Inhibition Of Tau Aggregation Research Articles

Discovery of Azaindolin-2-One as a Dual Inhibitor of GSK3β and Tau Aggregation with Potential Neuroprotective Activity.

The inhibition of glycogen synthase kinase 3β (GSK3β) activity through pharmacological intervention represents a promising approach for treating challenging neurodegenerative disorders like Alzheimer’s disease. Similarly, abnormal tau aggregate accumulation in neurons is a hallmark of various neurodegenerative diseases. We introduced new dual GSK3β/tau aggregation inhibitors due to the excellent clinical outcome of multitarget drugs. Compound (E)-2f stands out among the synthesized inhibitors as a promising GSK3β inhibitor (IC50 1.7 µM) with a pronounced tau anti-aggregation effect in a cell-based model of tauopathy. Concurrently, (E)-2f was demonstrated to be non-toxic to normal cells, making it a promising neuroprotective lead compound that needs further investigation.

Ginsenoside Rf inhibits human tau proteotoxicity and causes specific LncRNA, miRNA and mRNA expression changes in Caenorhabditis elegans model of tauopathy

Under pathological conditions, human tau (htau) hyperphosphorylation promotes formation of proteotoxic intracellular amyloid aggregates that may underlie neurodegenerative diseases known as tauopathies, prompting researchers to develop treatments that inhibit htau aggregation as a promising therapeutic strategy. Ginsenosides, the main active constituents of Panax ginseng C. A. Meyer (ginseng), appear to inhibit tau aggregation and disassociation in tauopathy models, although their active components and molecular mechanisms are unknown. Here, we used a novel Caenorhabditis elegans (C. elegans) tauopathy model to identify ginsenoside monomers which may repress htau proteotoxicity. Our findings indicated that ginsenoside Rf prevented tau aggregation and reversed abnormal tau aggregation-induced phenotypes and alleviated neurodegeneration in worms. Notably, deep RNA-seq analysis of ginsenoside Rf-treated and untreated worms with tauopathy revealed that ginsenoside Rf altered expression levels of 24 up- and 36 down-regulated lncRNA transcripts, 32 up- and 22 down-regulated miRNAs and 65 up- and 30 down-regulated mRNA transcripts. Based on GO and KEGG pathway annotation analyses, identified mRNAs, miRNAs and lncRNAs-associated gene targets were functionally related to neuron-related terms (e.g., neuron development, axon and motor neuron axon guidance) and longevity regulating pathways. Importantly, RT-qRCR results suggested that 6 miRNAs (miR-786, miR-2208b, miR-34, miR-241, miR-247 and miR-4805), 8 lncRNAs (MSTRG.20812.2, MSTRG.22617.2, MSTRG.28210.13, MSTRG.5728.12, MSTRG.29708.1, MSTRG.3342.25, MSTRG.3342.31 and MSTRG.8841.8) and 7 mRNAs (nas-33, math-28, T14B4.19, col-17, rol-6, sqt-1 and irg-4) were potential targets of ginsenoside Rf inhibition of tauopathy. These results partially explain mechanisms underlying ginsenoside Rf-associated alleviation of htau proteotoxicity and will guide future strategies to discover potential therapeutic targets for preventing and alleviating tauopathies.

Structure-activity relationship of carbon nitride dots in inhibiting Tau aggregation

Due to the numerous failed clinical trials of anti-amyloid drugs, microtubule associated protein tau (MAPT) now stands out as one of the most promising targets for AD therapy. In this study, we report for the first time the structure-dependent MAPT aggregation inhibition of carbon nitride dots (CNDs). CNDs have exhibited great promise as a potential treatment of Alzheimer's disease (AD) by inhibiting the aggregation of MAPT. In order to elucidate its structure-activity relationship, CNDs were separated via column chromatography and five fractions with different structures were obtained that were characterized by multiple spectroscopy methods. The increase of surface hydrophilic functional groups is consistent with the increase of polarity from fraction 1 to 5. Particle sizes (1–2 nm) and zeta potentials (∼-20 mV) are similar among five fractions. With the increase of polarity from fraction 1 to 5, their MAPT aggregation inhibition capacity was weakened. This suggests hydrophobic interactions between CNDs and MAPT, validated via molecular dynamics simulations. With a zebrafish blood-brain barrier (BBB) model, CNDs were observed to cross the BBB through passive diffusion. CNDs were also found to inhibit the generation of multiple reactive oxygen species, which is an important contributor to AD pathogenesis.

Drug delivery of memantine with carbon dots for Alzheimer’s disease: blood–brain barrier penetration and inhibition of tau aggregation

Neurofibrillary tangle, composed of aggregated tau protein, is a pathological hallmark of Alzheimer’s disease (AD). The inhibition of tau aggregation is therefore an important direction for AD drug discovery. In this work, we explored the efficacy of two types of carbon dots in targeting tau aggregation, as versatile nano-carriers for the development of carbon dots (CDs)-based AD therapy. We carried out synthesis, biophysical and biochemical characterizations of two types of CDs, namely, carbon nitride dots (CNDs) and black carbon dots (B-CDs). CDs, which are biocompatible and non-toxic, were successfully conjugated with memantine hydrochloride (MH) through EDC/NHS mediated amidation reactions followed by systematic characterizations using various biophysical techniques including UV–vis spectroscopy (UV–vis), photoluminescence (PL), Fourier-transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), mass spectrometry (MS), Transmission electron microscopy (TEM) and atomic force microscopy (AFM). The surface diversity along with small particle sizes of CDs allowed facile delivery of MH across the blood–brain barrier (BBB), as demonstrated using a zebrafish in vivo model. The tau aggregation inhibition experiments were conducted using the thioflavin-T (ThT) assay to identify the most effective inhibitor. The kinetics and magnitude of tau aggregation were measured in the presence of CDs, which demonstrates that both B-CDs-MH and B-CDs alone are the most effective inhibitors of tau aggregation with IC50 values of 1.5 ± 0.3 and 1.6 ± 1.5 μg/mL, respectively. Taken together, our findings hold therapeutic significance to enhance the efficient delivery of MH to target AD pathology in the brain for improved efficacy.

Structure-Activity Relationships of Novel Tau Ligands: Passive Fibril Binders and Active Aggregation Inhibitors.

Intrinsically disordered proteins (IDPs) are core components of many biological processes and are central players in several pathologies. Despite being important drug targets, attempts to design small-molecule ligands that would help understand and attenuate their behavior are frustrated by the structural diversity exhibited by these flexible proteins. To accommodate the dynamic nature of IDPs, we developed a procedure that efficiently identifies active small-molecule ligands for disordered proteins. By exploring the chemical space around these ligands, we refined their effect on aggregation and identified molecular features critical for activity and affinity. Notably, the discovery of this new family of disordered protein ligands was achieved more quickly and with less expense than conventional high-throughput screening (HTS) or docking alone would have allowed. The resulting ligands include tau aggregation inhibitors as well as at least one compound that binds fibrils potently but does not appear to perturb the extent of kinetics of aggregation.

Shikonin inhibits tau aggregation and phase separation and improves the viability of SH-SY5Y cells after exposure to tau oligomers

The pathological aggregation of tau, a microtubule-associated protein, is the chief feature of Alzheimer’s disease and related tauopathies. We investigated the potential of shikonin, a naphthoquinone from Lithospermum erythrorhizon, to inhibit tau aggregation. Shikonin reduced the initial aggregation rate of tau. The heparin-induced oligomerization and aggregation of tau were also arrested by shikonin, with a half-maximal inhibitory concentration of 1.20 ± 0.04 μM and 1.0 ± 0.3 μM, respectively.

A covalent strategy to target intrinsically disordered proteins: Discovery of novel tau aggregation inhibitors

Intrinsically disordered proteins (IDPs) play important roles in disease pathologies; however, their lack of defined stable 3D structures make traditional drug design strategies typically less effective against these targets. Based on promising results of targeted covalent inhibitors (TCIs) on challenging targets, we have developed a covalent design strategy targeting IDPs. As a model system we chose tau, an endogenous IDP of the central nervous system that is associated with severe neurodegenerative diseases via its aggregation. First, we mapped the tractability of available cysteines in tau and prioritized suitable warheads. Next, we introduced the selected vinylsulfone warhead to the non-covalent scaffolds of potential tau aggregation inhibitors. The designed covalent tau binders were synthesized and tested in aggregation models, and inhibited tau aggregation effectively. Our results revealed the usefulness of the covalent design strategy against therapeutically relevant IDP targets and provided promising candidates for the treatment of tauopathies.

Crossed cerebellar diaschisis in Alzheimer's disease.

Crossed cerebellar diaschisis (CCD) is characterized by hypometabolism and hypoperfusion on molecular imaging in the cerebellum due to a supratentorial lesion on the contralateral side. CCD is a well-established phenomenon in acute or subacute conditions such as infarction but it has been less well described in chronic conditions such as neurodegenerative dementias. Here, we investigate CCD in a large sample of 830 people meeting research criteria for Alzheimer's disease (AD) using [18F]fluorodeoxyglucose-positron emission tomography (FDG-PET). This study is based on FDG-PET data collected at baseline as part of two large-scale Phase III clinical trials of a novel tau aggregation inhibitor medication, methylthioninium in mild to moderate AD participants. Quantification of FDG-PET hypometabolism was carried out using standardized uptake value ratio (SUVR), with the pons as the comparison region. SUVR was compared in different regions of interest between the right and left hemispheres of the brain and cerebellum in people with mild AD (Mini-Mental State Examination score ≥ 20). Comparison of SUVR in different brain regions demonstrated significant differences in the temporal, occipital and cerebellar cortices. Right and left asymmetry was noted with lower SUVR in the left temporal and occipital regions, whereas SUVR was lower in the right side of the cerebellum. Here, we found robust evidence of CCD in a large sample of people with AD, a chronic neurodegenerative condition. The presence of this phenomenon in AD opens up a new avenue of research in AD pathogenesis and has the potential to change future diagnostic and therapeutic strategies.

A novel D-amino acid peptide with therapeutic potential (ISAD1) inhibits aggregation of neurotoxic disease-relevant mutant Tau and prevents Tau toxicity in vitro

BackgroundAlzheimer’s disease (AD), the most common form of dementia, is a progressive neurodegenerative disorder that mainly affects older adults. One of the pathological hallmarks of AD is abnormally aggregated Tau protein that forms fibrillar deposits in the brain. In AD, Tau pathology correlates strongly with clinical symptoms, cognitive dysfunction, and neuronal death.MethodsWe aimed to develop novel therapeutic D-amino acid peptides as Tau fibrillization inhibitors. It has been previously demonstrated that D-amino acid peptides are protease stable and less immunogenic than L-peptides, and these characteristics may render them suitable for in vivo applications. Using a phage display procedure against wild type full-length Tau (TauFL), we selected a novel Tau binding L-peptide and synthesized its D-amino acid version ISAD1 and its retro inversed form, ISAD1rev, respectively.ResultsWhile ISAD1rev inhibited Tau aggregation only moderately, ISAD1 bound to Tau in the aggregation-prone PHF6 region and inhibited fibrillization of TauFL, disease-associated mutant full-length Tau (TauFLΔK, TauFL-A152T, TauFL-P301L), and pro-aggregant repeat domain Tau mutant (TauRDΔK). ISAD1 and ISAD1rev induced the formation of large high molecular weight TauFL and TauRDΔK oligomers that lack proper Thioflavin-positive β-sheet conformation even at lower concentrations. In silico modeling of ISAD1 Tau interaction at the PHF6 site revealed a binding mode similar to those known for other PHF6 binding peptides. Cell culture experiments demonstrated that ISAD1 and its inverse form are taken up by N2a-TauRDΔK cells efficiently and prevent cytotoxicity of externally added Tau fibrils as well as of internally expressed TauRDΔK.ConclusionsISAD1 and related peptides may be suitable for therapy development of AD by promoting off-pathway assembly of Tau, thus preventing its toxicity.

Engineering a pathological tau-targeted nanochaperone for selective and synergetic inhibition of tau pathology in Alzheimer's Disease

Tau-targeting therapies for Alzheimer’s disease (AD) have received increasing attention in recent years, due to the clinical failure of various anti-amyloid β drugs and closer association between tau burden and cognitive impairments in AD patients. The intracellular tau aggregation is recognized as a vital event in tau pathway of AD progression. However, current developed tau aggregation inhibitors are still facing serious challenges on how to access the cytosol of cells where tau protein mainly located and differentiate pathological tau from normal tau. Herein, we report a customized biomimetic nanochaperone for targeting intracellular pathological tau, serving as a novel anti-tau therapy of AD. This nanochaperone was tailored by engineering VQIINK hexapeptide derived from tau protein onto the surface of self-assembly micelle equipped with chaperone-like hydrophobic microdomains and confined spaces. With the lysosomal microenvironment responsiveness of unique surface structure, the Tau-nChap can escape from lysosome and enter into cytoplasm to overcome the intracellular tau location challenge. More importantly, the cytosol-arrived nanochaperone can selectively capture pathological tau without interfering with normal tau and potently inhibit their aggregation through the synergic effect of tau-recognizing peptide and confined hydrophobic microdomain on surface. As a result, the tau-targeting nanochaperone rescues neuronal cells from tau-mediated neurotoxicity and ameliorates cognitive deficits of AD model mice. Therefore, we believe that this custom-built tau-nanochaperone represents a promising weapon for combating AD and provides valuable insights for the treatment of other neurodegenerative disorders induced by specific pathogenic proteins including Parkinson’s disease and Huntington’s disease.

Three-repeat and four-repeat tau isoforms form different oligomers.

Different tauopathies are characterized by the isoform-specific composition of the aggregates found in the brain and by structurally distinct tau strains. Although tau oligomers have been implicated as important neurotoxic species, little is known about how the primary structures of the six human tau isoforms affect tau oligomerization because the oligomers are metastable and difficult to analyze. To address this knowledge gap, here, we analyzed the initial oligomers formed by the six tau isoforms in the absence of posttranslational modifications or other manipulations using dot blots probed by an oligomer-specific antibody, native-PAGE/western blots, photo-induced cross-linking of unmodified proteins, mass-spectrometry, and ion-mobility spectroscopy. We found that under these conditions, three-repeat (3R) isoforms are more prone than four-repeat (4R) isoforms to form oligomers. We also tested whether known inhibitors of tau aggregation affect its oligomerization using three small molecules representing different classes of tau aggregation inhibitors, Methylene Blue (MB), the molecular tweezer CLR01, and the all-D peptide TLKIVW, for their ability to inhibit or modulate the oligomerization of the six tau isoforms. Unlike their reported inhibitory effect on tau fibrillation, the inhibitors had little or no effect on the initial oligomerization. Our study provides novel insight into the primary-quaternary structure relationship of human tau and suggests that 3R-tau oligomers may be an important target for future development of compounds targeting pathological tau assemblies.

Hydromethylthionine enhancement of central cholinergic signalling is blocked by rivastigmine and memantine.

The prevention of tau protein aggregations is a therapeutic goal for the treatment of Alzheimer's disease (AD), and hydromethylthionine (HMT) (also known as leucomethylthioninium-mesylate [LMTM]), is a potent inhibitor of tau aggregation in vitro and in vivo. In two Phase 3clinical trials in AD, HMT had greater pharmacological activity on clinical endpoints in patients not receiving approved symptomatic treatments for AD (acetylcholinesterase (AChE) inhibitors and/or memantine) despite different mechanisms of action. To investigate this drug interaction in an animal model, we used tau-transgenic L1 and wild-type NMRI mice treated with rivastigmine or memantine prior to adding HMT, and measured changes in hippocampal acetylcholine (ACh) by microdialysis. HMT given alone doubled hippocampal ACh levels in both mouse lines and increased stimulated ACh release induced by exploration of the open field or by infusion of scopolamine. Rivastigmine increased ACh release in both mouse lines, whereas memantine was more active in tau-transgenic L1mice. Importantly, our study revealed a negative interaction between HMT and symptomatic AD drugs: the HMT effect was completely eliminated in mice that had been pre-treated with either rivastigmine or memantine. Rivastigmine was found to inhibit AChE, whereas HMT and memantine had no effects on AChE or on choline acetyltransferase (ChAT). The interactions observed in this study demonstrate that HMT enhances cholinergic activity in mouse brain by a mechanism of action unrelated to AChE inhibition. Our findings establish that the drug interaction that was first observed clinically has a neuropharmacological basis and is not restricted to animals with tau aggregation pathology. Given the importance of the cholinergic system for memory function, the potential for commonly used AD drugs to interfere with the treatment effects of disease-modifying drugs needs to be taken into account in the design of clinical trials.

Cannabidiol Inhibits Tau Aggregation In Vitro

A hallmark of Alzheimer’s disease (AD) is the accumulation of tau protein in the brain. Compelling evidence indicates that the presence of tau aggregates causes irreversible neuronal destruction, eventually leading to synaptic loss. So far, the inhibition of tau aggregation has been recognized as one of the most effective therapeutic strategies. Cannabidiol (CBD), a major component found in Cannabis sativa L., has antioxidant activities as well as numerous neuroprotective features. Therefore, we hypothesize that CBD may serve as a potent substance to hamper tau aggregation in AD. In this study, we aim to investigate the CBD effect on the aggregation of recombinant human tau protein 1N/4R isoform using biochemical methods in vitro and in silico. Using Thioflavin T (ThT) assay, circular dichroism (CD), and atomic force microscopy (AFM), we demonstrated that CBD can suppress tau fibrils formation. Moreover, by quenching assay, docking, and job’s plot, we further demonstrated that one molecule of CBD interacts with one molecule of tau protein through a spontaneous binding. Experiments performed by quenching assay, docking, and Thioflavin T assay further established that the main forces are hydrogen Van der Waals and some non-negligible hydrophobic forces, affecting the lag phase of tau protein kinetics. Taken together, this study provides new insights about a natural substance, CBD, for tau therapy which may offer new hope for the treatment of AD.

Development of D‐enantiomeric peptides as tau aggregation inhibitors directed against the hexapeptide motif PHF6* of tau

AbstractBackgroundTwo hexapeptide motifs within Tau, designated PHF6* (275VQIINK280) and PHF6 (306VQIVYK311), are known to drive Tau aggregation, making them attractive targets for the development of Tau aggregation inhibitors for therapeutic options.MethodEmploying mirror‐image phage display with a large peptide library, we have identified PHF6* fibril binding peptides consisting of D‐enantiomeric amino acids. D‐enantiomeric peptides are extremely protease stable and considerably less immunogenic than L‐peptides, and the suitability of D‐peptides for therapeutic purposes has already been demonstrated. The most promising D‐peptide and its ability to inhibit Tau aggregation was investigated using various biochemical and biophysical methods, and compared to the performance of PHF6‐ or other PHF6*‐binding peptides selected earlier by our and other groups.ResultMMD3 and MMD3rev inhibited PHF6*, TauRDΔK280 and full‐length Tau fibrillization in vitro. Our data demonstrated that the peptides prevent the formation of Tau ß‐sheet‐rich fibrils by formation of large amorphous aggregates. The binding mode of MMD3 and MMD3rev to PHF6* fibrils was investigated by in silico modeling.ConclusionOur peptides were able to penetrate cells and might be interesting for therapeutic and diagnostic applications in AD research.

Identification of pharmacodynamics readouts through acute dosing of a small molecule inhibitor of tau self‐association in a tauopathy mouse model

AbstractBackgroundThe accumulation of tau aggregates correlates with and facilitates disease progression in pure and mixed tauopathies such as Progressive Supranuclear Palsy and Alzheimer’s disease. However, a disease‐modifying‐therapeutic targeting tau aggregation has not yet shown success in the clinic. Our target approach is highly differentiated from other programs that are focused on inhibiting tau aggregation and all downstream cascades. We have selected and optimized small molecules capable of targeting tau self‐association (TSA), the first and rate‐limiting step in tau aggregation. We have demonstrated the efficacy of our lead TSA inhibitor in two mouse models of tauopathy (htau mice; tau‐P301L‐JNPL3 mice) in preventive and therapeutic studies. Treatment with the lead TSA inhibitor blocked tau self‐association and formation of large tau aggregates, reduced overall tau accumulation, and rescued impaired motor function in aged mice with pre‐existing tau aggregates. IND‐enabling studies for the lead TSA inhibitor are completing, and a FDA submission is planned for Q1 2022. In this study, we aim to identify acute measures of efficacy in aged homozygous female tau‐P301L‐JNPL3 mice that will be used to model Pharmacokinetics (PK)/Pharmacodynamics (PD) relationship of our TSA inhibitors.MethodsAcute dose range‐finding (ADRF) study was performed using multiple routes of administration. C57BL/6 mice (n=30) were either administered vehicle or a single dose of a selected TSA inhibitor ranging from 2 to 200 mg/kg (n = 3 mice/gender/group). Biospecimens were collected at Cmax. PK studies in C57BL/6 mice will be performed with and without fasting to determine the food effects on bioavailability.ResultsSerum and brain homogenate samples from the ADRF study are being analyzed to evaluate TSA inhibitor concentrations by LC‐MS/MS. These results will be used to select the dose and route of administration for PK studies in 9 months‐old homozygous female JNPL3 mice. To determine the selective dose for PD evaluations, PK studies will be performed using three doses. Some of the PD measures that will be evaluated are tau self‐association, tau hyperphosphorylation, and motor behavior.ConclusionsUnderstanding PK/PD relationship will be crucial for determining appropriate dosing and target engagement biomarkers in subsequent Phase‐1b studies.

In vivo validation of therapeutic efficacy of a small molecule lead targeting tau self‐association in JNPL3 mice

AbstractBackgroundSmall molecules were screened and optimized for inhibiting tau self‐association, the initial step in the formation of soluble toxic tau oligomers and larger insoluble aggregates. This approach is differentiated from other tau aggregation inhibitor programs that have largely focused on inhibiting formation and/or dissociating large and relatively inert fibrils which could generate toxic tau oligomer seeds. Small molecules inhibitors are agnostic to specific post‐translational modifications, conformational changes or strains of tau aggregates, and have superior access to their target in the CNS relative to immunotherapeutic approaches. We have previously shown in vivo efficacy of a lead compound in preventive studies in htau (Davidowitz et al., 2020) and human tau P301L JNPL3 mouse models of tauopathy. The tau P301L mutation is the most common FTDP‐17 mutation and causes tau aggregation and motor deficits with age (Lewis et al., 2000).MethodBlinded studies of homozygous female tau 4R/0N P301L JNPL3 mice were performed at an independent laboratory where mice were treated for five months by administration of our lead compound in feed from 7 to 12 months‐of‐age. The four groups of mice in the study were baseline (7 months; n=20), vehicle, and two treatment groups (40 and 80 mg/kg doses; n=25 each). To quantify levels of Sarkosyl‐insoluble tau, self‐associated tau, and phosphorylated tau in the treated and control groups of mice ELISAs were performed (Acker et al., 2012; Forest et al., 2019). Rotarod and open field studies were performed to evaluate motor behavior.ResultEfficacy was shown in the cortex of treated mice that had reduced levels of self‐associated tau below baseline, baseline levels of insoluble tau aggregates, and reduced levels of aberrantly phosphorylated tau with statistical significance. In the hindbrain there was reduction of insoluble tau aggregates in the 40 mg/kg group compared to vehicle. Treatment groups showed improved Rotarod performance compared to baseline and vehicle groups.ConclusionTreatment with the lead compound reduced tau aggregation and improved motor behavior in aged JNPL3 mice and support preclinical development of the lead for clinical studies.

Discovery of 1-(phenylsulfonyl)-1H-indole-based multifunctional ligands targeting cholinesterases and 5-HT6 receptor with anti-aggregation properties against amyloid-beta and tau.

Multifunctional ligands as an essential variant of polypharmacology are promising candidates for the treatment of multi-factorial diseases like Alzheimer's disease. Based on clinical evidence and following the paradigm of multifunctional ligands we have rationally designed and synthesized a series of compounds targeting processes involved in the development of the disease. The biological evaluation led to the discovery of two compounds with favorable pharmacological characteristics and ADMET profile. Compounds 17 and 35 are 5-HT6R antagonists (Ki=13nM and Ki=15nM respectively) and cholinesterase inhibitors with distinct mechanisms of enzyme inhibition. Compound 17, a tacrine derivative is a reversible inhibitor of acetyl- and butyrylcholinesterase (IC50=8nM and IC50=24nM respectively), while compound 35 with rivastigmine-derived phenyl N-ethyl-N-methylcarbamate fragment is a selective, pseudo-irreversible inhibitor of butyrylcholinesterase (IC50=455nM). Both compounds inhibit aggregation of amyloid β invitro (75% for compound 17 and 68% for 35at 10μM) moreover, compound 35 is a potent tau aggregation inhibitor in cellulo (79%). In ADMET invitro studies both compounds showed acceptable metabolic stability on mouse liver microsomes (28% and 60% for compound 17 and 35 respectively), no or little effect on CYP3A4 and 2D6 up to a concentration of 10μM and lack of toxicity on HepG2 cell line (IC50 values of 80 and 21μM, for 17 and 35 respectively). Based on the pharmacological characteristics and favorable pharmacokinetic properties, we propose compounds 17 and 35 as an excellent starting point for further optimization and in-depth biological studies.

Structural study of the recognition mechanism of tau antibody Tau2r3 with the key sequence (VQIINK) in tau aggregation

One of the histopathological features of Alzheimer's disease (AD) is higher order neurofibrillary tangles formed by abnormally aggregated tau protein. The sequence 275VQIINK280 in the microtubule-binding domain of tau plays a key role in tau aggregation. Therefore, an aggregation inhibitor targeting the VQIINK region in tau may be an effective therapeutic agent for AD. We have previously shown that the Fab domain (Fab2r3) of a tau antibody that recognizes the VQIINK sequence can inhibit tau aggregation, and we have determined the tertiary structure of the Fab2r3-VQIINK complex. In this report, we determined the tertiary structure of apo Fab2r3 and analyzed differences in the structures of apo Fab2r3 and Fab2r3-VQIINK to examine the ligand recognition mechanism of Fab2r3. In comparison with the Fab2r3-VQIINK structure, there were large differences in the arrangement of the constant and variable domains in apo Fab2r3. Remarkable structural changes were especially observed in the H3 and L3 loop regions of the complementarity determining regions (CDRs) in apo Fab2r3 and the Fab2r3-VQIINK complex. These structural differences in CDRs suggest that formation of hydrophobic pockets suitable for the antigen is important for antigen recognition by tau antibodies.

Arginine and Arginine-Rich Peptides as Modulators of Protein Aggregation and Cytotoxicity Associated With Alzheimer's Disease.

A substantial body of evidence indicates cationic, arginine-rich peptides (CARPs) are effective therapeutic compounds for a range of neurodegenerative pathologies, with beneficial effects including the reduction of excitotoxic cell death and mitochondrial dysfunction. CARPs, therefore, represent an emergent class of promising neurotherapeutics with multimodal mechanisms of action. Arginine itself is a known chaotrope, able to prevent misfolding and aggregation of proteins. The putative role of proteopathies in chronic neurodegenerative diseases such as Alzheimer’s disease (AD) warrants investigation into whether CARPs could also prevent the aggregation and cytotoxicity of amyloidogenic proteins, particularly amyloid-beta and tau. While monomeric arginine is well-established as an inhibitor of protein aggregation in solution, no studies have comprehensively discussed the anti-aggregatory properties of arginine and CARPs on proteins associated with neurodegenerative disease. Here, we review the structural, physicochemical, and self-associative properties of arginine and the guanidinium moiety, to explore the mechanisms underlying the modulation of protein aggregation by monomeric and multimeric arginine molecules. Arginine-rich peptide-based inhibitors of amyloid-beta and tau aggregation are discussed, as well as further modulatory roles which could reduce proteopathic cytotoxicity, in the context of therapeutic development for AD.

Neuroprotective treatment of tauopathies

Tau pathology is now considered to be the main cause of awide spectrum of neurodegenerative diseases, which are collectively referred to as tauopathies. These include primary tauopathies, in which tau plays the main role in the pathogenesis as well as secondary tauopathies, such as Alzheimer's disease, in which amyloid beta also plays a substantial role in the disease process in addition to the tau pathology. Primary tauopathies include progressive supranuclear palsy, corticobasal degeneration, Pick's disease and rare hereditary tauopathies, which are referred to as frontotemporal lobar degeneration with microtubule-associated protein tau (MAPT) mutation. Tauopathies differ from each other pathologically by the affected brain regions and cell types as well as by the biochemical characteristics of the aggregated tau protein. Various tau-centered neuroprotective treatment approaches are currently in preclinical and clinical development. They target different mechanisms, including the reduction of tau expression, inhibition of tau aggregation, dissolution of tau aggregates, improvement of cellular mechanisms to eliminate toxic tau species, stabilization of microtubules and prevention of intercellular tau spreading. This review article gives an overview of tauopathies and the current concepts for the development of disease-modifying treatment.