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Genome-wide association studies have successfully identified many genetic risk loci for dementia, but exact biological mechanisms through which genetic risk factors contribute to dementia remains unclear. Integrating CSF proteomic data with dementia risk loci could reveal intermediate molecular pathways connecting genetic variance to the development of dementia. We tested to what extent effects of known dementia risk loci can be observed in CSF levels of 665 proteins [proximity extension-based (PEA) immunoassays] in a deeply-phenotyped mixed memory clinic cohort [ n = 502, mean age (standard deviation, SD) = 64.1 (8.7) years, 181 female (35.4%)], including patients with Alzheimer's disease (AD, n = 213), dementia with Lewy bodies (DLB, n = 50) and frontotemporal dementia (FTD, n = 93), and controls (n = 146). Validation was assessed in independent cohorts (n = 99 PEA platform, n = 198, mass reaction monitoring-targeted mass spectroscopy and multiplex assay). We performed additional analyses stratified according to diagnostic status (AD, DLB, FTD and controls separately), to explore whether associations between CSF proteins and genetic variants were specific to disease or not. We identified four AD risk loci as protein quantitative trait loci (pQTL): CR1 -CR2 (rs3818361, P = 1.65 × 10−8), ZCWPW1 -PILRB (rs1476679, P = 2.73 × 10−32), CTSH -CTSH (rs3784539, P = 2.88 × 10−24) and HESX1 -RETN (rs186108507, P = 8.39 × 10−8), of which the first three pQTLs showed direct replication in the independent cohorts. We identified one AD-specific association between a rare genetic variant of TREM2 and CSF IL6 levels (rs75932628, P = 3.90 × 10−7). DLB risk locus GBA showed positive trans effects on seven inter-related CSF levels in DLB patients only. No pQTLs were identified for FTD loci, either for the total sample as for analyses performed within FTD only. Protein QTL variants were involved in the immune system, highlighting the importance of this system in the pathophysiology of dementia. We further identified pQTLs in stratified analyses for AD and DLB, hinting at disease-specific pQTLs in dementia. Dissecting the contribution of risk loci to neurobiological processes aids in understanding disease mechanisms underlying dementia. [ABSTRACT FROM AUTHOR]

Autosomal dominant Alzheimer's disease (ADAD) offers a unique opportunity to study pathophysiological changes in a relatively young population with few comorbidities. A comprehensive investigation of proteome changes occurring in ADAD could provide valuable insights into AD-related biological mechanisms and uncover novel biomarkers and therapeutic targets. Furthermore, ADAD might serve as a model for sporadic AD, but in-depth proteome comparisons are lacking. We aimed to identify dysregulated CSF proteins in ADAD and determine the degree of overlap with sporadic AD. We measured 1472 proteins in CSF of PSEN1 or APP mutation carriers (n = 22) and age- and sex-matched controls (n = 20) from the Amsterdam Dementia Cohort using proximity extension-based immunoassays (PEA). We compared protein abundance between groups with two-sided t -tests and identified enriched biological pathways. Using the same protein panels in paired plasma samples, we investigated correlations between CSF proteins and their plasma counterparts. Finally, we compared our results with recently published PEA data from an international cohort of sporadic AD (n = 230) and non-AD dementias (n = 301). All statistical analyses were false discovery rate-corrected. We detected 66 differentially abundant CSF proteins (65 increased, 1 decreased) in ADAD compared to controls (q < 0.05). The most strongly upregulated proteins (fold change >1.8) were related to immunity (CHIT1, ITGB2, SMOC2), cytoskeletal structure (MAPT, NEFL) and tissue remodelling (TMSB10, MMP-10). Significant CSF-plasma correlations were found for the upregulated proteins SMOC2 and LILR1B. Of the 66 differentially expressed proteins, 36 had been measured previously in the sporadic dementias cohort, 34 of which (94%) were also significantly upregulated in sporadic AD, with a strong correlation between the fold changes of these proteins in both cohorts (rs = 0.730, P < 0.001). Twenty-nine of the 36 proteins (81%) were also upregulated among non-AD patients with suspected AD co-pathology. This CSF proteomics study demonstrates substantial biochemical similarities between ADAD and sporadic AD, suggesting involvement of the same biological processes. Besides known AD-related proteins, we identified several relatively novel proteins, such as TMSB10, MMP-10 and SMOC2, which have potential as novel biomarkers. With shared pathophysiological CSF changes, ADAD study findings might be translatable to sporadic AD, which could greatly expedite therapy development. [ABSTRACT FROM AUTHOR]

Background: Several assays reliably measure cerebrospinal fluid (CSF) total tau (t‐tau) protein for Alzheimer's diseases (AD) diagnosis. However, CSF sampling is invasive and hampers availability of biomarker testing in AD. Therefore, less invasive blood t‐tau assays are needed, preferably on different platforms to enable implementation in different clinical settings. Point‐of‐care (POC) platforms enable the development of practical and easily implemented assays. Here, we aim to develop and compare two blood t‐tau POC‐assays, (1)on the Ella microfluidics platform and (2)using upconversion‐nanoparticle based lateral flow (UCNP‐LF), for use in AD diagnosis. Method: The prototype assays were developed with the same pair of antibodies (capture‐mAb: ADx205 and detector‐mAb: ADx204 having epitopes in respectively the regions aa 194‐204 and 1‐20) and were calibrated using a recombinant protein (2N4R tau). For the Ella‐assay, we validated the analytical sensitivity, precision (4 different runs) and parallelism. The acceptance range was set to <15% coefficient of variation (CV) for precision and 85‐115% of slope‐accuracy for parallelism. Detectability of t‐tau was assessed using 20 plasma samples, including 10 AD‐dementia patients (age: 64±9y, 60%F) and 10 controls (age: 67±8y, 50%F). For the UCNP‐LF assay, spiked buffer and serum were used for preliminary performance testing. Result: For the Ella assay, we determined a limit of detection (LoD) of 3 pg/mL based on mean signal of 16 blanks plus 10x SD (figure1). The assay quantified t‐tau in the clinical samples above assay blank with an average intra‐assay CV of 5.3%. Average intra‐ and inter‐assay CV for QC plasma samples were 8% and 11.4%. The assay showed robust mean parallelism response of (95%) (figure 2). Median t‐tau concentration in AD samples was 36.9 pg/mL vs 8 pg/mL in controls. The prototype UCNP‐LF assay showed a LoD of 92 pg/mL, based upon spiked buffer with the recombinant protein, and 102 pg/mL for spiked serum (figure3). The UCNP‐LF assay has a read‐out time of 30 minutes. Conclusion: The prototype assays showed promising preliminary performance. Next, a technology comparison using Bland‐Altman graphs on samples of at least two different clinical groups, would be measured using a reference immunoassay platform, such as Simoa. [ABSTRACT FROM AUTHOR]

Diagnosis of Frontotemporal dementia (FTD) and the specific underlying neuropathologies (frontotemporal lobar degeneration; FTLD- Tau and FTLD-TDP) is challenging, and thus fluid biomarkers are needed to improve diagnostic accuracy. We used proximity extension assays to analyze 665 proteins in cerebrospinal fluid (CSF) samples from a multicenter cohort including patients with FTD (n = 189), Alzheimer's Disease dementia (AD; n = 232), and cognitively unimpaired individuals (n = 196). In a subset, FTLD neuropathology was determined based on phenotype or genotype (FTLD-Tau = 87 and FTLD-TDP = 68). Forty three proteins were differentially regulated in FTD compared to controls and AD, reflecting axon development, regulation of synapse assembly, and cell-cell adhesion mediator activity pathways. Classification analysis identified a 14- and 13-CSF protein panel that discriminated FTD from controls (AUC: 0.96) or AD (AUC: 0.91). Custom multiplex panels confirmed the highly accurate discrimination between FTD and controls (AUCs > 0.96) or AD (AUCs > 0.88) in three validation cohorts, including one with autopsy confirmation (AUCs > 0.90). Six proteins were differentially regulated between FTLD-TDP and FTLD-Tau, but no reproducible classification model could be generated (AUC: 0.80). Overall, this study introduces novel FTD-specific biomarker panels with potential use in diagnostic setting.

Introduction: Our previous antibody-based cerebrospinal fluid (CSF) proteomics study showed that Thimet oligopeptidase (THOP1), an amyloid beta (Aβ) neuropeptidase, was increased in mild cognitive impairment with amyloid pathology (MCI-Aβ+) and Alzheimer's disease (AD) dementia compared with controls and dementia with Lewy bodies (DLB), highlighting the potential of CSF THOP1 as an early specific biomarker for AD. We aimed to develop THOP1 immunoassays for large-scale analysis and validate our proteomics findings in two independent cohorts., Methods: We developed in-house CSF THOP1 immunoassays on automated Ella and Simoa platforms. The performance of the different assays were compared using Passing-Bablok regression analysis in a subset of CSF samples from the discovery cohort ( n = 72). Clinical validation was performed in two independent cohorts (cohort 1: n = 200; cohort 2: n = 165) using the Ella platform., Results: THOP1 concentrations moderately correlated between proteomics analysis and our novel assays ( Rho  > 0.580). In both validation cohorts, CSF THOP1 was increased in MCI-Aβ+ (>1.3-fold) and AD (>1.2-fold) compared with controls; and between MCI-Aβ+ and DLB (>1.2-fold). Higher THOP1 concentrations were detected in AD compared with DLB only when both cohorts were analyzed together. In both cohorts, THOP1 correlated with CSF total tau (t-tau), phosphorylated tau (p-tau), and Aβ40 ( Rho  > 0.540) but not Aβ42., Discussion: Validation of our proteomics findings underpins the potential of CSF THOP1 as an early specific biomarker associated with AD pathology. The use of antibody-based platforms in both the discovery and validation phases facilitated the translation of proteomics findings, providing an additional workflow that may accelerate the development of biofluid-based biomarkers., Competing Interests: Y.S.H., K.B., D.R., A.W.L., and M.C. report no conflicts of interest. D.A. participated in advisory boards from Fujirebio‐Europe and Roche Diagnostics and received speaker honoraria from Fujirebio‐Europe, Roche Diagnostics, Nutricia, Krka Farmacéutica S.L., Zambon S.A.U., and Esteve Pharmaceuticals S.A., D.A. declares a filed patent application (WO2019175379 A1 Markers of synaptopathy in neurodegenerative disease). A.L. participated in advisory boards from Fujirebio‐Europe, Grifols, Eisai, Novartis, Roche Diagnostics, Otsuka Pharmaceutical, Nutricia, Zambón S.A.U., and Biogen, and received speaker honoraria from Eli Lilly, Biogen, KRKA, and Zambon. A.L. declares a filed patent application (WO2019175379 A1 Markers of synaptopathy in neurodegenerative disease). Research programs of W.F. have been funded by ZonMW, NWO, EU‐FP7, EU‐JPND, Alzheimer Nederland, Hersenstichting CardioVascular Onderzoek Nederland, Health∼Holland, Topsector Life Sciences & Health, stichting Dioraphte, Gieskes‐Strijbis fonds, stichting Equilibrio, Edwin Bouw fonds, Pasman stichting, stichting Alzheimer & Neuropsychiatrie Foundation, Philips, Biogen MA Inc, Novartis‐NL, Life‐MI, AVID, Roche BV, Fujifilm, Eisai, and Combinostics. W.F. holds the Pasman chair. W.F. is recipient of ABOARD, which is a public‐private partnership receiving funding from ZonMW (#73305095007) and Health∼Holland, Topsector Life Sciences & Health (PPP‐allowance; #LSHM20106). W.F. has been an invited speaker at Biogen MA Inc, Danone, Eisai, WebMD Neurology (Medscape), NovoNordisk, Springer Healthcare, NovoNordisk, and European Brain Council. W.F. is a consultant to Oxford Health Policy Forum CIC, Roche, and Biogen MA Inc. W.F. participated in advisory boards of Biogen MA Inc, Roche, and Eli Lilly. All funding is paid to her institution. W.F. is member of the steering committee of PAVE and Think Brain Health. W.F. was associate editor of Alzheimer's Research & Therapy in 2020/2021. W.F. is associate editor at Brain. C.T. has a collaboration contract with ADx Neurosciences, Quanterix, and Eli Lilly, and performed contract research or received grants from AC‐Immune, Axon Neurosciences, Bioconnect, Bioorchestra, Brainstorm Therapeutics, Celgene, EIP Pharma, Eisai, Grifols, Novo Nordisk, PeopleBio, Roche, Toyama, and Vivoryon. C.T. serves on editorial boards of Medidact Neurologie/Springer, Alzheimer's Research & Therapy, and Neurology: Neuroimmunology & Neuroinflammation and is editor of a Neuromethods book (Springer). She had speaker contracts for Roche, Grifols, and Novo Nordisk. Author disclosures are available in the supporting information., (© 2023 The Authors. Alzheimer's & Dementia: Diagnosis, Assessment & Disease Monitoring published by Wiley Periodicals, LLC on behalf of Alzheimer's Association.)

The field of neurological diseases strongly needs biomarkers for early diagnosis and optimal stratification of patients in clinical trials or to monitor disease progression. Cerebrospinal fluid (CSF) is one of the main sources for the identification of novel protein biomarkers for neurological diseases. Despite the enormous efforts employed to identify novel CSF biomarkers, the high variability observed across different studies has hampered their validation and implementation in clinical practice. Such variability is partly caused by the effect of different pre-analytical confounding factors on protein stability, highlighting the importance to develop and comply with standardized operating procedures. In this chapter, we describe the international consensus pre-analytical guidelines for CSF processing and biobanking that have been established during the last decade, with a special focus on the influence of pre-analytical confounders on the global CSF proteome. In addition, we provide novel results on the influence of different delayed storage and freeze/thaw conditions on the CSF proteome using two novel large multiplex protein arrays (SOMAscan and Olink). Compliance to consensus guidelines will likely facilitate the successful development and implementation of CSF protein biomarkers in both research and clinical settings, ultimately facilitating the successful development of disease-modifying therapies.