Site-Specific Cerebrospinal Fluid Tau Hyperphosphorylation in Response to Alzheimer’s Disease Brain Pathology: Not All Tau Phospho-Sites are Hyperphosphorylated

Abstract

Background: Understanding patterns of association between CSF phosphorylated tau (p-tau) species and clinical disease severity will aid Alzheimer’s disease (AD) diagnosis and treatment. Objective: To evaluate changes in tau phosphorylation ratios to brain imaging (amyloid PET, [18F]GTP1 PET, and MRI) and cognition across clinical stages of AD in two different cohorts. Methods: A mass spectrometry (MS)-based method was used to evaluate the relationship between p-tau/tau phosphorylation ratios on 11 sites in CSF and AD pathology measured by tau PET ([18F]GTP1) and amyloid PET ([18F]florbetapir or [18F]florbetaben). Cohort A included cognitively normal-amyloid negative (n = 6) and positive (n = 5) individuals, and amyloid positive prodromal (n = 13), mild (n = 12), and moderate AD patients (n = 10); and Cohort B included amyloid positive prodromal (n = 24) and mild (n = 40) AD patients. Results: In this cross-sectional analysis, we identified clusters of phosphosites with different profiles of phosphorylation ratios across stages of disease. Eight of 11 investigated sites were hyperphosphorylated and associated with the SUVR measures from [18F]GTP1 and amyloid PET. Novel sites 111, 153, and 208 may be relevant biomarkers for AD diagnosis to complement tau hyperphosphorylation measures on previously established sites 181, 205, 217, and 231. Hypophosphorylation was detected on residues 175, 199, and 202, and was inversely associated with [18F]GTP1 and amyloid PET. Conclusion: Hyperphosphorylated and hypophosphorylated forms of tau are associated with AD pathologies, and due to their different site-specific profiles, they may be used in combination to assist with staging of disease.

CSF tau phosphorylation occupancies at T217 and T205 represent improved biomarkers of amyloid and tau pathology in Alzheimer’s disease

“…CSF tau was immunopurified and digested, then phosphorylated and nonphosphorylated peptides were quantified using high-resolution MS (HRMS) as previously described and detailed in Appendix 1 (ref. 25 ). Briefly, tau was immunopurified by incubating 450 µl of CSF with tau1 and HJ8.5 antibodies covalently attached to Sepharose beads at room temperature for 4 h. Immunopurified tau was digested for 16 h at 37 °C with 400 ng of trypsin (Promega).…”

“…In contrast, mass spectrometry (MS) enables simultaneous measurement of nonphosphorylated and phosphorylated tau species and has high specificity. MS has identified p-tau217, p-tau205 and a species of the microtubule-binding region of tau (MTBR-tau243) as candidate AD biomarkers 23 , 24 , revealed the degree of phosphorylation at different tau sites in response to AD 23 , 25 and enabled comparisons between CSF p-tau species that demonstrate the order in which sites are phosphorylated over the course of AD 26 .…”

Baseline [18F]GTP1 tau PET imaging is associated with subsequent cognitive decline in Alzheimer’s disease

“…Baseline and longitudinal data from an observational study evaluating longitudinal change in [ 18 F] GTP1 tau PET in CN and AD participants (GN30009; NCT02640092) were analyzed. Baseline data from this study have been included elsewhere [3,[21][22][23].…”

CSF tau phosphorylation at T217 and T205 are improved biomarkers of amyloid and tau pathology in Alzheimer disease

“…CSF tau was immunopuri ed and digested, then phosphorylated and non-phosphorylated peptides were quanti ed using high resolution mass spectrometry as previously described and detailed in Appendix 1 25 . Brie y, tau was immunopuri ed by incubating 450 µl CSF with Tau1 and HJ8.5 antibodies covalently attached to sepharose beads at room temperature for 4 hours.…”

“…In contrast, mass spectrometry enables simultaneous measurement of non-phosphorylated and phosphorylated tau species and has high speci city. Mass spectrometry has identi ed p-tau217, p-tau205, and a species of the microtubule binding region of tau (MTBR-tau243) as candidate AD biomarkers 23,24 , revealed the degree of phosphorylation at different tau sites in response to AD 23,25 , and enabled comparisons between CSF ptau species that demonstrate the order in which sites are phosphorylated over the course of AD 26 .…”