Search

Background: The lack of early molecular biomarkers in sporadic behavioural variant of frontotemporal dementia (bvFTD) and its clinical overlap with primary psychiatric disorders (PPD) and Alzheimer's Disease (AD) hampers its diagnostic distinction. Synaptic dysfunction is an early feature in bvFTD and identification of specific biomarkers might improve its diagnostic accuracy. We aimed to assess the diagnostic performance of synaptic biomarkers in bvFTD compared to PPD, AD and controls, and its association with social cognition, cognitive performance and disease severity. Method: Participants with probable bvFTD (n = 60), PPD (n = 60), biomarker confirmed AD (n = 60), and cognitively normal AD biomarker negative controls (n = 58) with available CSF, (social) cognitive tests, FTLD‐CDR, and Ekman Faces Test score were included (Table 1). CSF synaptosomal associated protein 25 (SNAP25), neurogranin (Ng), neuronal pentraxin 2 (NPTX2) were measured, along with neurofilament light (NfL), and compared between groups using regression models adjusted for age and sex. Diagnostic accuracy was assessed using ROC analysis and Spearman's correlation analysis for correlation with cognitive measures was performed. Result: NPTX2 was significantly lower in bvFTD patients compared to PPD and controls (P<0.001). NfL was higher in bvFTD patients compared to PPD, AD and controls (P<0.001) and showed the highest diagnostic accuracy between bvFTD and PPD (AUC: 0.95, CI: 0.91,0.99, sensitivity: 97% specificity: 82%). Between bvFTD versus AD, SNAP25 showed the highest diagnostic accuracy (AUC: 0.78 CI: 0.70, 0.86, sensitivity: 63% specificity: 83%). The selected biomarker panel to differentiate bvFTD from PPD consisted of NfL and NPTX2 and had the highest AUC among the measured biomarkers (AUC: 0.96, CI: 0.93, 0.99, sensitivity: 95%, specificity: 85%), while that for bvFTD and AD consisted of NfL and Ng (AUC: 0.82, CI: 0.75, 0.90, sensitivity: 63% specificity: 92%). Among cognitive scores of bvFTD patients, SNAP25 correlated significantly with visuospatial ability (r = 0.37, P<0.05). There was no correlation between synaptic markers and the Ekman 60 faces test or FTLD‐CDR scores of disease severity. Conclusion: Together with NfL, synaptic biomarkers NPTX2 and Ng have added value for the differential diagnosis of bvFTD versus PPD and AD respectively. However, there was almost no association of the CSF biomarkers with cognitive performance of bvFTD and PPD patients. [ABSTRACT FROM AUTHOR]

Background: Recent studies reported the potential use of elevated CSF β‐synuclein levels as an AD‐specific synaptic biomarker. These results were obtained with a sandwich ELISA where one of the antibodies recognizes both alpha‐and β‐synuclein1. Blood contamination can affect alpha‐synuclein concentrations in CSF, which is not observed for β‐synuclein2. Therefore, we aimed to develop a β‐synuclein‐specific assay to avoid possible interference with alpha‐synuclein. Methods: Two novel sandwich CSF ELISA assays were established using only β‐specific synuclein monoclonal antibodies: a mid‐region and C‐terminal assay. Both assays use a monoclonal capture antibody that maps to C‐terminal. The mid‐region assay involves a detection antibody mapping to an epitope that overlaps with the SRM peptide used for CSF mass‐spectrometry quantification3. The C‐terminal assay includes a detection antibody targeting a less well‐defined C‐terminal epitope. Forty‐four remnant routine CSF samples were analyzed in duplicate (22 individuals with AD biomarker profile based on pTau181 and amyloid‐beta(1‐42), age70±7 years, 38%F and 22 control profiles, age70±7, 38%F). Results: The mid‐region assay showed a precision of 4%CV in clinical samples and a sensitivity of 1.25 pg/ml compared to 10%CV and 4.22 pg/ml with the C‐terminal assay (Fig. 1). CSF β‐synuclein levels were significantly elevated in patients with an AD CSF‐profile compared to individuals with negative‐AD CSF‐profiles, with AUCs of 0.77 [95%CI: 0.62‐0.90] for the C‐terminal assay and 0.71 [0.55‐0.87] for the mid‐region assay (Fig. 2). The assays showed comparable (Delong's P = 0.53) clinical performance and their results correlated (r = 0.61, P = <0.001)(Fig. 2). Both C‐terminal and mid‐region β‐synuclein associated with CSF pTau181 (Spearman's rho = 0.58 (p<0.001), rho = 0.51 (P = <0.001), respectively), and did not correlate with CSF Abeta(1‐42) (Fig. 3). Both assays are specific for recombinant beta‐synuclein, but the C‐terminal assay showed higher selectivity for β‐synuclein compared to the mid‐region assay, as increasing signals were observed with the latter with increasing concentrations of spiked recombinant alpha/gamma‐synuclein. Conclusion: The two novel β‐synuclein assays show clinically relevant changes in relation to AD CSF biomarker positivity. Both assays are specific to β‐synuclein. The assays will be transferred to SIMOA, which provides a wider dynamic range, and thus we will further assess the selectivity in plasma/serum. [ABSTRACT FROM AUTHOR]

Background: A hallmark of neurodegenerative disorders is synaptic dysfunction and degeneration. This makes the presynaptic protein synaptosomal‐associated protein 25 (SNAP‐25) and the Vesicle‐associated membrane protein 2 (VAMP‐2) targets of interest and importance. Previously, these proteins have only been measurable in cerebrospinal fluid (CSF) and are increased in Alzheimer's disease (AD) patients. Considering the advancement of blood‐based detection of phosphorylated tau (p‐tau), neurofilament light (NfL) and amyloid‐β (Aβ42/40), a measure of synaptic degeneration in blood would be important for disease prognosis and clinical trial outcome. In this study, we describe results from novel single molecular arrays (Simoa) for blood SNAP‐25 and VAMP‐2. Method: Prototype Simoa assays for N‐terminal SNAP‐25 and VAMP‐2 were developed as more sensitive versions of previously validated in‐house CSF assays with diluent suited for both plasma and CSF. We examined 110 participants (54 AD and 56 healthy controls) from the Swedish BioFINDER pilot study. The mean age of patients was 75.9 years (7.1 SD) and there was an even distribution of sex. We examined group differences by the Wilcoxon rank sum test and associations between these two blood biomarkers and Aβ‐PET, tau‐PET, MRI and MMSE were determined by linear regression, adjusting for age and sex. Result: SNAP‐25 was significantly increased in patients with AD (mean [SD], 0.81 pg/mL [0.27]; p < 0.001) as compared to controls (0.60 pg/mL [0.22]). SNAP‐25 distinguished AD patients from controls with an area under the curve of 0.74 (95% CI = 0.65‐0.83). SNAP‐25 showed association with Aβ PET (β = 0.10, p = 0.007) but not tau PET. SNAP‐25 associated with cortical thickness (β = ‐0.05, p = 0.009) and MMSE (β = ‐6.69, p = 0.001). No significant associations were found when analysing the AD group alone. VAMP‐2 showed no significant association with any of the mentioned parameters above. Conclusion: This novel ultra‐sensitive immunoassay demonstrates an increase of plasma SNAP‐25 in AD patients, and a significant relationship with cognition and cortical atrophy. Further studies will verify these changes in independent cohorts and explore the relationship between plasma SNAP‐25 and VAMP‐2 with synaptic vesicle glycoprotein 2A (SV2A) PET in a broad range of neurodegenerative disorders. [ABSTRACT FROM AUTHOR]

Background: Incorporation of biomarkers into the diagnosis of AD has shifted its disease definition from clinical dementia (on average 7 years) into a biological change that may span two decades of a human life. Synaptic biomarkers and neurofilament light (NfL) have shown some potential to predict stage and disease course, however few studies have combined these markers in larger sample sets. Method: Negative controls, preclinical subjects (n=90), and subjects with MCI (n=57) and mild AD (n=46) from the longitudinal observational study at the UCSD Shirley‐Marcos ADRC (UCSD cohort) were tested for their biomarker levels. One molecular/synaptic marker, neuronal pentraxin 2 (NPTX2), is quantified in CSF with an ELISA containing two monoclonal antibodies (mAbs). We describe a newly developed NfL assay based on two mAbs that specifically targets the C‐terminus of coil 2a from human NfL. A third marker is synaptobrevin 2 (VAMP2), an essential component of the synaptic vesicle. Classical biomarkers such as t‐tau, β‐amyloid42 and synaptic markers SNAP25 and neurogranin were available for this sample set. Result: As expected, CSF NfL levels increase gradually over the course of the disease, while VAMP2, as well as SNAP25 and neurogranin, shows a significant change in a stage of AD characterized by low Aβ42 and normal CSF tau and CSF NfL levels, thus suggesting that synaptic changes may precede overt neuronal (axonal) damage. CSF NPTX2 is a robust biochemical reflection of cognitive decline, as defined by California Verbal Learning Tests and Mattis Dementia Rating Scale and in contrast to all other fluid markers, it is not age‐dependent in control individuals. Conclusion: Exploring all these biochemical markers in observational longitudinal studies suggests changes might delineate different stages in the disease process: (1) intracellular synaptic markers such as VAMP2, SNAP25 and neurogranin precede neuronal degeneration as defined by increased CSF tau and CSF NfL levels, while NPTX2 only begins to decrease towards late clinical stages, showing a robust correlation to cognitive decline. When further verified and/or validated in other well‐designed longitudinal studies, such prognostic biomarkers might define specific stages and predict the onset of specific pathophysiological progression during the long course of the disease. [ABSTRACT FROM AUTHOR]

Background: An early objective cerebrospinal fluid (CSF) marker of cognitive decline in preclinical Alzheimer's disease (AD) would be a useful addition to the current AD biomarker arsenal. We recently identified VAMP2 as a potential CSF marker of AD‐related synapse loss. The aim of this study was 1) to evaluate the regional brain expression of VAMP2 in human postmortem tissue and 2) to assess the correlation of CSF VAMP2 levels with cognitive performance in cognitively normal controls and AD patients. Method: Paraffin fixed brain sections from hippocampus, frontal, posterior‐cingulate and temporal inferior cortices from AD cases (n=10) and non‐pathological controls (n=10) were stained with anti‐VAMP2 (Cell Signaling) and an HRP‐conjugated (Dako) secondary antibody. VAMP2 was quantified using MATLAB software (MA, USA). CSF from the Sant Pau Initiative in Neurodegeneration (SPIN) cohort was selected from cognitively normal controls (n=68), preclinical AD stages 1 (n=31) and 2 (n=8), prodromal AD (n=80) and AD dementia patients (n=39). Cognitive performance was evaluated using the total score in the Free and Cued Selective Reminding Test (Buschke et al). We quantified VAMP2 in CSF by Single MOlecule Array (SIMOA). Result: In control and AD brains, VAMP2 showed a neuropil‐like expression pattern consistent with a synapse marker and was widely expressed in cortical and subcortical regions (hippocampus>frontal cortex>cingulate cortex>temporal cortex). VAMP2 expression in AD brains was reduced 0.6‐fold compared to controls in the hippocampus (p=0.01) but comparable to controls in all other regions tested (all p>0.1). Reduced hippocampal VAMP2 levels correlated with increased Braak (r2=0.4, p=0.006) and CERAD (r2=0.3, p=0.03) staging. In CSF, VAMP2 levels were decreased compared to controls in preclinical AD and sequentially elevated over the course of AD, plateauing at the prodromal stage. Worse episodic memory was associated with increased CSF VAMP2 in controls (r2=0.09, p=0.009). In AD, the best fit was a nonlinear model (r2=0.07, p=0.0009) in which worse episodic memory was associated with increased CSF VAMP2 in preclinical AD but decreased VAMP2 in clinical AD. Conclusion: These data support VAMP2 as a promising CSF marker of selective hippocampal synapse loss and episodic memory performance in Alzheimer's disease and healthy controls. [ABSTRACT FROM AUTHOR]

Background: Synapse loss, a pathological hallmark of Alzheimer's disease (AD), is quantified in the brain using synaptophysin in western blot and immunohistochemistry, while the in vivo imaging compound 11C‐UCB‐J targets synaptic vesicle glycoprotein 2A (SV2A). Both proteins are characteristic of synaptic vesicles (SVs) and are found in the majority of synapses. Since synaptophysin is not stable in CSF or serum, it is not a suitable marker to monitor synaptic integrity in fluids (Schlaf et al., 1998). Vesicle‐associated membrane protein 2 (VAMP2), another SV component critical for vesicular trafficking, was identified as potential fluid surrogate for synapse loss using relative quantification (Lleo et al., 2019). Here, we have developed a new digital immunoassay for VAMP2 and evaluated its CSF profile in 226 samples from the AD continuum. For comparison the more established post‐synaptic marker Neurogranin (Ng) was included, which is expressed in dendrites of excitatory glutaminergic neurons. Method: The immunoassay (Simoa) for VAMP2 was established using a commercial rabbit mAb and a novel high‐affinity mouse mAb 15E4. CSF was measured in duplicate on the HD‐1 analyzer across cognitively normal controls (n=68), preclinical AD stage 1 (Amyloidosis +, n=31), preclinical AD stage 2 (Amyloidosis +, neurodegeneration +, n=8), prodromal AD (n=80) and AD dementia (n=39). Ng was quantified with ELISA (EUROIMMUN). Result: The assay was able to quantify VAMP2 in all CSF samples with an average CV below 10%. Three quality control samples had an inter‐/intra‐assay variability below 10%. VAMP2 levels correlated strongly with Ng in all AD stages (r=0.78‐0.96, p<0.0001). VAMP2 (p<0.0001) and Ng (p<0.0001) were decreased by 40% in preclinical AD stage 1. They were both elevated in the prodromal (VAMP2 25%, p=0.0009; Ng 70%, p<0.0001) and dementia group (VAMP2 25%, p=0.014; Ng 50%, p<0.0001). Conclusion: A novel immunoassay for VAMP2 is capable of detecting changes in different AD stages. CSF changes in VAMP2 and Ng precede clinical symptoms and CSF neurodegeneration markers in the AD continuum. In line with observations that CSF‐tau and CSF‐Aβ abnormality precede plaque and tangle load via PET imaging, future longitudinal studies relating synaptic fluid markers with synaptic PET ligands (i.e. 11C‐UCB‐J) could extend these findings. [ABSTRACT FROM AUTHOR]