Accurate diagnosis and monitoring of neurodegenerative diseases require reliable biomarkers. Cerebrospinal fluid (CSF) proteins are promising candidates for reflecting brain pathology; however, their diagnostic utility may be compromised by natural variability between individuals, weakening their association with disease. Here, we measured the levels of 69 pre-selected proteins in cerebrospinal fluid using antibody-based suspension bead array technology in a multi-disease cohort of 499 individuals with neurodegenerative disorders including Alzheimer’s disease (AD), behavioral variant frontotemporal dementia, primary progressive aphasias, amyotrophic lateral sclerosis (ALS), corticobasal syndrome, primary supranuclear palsy, along with healthy controls. We identify significant inter-individual variability in overall CSF levels of brain-derived proteins, which could not be attributed to specific disease associations. Using linear modelling, we show that adjusting for median CSF levels of brain-derived proteins increases the diagnostic accuracy of proteins previously identified as altered in CSF in the context of neurodegenerative disorders. We further demonstrate a simplified approach for the adjustment using pairs of correlated proteins with opposite alteration in the diseases. With this approach, the proteins adjust for each other and further increase the biomarker performance through additive effect. When comparing the diseases, two proteins—neurofilament medium and myelin basic protein—showed increased levels in ALS compared to other diseases, and neurogranin showed a specific increase in AD. Several other proteins showed similar trends across the studied diseases, indicating that these proteins likely reflect shared processes related to neurodegeneration. Overall, our findings suggest that accounting for inter-individual variability is crucial in future studies to improve the identification and performance of relevant biomarkers. Importantly, we highlight the need for multi-disease studies to identify disease-specific biomarkers.

Alzheimer's disease and related dementias have a multifactorial aetiology and heterogeneous biology. The current study aims to identify different biological signatures in a deeply phenotyped memory clinic patient population. In this cross-sectional study, we analysed 49 pre-specified proteins using a multiplex antibody-based suspension bead array in 278 CSF samples from the real-world research database and biobank at the Karolinska University Hospital Memory Clinic, Solna, Sweden. Patients with a clinical diagnosis of subjective cognitive decline (N = 151), mild cognitive impairment (N = 61), Alzheimer's disease (N = 47), or other diagnoses (N = 19; vascular dementias, alcohol-related dementia, unspecified dementias, or other amnesias) were included. Principal component analyses were performed, and resulting principal components (PCs) were tested for associations with clinical variables and Alzheimer's disease biomarkers (CSF biomarkers beta-amyloid 42, beta-amyloid 42/40, phosphorylated tau 181, phosphorylated tau 181/beta-amyloid 42). PC 1 (explaining 52% of the variance between patients) was associated with the clinical Alzheimer's disease CSF biomarkers beta-amyloid 42, phosphorylated tau 181, and total tau but not with Alzheimer's disease-related neurodegeneration imaging markers, cognitive performance, or clinical diagnosis. PC 2 (explaining 9% of the variance) displayed an inflammatory profile with high contributions of chitinase 3 like 1 (CHI3L1) and triggering receptor expressed on myeloid cells 2 (TREM2) and significant correlation to CSF free light chain kappa. In contrast to PC 1, PC 3 (explaining 5% of the variance) showed associations with all the clinical Alzheimer's disease CSF biomarkers, the imaging markers, cognitive impairment and clinical diagnosis. Serpin family A member 3 (SERPINA3), chitinase 1 (CHIT1), and neuronal pentraxin 2 (NPTX2) contributed most to PC 3. PC 4 (explaining 4% of the variance) exhibited an inflammatory profile distinct from PC 2, with the largest contributions from TREM2, leucine-rich alpha-2-glycoprotein 1 (LRG1) and complement C9. The component was associated with peripheral inflammation. We found that CSF protein profiles in a memory clinic cohort reflect molecular differences across diagnostic groups. Our results emphasize that real-world memory clinic patients can have different ongoing biological processes despite receiving the same diagnosis. In the future, this information could be utilized to identify patient endotypes and uncover precision biomarkers and novel therapeutic targets.

Background: Autoantibodies are found in up to 80 % of patients with idiopathic inflammatory myopathies (IIM) and are associated with distinct clinical phenotypes. Autoantibodies targeting cytosolic 5′-nucleotidase 1A (anti-NT5C1A) are currently the only known serum biomarker for the subgroup inclusion body myositis (IBM), although detected even in other autoimmune diseases. The aim of the study was to identify new autoimmune targets in IIM. Methods: In a first cross-sectional exploratory study, samples from 219 IIM (108 Polymyositis (PM), 80 Dermatomyositis (DM) and 31 IBM) patients, 349 Systemic Lupus Erythematosus (SLE) patients and 306 population controls were screened for IgG reactivity against a panel of 357 proteins using an antigen bead array. All samples were identified in the local biobank of the Rheumatology clinic, Karolinska University Hospital. Positive hits for the IBM subgroup were then validated in an independent larger cohort of 287 patients with IBM followed at nine European rheumatological or neurological centers. IBM serum samples were explored by antigen bead array and results validated by Western blot. As controls, sera from 29 patients with PM and 30 with DM, HLA-matched with the Swedish IBM cohort, were included. Demographics, laboratory, clinical, and muscle biopsy data of the IBM cohort was retrieved. Results: In the exploratory study, IgG reactivity towards NADH dehydrogenase 1 α subcomplex 11 (NDUFA11), a subunit of the membrane-bound mitochondrial respiratory chain complex I, was discovered with higher frequency in the IBM (9.7 %) than PM (2.8 %) and DM samples (1.3 %), although the difference was not statistically significant. Anti-NDUFA11 IgG was also found in 1.4 % of SLE and 2.0 % of population control samples. In the validation study, anti-NDUFA11 autoantibodies were detected in 10/287 IBM patients (3.5 %), 0/29 p.m. and 0/30 DM patients. Reactivity against NDUFA11 could be confirmed by Western blot. No statistically significant differences were found between patients with and without anti-NDUFA11 antibodies when comparing clinical, laboratory and histological data. However, we observed a trend of higher frequency of distal lower extremity muscle weakness, ragged red fibers and higher CK levels at time of diagnosis in the anti-NDUFA11 positive group. Co-existence of anti-NDUFA11 and anti-NT5C1A antibodies was not observed in any IBM patient. Conclusion: Our results reveal a new autoimmune target in the mitochondrial respiratory chain complex I that might be specifically associated with IBM. This is of particular interest as mitochondrial abnormalities are known histological findings in muscle biopsies of IBM patients.

Background: Amyloid and tau aggregates are considered to cause neurodegeneration and consequently cognitive decline in individuals with Alzheimer’s disease (AD). Here, we explore the potential of cerebrospinal fluid (CSF) proteins to reflect AD pathology and cognitive decline, aiming to identify potential biomarkers for monitoring outcomes of disease-modifying therapies targeting these aggregates. Method: We used a multiplex antibody-based suspension bead array to measure the levels of 49 proteins in CSF from the Swedish GEDOC memory clinic cohort at the Karolinska University Hospital. The cohort comprised 148 amyloid- and tau-negative individuals (A-T-) and 65 amyloid- and tau-positive individuals (A+T+). An independent sample set of 26 A-T- and 26 A+T+ individuals from the Amsterdam Dementia Cohort was used for validation. The measured proteins were clustered based on their correlation to CSF amyloid beta peptides, tau and NfL levels. Further, we used support vector machine modelling to identify protein pairs, matched based on their cluster origin, that reflect AD pathology and cognitive decline with improved performance compared to single proteins. Results: The protein-clustering revealed 11 proteins strongly correlated to t-tau and p-tau (tau-associated group), including mainly synaptic proteins previously found elevated in AD such as NRGN, GAP43 and SNCB. Another 16 proteins showed predominant correlation with Aβ42 (amyloid-associated group), including PTPRN2, NCAN and CHL1. Support vector machine modelling revealed that proteins from the two groups combined in pairs discriminated A-T- from A+T+ individuals with higher accuracy compared to single proteins, as well as compared to protein pairs composed of proteins originating from the same group. Moreover, combining the proteins from different groups in ratios (tau-associated protein/amyloid-associated protein) significantly increased their correlation to cognitive decline measured with cognitive scores. The results were validated in an independent cohort. Conclusions: Combining brain-derived proteins in pairs largely enhanced their capacity to discriminate between AD pathology-affected and unaffected individuals and increased their correlation to cognitive decline, potentially due to adjustment of inter-individual variability. With these results, we highlight the potential of protein pairs to monitor neurodegeneration and thereby possibly the efficacy of AD disease-modifying therapies.

Background: The cervicovaginal epithelial barrier is crucial for defending the female reproductive tract against sexually transmitted infections. Hormones, specifically estradiol and progesterone, along with their respective receptor expressions, play an important role in modulating this barrier. However, the influence of estradiol and progesterone on gene and protein expression in the ectocervical mucosa of naturally cycling women is not well understood. Methods: Mucosal and blood samples were collected from Kenyan female sex workers at high risk of sexually transmitted infections. All samples were obtained at two time points, separated by two weeks, aiming for the follicular and luteal phases of the menstrual cycle. Ectocervical tissue biopsies were analyzed by RNA-sequencing and in situ immunofluorescence staining, cervicovaginal lavage samples (CVL) were evaluated using protein profiling, and plasma samples were analyzed for hormone levels. Results: Unsupervised clustering of RNA-sequencing data was performed using Weighted gene co-expression network analysis (WGCNA). In the follicular phase, estradiol levels positively correlated with a gene module representing epithelial structure and function, and negatively correlated with a gene module representing cell cycle regulation. These correlations were confirmed using regression analysis including adjustment for bacterial vaginosis status. Using WGCNA, no gene module correlated with progesterone levels in the follicular phase. In the luteal phase, no gene module correlated with either estradiol or progesterone levels. Protein profiling on CVL revealed that higher levels of estradiol during the follicular phase correlated with increased expression of epithelial barrier integrity markers, including DSG1. This contrasted to the limited correlations of protein expression with estradiol levels in the luteal phase. In situ imaging analysis confirmed that higher estradiol levels during the follicular phase correlated with increased DSG1 expression. Conclusion: We demonstrate that estradiol levels positively correlate with specific markers of ectocervical epithelial structure and function, particularly DSG1, during the follicular phase of the menstrual cycle. Neither progesterone levels during the follicular phase nor estradiol and progesterone levels during the luteal phase correlated with any specific sets of gene markers. These findings align with the expression of estradiol and progesterone receptors in the ectocervical epithelium during these menstrual phases.

Objective: The objective of this study was to discover autoantibodies to non-modified proteins associated with the presence/absence of ACPAs in RA. Methods: The autoantibody repertoire of 80 ACPA-negative and 80 ACPA-positive RA subjects from the Swedish population-based Epidemiological Investigation of RA (EIRA) cohort was screened using a suspension bead array built on protein fragments earlier described as autoimmunity targets. Four autoantibodies positive in the initial screening were validated in another set of EIRA samples containing 317 ACPA-positive, 302 ACPA-negative and 372 age- and sex-matched controls. The relationship between the four autoantibodies and lung abnormalities on high-resolution CT (HRCT) was examined in 93 early-RA patients from the LURA cohort. Association between the autoantibodies, smoking and MHC class II alleles was assessed by logistic regression analysis. Results: Anti-ANOS1 and anti-MURC IgG levels were associated with ACPA-positive status [odds ratio (OR) = 3.02; 95% CI 1.87-4.89; and OR = 1.86; 95% CI 1.16-2.97, respectively] and increased in ACPA-positive patients compared with controls. Anti-ANOS1 IgG was associated with smoking habit (OR = 2.11; 95% CI 1.22-3.69) and anti-MURC IgG with the presence of the MHC class II 'shared-epitope' genes (OR = 1.95; 95% CI 1.11-3.46). Anti-TSPYL4 IgG was associated with being ACPA negative (OR = 0.41; 95% CI 0.19-0.89). Anti-TSPYL4 IgG and anti-MAP2K6 IgG levels were increased in the ACPA-negative patients compared with controls. Presence of anti-MAP2K6 IgG and anti-TSPYL4 IgG correlated negatively with HRCT-defined lung abnormalities. Conclusion: These four autoantibodies may be useful in diagnostics and in predicting clinical phenotypes of RA.

Autoantibodies have been shown to be implied in COVID-19 but the emerging autoantibody repertoire remains largely unexplored. We investigated the new-onset autoantibody repertoire in 525 healthcare workers and hospitalized COVID-19 patients at five time points over a 16-month period in 2020 and 2021 using proteome-wide and targeted protein and peptide arrays. Our results show that prevalent new-onset autoantibodies against a wide range of antigens emerged following SARS-CoV-2 infection in relation to pre-infectious baseline samples and remained elevated for at least 12 months. We found an increased prevalence of new-onset autoantibodies after severe COVID-19 and demonstrated associations between distinct new-onset autoantibodies and neuropsychiatric symptoms post-COVID-19. Using epitope mapping, we determined the main epitopes of selected new-onset autoantibodies, validated them in independent cohorts of neuro-COVID and pre-pandemic healthy controls, and identified sequence similarities suggestive of molecular mimicry between main epitopes and the conserved fusion peptide of the SARS-CoV-2 Spike glycoprotein. Our work describes the complexity and dynamics of the autoantibody repertoire emerging with COVID-19 and supports the need for continued analysis of the new-onset autoantibody repertoire to elucidate the mechanisms of the post-COVID-19 condition.

Introduction Serological surveys of the prevalence of SARS-CoV-2 are instrumental to understanding the course of the COVID-19 epidemic. We evaluate the seroprevalence of SARS-CoV-2 among young adult Finnish females residing in 25 communities all over Finland from 2020 until 2022. Methods Between 1st March 2020 and 30th June 2022, 3589 blood samples were collected from 3583 women born in 1992–95 when aged 25 or 28 years old attending the follow-up of an ongoing population-based trial of cervical screening strategies. The crude and population standardized SARS-CoV-2 seroprevalence was measured using nucleocapsid (induced by infection) and spike wild-type (WT) protein (induced both by infection and by vaccination) antigens over time and stratified by place of residence (inside or outside the Helsinki metropolitan region). Results During 2020 (before vaccinations), spike-WT and nucleocapsid IgG antibodies followed each other closely, at very low levels (<5%). Spike-WT seropositivity increased rapidly concomitant with mass vaccinations in 2021 and reached 96.3% in the 2nd quartile of 2022. Antibodies to nucleocapsid IgG remained relatively infrequent throughput 2020–2021, increasing rapidly in the 1st and 2nd quartiles of 2022 (to 19.7% and 56.6% respectively). The nucleocapsid IgG seropositivity increased more profoundly in participants residing in the Helsinki metropolitan region (4.5%, 8.4% and 43.9% in 2020, 2021 and 2022 respectively) compared to those residing in communities outside the capital region (4.5%, 4.3% and 34.7%). Conclusions Low SARS-CoV-2 infection-related seroprevalence during 2020–2021 suggest a comparatively successful infection control. Antibodies to the SARS-CoV-2 WT spike protein became extremely common among young women by the end of 2021, in line with the high uptake of SARS-CoV-2 vaccination. Finally, the rapid increase of seroprevalences to the SARS-CoV-2 nucleocapsid protein during the first and second quartile of 2022, imply a high incidence of infections with SARS-CoV-2 variants able to escape vaccine-induced protection.

Background: Plasma biomarkers reflecting the pathology of frontotemporal dementia would add significant value to clinical practice, to the design and implementation of treatment trials as well as our understanding of disease mechanisms. The aim of this study was to explore the levels of multiple plasma proteins in individuals from families with genetic frontotemporal dementia. Methods: Blood samples from 693 participants in the GENetic Frontotemporal Dementia Initiative study were analysed using a multiplexed antibody array targeting 158 proteins. Results: We found 13 elevated proteins in symptomatic mutation carriers, when comparing plasma levels from people diagnosed with genetic FTD to healthy non-mutation controls and 10 proteins that were elevated compared to presymptomatic mutation carriers. Conclusion: We identified plasma proteins with altered levels in symptomatic mutation carriers compared to non-carrier controls as well as to presymptomatic mutation carriers. Further investigations are needed to elucidate their potential as fluid biomarkers of the disease process.