Inhibition of O-GlcNAcase leads to elevation of O-GlcNAc tau and reduction of tauopathy and cerebrospinal fluid tau in rTg4510 mice

Nicholas B Hastings,Julie Lee,Cathy Ruey-Ruey Huang,J Fred Hess,Lynn A Hyde ,Richard Hargreaves,Diane E Grotz ,Michaela C Stanton ,Diane Levitan,Maureen Laverty,Harold G Selnick ,Kwok-Lam Karen Hong,Brent D Butts,Steve Lu ,Xuesong Ouyang,David J Vocadlo,Xiaohai Wang,Joseph Duffy ,Mandy J Maguire ,Lixin Song,Giuseppe Terracina,Ganfeng Wang,Veeravan Mahadomrongkul,Ernest J Mceachern,Thomas W Rosahl,Eric M Parker,Lili Zhang

doi:10.1186/s13024-017-0181-0

Abstract

BackgroundHyperphosphorylation of microtubule-associated protein tau is a distinct feature of neurofibrillary tangles (NFTs) that are the hallmark of neurodegenerative tauopathies. O-GlcNAcylation is a lesser known post-translational modification of tau that involves the addition of N-acetylglucosamine onto serine and threonine residues. Inhibition of O-GlcNAcase (OGA), the enzyme responsible for the removal of O-GlcNAc modification, has been shown to reduce tau pathology in several transgenic models. Clarifying the underlying mechanism by which OGA inhibition leads to the reduction of pathological tau and identifying translatable measures to guide human dosing and efficacy determination would significantly facilitate the clinical development of OGA inhibitors for the treatment of tauopathies.MethodsGenetic and pharmacological approaches are used to evaluate the pharmacodynamic response of OGA inhibition. A panel of quantitative biochemical assays is established to assess the effect of OGA inhibition on pathological tau reduction. A “click” chemistry labeling method is developed for the detection of O-GlcNAcylated tau.ResultsSubstantial (>80%) OGA inhibition is required to observe a measurable increase in O-GlcNAcylated proteins in the brain. Sustained and substantial OGA inhibition via chronic treatment with Thiamet G leads to a significant reduction of aggregated tau and several phosphorylated tau species in the insoluble fraction of rTg4510 mouse brain and total tau in cerebrospinal fluid (CSF). O-GlcNAcylated tau is elevated by Thiamet G treatment and is found primarily in the soluble 55 kD tau species, but not in the insoluble 64 kD tau species thought as the pathological entity.ConclusionThe present study demonstrates that chronic inhibition of OGA reduces pathological tau in the brain and total tau in the CSF of rTg4510 mice, most likely by directly increasing O-GlcNAcylation of tau and thereby maintaining tau in the soluble, non-toxic form by reducing tau aggregation and the accompanying panoply of deleterious post-translational modifications. These results clarify some conflicting observations regarding the effects and mechanism of OGA inhibition on tau pathology, provide pharmacodynamic tools to guide human dosing and identify CSF total tau as a potential translational biomarker. Therefore, this study provides additional support to develop OGA inhibitors as a treatment for Alzheimer’s disease and other neurodegenerative tauopathies.

Highlights

Hyperphosphorylation of microtubule-associated protein tau is a distinct feature of neurofibrillary tangles (NFTs) that are the hallmark of neurodegenerative tauopathies
The present study demonstrates that chronic inhibition of OGA reduces pathological tau in the brain and total tau in the cerebrospinal fluid (CSF) of rTg4510 mice, most likely by directly increasing O-GlcNAcylation of tau and thereby maintaining tau in the soluble, non-toxic form by reducing tau aggregation and the accompanying panoply of deleterious post-translational modifications
Effect of OGA inhibition on O-protein levels A sensitive, quantitative, high throughput MesoScale Diagnostics (MSD) sandwich immunoassay was developed to examine the effect of OGA inhibition on global protein O-GlcNAcylation

Summary

Introduction

Hyperphosphorylation of microtubule-associated protein tau is a distinct feature of neurofibrillary tangles (NFTs) that are the hallmark of neurodegenerative tauopathies. Dominant tau mutations have been identified that are associated with aggressive tauopathies, including frontotemporal dementia with Parkinsonism on chromosome 17 and PSP [3, 4] These findings support a pathogenic role of tau in neurodegeneration and lead to the therapeutic hypothesis that reduction of tau pathology may be a viable approach to slow down the progression of diseases involving tauopathy. Protein O-GlcNAcylation is a reversible posttranslational modification involving addition of a single N-acetylglucosamine (O-GlcNAc) moiety onto the hydroxyl group of serine and threonine residues (reviewed by [5]). This modification is regulated by two enzymes in mammalian cells. Many cytoplasmic and nuclear proteins are subject to O-GlcNAcylation, and because this modification occurs on serine and threonine residues, it can potentially modulate protein phosphorylation directly on the same sites or indirectly on proximal sites [5]

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