Total CSF α-synuclein is lower in de novo Parkinson patients than in healthy subjects
Highlights
► We established a cohort comprising 78 drug-naïve PD and 48 healthy subjects (HC). ► We analyzed the samples by two different enzyme-linked immunoabsorbent assays (ELISA). ► One assay represents a renewable antibody-based commercial test. ► We show a significant decrease in total CSF α-synuclein values in drug-naïve PD patients versus HC.
Introduction
The quantification of α-synuclein in cerebrospinal fluid (CSF) was recently identified as a biomarker candidate for Parkinson's disease (PD) and α-synuclein-related diseases [17]. Importantly, we recently determined that the presence of α-synuclein in human CSF is the result of neuronal metabolism in the central nervous system (rather than of filtration from peripheral sources) [10]. Several (but not all) studies reported decreased total α-synuclein values in advanced PD, dementia with Lewy bodies and multiple system atrophy {reviewed in: [7]; decrease reported in: [2], [3], [4], [8], [9], [11], [15], [17], [19]; no change published in: [12], [14]}. A 2nd generation, sandwich enzyme-linked immunoabsorbent assay (ELISA)-based platform for α-synuclein quantification, which we have previously used in five independent cohort studies including one with autopsy validation, revealed a significant separation of patients with α-synuclein-linked disorders from neurological controls [8].
However, three unanswered issues arose from these investigations concerning CSF α-synuclein: (1) the unknown concentration of the analyte in age-matched, neurologically healthy subjects; (2) the possibility of a pharmacological treatment effect on its values (such as those mediated by dopaminergic agents via changes in chorioid plexus function); and (3) the wider applicability of these results, given that data were generated by either non-renewable systems, or by assays that were not yet widely available, or both.
We therefore established a new, single center-based, longitudinal cohort study of drug-naïve PD patients and healthy controls, where the study protocol sees the routine collection of CSF at the time of enrolment (and during future follow up visits) in accordance with previously validated, standardized operating procedures [7]. We hypothesized that CSF α-synuclein is reduced in early, drug-naïve PD patients when compared with age-matched, healthy control subjects. We also pursued – in collaboration with an industry-based assay development team – the transformation of our original ELISA system into a 3rd generation assay, i.e., a commercially available platform that employs continuously renewable, monoclonal antibodies. We anticipated that the two assay systems (2nd vs. 3rd generation) would demonstrate acceptable correlation efficiency.
Section snippets
Subjects and methods
Study participants: We established a new cohort of recently diagnosed, drug-naïve patients with PD and healthy controls as part of a longitudinal follow-up study (“DeNoPa cohort”). A full description of the cohort and its entire study design will be published elsewhere. Between 2009 and 2011, CSF specimens were collected during baseline visits from 130 study participants at the Paracelsus-Elena Klinik in Kassel, Germany (Table 1). Ethics approval of the study, diagnostic criteria employed in
Results
Each study participant was subjected to routine lumbar puncture to examine α-synuclein, hemoglobin and total protein concentrations in untreated persons with early PD (n = 78) and age-matched controls (n = 48; Table 1). A new assay to determine α-synuclein was developed using renewable components. It encompasses the pairing of a previously developed monoclonal antibody (mAb) to human α-synuclein, which was raised in mice [epitope, amino acids (aa 103–107), with a new mAb raised in rabbits (epitope,
Discussion
A decrease in the concentration of total CSF α-synuclein was shown in several (but not all) recently published studies of PD patients {reviewed in: [7]}. The discrepant findings between positive and negative studies are currently best explained by the different technological platforms used, variable antibody characteristics and different sample processing protocols [7]. Furthermore, the clinical severity of PD subjects analyzed varied considerably between these investigations. Last but not
Conclusion
To our knowledge, we show for the first time that total CSF α-synuclein values are reduced in early, drug-naïve PD patients (mean, 1339.5 pg/ml) when compared to healthy subjects (1504.0 pg/ml). The relatively low AUC value generated by our results, which is mainly caused by low specificity of CSF α-synuclein as a laboratory marker for the diagnosis of PD in a ‘receiver operating curve’-based model, reflect a substantial overlap of individual concentrations for CSF α-synuclein among our study
Author contributions
(a) Study concept, assay development, experimental design; (b) acquisition of data; (c) analysis and interpretation; (d) critical revision of the manuscript for important intellectual content; (e) study supervision; (f) statistical analysis. Brit Mollenhauer (a, b, c, e); Ellen Trautmann (f, c, d), Peggy Taylor (a, b, d), Paul Manninger (b, d), Friederike Sixel-Döring (a, b, c), Jens Ebentheuer (b, d), Claudia Trenkwalder (a, c, e), Michael G. Schlossmacher (a, c, d, e).
Conflict of interest statement
BM was supported by the Michael J. Fox Foundation for Parkinson's Research, the American Parkinson's Disease Association and the Stifterverband für die Deutsche Wissenschaft (Dr. Werner Jackstädt-Stipendium); grants from TEVA-Pharma, Desitin, Boehringer Ingelheim and received honoraria for presentations or reimbursements for travel/meeting expenses from Boehringer-Ingelheim, Novartis, GlaxoSmithKline, Orion Pharma, TEVA and Bayer Schering Pharma AG. ET, JE and PM have no conflict of interest to
Acknowledgements
This study was supported by the Michael J Fox Foundation for Parkinson's Research and TEVA-Pharma. We thank our study subjects for their participation and support.
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Both authors contributed equally.