogenous reporter peptides with MS, and activity-based protein profiling. In the present article, we summarize and discuss the current status of proteomic techniques to identify tumour-specific protease-substrate interactions for functional protease profiling. Thereby, we focus on the potential diagnostic use of the respective approaches.
基于质谱（MS）的临床蛋白质组学分析旨在揭示临床标本中质量谱的特定变化，这些变化对于诊断各种疾病（包括癌症）的检测和分类具有诊断价值。然而，尽管该领域取得了实质性进展，但临床蛋白质组学分析方法迄今尚未成熟为常规诊断应用。它们的局限性主要与高丰度蛋白及其被多种内源性蛋白酶复杂处理有关，因此难以进行严格的标准化。MS 偏向于检测低分子量肽。具体来说，在血清标本中，蛋白质经蛋白水解降解的特定片段适用于 MS 分析。众所周知，蛋白酶参与肿瘤的进展，肿瘤特异性蛋白酶可能是由于侵袭性进展和转移而释放到血液中。因此，测定恶性疾病患者临床标本中的蛋白酶活性可以提供诊断和治疗选择。在复杂的生物样本中鉴定肿瘤蛋白酶的特定底物具有挑战性，但蛋白酶/底物相互作用的蛋白质组学筛选目前正在取得令人印象深刻的进展。这些蛋白质组学筛选包括基于肽的文库、与 MS 结合的差异同位素标记、定量分析蛋白水解产生的新 N 末端、用 MS 监测外源性报告肽的降解以及基于活性的蛋白质谱分析。在本文中，我们总结和讨论了用于鉴定肿瘤特异性蛋白酶-底物相互作用的蛋白质组学技术的现状，以进行功能性蛋白酶分析。因此，我们重点讨论了各自方法的潜在诊断用途。
