Biomarker-Based Approach to α-Synucleinopathies: Lessons from Neuropathology

Abstract

Recently, proposals have attempted to reclassify Lewy body diseases in vivo by merging the long-established clinicopathological entities of Parkinson's disease (PD), Parkinson's disease dementia (PDD), and dementia with Lewy bodies (DLB), and some also to include rapid eye movement (REM)-sleep behavior disorder (RBD). A position paper¹ and a personal view paper² proposed biomarker-based staging and classification of these conditions. As both papers emphasize,^{1, 2} clinical diagnosis has challenges and limitations including early diagnosis that reflects the pathogenesis and clinical staging hindered by disease heterogeneity even within the same proteinopathy types. The suggested staging system is founded on the idea that diagnoses should rely on biomarkers, independent of the clinical syndrome, and the term neuronal α-synuclein disease (NSD) is proposed to redefine these conditions.¹ Together with the paper on biological classification² it was proposed that the detection of α-synuclein in cerebrospinal fluid (CSF) or skin and dopaminergic dysfunction assessed via positron emission tomography (PET) or single photon emission computed tomography (SPECT) possess the necessary sensitivity and specificity to identify the gold standard neuropathological alterations associated with Lewy body diseases.^{1, 2} We welcome these initial attempts to redefine these diseases incorporating biological constructs, particularly for the early and in vivo diagnosis of these disorders. Imaging and biofluid biomarkers for neurodegeneration have potential advantages in assessing disease presence and progression during life and have made major contributions in the research setting, most notably in enriching clinical trials for Alzheimer's disease (AD). Biology-based disease definition and classification has always been a central focus of neuropathology. As noted in the original publications,^{1, 2} this initial research framework will require much work to fill gaps in technology and knowledge, validate, and improve as we attempt transition from research biomarkers to disease surrogates. For example, currently α-synuclein seeding amplification assays (SAA) lack sensitivity and specificity for brain region and cell type, features known from neuropathology to be critically important to clinical outcomes. Our goal is to share the collective experience of our international group of neuropathology experts by suggesting future research priorities to further improve the proposed research frameworks.

The first description of Lewy bodies detectable on hematoxylin and eosin staining³ and glial cytoplasmic inclusions in multiple system atrophy (MSA), observed first using Gallyas silver staining⁴ was followed by the discovery of the central role of α-synuclein, which linked these diseases together as α-synucleinopathies.⁵ Application of various anti-α-synuclein antibodies used in immunohistochemistry^6-8 revealed a wide range of cytopathologies beyond the classical Lewy bodies, diffusely distributed in neuronal processes and the perikarya, and beyond that, in astrocytes^9-12 and oligodendroglia.^{11, 13} In MSA, the pathognomonic glial cytoplasmic inclusions (“Papp-Lantos bodies”)⁴ are accompanied by neuronal cytoplasmic and nuclear inclusions.¹⁴ New subtypes of MSA, where neuronal α-synuclein pathology in the limbic system is a predominant feature, have also been recognized.^{15, 16}

Several genes associated with the clinical features of PD are unaccompanied by Lewy bodies on neuropathological examination. Other, not PD-related mutations and genetic conditions, including those in PRNP (ie, genetic prion disease),¹⁷ APP,¹⁸ PSEN 1, PSEN 2,¹⁹ and trisomy 21²⁰ (ie, AD-related neuropathology), infantile neurodegenerative disorders,²¹ or neurodegeneration with brain iron accumulation,²² can also show Lewy body or other types of α-synuclein pathology.

From a neuropathological viewpoint, two major categories of conditions with α-synuclein pathology can be distinguished: those where α-synuclein pathology is consistently versus inconsistently detected (Table 1). One of the latter conditions is AD. The combination of tau and α-synuclein pathology can be a foundation for disease diversity, for example, in DLB,²³ but also in AD, and that along with other factors (genetic, environmental, etc) might reflect many distinct “biological” associations, warranting unique approaches to clinical diagnosis, prevention, and therapy to aid in precision medicine. Indeed, individuals with AD and α-synuclein pathology might not necessarily have dopaminergic alterations. Even within parkinsonian disorders, many concomitant clinicopathological diagnoses exist.²⁴

Novel approaches to diagnosis of α-synucleinopathy using peripheral tissues have emerged, but important caveats remain. α-Synuclein pathology is found in peripheral organs, which have been implicated as a site of initiation.²⁵ Because some studies did not find α-synuclein pathology in the periphery without brain pathology,^26-28 in contrast to another study,²⁹ current autopsy studies do not consistently support this position. Studies on peripheral organs may show variable results depending on the sampling, the processing methods, antibodies used, and type of cohorts evaluated, including whether autopsy confirmation was included. Furthermore, the clinical implications of α-synuclein detected in nasal swab³⁰ or postmortem in the retina³¹ need to be explored.

α-Synuclein SAA has been established not only in CSF, but also in peripheral tissues such as skin. Meta-analysis on studies, most of which did not include autopsy confirmation, have shown that the pooled sensitivity and specificity to differentiate α-synucleinopathies from controls using CSF samples and SAA is high, and that overall the biological samples tested to date, the CSF and skin α-synuclein SAA have demonstrated the best diagnostic performance.^32-34 Current CSF SAA show that α-synuclein aggregates are detectable only after α-synuclein pathology reaches a threshold in the brain. In particular, SAA CSF is less sensitive to detect α-synuclein pathology restricted to the brainstem or amygdala.^35-38 Furthermore, several conditions with α-synuclein pathology defined by neuropathology are yet to be examined using in vivo α-synuclein SAA biomarkers and the most reliable method needs to be validated (Table 1).

The integration of blood-based molecular findings⁴² complemented by tissue-based bulk, single cell,⁶⁷ spatial transcriptomics,⁶⁸ proteomics,⁴⁴ imaging to detect α-synuclein in vivo,⁶⁹ genetic studies⁷⁰ reflecting pathogenic scenarios of disease, observations on the spectrum of biochemical modifications⁷¹ of the protein used as the marker of pathology (ie, α-synuclein), and anatomical overlap with other proteins⁴⁰ with potential effect on seeding⁴⁵ will pave the path for stratified medicine. Artificial intelligence-based methods for the image analysis⁷² or for the detection of novel histological subtypes of disease⁷³ with clinical relevance are also expected to expand.

We believe that these tasks need to be addressed with harmonized SAA methods and human tissue-based studies with expanded expertise, for example, as performed for the validation and the Food and Drug Administration approval of the amyloid PET tracers.⁷⁴ Although biomarkers have already contributed to clinical trial successes, there is much research to be done to ensure their appropriate and effective use in clinical settings. The neuropathology community is eager to work with our clinical and neuroimaging colleagues to achieve this important goal.

1. Research project: A. Conception, B. Organization, C. Execution. (2) Manuscript: A. Writing of first draft, B. Review and critique.

G.G.K.: 1A, 1B, 1C, 2A

L.T.G: 1A, 2B

G.H.: 1A, 2B

I.A.: 1A, 2B

B.N.D.: 1A, 2B

S.M.: 1A, 2B

S.L.F.: 1A, 2B

I.MV.: 1A, 2B

H.T.: 1A, 2B

T.K.: 1A, 2B

K.Y.: 1A, 2B

Z.J.: 1A, 2B

S.S.: 1A, 2B

P.T.N.: 1A, 2B

S.G.: 1A, 2B

J.AA.: 1A, 2B

G.E.S.: 1A, 2B

V.R.P.: 1A, 2B

M.T.: 1A, 2B

K.W.: 1A, 2B

T.U.: 1A, 2B

M.Y.: 1A, 2B

Y.S.: 1A, 2B

J.K.: 1A, 2B

R.D.R.: 1A, 2B

E.G.: 1A, 2B

J.A.: 1A, 2B

J.F.C.: 1A, 2B

W.W.S.: 1A, 2B

J.E.D.: 1A, 2B

C.D.K.: 1A, 2B

J.W.: 1A, 2B

D.M.: 1A, 2B

C.S.: 1A, 2B

E.B.L.: 1A, 2B

M.M.: 1A, 2B

C.L.W.: 1A, 2B

A.C.M.: 1A, 2B

D.R.T.: 1A, 2B

K.J.: 1A, 2B

B.G.: 1A, 2B

I.R.A.M.: 1A, 2B

D.W.D.: 1A, 1B, 2B

T.G.B.: 1A, 1B, 1C, 2A, 2B

G.G.K. has served as an advisor for Parexel in 2023; received a royalty for 5G4 synuclein antibody and publishing royalties from Wiley, Cambridge University Press, and Elsevier; and received grants from Edmond J Safra Philanthropic Foundation, Rossy Family Foundation, Krembil Foundation, MSA Coalition, The Michael J. Fox Foundation, Parkinson Canada, National Institutes of Health (NIH), and Canada Foundation for Innovation. L.T.G. provides consultancy for Otsuka Speakers Bureau; serves on the Medical and Scientific Advisory Group for The Alzheimer Association; received honoraria from Medscape and Guidapoint Global, and Evidera; received grants from the NIH, Rainwater Charitable Foundation, and Weill Neurosciences Institute; and is funded by NIH K240534305 and R01AG075802. G.H. declares stock ownership in Cochlear and NIB holdings; has done committee work for the National Health and Medical Research Council (NHMRC) and Australian Department of Health and Aged Care; is employed at the University of Sydney; has received honoraria from Aligning Science Across Parkinson's, Gordon Conference, MDS, Viertel Symposium; has received royalties from Academic press, Elsevier and Oxford University Press; and has received grants from NHMRC, Medical Research Future Fund of Australia, NIH, Aligning Science Across Parkinson's, Defeat MSA Canada and Defeat MSA Australia-New Zealand, The Michael J. Fox Foundation, Shake-it-up Australia, and University of Sydney. B.D. had received funding from grants from the NIH under award numbers (P30AG072972, R01AG050782, U01AG061357-S1, U24NS133949, R01AG073474, RF1NS130659, U24AG072122-03S1 R01AG052132, R01AG056519, and R01AG062517), the Noyce Foundation, and funding from a residual class settlement funds in the matter of April Krueger vs. Wyeth, case no. 03-cv-2496 (United States District Court, Southern District of California). The views and opinions expressed in this manuscript are those of the author and do not necessarily reflect the official policy or position of any public health agency of California or of the United States government. S.L.F. received funding from the NHMRC, Australia. S.S. declares research funding from the NIH, the Bluefield Project to Cure FTD, the Rainwater Charitable Foundation, and consultation fees from Techspert.io. S.G. is Scientific Director of the Parkinson's UK Brain Bank at Imperial College London, funded by a Parkinson's United Kingdom grant. M.T. declares honoraria from Igaku Shoin, Eisai, Pfizer, Kyowa Kirin, Biogen Japan, and AbbVie; and grants from AMED, JSPS KAKENHI, intramural fund from NCNP, Health and Labour Sciences Research grants from the Ministry of Health, Labour, and Welfare, Japan. M.Y. declares grants from AMED. J.K. declares employment by the University of Pittsburgh and University of Pittsburgh Physicians and grant funding from National Institute on Aging, National Institute of Mental Health, National Institute of Neurological Disorders and Stroke, Chuck Noll Foundation, Pittsburgh Foundation, and Richard King Mellon Foundation. W.W.S. declares grant support from the NIH, Tau Consortium, Bluefield Project to Cure FTD, and the Chan-Zuckerberg Initiative; has received consulting fees from Biogen, Athenaeum Consulting, and Guidepoint Global Consulting; has received speaker honoraria from Verge Genomics; and is filing Patent Application no. PCT/US2021/53031 in the United States and the European Union. C.D.K. declares grants from NIH, US DoD, Chan-Zuckerberg Initiative, and Allen Brain Institute. J.W. has served as a paid collaborator on The Michael J. Fox Aligning Science Across Parkinson's (ASAP) grant (PI: Michael Schlossmacher). C.S. declares grant funding from NIH (grant numbers: 1U19AG074862-01A1; 1R01NS118183-01; 1U54AG076040-01: NIHR PR-ST-0614). E.B.L. declares funding from NIH and the Delaware Community Foundation; served as a consultant for Lilly and WaveBreak Therapeutics; served on advisory boards for academic Alzheimer's Disease Research Centers and the Rainwater Foundation; filed for a patent related to small molecule VCP activators; and received honoraria from the University of Auckland, University of Oslo, the Alzheimer's Drug Discovery Foundation/The Association for Frontotemporal Degeneration, NIH, Seoul University, Columbia University, and Stanford University. M.N. declares grant funding from the Deutsche Forschungsgemeinschaft and Alzheimer Forschung Initiative (Germany); serves on the advisory board of the Breuer Foundation (Germany). C.L.W. declares consultancies with Banner SunHealth Research Institute, Sun City, Arizona, United States (consultant on NIH-funded grant); is employed as a fulltime faculty member, Department of Pathology, University of Texas Southwestern Medical Center, Dallas, Texas, United States; received honoraria from NIH for grant review service; and grants from the National Institutes of Health, Texas Alzheimer's Research and Care Consortium. D.R.T. declares advisory boards memberships with the Alzheimer Forschung Initiative (Germany), and Luxembourg National Research Fund (Luxembourg); editorial board memberships with Acta Neuropathologica, Journal of Neuroinflammation, and Brain; industry collaborations with Novartis (Switzerland), GE-Healthcare (United Kingdom); and grants from Fond Wetenschappelijk Onderzoek (FWO; Flanders; G065721N), Stichting Alzheimer Onderzoek (SAO/FRA; Belgium; SAO/FRA 2020/017, 2023/0009), KU-Leuven Onderzoeksraad (Belgium; C14/22/132; C3/20/057; Opening the Future), and Alzheimer Association (USA; 22-AAIIA-963171). K.J. declares funding from the Society for the Promotion of Research in Experimental Neurology, Vienna, Austria. I.M. declares salaried employment with Vancouver Coastal Health; has received honoraria from the Japanese Society for Dementia Research Conference (plenary speaker); royalties from a patent license with Athena and AviadoBio; the United States patent 12/302.691 “Detecting and Treating Dementia”; and grants from NIH, the Canadian Institutes of Health Research, and the Alzheimer's Society of Canada. T.G.B. declares research funding from the NIH, The Michael J. Fox Foundation, and the State of Arizona; received research contracts funding from Life Molecular Imaging and Meilleur Technologies; received paid consultancy for Biogen and Aprinoia Therapeutics; and has stock options with Vivid Genomics; and received honoraria from the NIH, Mayo Clinic, Stanford University, and the International Movement Disorders Association. I.A., S.M., I.M.V., H.T., T.K., K.Y., Z.J., P.T.N., J.A.A., G.E.S.,V.R.P., K.W., T.U., Y.S., R.D.R., E.G., J.A., J.F.C., J.E.D., D.M., A.C.M., B.G., and D.W.D. have nothing to declare.