Role of Tau Protein in Neurodegenerative Diseases and Development of Its Targeted Drugs: A Literature Review.

Tau protein is a microtubule-associated protein that is widely distributed in the central nervous system and maintains and regulates neuronal morphology and function. Tau protein aggregates abnormally and forms neurofibrillary tangles in neurodegenerative diseases, disrupting the structure and function of neurons and leading to neuronal death, which triggers the initiation and progression of neurological disorders. The aggregation of tau protein in neurodegenerative diseases is associated with post-translational modifications, which may affect the hydrophilicity, spatial conformation, and stability of tau protein, promoting tau protein aggregation and the formation of neurofibrillary tangles. Therefore, studying the role of tau protein in neurodegenerative diseases and the mechanism of aberrant aggregation is important for understanding the mechanism of neurodegenerative diseases and finding therapeutic approaches. This review describes the possible mechanisms by which tau protein promotes neurodegenerative diseases, the post-translational modifications of tau protein and associated influencing factors, and the current status of drug discovery and development related to tau protein, which may contribute to the development of new therapeutic approaches to alleviate or treat neurodegenerative diseases.

Pharmacological inhibition of α-synuclein aggregation within liquid condensates.

Aggregated forms of α-synuclein constitute the major component of Lewy bodies, the proteinaceous aggregates characteristic of Parkinson's disease. Emerging evidence suggests that α-synuclein aggregation may occur within liquid condensates formed through phase separation. This mechanism of aggregation creates new challenges and opportunities for drug discovery for Parkinson's disease, which is otherwise still incurable. Here we show that the condensation-driven aggregation pathway of α-synuclein can be inhibited using small molecules. We report that the aminosterol claramine stabilizes α-synuclein condensates and inhibits α-synuclein aggregation within the condensates both in vitro and in a Caenorhabditis elegans model of Parkinson's disease. By using a chemical kinetics approach, we show that the mechanism of action of claramine is to inhibit primary nucleation within the condensates. These results illustrate a possible therapeutic route based on the inhibition of protein aggregation within condensates, a phenomenon likely to be relevant in other neurodegenerative disorders.

Linalool acts as a chemical chaperone by inhibiting amyloid-ß aggregation.

Linalool is a neuroprotective monoterpene found in essential oils from aromatic plants. Linalool's effectiveness in AD animal models has been established previously, but its mechanisms of action remain unclear. Therefore, this study aims to investigate whether linalool binds directly to the amyloid beta (Aß) fibrils to understand it's role in preventing neurodegeneration. The anti-aggregation ability of Linalool was determined using Dithiothreitol (DTT), and thermal aggregation assays followed by Thioflavin T (ThT) binding assay. AD animals were treated with Linalool, and Thioflavin T staining was used to check the binding of linalool to Aß fibrils in rat brain tissue sections. Preliminary studies revealed the anti-aggregation potential of linalool under the thermal and chemical stimulus. Further, in ThT binding assay Linalool inhibited Aß aggregation, binding directly to Aß fibrils. The reduced fluorescence intensity of ThT in AD brain tissues following linalool administration, highlights its neuroprotective potential as a therapeutic agent for AD.

Biological effects and mechanism of ß-amyloid aggregation inhibition by penetrable recombinant human HspB5-ACD structural domain protein.

Alzheimer's disease (AD) is a global medical challenge. Studies have shown that neurotoxicity caused by pathological aggregation of ß-amyloid (Aß) is an important factor leading to AD. Therefore, inhibiting the pathological aggregation of Aß is the key to treating AD. The recombinant human HspB5-ACD structural domain protein (AHspB5) prepared by our group in the previous period has been shown to have anti-amyloid aggregation effects, but its inability to penetrate biological membranes has limited its development. In this study, we prepared a recombinant fusion protein (T-AHspB5) of TAT and AHspB5. In vitro experiments showed that T-AHspB5 inhibited the formation of Aß1-42 protofibrils and had the ability to penetrate the blood-brain barrier; in cellular experiments, T-AHspB5 prevented Aß1-42-induced oxidative stress damage, apoptosis, and inflammatory responses in neuronal cells, and its mechanism of action was related to microglia activation and mitochondria-dependent apoptotic pathway. In animal experiments, T-AHspB5 improved memory and cognitive dysfunction and inhibited pathological changes of AD in APP/PS1 mice. In conclusion, this paper is expected to reveal the intervention mechanism and biological effect of T-AHspB5 on pathological aggregation of Aß1-42, provide a new pathway for the treatment of AD, and lay the foundation for the future development and application of T-AHspB5.

Cyclic-NDGA Effectively Inhibits Human γ-Synuclein Fibrillation, Forms Nontoxic Off-Pathway Species, and Disintegrates Preformed Mature Fibrils.

Parkinson's disease arises from protein misfolding, aggregation, and fibrillation and is characterized by LB (Lewy body) deposits, which contain the protein α-synuclein (α-syn) as their major component. Another synuclein, γ-synuclein (γ-syn), coexists with α-syn in Lewy bodies and is also implicated in various types of cancers, especially breast cancer. It is known to seed α-syn fibrillation after its oxidation at methionine residue, thereby contributing in synucleinopathy. Despite its involvement in synucleinopathy, the search for small molecule inhibitors and modulators of γ-syn fibrillation remains largely unexplored. This work reveals the modulatory properties of cyclic-nordihydroguaiaretic acid (cNDGA), a natural polyphenol, on the structural and aggregational properties of human γ-syn employing various biophysical and structural tools, namely, thioflavin T (ThT) fluorescence, Rayleigh light scattering, 8-anilinonaphthalene-1-sulfonic acid binding, far-UV circular dichroism (CD), Fourier transform infrared spectroscopy (FTIR) spectroscopy, atomic force microscopy, ITC, molecular docking, and MTT-toxicity assay. cNDGA was observed to modulate the fibrillation of γ-syn to form off-pathway amorphous species that are nontoxic in nature at as low as 75 µM concentration. The modulation is dependent on oxidizing conditions, with cNDGA weakly interacting (Kd ∼10-5 M) with the residues at the N-terminal of γ-syn protein as investigated by isothermal titration calorimetry and molecular docking, respectively. Increasing cNDGA concentration results in an increased recovery of monomeric γ-syn as shown by sodium dodecyl sulfate and native-polyacrylamide gel electrophoresis. The retention of native structural properties of γ-syn in the presence of cNDGA was further confirmed by far-UV CD and FTIR. In addition, cNDGA is most effective in suppression of fibrillation when added at the beginning of the fibrillation kinetics and is also capable of disintegrating the preformed mature fibrils. These findings could, therefore, pave the ways for further exploring cNDGA as a potential therapeutic against γ-synucleinopathies.

Protein mimetic 2D FAST rescues alpha synuclein aggregation mediated early and post disease Parkinson's phenotypes.

Abberent protein-protein interactions potentiate many diseases and one example is the toxic, self-assembly of α-Synuclein in the dopaminergic neurons of patients with Parkinson's disease; therefore, a potential therapeutic strategy is the small molecule modulation of α-Synuclein aggregation. In this work, we develop an Oligopyridylamide based 2-dimensional Fragment-Assisted Structure-based Technique to identify antagonists of α-Synuclein aggregation. The technique utilizes a fragment-based screening of an extensive array of non-proteinogenic side chains in Oligopyridylamides, leading to the identification of NS132 as an antagonist of the multiple facets of α-Synuclein aggregation. We further identify a more cell permeable analog (NS163) without sacrificing activity. Oligopyridylamides rescue α-Synuclein aggregation mediated Parkinson's disease phenotypes in dopaminergic neurons in early and post disease Caenorhabditis elegans models. We forsee tremendous potential in our technique to identify lead therapeutics for Parkinson's disease and other diseases as it is expandable to other oligoamide scaffolds and a larger array of side chains.

The mechanistic interaction, aggregation and neurotoxicity of α-synuclein after interaction with glycyrrhizic acid: Modulation of synucleinopathies.

This article reveals the binding mechanism between glycyrrhizic acid (GA) and α-synuclein to may provide further information for the modulation of synucleinopathies using bioactive compounds. Therefore, the inhibitory activities of GA against α-synuclein aggregation and induced neurotoxicity were evaluated using different assays. Results showed that α-synuclein-GA binding was mediated by intermolecular hydrogen bonds leading to the formation of a slightly folded complex. Theoretical studies revealed that GA binds to the N-terminal domain of α-synuclein and triggers a compact structure around a major part of the N-terminal and the NAC regions along with fluctuations in the C-terminal domain, which are prerequisites for the inhibition of α-synuclein aggregation. Then, the cellular assays showed that GA as a potential small molecule can inhibit the oligomerization of α-synuclein and relevant neurotoxicity through modulation of neural viability, membrane leakage, and ROS formation in a concentration-dependent manner. As a result, the primary mechanism of GA's anti-aggregation and neuroprotective activities is the reorganized α-synuclein structure and fluctuating C-terminal domain, which promotes long-range transient intramolecular contacts between the N-terminal and the C-terminal domain.

Computer aided therapeutic tripeptide design, in alleviating the pathogenic proclivities of nocuous α-synuclein fibrils.

Parkinson's disorder (PD) exacerbates neuronal degeneration of motor nerves, thereby effectuating uncoordinated movements and tremors. Aberrant alpha-synuclein (α-syn) is culpable of triggering PD, wherein cytotoxic amyloid aggregates of α-syn get deposited in motor neurons to instigate neuro-degeneration. Amyloid aggregates, typically rich in beta sheets are cardinal targets to mitigate their neurotoxic effects. In this analysis, owing to their interaction specificity, we formulated an efficacious tripeptide out of the aggregation-prone region of α-syn protein. With the help of a proficient computational pipeline, systematic peptide shortening and an adept molecular simulation platform, we formulated a tripeptide, VAV from α-syn structure based hexapeptide KISVRV. Indeed, the VAV tripeptide was able to effectively mitigate the α-syn amyloid fibrils' dynamic rate of beta-sheet formation. Additional trajectory analyses of the VAV- α-syn complex indicated that, upon its dynamic interaction, VAV efficiently altered the distinct pathogenic structural dynamics of α-syn, further advocating its potential in alleviating aberrant α-syn's amyloidogenic proclivities. Consistent findings from various computational analyses have led us to surmise that VAV could potentially re-alter the pathogenic conformational orientation of α-syn, essential to mitigate its cytotoxicity. Hence, VAV tripeptide could be an efficacious therapeutic candidate to efficiently ameliorate aberrant α-syn amyloid mediated neurotoxicity, eventually attenuating the nocuous effects of PD.Communicated by Ramaswamy H. Sarma.

De novo designed protein inhibitors of amyloid aggregation and seeding.

Neurodegenerative diseases are characterized by the pathologic accumulation of aggregated proteins. Known as amyloid, these fibrillar aggregates include proteins such as tau and amyloid-ß (Aß) in Alzheimer's disease (AD) and alpha-synuclein (αSyn) in Parkinson's disease (PD). The development and spread of amyloid fibrils within the brain correlates with disease onset and progression, and inhibiting amyloid formation is a possible route toward therapeutic development. Recent advances have enabled the determination of amyloid fibril structures to atomic-level resolution, improving the possibility of structure-based inhibitor design. In this work, we use these amyloid structures to design inhibitors that bind to the ends of fibrils, "capping" them so as to prevent further growth. Using de novo protein design, we develop a library of miniprotein inhibitors of 35 to 48 residues that target the amyloid structures of tau, Aß, and αSyn. Biophysical characterization of top in silico designed inhibitors shows they form stable folds, have no sequence similarity to naturally occurring proteins, and specifically prevent the aggregation of their targeted amyloid-prone proteins in vitro. The inhibitors also prevent the seeded aggregation and toxicity of fibrils in cells. In vivo evaluation reveals their ability to reduce aggregation and rescue motor deficits in Caenorhabditis elegans models of PD and AD.

α-Crystallin chaperone mimetic drugs inhibit lens γ-crystallin aggregation: Potential role for cataract prevention.

Γ-Crystallins play a major role in age-related lens transparency. Their destabilization by mutations and physical chemical insults are associated with cataract formation. Therefore, drugs that increase their stability should have anticataract properties. To this end, we screened 2560 Federal Drug Agency-approved drugs and natural compounds for their ability to suppress or worsen H2O2 and/or heat-mediated aggregation of bovine γ-crystallins. The top two drugs, closantel (C), an antihelminthic drug, and gambogic acid (G), a xanthonoid, attenuated thermal-induced protein unfolding and aggregation as shown by turbidimetry fluorescence spectroscopy dynamic light scattering and electron microscopy of human or mouse recombinant crystallins. Furthermore, binding studies using fluorescence inhibition and hydrophobic pocket-binding molecule bis-8-anilino-1-naphthalene sulfonic acid revealed static binding of C and G to hydrophobic sites with medium-to-low affinity. Molecular docking to HγD and other γ-crystallins revealed two binding sites, one in the "NC pocket" (residues 50-150) of HγD and one spanning the "NC tail" (residues 56-61 to 168-174 in the C-terminal domain). Multiple binding sites overlap with those of the protective mini αA-crystallin chaperone MAC peptide. Mechanistic studies using bis-8-anilino-1-naphthalene sulfonic acid as a proxy drug showed that it bound to MAC sites, improved Tm of both H2O2 oxidized and native human gamma D, and suppressed turbidity of oxidized HγD, most likely by trapping exposed hydrophobic sites. The extent to which these drugs act as α-crystallin mimetics and reduce cataract progression remains to be demonstrated. This study provides initial insights into binding properties of C and G to γ-crystallins.

The small aromatic compound SynuClean-D inhibits the aggregation and seeded polymerization of multiple α-synuclein strains.

Parkinson's disease is a neurodegenerative disorder characterized by the loss of dopaminergic neurons in the substantia nigra, as well as the accumulation of intraneuronal proteinaceous inclusions known as Lewy bodies and Lewy neurites. The major protein component of Lewy inclusions is the intrinsically disordered protein α-synuclein (α-Syn), which can adopt diverse amyloid structures. Different conformational strains of α-Syn have been proposed to be related to the onset of distinct synucleinopathies; however, how specific amyloid fibrils cause distinctive pathological traits is not clear. Here, we generated three different α-Syn amyloid conformations at different pH and salt concentrations and analyzed the activity of SynuClean-D (SC-D), a small aromatic molecule, on these strains. We show that incubation of α-Syn with SC-D reduced the formation of aggregates and the seeded polymerization of α-Syn in all cases. Moreover, we found that SC-D exhibited a general fibril disaggregation activity. Finally, we demonstrate that treatment with SC-D also reduced strain-specific intracellular accumulation of phosphorylated α-Syn inclusions. Taken together, we conclude that SC-D may be a promising hit compound to inhibit polymorphic α-Syn aggregation.

A review on prevention of glycation of proteins: Potential therapeutic substances to mitigate the severity of diabetes complications.

Non-enzymatic reaction involving carbonyl of reducing sugars and amino groups in proteins produces advanced glycation end products (AGEs). AGE accumulation in vivo is a crucial factor in the progression of metabolic and pathophysiological mechanisms like obesity, diabetes, coronary artery disease, neurological disorders, and chronic renal failure. The body's own defense mechanism, synthetic inhibitors, and natural inhibitors can all help to prevent the glycation of proteins. Synthetic inhibitors have the potential to suppress the glycation of proteins through a variety of pathways. They could avoid Amadori product development by tampering with the addition of sugars to the proteins. Besides which, the free radical scavenging and blocking crosslink formation could be another mechanism behind their anti-glycation properties. In comparison with synthetic substances, naturally occurring plant products have been found to be comparatively non-toxic, cheap, and usable in an ingestible form. This review gives a brief introduction of the Maillard reaction; formation, characterization and pathology related to AGEs, potential therapeutic approaches against glycation, natural and synthetic inhibitors of glycation and their probable mechanism of action. The scientific community could get benefit from the combined knowledge about important molecules, which will further guide to the design and development of new pharmaceutical compounds.

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.

Diplazium esculentum (Retz.) Sw. reduces BACE-1 activities and amyloid peptides accumulation in Drosophila models of Alzheimer's disease.

Alzheimer's disease (AD), one type of dementia, is a complex disease affecting people globally with limited drug treatment. Thus, natural products are currently of interest as promising candidates because of their cost-effectiveness and multi-target abilities. Diplazium esculentum (Retz.) Sw., an edible fern, inhibited acetylcholinesterase in vitro, inferring that it might be a promising candidate for AD treatment by supporting cholinergic neurons. However, evidence demonstrating anti-AD properties of this edible plant via inhibiting of neurotoxic peptides production, amyloid beta (Aß), both in vitro and in vivo is lacking. Thus, the anti-AD properties of D. esculentum extract both in vitro and in Drosophila models of Aß-mediated toxicity were elucidated. Findings showed that an ethanolic extract exhibited high phenolics and flavonoids, contributing to antioxidant and inhibitory activities against AD-related enzymes. Notably, the extract acted as a BACE-1 blocker and reduced amyloid beta 42 (Aß42) peptides in Drosophila models, resulting in improved locomotor behaviors. Information gained from this study suggested that D. esculentum showed potential for AD amelioration and prevention. Further investigations in vertebrates or humans are required to determine the effective doses of D. esculentum against AD, particularly via amyloidogenic pathway.

Ca2+ administration prevents α-synuclein proteotoxicity by stimulating calcineurin-dependent lysosomal proteolysis.

The capacity of a cell to maintain proteostasis progressively declines during aging. Virtually all age-associated neurodegenerative disorders associated with aggregation of neurotoxic proteins are linked to defects in the cellular proteostasis network, including insufficient lysosomal hydrolysis. Here, we report that proteotoxicity in yeast and Drosophila models for Parkinson's disease can be prevented by increasing the bioavailability of Ca2+, which adjusts intracellular Ca2+ handling and boosts lysosomal proteolysis. Heterologous expression of human α-synuclein (αSyn), a protein critically linked to Parkinson's disease, selectively increases total cellular Ca2+ content, while the levels of manganese and iron remain unchanged. Disrupted Ca2+ homeostasis results in inhibition of the lysosomal protease cathepsin D and triggers premature cellular and organismal death. External administration of Ca2+ reduces αSyn oligomerization, stimulates cathepsin D activity and in consequence restores survival, which critically depends on the Ca2+/calmodulin-dependent phosphatase calcineurin. In flies, increasing the availability of Ca2+ discloses a neuroprotective role of αSyn upon manganese overload. In sum, we establish a molecular interplay between cathepsin D and calcineurin that can be activated by Ca2+ administration to counteract αSyn proteotoxicity.

Aberrant enteric neuromuscular system and dysbiosis in amyotrophic lateral sclerosis.

Amyotrophic Lateral Sclerosis is a neuromuscular disease characterized by the progressive death of motor neurons and muscle atrophy. The gastrointestinal symptoms in ALS patients were largely ignored or underestimated. The relationship between the enteric neuromuscular system and microbiome in ALS progression is unknown. We performed longitudinal studies on the enteric neuron system (ENS) and microbiome in the ALS human-SOD1G93A (Superoxide Dismutase 1) transgenic mice. We treated age-matched wild-type and ALS mice with butyrate or antibiotics to investigate the microbiome and neuromuscular functions. We examined intestinal mobility, microbiome, an ENS marker GFAP (Glial Fibrillary Acidic Protein), a smooth muscle marker (SMMHC, Smooth Muscle Myosin Heavy Chain), and human colonoids. The distribution of human-G93A-SOD1 protein was tested as an indicator of ALS progression. At 2-month-old before ALS onset, SOD1G93A mice had significantly lower intestinal mobility, decreased grip strength, and reduced time in the rotarod. We observed increased GFAP and decreased SMMHC expression. These changes correlated with consistent increased aggregation of mutated SOD1G93A in the colon, small intestine, and spinal cord. Butyrate or antibiotics treated SOD1G93A mice had a significantly longer latency to fall in the rotarod test, reduced SOD1G93A aggregation, and enhanced enteric neuromuscular function. Feces from 2-month-old SOD1G93A mice significantly enhanced SOD1G93A aggregation in human colonoids transfected with a SOD1G93A-GFP plasmid. Longitudinal studies of microbiome data further showed the altered bacterial community related to autoimmunity (e.g., Clostridium sp. ASF502, Lachnospiraceae bacterium A4), inflammation (e.g., Enterohabdus Muris,), and metabolism (e.g., Desulfovibrio fairfieldensis) at 1- and 2-month-old SOD1G93A mice, suggesting the early microbial contribution to the pathological changes. We have demonstrated a novel link between the microbiome, hSOD1G93A aggregation, and intestinal mobility. Dysbiosis occurred at the early stage of the ALS mice before observed mutated-SOD1 aggregation and dysfunction of ENS. Manipulating the microbiome improves the muscle performance of SOD1G93A mice. We provide insights into the fundamentals of intestinal neuromuscular function and microbiome in ALS.

(De)stabilization of Alpha-Synuclein Fibrillary Aggregation by Charged and Uncharged Surfactants.

Parkinson's disease (PD) is the second most common neurodegenerative disorder. An important hallmark of PD involves the pathological aggregation of proteins in structures known as Lewy bodies. The major component of these proteinaceous inclusions is alpha (α)-synuclein. In different conditions, α-synuclein can assume conformations rich in either α-helix or ß-sheets. The mechanisms of α-synuclein misfolding, aggregation, and fibrillation remain unknown, but it is thought that ß-sheet conformation of α-synuclein is responsible for its associated toxic mechanisms. To gain fundamental insights into the process of α-synuclein misfolding and aggregation, the secondary structure of this protein in the presence of charged and non-charged surfactant solutions was characterized. The selected surfactants were (anionic) sodium dodecyl sulphate (SDS), (cationic) cetyltrimethylammonium chloride (CTAC), and (uncharged) octyl ß-D-glucopyranoside (OG). The effect of surfactants in α-synuclein misfolding was assessed by ultra-structural analyses, in vitro aggregation assays, and secondary structure analyses. The α-synuclein aggregation in the presence of negatively charged SDS suggests that SDS-monomer complexes stimulate the aggregation process. A reduction in the electrostatic repulsion between N- and C-terminal and in the hydrophobic interactions between the NAC (non-amyloid beta component) region and the C-terminal seems to be important to undergo aggregation. Fourier transform infrared spectroscopy (FTIR) measurements show that ß-sheet structures comprise the assembly of the fibrils.

An in vivo Caenorhabditis elegans model for therapeutic research in human prion diseases.

Human prion diseases are fatal neurodegenerative disorders that include sporadic, infectious and genetic forms. Inherited Creutzfeldt-Jakob disease due to the E200K mutation of the prion protein-coding gene is the most common form of genetic prion disease. The phenotype resembles that of sporadic Creutzfeldt-Jakob disease at both the clinical and pathological levels, with a median disease duration of 4 months. To date, there is no available treatment for delaying the occurrence or slowing the progression of human prion diseases. Existing in vivo models do not allow high-throughput approaches that may facilitate the discovery of compounds targeting pathological assemblies of human prion protein or their effects on neuronal survival. Here, we generated a genetic model in the nematode Caenorhabditis elegans, which is devoid of any homologue of the prion protein, by expressing human prion protein with the E200K mutation in the mechanosensitive neuronal system. Expression of E200K prion protein induced a specific behavioural pattern and neurodegeneration of green fluorescent protein-expressing mechanosensitive neurons, in addition to the formation of intraneuronal inclusions associated with the accumulation of a protease-resistant form of the prion protein. We demonstrated that this experimental system is a powerful tool for investigating the efficacy of anti-prion compounds on both prion-induced neurodegeneration and prion protein misfolding, as well as in the context of human prion protein. Within a library of 320 compounds that have been approved for human use and cross the blood-brain barrier, we identified five molecules that were active against the aggregation of the E200K prion protein and the neurodegeneration it induced in transgenic animals. This model breaks a technological limitation in prion therapeutic research and provides a key tool to study the deleterious effects of misfolded prion protein in a well-described neuronal system.

Thioridazine reverts the phenotype in cellular and Drosophila models of amyotrophic lateral sclerosis by enhancing TDP-43 aggregate clearance.

Brain inclusions mainly composed of misfolded and aggregated TAR DNA binding protein 43 (TDP-43), are characteristic hallmarks of amyotrophic lateral sclerosis (ALS). Irrespective of the role played by the inclusions, their reduction represents an important therapeutic pathway that is worth exploring. Their removal can either lead to the recovery of TDP-43 function by removing the self-templating conformers that sequester the protein in the inclusions, and/or eliminate any potential intrinsic toxicity of the aggregates. The search for curative therapies has been hampered by the lack of ALS models for use in high-throughput screening. We adapted, optimised, and extensively characterised our previous ALS cellular model for such use. The model demonstrated efficient aggregation of endogenous TDP-43, and concomitant loss of its splicing regulation function. We provided a proof-of-principle for its eventual use in high-throughput screening using compounds of the tricyclic family and showed that recovery of TDP-43 function can be achieved by the enhanced removal of TDP-43 aggregates by these compounds. We observed that the degradation of the aggregates occurs independent of the autophagy pathway beyond autophagosome-lysosome fusion, but requires a functional proteasome pathway. The in vivo translational effect of the cellular model was tested with two of these compounds in a Drosophila model expressing a construct analogous to the cellular model, where thioridazine significantly improved the locomotive defect. Our findings have important implications as thioridazine cleared TDP-43 aggregates and recovered TDP-43 functionality. This study also highlights the importance of a two-stage, in vitro and in vivo model system to cross-check the search for small molecules that can clear TDP-43 aggregates in TDP-43 proteinopathies.

Effects of Aß-derived peptide fragments on fibrillogenesis of Aß.

Amyloid ß (Aß) peptide aggregation plays a central role in Alzheimer's disease (AD) etiology. AD drug candidates have included small molecules or peptides directed towards inhibition of Aß fibrillogenesis. Although some Aß-derived peptide fragments suppress Aß fibril growth, comprehensive analysis of inhibitory potencies of peptide fragments along the whole Aß sequence has not been reported. The aim of this work is (a) to identify the region(s) of Aß with highest propensities for aggregation and (b) to use those fragments to inhibit Aß fibrillogenesis. Structural and aggregation properties of the parent Aß1-42 peptide and seven overlapping peptide fragments have been studied, i.e. Aß1-10 (P1), Aß6-15 (P2), Aß11-20 (P3), Aß16-25 (P4), Aß21-30 (P5), Aß26-36 (P6), and Aß31-42 (P7). Structural transitions of the peptides in aqueous buffer have been monitored by circular dichroism and Fourier transform infrared spectroscopy. Aggregation and fibrillogenesis were analyzed by light scattering and thioflavin-T fluorescence. The mode of peptide-peptide interactions was characterized by fluorescence resonance energy transfer. Three peptide fragments, P3, P6, and P7, exhibited exceptionally high propensity for ß-sheet formation and aggregation. Remarkably, only P3 and P6 exerted strong inhibitory effect on the aggregation of Aß1-42, whereas P7 and P2 displayed moderate inhibitory potency. It is proposed that P3 and P6 intercalate between Aß1-42 molecules and thereby inhibit Aß1-42 aggregation. These findings may facilitate therapeutic strategies of inhibition of Aß fibrillogenesis by Aß-derived peptides.