Mass Spectrometric Analysis of Cerebrospinal Fluid Ubiquitin in Alzheimer's Disease and Parkinsonian Disorders

Abstract

Purpose: Dysfunctional proteostasis, with decreased protein degradation and an accumulation of ubiquitin into aggregated protein inclusions, is a feature of neurodegenerative diseases. Identifying new potential biomarkers in cerebrospinal fluid (CSF) reflecting this process could contribute important information on pathophysiology.

Experimental design: A developed method combining SPE and PRM-MS is employed to monitor the concentration of ubiquitin in CSF from subjects with Alzheimer's disease (AD), Parkinson's disease (PD), and progressive supranuclear palsy (PSP). Four independent cross-sectional studies are conducted, studies 1-4, including controls (n = 86) and participants with AD (n = 60), PD (n = 15), and PSP (n = 11).

Results: The method shows a repeatability and intermediate precision not exceeding 6.1 and 7.9%, respectively. The determined LOD is 0.1 nm and the LOQ range between 0.625 and 80 nm. The CSF ubiquitin concentration is 1.2-1.5-fold higher in AD patients compared with controls in the three independent AD-control studies (Study 1, p < 0.001; Study 2, p < 0.001; and Study 3, p = 0.003). In the fourth study, there is no difference in PD or PSP, compared to controls.

Conclusion and clinical relevance: CSF ubiquitin may reflect dysfunctional proteostasis in AD. The described method can be used for further exploration of ubiquitin as a potential biomarker in neurodegenerative diseases.

Keywords: alzheimer's disease; biomarker; parkinson's disease; progressive supranuclear palsy; ubiquitin.

Figure 1

Ubiquitin concentrations in AD core CSF biomarker profile samples. Two cross‐sectional studies were performed including subjects selected by their AD core CSF biomarker profile. A) Study 1 included subjects designated as controls (n = 15) and AD (n = 9). B) Study 2 included subjects designated as controls (n = 15) and AD (n = 14). The groups were compared using Wilcoxon rank‐sum test.

Figure 2

CSF ubiquitin concentrations in participants with AD, PD, and PSP. Two cross‐sectional studies were performed including clinically characterized participants. The participants with AD met the AD core CSF biomarker criteria suggesting AD. A) Study 3 included healthy volunteers (controls, n = 45) and participants with AD (n = 37). The groups were compared using Wilcoxon rank‐sum test. B) Study 4 included a control group (n = 11), participants with PD (n = 15) and PSP (n = 11). The groups were compared using Kruskal–Wallis with Dunn's multiple comparison test, comparing all pairs.

Figure 3

Correlations of CSF ubiquitin with AD core CSF biomarker concentrations. Correlations were calculated between the concentration of ubiquitin and the AD core CSF biomarkers; T‐tau, P‐tau₁₈₁, or the 42 amino acid long Aβ_1–42. Indicated are Spearman's ρ and the p‐value. The scatter plots show the concentrations in controls (n = 45), A–C, and in participants with AD (n = 37), D–F, from Study 3.