Tissue-enhanced proteomic analysis of plasma samples reveals new mechanistic biomarker candidates for the stratification of Amyotrophic lateral sclerosis patients
Creators: name: Zubiri, Irene
Publisher: name: F1000Research
Year: 2016

description: Background.<br>Early diagnosis and patient stratification in ALS based on disease sub-type and progression rate requires the ability to measure objectively the molecular changes underlying the development of disease. The simultaneous analysis of diseased tissues and biological fluids using a powerful and revolutionary proteomic technique called TMTcalibrator™, enhances detection of low-abundant, tissue-derived proteins that reflect the central disease processes in the periphery. These are powerful biomarkers that should lead to a better clinical stratification of ALS patients.<br><br>Objectives.<br>To co-analyse the plasma and peripheral blood mononuclear cell proteomes from patients with different forms and rates of progression of ALS and equivalent mouse models to identify co-expressed proteins, which are differentially regulated at different stages of disease and may serve as useful stratification biomarkers.<br><br>Methods.<br>TMTcalibrator™ was employed to enhance detection of disease-associated plasma proteins. PBMC digests were used as the calibrator, spiked in a 1.75:1 excess to plasma samples from patients stratified according to progression rate and disease stage. Six patients with a slow progression rate (monthly slope of ALSRS-R from disease onset to last visit or progression rate to last visit, PRL&lt;0.5) and 6 patients with a fast progression rate (PRL&gt;1) were studied. For each patient, two plasma samples were included in the study from an early (ALSRS-R (/48) &gt;40) and a late disease stage (ALSRS-R (/48) &lt;30.) An equivalent study was performed in PBMCS/plasma from mouse models reproducing a similar fast and slow disease progression.<br><br>Results.<br>We have identified approximately 4500 unique proteins in each data set. We first analysed data by time point to identify 95 differentially regulated proteins between slow and fast progressors in early samples, and 107 significantly regulated proteins in late stage samples. When considering slow and fast progressing patients separately, 625 proteins (259 unique, 366 shared) were differentially expressed between early and late samples for the slow progressing patients and 525 (159 unique, 366 shared) for the fast progressing patients. Functional annotation was performed to extract relevant biological interpretation in the context of known mechanisms of ALS. In silico comparisons of these results with those obtained from the mouse models have also been performed to highlight those proteins that are detected as regulated in both species.<br> <br>Discussion and conclusions.<br>This is the deepest proteomic profiling yet undertaken on plasma from ALS patients. The TMTcalibrator™ technique has provided a unique opportunity to study the interface between cellular and fluid components of blood in unravelling of ALS pathology.  The parallel analysis of mouse models recapitulating a similar heterogeneity of the disease will provide a formidable working platform with translational potential and a dataset to support researchers working in ALS.<br> <br>Acknowledgements.<br>We are grateful to the Motor Neurone Disease Association and The Wellcome Trust for support.<br>