Cerebrospinal fluid and brain α-synuclein seed amplification in autopsy-confirmed Lewy body disease relates to the distribution of pathology

Objective To determine the sensitivity and specificity of α-synuclein seed amplification assay (αSyn-SAA) in antemortem and postmortem CSF and brain homogenate samples of autopsy-confirmed patients with a spectrum of Lewy-related pathology (LRP). Methods Antemortem CSF samples were examined from 119 subjects with standardized neuropathological examinations from OHSU and UCSD (56 additional postmortem CSF samples available). The assay was also applied to frontal cortex and amygdala tissue to determine if the results could be explained by a regional variation in the propensity for seed aggregation. Sensitivity, specificity, and assay kinetics were compared across pathology groups and clinical data was compared across αSyn-SAA positive and negative groups. Results Fifty-three LRP-individuals and 66 LRP+ individuals (neocortical (n=38), limbic (n=7), and amygdala-predominant (n=21)) were included. There was a sensitivity of 97.8% and specificity of 98.1% of the αSyn-SAA to identify patients with limbic/neocortical pathology from antemortem CSF. Sensitivity to detect amygdala-predominant pathology was only 14.3%. Postmortem CSF and brain tissue αSyn-SAA analyses showed a similar detection pattern, with higher positivity in samples from limbic/neocortical cases. Kinetic parameters of aggregation were significantly slower in amygdala-predominant cases compared to limbic and neocortical cases. Interpretation In this multicenter study of autopsy-confirmed subjects with a spectrum of Lewy-related pathology, we confirm that the αSyn-SAA using CSF and brain tissue reliably identifies α-synuclein seeds in patients with diffuse pathology and related cognitive symptoms. Pathological α-synuclein in the amygdala appears less likely to form detectable seeds, which may result from differences in abundance, conformation, or strains of α-synuclein. Summary for Social Media If Published 1. Twitter handles of the authors: none 2. Alpha-synuclein seed amplification assays have shown high sensitivity and specificity in clinically defined DLB and PD cohorts 3. It is less well known how well these assays detect synuclein seeds across a pathologically defined spectrum of Lewy body disease. Here we examine the ability of the αSyn-SAA to detect alpha-synuclein seeds in a multicenter cohort of autopsy-validated cases with a spectrum of Lewy body related pathology. 4. High sensitivity and specificity of the αSyn-SAA is confirmed in detecting alpha-synuclein seeds in spinal fluid and brain tissue in limbic and neocortical stage Lewy body stage pathology, but markedly decreased sensitivity is observed in detecting alpha-synuclein seeds in both spinal fluid and brain tissue in amygdala-predominant type Lewy body related pathology. A small number of these cases showed seeding capability from the amygdala that was not present in the frontal cortex, suggesting a topographic spread of alpha-synuclein seeds. 5. The current generation of αSyn-SAAs have a high sensitivity and specificity for detecting the most clinically relevant forms of Lewy body related pathology. Further study is needed to understand the differences in Lewy body related pathology between limbic/neocortical cases and amygdala-predominant cases that result in this difference in seeding capability. ### Competing Interest Statement Dr. Concha, Ms. Farris, and Mr. Ma are inventors on several patents related to PMCA technology (SAA) and are associated to Amprion Inc, a biotech company focused on the commercial utilization of SAA for diagnosis. All other authors have no conflicts of interest to disclose. ### Funding Statement This study was funded in part by the Alzheimer Disease Center Clinical Core at Oregon Health and Science University (PI: Kaye, eIRB 725, supported by NIH P30 AG008017, P30 AG066518) as well as the National Center for Advancing Translational Sciences (National Institutes of Health, Grant Award Number UL1TR002369). Amprion's efforts were funded in part by the Alzheimer's Drug Discovery Foundation (ADDF) Diagnostics Accelerator, as well as by the National Institute of Neurological Disorders and Stroke of the National Institutes of Health (Award Number U44NS111672). Moriah R. Arnold is funded through the Medical Scientist Training Program of Oregon Health & Science University (T32 GM 109835). Dr. Coughlin is funded by the National Institute of Neurological Disorders and Stroke of the National Institutes of Health (Award Number NS120038) and the National Institute on Aging (Award Number AG062429). Dr. Galasko is funded by the UCSD Shiley-Marcos ADRC (AG062429), and by the DLB Research Center of Excellence award from the Lewy Body Dementia Association. The funding sources had no role in the design and conduct of the study; collection, management, analysis, and interpretation of the data, preparation, review of approval of the manuscript, and decision to submit the manuscript for publication. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: eIRB 725 of Oregon Health and Science University ADRC gave ethical approval for this work. IRB 170957 of University of California San Diego ADRC gave ethical approval for this work. 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Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors.