Because of their abundance, resistance to proteolysis, rapid aggregation and neurotoxicity, N-terminally truncated and, in particular, pyroglutamate (pE)-modified Abeta peptides have been suggested as being important in the initiation of pathological cascades resulting in the development of Alzheimer's disease. We found that the N-terminal pE-formation is catalyzed by glutaminyl cyclase in vivo. Glutaminyl cyclase expression was upregulated in the cortices of individuals with Alzheimer's disease and correlated with the appearance of pE-modified Abeta. Oral application of a glutaminyl cyclase inhibitor resulted in reduced Abeta(3(pE)-42) burden in two different transgenic mouse models of Alzheimer's disease and in a new Drosophila model. Treatment of mice was accompanied by reductions in Abeta(x-40/42), diminished plaque formation and gliosis and improved performance in context memory and spatial learning tests. These observations are consistent with the hypothesis that Abeta(3(pE)-42) acts as a seed for Abeta aggregation by self-aggregation and co-aggregation with Abeta(1-40/42). Therefore, Abeta(3(pE)-40/42) peptides seem to represent Abeta forms with exceptional potency for disturbing neuronal function. The reduction of brain pE-Abeta by inhibition of glutaminyl cyclase offers a new therapeutic option for the treatment of Alzheimer's disease and provides implications for other amyloidoses, such as familial Danish dementia.

Extracellular plaques of β-amyloid (Aβ) and intraneuronal neurofibrillary tangles made from tau are the histopathological signatures of Alzheimer's disease (AD). Plaques comprise Aβ fibrils that assemble from monomeric and oligomeric intermediates, and are prognostic indicators of AD. Despite the significance of plaques to AD, oligomers are considered to be the principal toxic forms of Aβ 1,2 . Interestingly, many adverse responses to Aβ, such as cytotoxicity 3 , microtubule loss 4 , impaired memory and learning 5 , and neuritic degeneration 6 , are greatly amplified by tau expression. N-terminally truncated, pyroglutamylated (pE) forms of Aβ 7,8 are strongly associated with AD, are more toxic than Aβ 1-42 and Aβ , and have been proposed as initiators of AD Users may view, print, copy, download and text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: http://www.nature.com/authors/editorial_policies/license.html#terms * Correspondence: gsb4g@virginia.edu. **Correspondence: Hans-Ulrich.Demuth@probiodrug.de. J.M.N and S.S. contributed equally to the paper.Full Methods and relevant references will be available in the online Supplementary Information accompanying this paper at http:// www.nature.com/nature.Author Contributions: J.M.N. performed most of the biochemical and cell biological experiments; S.S. was the principal force behind the experiments involving hAPP SL /hQC and TBA2.1/tau KO mice, and was aided by B.H.-P., H.C.; A.S. and T.W. fractionated and analyzed human brain extracts; E.S., K.Y. and B.W. performed the peri-hippocampal injection experiments; A.H. and C.G.G. produced and characterized the M64 and M87 antibodies; R.R. and K.R. performed the electrophysiology experiments; A.A., W.J. and S.G. performed and analyzed the immunohistochemical experiments on TBA2.1 and Tau-KO/TBA2.1 mice; G.S.B. and H.-U.D. initiated and directed the project; G.S.B. was the principal writer of the paper; all of the authors participated in the design and analysis of experiments, and in editing of the paper. Fig. 2) to the oligomers. HHS Public AccessAt 5 μM peptide, 5% pE-Aβ aggregated faster than Aβ 3(pE)-42 or Aβ 1-42 alone based on thioflavin T fluorescence shifts 15 ( Supplementary Fig. 3). The OD 450 /OD 490 ratio for Aβ 3(pE)-42 rose and peaked more rapidly than for Aβ 1-42 , but peaked at an ~25% lower level. The fastest rise in the OD 450 /OD 490 ratio was for 5% pE-Aβ, which peaked similarly to Aβ 3(pE)-42 . Aβ 3(pE)-42 , Aβ 1-42 and 5% pE-Aβ thus oligomerized by different pathways.To test whether distinct biological activities were coupled to these oligomerization differences, we compared cytotoxicity of the peptides towards cultured neurons or glia using calcein-AM and fluorescence microscopy 16 . Twelve hours of Aβ 1-42 exposure had little effect on cell viability for wild type (WT) or tau knockout (KO) neurons, or WT glial cells (Fig. 1a). Contrastingly, most WT neurons died and detached from the substrate after exposur...

Synaptic dysfunction and loss caused by age-dependent accumulation of synaptotoxic beta amyloid (Abeta) 1–42 oligomers is proposed to underlie cognitive decline in Alzheimer's disease (AD). Alterations in membrane trafficking induced by Abeta oligomers mediates reduction in neuronal surface receptor expression that is the basis for inhibition of electrophysiological measures of synaptic plasticity and thus learning and memory. We have utilized phenotypic screens in mature, in vitro cultures of rat brain cells to identify small molecules which block or prevent the binding and effects of Abeta oligomers. Synthetic Abeta oligomers bind saturably to a single site on neuronal synapses and induce deficits in membrane trafficking in neuronal cultures with an EC50 that corresponds to its binding affinity. The therapeutic lead compounds we have found are pharmacological antagonists of Abeta oligomers, reducing the binding of Abeta oligomers to neurons in vitro, preventing spine loss in neurons and preventing and treating oligomer-induced deficits in membrane trafficking. These molecules are highly brain penetrant and prevent and restore cognitive deficits in mouse models of Alzheimer's disease. Counter-screening these compounds against a broad panel of potential CNS targets revealed they are highly potent and specific ligands of the sigma-2/PGRMC1 receptor. Brain concentrations of the compounds corresponding to greater than 80% receptor occupancy at the sigma-2/PGRMC1 receptor restore cognitive function in transgenic hAPP Swe/Ldn mice. These studies demonstrate that synthetic and human-derived Abeta oligomers act as pharmacologically-behaved ligands at neuronal receptors - i.e. they exhibit saturable binding to a target, they exert a functional effect related to their binding and their displacement by small molecule antagonists blocks their functional effect. The first-in-class small molecule receptor antagonists described here restore memory to normal in multiple AD models and sustain improvement long-term, representing a novel mechanism of action for disease-modifying Alzheimer's therapeutics.

Alzheimer’s disease is characterized by accumulation of amyloid plaques and tau aggregates in several cortical brain regions. Tau phosphorylation causes formation of neurofibrillary tangles and neuropil threads. Phosphorylation at tau Ser202/Thr205 is well characterized since labeling of this site is used to assign Braak stage based on occurrence of neurofibrillary tangles. Only little is known about the spatial and temporal phosphorylation profile of other phosphorylated tau (ptau) sites. Here, we investigate total tau and ptau at residues Tyr18, Ser199, Ser202/Thr205, Thr231, Ser262, Ser396, Ser422 as well as amyloid-β plaques in human brain tissue of AD patients and controls. Allo- and isocortical brain regions were evaluated applying rater-independent automated quantification based on digital image analysis. We found that the level of ptau at several residues, like Ser199, Ser202/Thr205, and Ser422 was similar in healthy controls and Braak stages I to IV but was increased in Braak stage V/VI throughout the entire isocortex and transentorhinal cortex. Quantification of ThioS-stained plaques showed a similar pattern. Only tau phosphorylation at Tyr18 and Thr231 was already significantly increased in the transentorhinal region at Braak stage III/IV and hence showed a progressive increase with increasing Braak stages. Additionally, the increase in phosphorylation relative to controls was highest at Tyr18, Thr231 and Ser199. By contrast, Ser396 tau and Ser262 tau showed only a weak phosphorylation in all analyzed brain regions and only minor progression. Our results suggest that the ptau burden in the isocortex is comparable between all analyzed ptau sites when using a quantitative approach while levels of ptau at Tyr18 or Thr231 in the transentorhinal region are different between all Braak stages. Hence these sites could be crucial in the pathogenesis of AD already at early stages and therefore represent putative novel therapeutic targets.Electronic supplementary materialThe online version of this article (10.1186/s40478-018-0557-6) contains supplementary material, which is available to authorized users.

Amyloid beta-peptide (Abeta) accumulation in specific brain regions is a pathological hallmark of Alzheimer's disease (AD). We have previously reported that a well-characterized acyl-coenzyme A: cholesterol acyltransferase (ACAT) inhibitor, CP-113,818, inhibits Abeta production in cell-based experiments. Here, we assessed the efficacy of CP-113,818 in reducing AD-like pathology in the brains of transgenic mice expressing human APP(751) containing the London (V717I) and Swedish (K670M/N671L) mutations. Two months of treatment with CP-113,818 reduced the accumulation of amyloid plaques by 88%-99% and membrane/insoluble Abeta levels by 83%-96%, while also decreasing brain cholesteryl-esters by 86%. Additionally, soluble Abeta(42) was reduced by 34% in brain homogenates. Spatial learning was slightly improved and correlated with decreased Abeta levels. In nontransgenic littermates, CP-113,818 also reduced ectodomain shedding of endogenous APP in the brain. Our results suggest that ACAT inhibition may be effective in the prevention and treatment of AD by inhibiting generation of the Abeta peptide.