Biomarker discovery in mass spectrometry-based urinary proteomics

Abstract

Urinary proteomics has become one of the most attractive topics in disease biomarker discovery. MS-based proteomic analysis has advanced continuously and emerged as a prominent tool in the field of clinical bioanalysis. However, only few protein biomarkers have made their way to validation and clinical practice. Biomarker discovery is challenged by many clinical and analytical factors including, but not limited to, the complexity of urine and the wide dynamic range of endogenous proteins in the sample. This article highlights promising technologies and strategies in the MS-based biomarker discovery process, including study design, sample preparation, protein quantification, instrumental platforms, and bioinformatics. Different proteomics approaches are discussed, and progresses in maximizing urinary proteome coverage and standardization are emphasized in this review. MS-based urinary proteomics has great potential in the development of noninvasive diagnostic assays in the future, which will require collaborative efforts between analytical scientists, systems biologists, and clinicians.

Keywords: Biomarker; Mass spectrometry; Proteomics; Urine.

Fig. 1

Web of Science search results of the yearly trend in publishing frequency for general “proteom*” biomarker papers, and those including “urin*”

Fig. 2

Flowchart of the multi-fractionation procedures for urinary proteomic analysis (left) and Venn diagram of the detected 3429 proteins (right), adapted and modified from Santucci et al. [49] with permission.

Fig. 3

A representative workflow of quantitative urine proteomics using isobaric labeling. Urine samples are collected from human patients (control and disease groups) and centrifuged to remove particulates and cell debris. Protein fraction from each urine sample is obtained with molecular weight cut-off filter. Total protein concentration is measured with bicinchoninic acid (BCA) assay for normalization. After protein digestion and desalting, each sample is labeled by different channels of isobaric tags and combined after labeling reaction. Offline SCX LC is often performed to fractionate samples before LC-MS/MS analysis. The intensities of reporter ions are used for relative quantification.