Plasma p-tau212 antemortem diagnostic performance and prediction of autopsy verification of Alzheimer's disease neuropathology.

Blood phosphorylated tau (p-tau) biomarkers, including p-tau217, show high associations with Alzheimer's disease (AD) neuropathologic change and clinical stage. Certain plasma p-tau217 assays recognize tau forms phosphorylated additionally at threonine-212, but the contribution of p-tau212 alone to AD is unknown. We developed a blood-based immunoassay that is specific to p-tau212 without cross-reactivity to p-tau217. Here, we examined the diagnostic utility of plasma p-tau212. In five cohorts (n = 388 participants), plasma p-tau212 showed high performances for AD diagnosis and for the detection of both amyloid and tau pathology, including at autopsy as well as in memory clinic populations. The diagnostic accuracy and fold changes of plasma p-tau212 were similar to those for p-tau217 but higher than p-tau181 and p-tau231. Immunofluorescent staining of brain tissue slices showed prominent p-tau212 reactivity in neurofibrillary tangles that co-localized with p-tau217 and p-tau202/205. These findings support plasma p-tau212 as a peripherally accessible biomarker of AD pathophysiology.

The Association of Selected GWAS Reported AD Risk Loci with CSF Biomarker Levels and Cognitive Decline in Slovenian Patients.

Alzheimer's disease (AD) is the most common neurodegenerative disease, with a complex genetic background. Apart from rare, familial cases, a combination of multiple risk loci contributes to the susceptibility of the disease. Genome-wide association studies (GWAS) have identified numerous AD risk loci. Changes in cerebrospinal fluid (CSF) biomarkers and imaging techniques can detect AD-related brain changes before the onset of clinical symptoms, even in the presence of preclinical mild cognitive impairment. In this study, we aimed to assess the associations between SNPs in well-established GWAS AD risk loci and CSF biomarker levels or cognitive test results in Slovenian patients with cognitive decline. The study included 82 AD patients, 28 MCI patients with pathological CSF biomarker levels and 35 MCI patients with normal CSF biomarker levels. Carriers of at least one polymorphic TOMM40 rs157581 C allele had lower Aß42 (p = 0.033) and higher total tau (p = 0.032) and p-tau181 levels (p = 0.034). Carriers of at least one polymorphic T allele in SORCS1 rs1358030 had lower total tau (p = 0.019), while polymorphic SORCS1 rs1416406 allele was associated with lower total tau (p = 0.013) and p-tau181 (p = 0.036). In addition, carriers of at least one polymorphic T allele in BCHE rs1803274 had lower cognitive test scores (p = 0.029). The study findings may contribute to the identification of genetic markers associated with AD and MCI and provide insights into early disease diagnostics.

Genetic Polymorphisms in Oxidative Stress and Inflammatory Pathways as Potential Biomarkers in Alzheimer's Disease and Dementia.

Oxidative stress and neuroinflammation are important processes involved in Alzheimer's disease (AD) and mild cognitive impairment (MCI). Numerous risk factors, including genetic background, can affect the complex interplay between those mechanisms in the aging brain and can also affect typical AD hallmarks: amyloid plaques and neurofibrillary tangles. Our aim was to evaluate the association of polymorphisms in oxidative stress- and inflammation-related genes with cerebrospinal fluid (CSF) biomarker levels and cognitive test results. The study included 54 AD patients, 14 MCI patients with pathological CSF biomarker levels, 20 MCI patients with normal CSF biomarker levels and 62 controls. Carriers of two polymorphic IL1B rs16944 alleles had higher CSF Aß1-42 levels (p = 0.025), while carriers of at least one polymorphic NFE2L2 rs35652124 allele had lower CSF Aß1-42 levels (p = 0.040). Association with IL1B rs16944 remained significant in the AD group (p = 0.029). Additionally, MIR146A rs2910164 was associated with Aß42/40 ratio (p = 0.043) in AD. Significant associations with cognitive test scores were observed for CAT rs1001179 (p = 0.022), GSTP1 rs1138272 (p = 0.005), KEAP1 rs1048290 and rs9676881 (both p = 0.019), as well as NFE2L2 rs35652124 (p = 0.030). In the AD group, IL1B rs1071676 (p = 0.004), KEAP1 rs1048290 and rs9676881 (both p = 0.035) remained associated with cognitive scores. Polymorphisms in antioxidative and inflammation genes might be associated with CSF biomarkers and cognitive test scores and could serve as additional biomarkers contributing to early diagnosis of dementia.

Plasma p-tau212: antemortem diagnostic performance and prediction of autopsy verification of Alzheimer's disease neuropathology.

Blood phosphorylated tau (p-tau) biomarkers, including p-tau217, show high associations with Alzheimer's disease (AD) neuropathologic change and clinical stage. Certain plasma p-tau217 assays recognize tau forms phosphorylated additionally at threonine-212, but the contribution of p-tau212 alone to AD is unknown. We developed a blood-based immunoassay that is specific to p-tau212 without cross-reactivity to p-tau217. Thereafter, we examined the diagnostic utility of plasma p-tau212. In five cohorts (n=388 participants), plasma p-tau212 showed high performances for AD diagnosis and for the detection of both amyloid and tau pathology, including at autopsy as well as in memory clinic populations. The diagnostic accuracy and fold changes of plasma p-tau212 were similar to those for p-tau217 but higher than p-tau181 and p-tau231. Immunofluorescent staining of brain tissue slices showed prominent p-tau212 reactivity in neurofibrillary tangles that co-localized with p-tau217 and p-tau202/205. These findings support plasma p-tau212 as a novel peripherally accessible biomarker of AD pathophysiology.

Diagnostic value of kappa free light chain index in patients with primary progressive multiple sclerosis - a multicentre study.

Background: Kappa free light chains (κ-FLC) in the cerebrospinal fluid (CSF) are an emerging biomarker in multiple sclerosis (MS). Objective: To investigate whether κ-FLC index has similar diagnostic value in patients with primary progressive multiple sclerosis (PPMS) compared to oligoclonal bands (OCB). Methods: Patients with PPMS were recruited through 11 MS centres across 7 countries. κ-FLC were measured by immunonephelometry/-turbidimetry. OCB were determined by isoelectric focusing and immunofixation. Results: A total of 174 patients (mean age of 52±11 years, 51% males) were included. κ-FLC index using a cut-off of 6.1 was positive in 161 (93%) and OCB in 153 (88%) patients. Conclusion: κ-FLC index shows similar diagnostic sensitivity than OCB in PPMS.

Identification and validation of Alzheimer's disease-related metabolic brain pattern in biomarker confirmed Alzheimer's dementia patients.

Metabolic brain biomarkers have been incorporated in various diagnostic guidelines of neurodegenerative diseases, recently. To improve their diagnostic accuracy a biologically and clinically homogeneous sample is needed for their identification. Alzheimer's disease-related pattern (ADRP) has been identified previously in cohorts of clinically diagnosed patients with dementia due to Alzheimer's disease (AD), meaning that its diagnostic accuracy might have been reduced due to common clinical misdiagnosis. In our study, we aimed to identify ADRP in a cohort of AD patients with CSF confirmed diagnosis, validate it in large out-of-sample cohorts and explore its relationship with patients' clinical status. For identification we analyzed 2-[18F]FDG PET brain scans of 20 AD patients and 20 normal controls (NCs). For validation, 2-[18F]FDG PET scans from 261 individuals with AD, behavioral variant of frontotemporal dementia, mild cognitive impairment and NC were analyzed. We identified an ADRP that is characterized by relatively reduced metabolic activity in temporoparietal cortices, posterior cingulate and precuneus which co-varied with relatively increased metabolic activity in the cerebellum. ADRP expression significantly differentiated AD from NC (AUC = 0.95) and other dementia types (AUC = 0.76-0.85) and its expression correlated with clinical measures of global cognition and neuropsychological indices in all cohorts.

N-terminal and mid-region tau fragments as fluid biomarkers in neurological diseases.

Brain-derived tau secreted into CSF and blood consists of different N-terminal and mid-domain fragments, which may have a differential temporal course and thus, biomarker potential across the Alzheimer's disease continuum or in other neurological diseases. While current clinically validated total tau assays target mid-domain epitopes, comparison of these assays with new biomarkers targeting N-terminal epitopes using the same analytical platform may be important to increase the understanding of tau pathophysiology. We developed three total tau immunoassays targeting specific N-terminal (NTA and NTB total tau) or mid-region (MR total tau) epitopes, using single molecule array technology. After analytical validation, the diagnostic performance of these biomarkers was evaluated in CSF and compared with the Innotest total tau (and as proof of concept, with N-p-tau181 and N-p-tau217) in three clinical cohorts (n = 342 total). The cohorts included participants across the Alzheimer's disease continuum (n = 276), other dementias (n = 22), Creutzfeldt-Jakob disease (n = 24), acute neurological disorders (n = 18) and progressive supranuclear palsy (n = 22). Furthermore, we evaluated all three new total tau biomarkers in plasma (n = 44) and replicated promising findings with NTA total tau in another clinical cohort (n = 50). In CSF, all total tau biomarkers were increased in Alzheimer's disease compared with controls (P < 0.0001) and correlated with each other (rs = 0.53-0.95). NTA and NTB total tau, but not other total tau assays, distinguished amyloid-positive and amyloid-negative mild cognitive impairment with high accuracies (AUCs 84% and 82%, P < 0.001) matching N-p-tau217 (AUC 83%; DeLong test P = 0.93 and 0.88). All total tau assays were excellent in differentiating Alzheimer's disease from other dementias (P < 0.001, AUCs 89-100%). In Creutzfeldt-Jakob disease and acute neurological disorders, N-terminal total tau biomarkers had significantly higher fold changes versus controls in CSF (45-133-fold increase) than Innotest or MR total tau (11-42-fold increase, P < 0.0001 for all). In progressive supranuclear palsy, CSF concentrations of all total tau biomarkers were similar to those in controls. Plasma NTA total tau concentrations were increased in Alzheimer's disease compared with controls in two independent cohorts (P = 0.0056 and 0.0033), while Quanterix total tau performed poorly (P = 0.55 and 0.44). Taken together, N-terminal-directed CSF total tau biomarkers increase ahead of standard total tau alternatives in the Alzheimer's disease continuum, increase to higher degrees in Creutzfeldt-Jakob disease and acute neurological diseases and show better potential than Quanterix total tau as Alzheimer's disease blood biomarkers. For progressive supranuclear palsy, other tau biomarkers should continue to be investigated.

Humoral immune response in convalescent COVID-19 people with multiple sclerosis treated with high-efficacy disease-modifying therapies: A multicenter, case-control study.

AIM: To determine the influence of high-efficacy disease modifying therapy (DMT) on the development of IgG SARS-CoV-2 antibody response in COVID-19 convalescent people with multiple sclerosis (pwMS). METHODS: Seventy-four pwMS taking high-efficacy DMTs (specifically natalizumab, fingolimod, alemtuzumab, ocrelizumab, cladribine and ublituximab) and diagnosed with COVID-19 and 44 healthy persons (HC) were enrolled. SARS-CoV2 antibodies were tested with Elecsys® Anti-SARSCoV-2 S assay. RESULTS: pwMS taking high-efficacy DMTs had a significantly higher chance of having negative titer of SARS-CoV2 antibodies compared to healthy controls (33 negative pwMS [44.6%] compared to one negative HC [2.3%], p < 0.001). pwMS taking B-cell depleting therapy (ocrelizumab and ublituximab) had a significantly higher chance of having negative titer of SARS-CoV2 antibodies compared to pwMS on all other DMTs (29 negative pwMS on B-cell therapy [64.4%] compared to four negative pwMS on all other DMTs [13.8%], p < 0.001). Out of other DMTs, two (33.3%) pwMS taking fingolimod and two (16.7%) pwMS taking cladribine failed to develop IgG SARS-COV-2 antibodies. B-cell depleting therapy independently predicted negative titer of IgG SARS-CoV-2 antibody (Exp[B] =0.014, 95%CI 0.002-0.110, p < 0.001). CONCLUSIONS: A significant proportion of convalescent COVID-19 pwMS on high-efficacy DMTs will not develop IgG SARS-CoV-2 antibodies. B-cell depleting therapies independently predict negative and low titer of IgG SARS-CoV-2 antibody.

Head-to-head comparison of clinical performance of CSF phospho-tau T181 and T217 biomarkers for Alzheimer's disease diagnosis.

INTRODUCTION: Phosphorylated tau (p-tau) in cerebrospinal fluid (CSF) is an established Alzheimer's disease (AD) biomarker. Novel immunoassays targeting N-terminal and mid-region p-tau181 and p-tau217 fragments are available, but head-to-head comparison in clinical settings is lacking. METHODS: N-terminal-directed p-tau217 (N-p-tau217), N-terminal-directed p-tau181 (N-p-tau181), and standard mid-region p-tau181 (Mid-p-tau181) biomarkers in CSF were evaluated in three cohorts (n = 503) to assess diagnostic performance, concordance, and associations with amyloid beta (Aß). RESULTS: CSF N-p-tau217 and N-p-tau181 had better concordance (88.2%) than either with Mid-p-tau181 (79.7%-82.7%). N-p-tau217 and N-p-tau181 were significantly increased in early mild cognitive impairment (MCI)-AD (A+T-N-) without changes in Mid-p-tau181 until AD-dementia. N-p-tau217 and N-p-tau181 identified Aß pathophysiology (area under the curve [AUC] = 94.8%-97.1%) and distinguished MCI-AD from non-AD MCI (AUC = 82.6%-90.5%) signficantly better than Mid-p-tau181 (AUC = 91.2% and 70.6%, respectively). P-tau biomarkers equally differentiated AD from non-AD dementia (AUC = 99.1%-99.8%). DISCUSSION: N-p-tau217 and N-p-tau181 could improve diagnostic accuracy in prodromal-AD and clinical trial recruitment as both identify Aß pathophysiology and differentiate early MCI-AD better than Mid-p-tau181.

Serum Glial Fibrillary Acidic Protein (GFAP) Is a Marker of Disease Severity in Frontotemporal Lobar Degeneration.

BACKGROUND: It is still unknown if serum glial fibrillary acidic protein (GFAP) is a useful marker in frontotemporal lobar degeneration (FTLD). OBJECTIVE: To assess the diagnostic and prognostic value of serum GFAP in a large cohort of patients with FTLD. METHODS: In this retrospective study, performed on 406 participants, we measured serum GFAP concentration with an ultrasensitive Single molecule array (Simoa) method in patients with FTLD, Alzheimer's disease (AD), and in cognitively unimpaired elderly controls. We assessed the role of GFAP as marker of disease severity by analyzing the correlation with clinical variables, neurophysiological data, and cross-sectional brain imaging. Moreover, we evaluated the role of serum GFAP as a prognostic marker of disease survival. RESULTS: We observed significantly higher levels of serum GFAP in patients with FTLD syndromes, except progressive supranuclear palsy, compared with healthy controls, but not compared with AD patients. In FTLD, serum GFAP levels correlated with measures of cognitive dysfunction and disease severity, and were associated with indirect measures of GABAergic deficit. Serum GFAP concentration was not a significant predictor of survival. CONCLUSION: Serum GFAP is increased in FTLD, correlates with cognition and GABAergic deficits, and thus shows promise as a biomarker of disease severity in FTLD.

Diagnostic and prognostic value of serum NfL and p-Tau181 in frontotemporal lobar degeneration.

OBJECTIVE: To assess the diagnostic and prognostic value of serum neurofilament light (NfL) and serum phospho-Tau181 (p-Tau181) in a large cohort of patients with frontotemporal lobar degeneration (FTLD). METHODS: In this retrospective study, performed on 417 participants, we analysed serum NfL and p-Tau181 concentrations with an ultrasensitive single molecule array (Simoa) approach. We assessed the diagnostic values of serum biomarkers in the differential diagnosis between FTLD, Alzheimer's disease (AD) and healthy ageing; their role as markers of disease severity assessing the correlation with clinical variables, cross-sectional brain imaging and neurophysiological data; their role as prognostic markers, considering their ability to predict survival probability in FTLD. RESULTS: We observed significantly higher levels of serum NfL in patients with FTLD syndromes, compared with healthy controls, and lower levels of p-Tau181 compared with patients with AD. Serum NfL concentrations showed a high accuracy in discriminating between FTLD and healthy controls (area under the curve (AUC): 0.86, p<0.001), while serum p-Tau181 showed high accuracy in differentiating FTLD from patients with AD (AUC: 0.93, p<0.001). In FTLD, serum NfL levels correlated with measures of cognitive function, disease severity and behavioural disturbances and were associated with frontotemporal atrophy and indirect measures of GABAergic deficit. Moreover, serum NfL concentrations were identified as the best predictors of survival probability. CONCLUSIONS: The assessment of serum NfL and p-Tau181 may provide a comprehensive view of FTLD, aiding in the differential diagnosis, in staging disease severity and in defining survival probability.

Plasma p-tau181 accurately predicts Alzheimer's disease pathology at least 8 years prior to post-mortem and improves the clinical characterisation of cognitive decline.

The neuropathological confirmation of amyloid-ß (Aß) plaques and tau neurofibrillary tangles (NFT) remains the gold standard for a definitive diagnosis of Alzheimer's disease (AD). Nowadays, the in vivo diagnosis of AD is greatly aided by both cerebrospinal fluid (CSF) and positron emission tomography (PET) biomarkers. Although highly accurate, their broad implementation is restricted by high cost, limited accessibility and invasiveness. We recently developed a high-performance, ultrasensitive immunoassay for the quantification of tau phosphorylated at threonine-181 (p-tau181) in plasma, which identifies AD pathophysiology with high accuracy. However, it remains unclear whether plasma p-tau181, measured years before the death, can predict the eventual neuropathological confirmation of AD, and successfully discriminates AD from non-AD dementia pathologies. We studied a unique cohort of 115 individuals with longitudinal blood collections with clinical evaluation at 8, 4 and 2 years prior to neuropathological assessment at death. The results demonstrate that plasma p-tau181 associates better with AD neuropathology and Braak staging than a clinical diagnosis 8 years before post-mortem. Moreover, while all patients had a diagnosis of AD dementia during life, plasma p-tau181 proved to discriminate AD from non-AD pathologies with high accuracy (AUC = 97.4%, 95% CI = 94.1-100%) even 8 years before death. Additionally, the longitudinal trajectory of plasma p-tau181 was assessed in all patients. We found that the main increases in plasma p-tau181 occurred between 8 and 4 years prior to death in patients with AD neuropathology and later plateauing. In contrast, non-AD pathologies and controls exhibited minor, albeit significant, increases in p-tau181 up until death. Overall, our study demonstrates that plasma p-tau181 is highly predictive and specific of AD neuropathology years before post-mortem examination. These data add further support for the use of plasma p-tau181 to aid clinical management in primary care and recruitment for clinical trials.

Intrathecal immunoglobulin synthesis: The potential value of an adjunct test.

BACKGROUND: Detection of cerebrospinal fluid (CSF) specific oligoclonal bands (OCB) supports the diagnosis of multiple sclerosis (MS), but the method is technically demanding and gives only qualitative information. Kappa free light chains (KFLC) quantification could represent a convenient alternative. We evaluated the diagnostic accuracy of OCB and KFLC in our cohort to further estimate the gain in diagnostic performance when combining both of them. METHODS: KFLC were measured in paired serum and CSF samples of 80 patients with MS and 50 patients with non-inflammatory neurological disorders. OCB were detected using an in-house alkaline phosphatase assay. Likelihood ratio (LR) was used to explore the benefit of the combined KFLC and OCB test. RESULTS: Sensitivity of KFLC index (≥5.3) and intrathecal KFLC fraction (≥10%) was 96% and 95% respectively, compared to 91% sensitivity of OCB assay. Specificity was 96% for intrathecal KFLC synthesis and 98% for OCB. Probability of MS in the absence of OCB was further reduced with concurrently normal KFLC index. CONCLUSIONS: Normal KFLC parameters allow confident exclusion of intrathecal inflammation, but probability of MS is greater with positive OCB. Use of KFLC as an adjunct test might be beneficial in specialized MS centers with larger pretest probability.

Validation of a quantitative cerebrospinal fluid alpha-synuclein assay in a European-wide interlaboratory study.

Decreased levels of alpha-synuclein (aSyn) in cerebrospinal fluid (CSF) in Parkinson's disease and related synucleinopathies have been reported, however, not consistently in all cross-sectional studies. To test the performance of one recently released human-specific enzyme-linked immunosorbent assay (ELISA) for the quantification of aSyn in CSF, we carried out a round robin trial with 18 participating laboratories trained in CSF ELISA analyses within the BIOMARKAPD project in the EU Joint Program - Neurodegenerative Disease Research. CSF samples (homogeneous aliquots from pools) and ELISA kits (one lot) were provided centrally and data reported back to one laboratory for data analysis. Our study showed that although factors such as preanalytical sample handling and lot-to-lot variability were minimized by our study design, we identified high variation in absolute values of CSF aSyn even when the same samples and same lots of assays were applied. We further demonstrate that although absolute concentrations differ between laboratories the quantitative results are comparable. With further standardization this assay may become an attractive tool for comparing aSyn measurements in diverse settings. Recommendations for further validation experiments and improvement of the interlaboratory results obtained are given.