ANALYSIS OF NEW POTENTIAL CSF BIOMARKERS FOR ALZHEIMER’S DISEASE BY PARALLEL REACTION MONITORING MASS SPECTROMETRY

Abstract

Targeted mass spectrometric analysis has the advantage of high resolution and sensitivity without the use of antibodies, but requires instrumentation capable of parallel reaction monitoring (PRM). The aim for this study was the development of a parallel reaction monitoring mass spectrometry (PRM-MS) assay consisting of a panel of potential protein biomarkers in cerebrospinal fluid (CSF). The 13 proteins in the assay (Cystatin C, beta-2-microglobulin, neurosecretory protein VGF, Chromogranin A, Secretogranin-2, APP, Lysozyme c, Neurexin-1, -2, and -3, neuronal pentraxin-1, neurofascin, and neurocan CP) were selected based on their presumed role in processes regarding excretory vesicle function, synaptic function and innate immunity. 100-μL-CSF-samples were subjected to digestion with trypsin and 2–3 peptides of each protein were monitored. Stable-isotope-labeled peptides were used as standards in the quantification. The coefficient of variation was below 15% for the vast majority of peptides investigated. The assay was subjected to evaluation with a sample set of 10 patients with Alzheimer's disease (AD) and 15 healthy subjects. Secretogranin-2 (P<0.005) and neurosecretory protein VGF (P<0.001) levels were lowered in the AD patient group. For chromogranin A the levels of two peptides were significantly lowered in AD (P<0.01), while a third peptide was not. The synaptic proteins neurexin-1 and neuronal pentraxin-1, as well as neurofascin were also significantly lowered in AD but at a lower confidence level (P<0.05). PRM-MS seems to be a feasible technique to validate potential protein and peptide markers in CSF. The differentially expressed proteins must now be validated in a larger cohort of patients with AD and other neurodegenerative disorders, and healthy control subjects.