A hybrid molecule that prohibits amyloid fibrils and alleviates neuronal toxicity induced by beta-amyloid (1-42)

Inhibition of oligomeric amyloid beta (Abeta) peptide or fibril formation has emerged as a major therapeutic target for developing new drugs for Alzheimer's disease. We focused on developing inhibitors by synthesizing hybrid molecules of ferulic acid and styryl benzene, which has been known as a fibril binder. Initially, cell-based assay was carried out to evaluate the effective compound. A selected effector, 1, alleviated the Abeta-induced neuronal toxicity in differentiated SH-SY5Y human neuroblastoma cells. The effector could also inhibit Abeta fibril formation, monitored by thioflavin T fluorescence intensity assay and transmitted electron microscopic images. A strong binding affinity of 1 to non-fibrous monomer-like Abeta, which was immobilized to a surface chip, was measured using a surface plasmon resonance technique. The data suggest that the effector shifts the equilibrium of multimeric Abeta, inhibiting the pathogenic oligomer or fibril formation.     

Self-assembly of beta-amyloid 42 is retarded by small molecular ligands at the stage of structural intermediates

Assemblyof the amyloid-beta peptide (Abeta) into fibrils and its deposition in distinct brain areas is considered responsible for the pathogenesis of Alzheimer's disease (AD). Thus, inhibition of fibril assembly is a potential strategy for therapeutic intervention. Electron cryomicroscopy was used to monitor the initial, native assembly structure of Abeta42. In addition to the known fibrillar intermediates, a nonfibrillar, polymeric sheet-like structure was identified. A temporary sequence of supramolecular structures was revealed with (i) polymeric Abeta42 sheets during the onset of assembly, inversely related to the appearance of (ii) fibril intermediates, which again are time-dependently replaced by (iii) mature fibrils. A cell-based primary screening assay was used to identify compounds that decrease Abeta42-induced toxicity. Hit compounds were further assayed for binding to Abeta42, radical scavenger activity, and their influence on the assembly structure of Abeta42. One compound, Ro 90-7501, was found to efficiently retard mature fibril formation, while extended polymeric Abeta42 sheets and fibrillar intermediates are accumulated. Ro 90-7501 may serve as a prototypic inhibitor for Abeta42 fibril formation and as a tool for studying the molecular mechanism of fibril assembly.     

Search for dual function inhibitors for Alzheimer's disease: synthesis and biological activity of acetylcholinesterase inhibitors of pyridinium-type and their Abeta fibril formation inhibition capacity

Alzheimer's disease (AD) represents the most common neurodegenerative disorder, which is expressed through decline of mental function. Current treatment approaches include acetylcholinesterase inhibitors and NMDA-receptor partial antagonists. The most explored recent approaches that are closely related to the pathogenesis of this disease based on formally articulated amyloid hypothesis are: Abeta fibril formation inhibitors, amyloid precursor protein, and secretase inhibitors. [Scarpini, E.; Scheltens, P.; Feldman, H. Lancet Neurol.2003, 2, 539] In view of the development of new AChE inhibitors as drugs capable of reducing the symptoms of AD, the capacity of newly synthesized AChE inhibitors of pyridinium-type to inhibit the AChE was examined and compared to those of other inhibitors of this type presented earlier. [Kapková, P.; Stiefl, N.; Sürig, U.; Engels, B.; Baumann, K.; Holzgrabe, U. Arch. Pharm. Pharm. Med. Chem.2003, 336, 523; Alptüzun, V.; Kapková, P.; Baumann, K.; Erciyas, E.; Holzgrabe, U. J. Pharm. Pharmacol.2003, 55, 1397] Furthermore, the anti-Abeta fibril formation property of AChE inhibitors of pyridinium- and bispyridinium-type was evaluated to expand their activity profile and to reveal potential additive pharmacological effects which may reinforce their therapeutic application besides their capacity of increasing acetylcholine levels. Abeta fibril formation studies were performed by means of thioflavin T fluorescence assay.     

Designing peptide inhibitors for oligomerization and toxicity of Alzheimer's beta-amyloid peptide

Convergent biochemical and genetic evidence suggests that the formation of beta-amyloid (Abeta) deposits in the brain is an important and, probably, seminal step in the development of Alzheimer's disease (AD). Recent studies support the hypothesis that Abeta soluble oligomers are the pathogenic species that prompt the disease. Inhibiting Abeta self-oligomerization could, therefore, provide a novel approach to treating the underlying cause of AD. Here, we designed potential peptide-based aggregation inhibitors containing Abeta amino acid sequences (KLVFF) from part of the binding region responsible for Abeta self-association (residues 16-20), with RG-/-GR residues added at their N- and C-terminal ends to aid solubility. Two such peptides (RGKLVFFGR, named OR1, and RGKLVFFGR-NH2, named OR2) were effective inhibitors of Abeta fibril formation, but only one of these peptides (OR2) inhibited Abeta oligomer formation. Interestingly, this same OR2 peptide was the only effective inhibitor of Abeta toxicity toward human neuroblastoma SH-SY5Y cells. Our data support the idea that Abeta oligomers are responsible for the cytotoxic effects of Abeta and identify a potential peptide inhibitor for further development as a novel therapy for AD.     

Abeta aggregation inhibitors. Part 1: Synthesis and biological activity of phenylazo benzenesulfonamides

Phenylazo benzenesulfonamides were designed and synthesized as beta-amyloid (Abeta40) fibril assembly inhibitors, and evaluated for inhibition of Abeta40 aggregation and neurotoxicity using rat cortical neurons. Compound 2 (LB-152) was the most potent compound in this study, and the para-NMe(2) group on the end of the phenylazo moiety may play an important role in preventing Abeta40 fibril formation. LB-152 provides a new lead for further development of potential beta-amyloid aggregation inhibitors to treat AD.     

Direct observation of Abeta amyloid fibril growth and inhibition

Amyloid fibril formation is a phenomenon common to many proteins and peptides, including amyloid beta (Abeta) peptide associated with Alzheimer's disease. To clarify the mechanism of fibril formation and to create inhibitors, real-time monitoring of fibril growth is essential. Here, seed-dependent amyloid fibril growth of Abeta(1-40) was visualized in real-time at the single fibril level using total internal reflection fluorescence microscopy (TIRFM) combined with the binding of thioflavin T, an amyloid-specific fluorescence dye. The clear image and remarkable length of the fibrils enabled an exact analysis of the rate of growth of individual fibrils, indicating that the fibril growth was a highly cooperative process extending the fibril ends at a constant rate. It has been known that Abeta amyloid formation is a stereospecific reaction and the stability is affected by l/d-amino acid replacement. Focusing on these aspects, we designed several analogues of Abeta(25-35), a cytotoxic fragment of Abeta(1-40), consisting of l and d-amino acid residues, and examined their inhibitory effects by TIRFM. Some chimeric Abeta(25-35) peptides inhibited the fibril growth of Abeta(25-35) strongly, although they could not inhibit the growth of Abeta(1-40). The results suggest that a more rational design of stereospecific inhibitors, combined with real-time monitoring of fibril growth, will be useful to invent a potent inhibitor preventing the amyloid fibril growth of Abeta(1-40) and other proteins.     

Practical assay and molecular mechanism of aggregation inhibitors of beta-amyloid.

Beta-Amyloid peptide (Abeta) is the main protein component of neuritic plaques in the brain of patients of Alzheimer's disease (AD), and its neurotoxicity would be exposed by the formation of aggregates. The aggregation inhibitors composed of an Abeta recognition element (KLVFF) and a hydrophilic moiety are evaluated by a novel fluorescence assay. These compounds inhibit growth of the model aggregates on the KLVFF immobilized surface. In addition, some compounds also possess disrupting activities of preformed aggregates. These compounds could be a key candidate for therapeutic drugs for AD by their novel molecular mechanisms.     

Salvianolic acid B inhibits Abeta fibril formation and disaggregates preformed fibrils and protects against Abeta-induced cytotoxicty

One of the major pathological features of Alzheimer's disease (AD) is the appearance of senile plaques characterized by extracellular aggregation of amyloid beta-peptide (Abeta) fibrils. Inhibition of Abeta fibril aggregation is therefore viewed as one possible method to halt the progression of AD. Salvianolic acid B (Sal B) is an active ingredient isolated from Salvia miltiorrhiza, a Chinese herbal medicine commonly used for the treatment of cardiovascular and cerebrovascular disorders. Recent findings show that Sal B prevents Abeta-induced cytotoxicity in a rat neural cell line. To understand the mechanism of Sal B-mediated neuroprotection, its effects on the inhibition of Abeta1-40 fibril formation and destabilization of the preformed Abeta1-40 fibrils were studied. The results were obtained using Thioflavin T fluorescence assay and Abeta aggregating immunoassay. We found that Sal B can inhibit fibril aggregation (IC(50): 1.54-5.37 microM) as well as destabilize preformed Abeta fibril (IC(50): 5.00-5.19 microM) in a dose- and time-dependent manner. Sal B is a better aggregation inhibitor than ferulic acid but less active than curcumin in the inhibition of Abeta1-40 aggregation. In electron microscope study, Sal B-treated Abeta1-40 fibrils are seen in various stages of shortening or wrinkling with numerous deformed aggregates of amorphous structure. Circular dichroism data indicate that Sal B dose dependently prevents the formation of beta-structured aggregates of Abeta1-40. Addition of preincubated Sal B with Abeta1-42 significantly reduces its cytotoxic effects on human neuroblastoma SH-SY5Y cells. These results suggest that Sal B has therapeutic potential in the treatment of AD, and warrant its study in animal models.     

The binding of resveratrol to monomer and fibril amyloid beta

As currently understood, Alzheimer’s disease (AD) is a chronic neurodegenerative disorder that is driven by the aggregation of amyloid beta (Aβ) protein. It has been shown that resveratrol (RES) may attenuate amyloid β peptide-induced toxicity, promote Aβ clearance and reduce senile plaques. However, it remains to be determined whether RES could interact directly with Aβ. The aim of the present study was to examine the direct binding of RES to monomer and fibril Aβ. Using surface plasmon resonance (SPR) and proton nuclear magnetic resonance (¹H NMR), our results identified the direct binding of RES to Aβ. The ability of RES to bind to both fibril and monomer Aβ(1–40 and 1–42) was further analyzed by SPR. The binding response of RES to fAβ(1–42) was higher than that to monomer Aβ(1–42), whereas the binding response of RES to fAβ(1–40) was lower than that to monomer Aβ(1–40). The K_D of RES for fibril Aβ(1–40 or 1–42) was higher than that for the corresponding monomer Aβ. Compared to the control compound Congo red (CR), the binding responses of RES to monomer Aβ(1–42) and Aβ(1–40) were stronger, but binding to fibril Aβ(1–42) was weaker, and the K_Ds of RES with both monomer and fibril Aβ(1–40) and Aβ(1–42) were higher than that of CR. When Aβ(1–40 or 1–42) was co-incubated with RES (50 μM), the thioflavin T fluorescence of the mixture was weakened, and the number and length of amyloid fibrils were decreased. Furthermore, the results of staining in consecutive brain slices from AD patients showed that RES (10⁻⁴ M) could stain senile plaques. These results indicated that RES could bind directly to Aβ in different states, which may provide new insight into the protective properties of RES against AD.     

Pentapeptide amides interfere with the aggregation of beta-amyloid peptide of Alzheimer's disease

Amyloid peptides (Abeta) play a central role in the pathogenesis of Alzheimer's disease (AD). The aggregation of Abeta molecules leads to fibril and plaque formation. Fibrillogenesis is at the same time a marker and an indirect cause of AD. Inhibition of the aggregation of Abeta could be a realistic therapy for the illness. Beta sheet breakers (BSBs) are one type of fibrillogenesis inhibitors. The first BSB peptides were designed by Tjernberg et al. (1996) and Soto et al. (1998). These pentapeptides have proved their efficiency in vitro and in vivo. In the present study, the effects of two pentapeptide amides are reported. These compounds were designed by using the C-terminal sequence of the amyloid peptide as a template. Biological assays were applied to demonstrate efficiency. Modes of action were studied by FT-IR spectroscopy and molecular modeling methods.     

Molecular mechanisms initiating amyloid beta-fibril formation in Alzheimer's disease

The deposition of aggregated amyloid beta-protein (Abeta) in the human brain is a major lesion in Alzheimer' disease (AD). The process of Abeta fibril formation is associated with a cascade of neuropathogenic events that induces brain neurodegeneration leading to the cognitive and behavioral decline characteristic of AD. Although a detailed knowledge of Abeta assembly is crucial for the development of new therapeutic approaches, our understanding of the molecular mechanisms underlying the initiation of Abeta fibril formation remains very incomplete. The genetic defects responsible for familial AD influence fibrillogenesis. In a majority of familial cases determined by amyloid precursor protein (APP) and presenilin (PS) mutations, a significant overproduction of Abeta and an increase in the Abeta42/Abeta40 ratio are observed. Recently, it was shown that the two main alloforms of Abeta have distinct biological activity and behaviour at the earliest stage of assembly. In vitro studies demonstrated that Abeta42 monomers, but not Abeta40, form initial and minimal structures (pentamer/hexamer units called paranuclei) that can oligomerize to larger forms. It is now apparent that Abeta oligomers and protofibrils are more neurotoxic than mature Abeta fibrils or amyloid plaques. The neurotoxicity of the prefibrillar aggregates appears to result from their ability to impair fundamental cellular processes by interacting with the cellular membrane, causing oxidative stress and increasing free Ca(2+) that eventually lead to apoptotic cell death.     

The molecular chaperone alphaB-crystallin enhances amyloid beta neurotoxicity.

Amyloid beta (Abeta) is a 40- to 42-residue peptide that is implicated in the pathogenesis of Alzheimer's Disease (AD). As a result of conformational changes, Abeta assembles into neurotoxic fibrils deposited as 'plaques' in the diseased brain. In AD brains, the small heat shock proteins (sHsps) alphaB-crystallin and Hsp27 occur at increased levels and colocalize with these plaques. In vitro, sHsps act as molecular chaperones that recognize unfolding peptides and prevent their aggregation. The presence of sHsps in AD brains may thus reflect an attempt to prevent amyloid fibril formation and toxicity. Here we report that alphaB-crystallin does indeed prevent in vitro fibril formation of Abeta(1-40). However, rather than protecting cultured neurons against Abeta(1-40) toxicity, alphaB-crystallin actually increases the toxic effect. This indicates that the interaction of alphaB-crystallin with conformationally altering Abeta(1-40) may keep the latter in a nonfibrillar, yet highly toxic form.     

Approaches to discovery and characterization of inhibitors of amyloid beta-peptide polymerization

Polymerization of the amyloid beta-peptide (Abeta) has been identified as a major feature of the pathogenesis of Alzheimer's disease (AD). Inhibition of the formation of these toxic polymers of Abeta has thus emerged as an approach to developing therapeutics for AD. Techniques for studying Abeta polymerization include the use of fibril nucleation and extension assays in a variety of formats. Detection of polymeric forms of Abeta has been achieved using turbidity, dye binding, light scattering and toxicity among other methods. Direct and indirect methods have been described for the measurement of binding affinities for Abeta fibrils. Imaging techniques include electron microscopy, X-ray diffraction and atomic force microscopy. These techniques have been used to characterize different classes of compounds that inhibit the formation of Abeta polymers. These compounds include dyes such as Congo Red, the antibiotic rifampicin, the anthracycline 4'-iodo-4'-deoxydoxorubicin, and a large variety of Abeta-derived peptides and modified peptides, among other reported inhibitors.     

Curcumin inhibits formation of amyloid beta oligomers and fibrils, binds plaques, and reduces amyloid in vivo

Alzheimer's disease (AD) involves amyloid beta (Abeta) accumulation, oxidative damage, and inflammation, and risk is reduced with increased antioxidant and anti-inflammatory consumption. The phenolic yellow curry pigment curcumin has potent anti-inflammatory and antioxidant activities and can suppress oxidative damage, inflammation, cognitive deficits, and amyloid accumulation. Since the molecular structure of curcumin suggested potential Abeta binding, we investigated whether its efficacy in AD models could be explained by effects on Abeta aggregation. Under aggregating conditions in vitro, curcumin inhibited aggregation (IC(50) = 0.8 microM) as well as disaggregated fibrillar Abeta40 (IC(50) = 1 microM), indicating favorable stoichiometry for inhibition. Curcumin was a better Abeta40 aggregation inhibitor than ibuprofen and naproxen, and prevented Abeta42 oligomer formation and toxicity between 0.1 and 1.0 microM. Under EM, curcumin decreased dose dependently Abeta fibril formation beginning with 0.125 microM. The effects of curcumin did not depend on Abeta sequence but on fibril-related conformation. AD and Tg2576 mice brain sections incubated with curcumin revealed preferential labeling of amyloid plaques. In vivo studies showed that curcumin injected peripherally into aged Tg mice crossed the blood-brain barrier and bound plaques. When fed to aged Tg2576 mice with advanced amyloid accumulation, curcumin labeled plaques and reduced amyloid levels and plaque burden. Hence, curcumin directly binds small beta-amyloid species to block aggregation and fibril formation in vitro and in vivo. These data suggest that low dose curcumin effectively disaggregates Abeta as well as prevents fibril and oligomer formation, supporting the rationale for curcumin use in clinical trials preventing or treating AD.     

Effect of different anti-Abeta antibodies on Abeta fibrillogenesis as assessed by atomic force microscopy

Extensive data suggest that the conversion of the amyloid-beta (Abeta) peptide from soluble to insoluble forms is a key factor in the pathogenesis of Alzheimer's disease (AD). In recent years, atomic force microscopy (AFM) has provided useful insights into the physicochemical processes involving Abeta morphology, and it can now be used to explore factors that either inhibit or promote fibrillogenesis. We used ex situ AFM to explore the impact of anti-Abeta antibodies directed against different domains of Abeta on fibril formation. For the AFM studies, two monoclonal antibodies (m3D6 and m266.2) were incubated in solution with Abeta(1-42) with a molar ratio of 1:10 (antibody to Abeta) over several days. Fibril formation was analyzed quantitatively by determining the number of fibrils per microm(2) and by aggregate size analysis. m3D6, which is directed against an N-terminal domain of Abeta (amino acid residues 1-5) slowed down fibril formation. However, m266.2, which is directed against the central domain of Abeta (amino acid residues 13-28) appeared to completely prevent the formation of fibrils over the course of the experiment. Inhibition of fibril formation by both antibodies was also confirmed by thioflavin-T (ThT) fluorescence experiments carried out with Abeta(1-40) incubated for five days. However, unlike AFM results, ThT did not differentiate between the samples incubated with m3D6 versus m266.2. These results indicate that AFM can be not only reliably used to study the effect of different molecules on Abeta aggregation, but that it can provide additional information such as the role of epitope specificity of antibodies as potential inhibitors of fibril formation.     

Amyloid beta-protein (Abeta)1-40 protects neurons from damage induced by Abeta1-42 in culture and in rat brain

Previously, we found that amyloid beta-protein (Abeta)1-42 exhibits neurotoxicity, while Abeta1-40 serves as an antioxidant molecule by quenching metal ions and inhibiting metal-mediated oxygen radical generation. Here, we show another neuroprotective action of nonamyloidogenic Abeta1-40 against Abeta1-42-induced neurotoxicity in culture and in vivo. Neuronal death was induced by Abeta1-42 at concentrations higher than 2 microm, which was prevented by concurrent treatment with Abeta1-40 in a dose-dependent manner. However, metal chelators did not prevent Abeta1-42-induced neuronal death. Circular dichroism spectroscopy showed that Abeta1-40 inhibited the beta-sheet transformation of Abeta1-42. Thioflavin-T assay and electron microscopy analysis revealed that Abeta1-40 inhibited the fibril formation of Abeta1-42. In contrast, Abeta1-16, Abeta25-35, and Abeta40-1 did not inhibit the fibril formation of Abeta1-42 nor prevent Abeta1-42-induced neuronal death. Abeta1-42 injection into the rat entorhinal cortex (EC) caused the hyperphosphorylation of tau on both sides of EC and hippocampus and increased the number of glial fibrillary acidic protein (GFAP)-positive astrocytes in the ipsilateral EC, which were prevented by the concurrent injection of Abeta1-40. These results indicate that Abeta1-40 protects neurons from Abeta1-42-induced neuronal damage in vitro and in vivo, not by sequestrating metals, but by inhibiting the beta-sheet transformation and fibril formation of Abeta1-42. Our data suggest a mechanism by which elevated Abeta1-42/Abeta1-40 ratio accelerates the development of Alzheimer's disease (AD) in familial AD.     

Studies on the in vitro assembly of a beta 1-40: implications for the search for a beta fibril formation inhibitors

The progressive deposition of the amyloid beta peptide (Abeta) in fibrillar form is a key feature in the development of the pathology in Alzheimer's disease (AD). We have characterized the time course of Abeta fibril formation using a variety of assays and under different experimental conditions. We describe in detail the morphological development of the Abeta polymerization process from pseudo-spherical structures and protofibrils to mature thioflavin-T-positive/Congo red-positive amyloid fibrils. Moreover, we structurally characterize the various polymorphic fibrillar assemblies using transmission electron microscopy and determine their mass using scanning transmission electron microscopy. These results provide the framework for future investigations into how target compounds may interfere with the polymerization process. Such substances might have a therapeutic potential in AD.     

Acetylcholinesterase triggers the aggregation of PrP 106-126

Acetylcholinesterase (AChE), a senile plaque component, promotes amyloid-beta-protein (Abeta) fibril formation in vitro. The presence of prion protein (PrP) in Alzheimer's disease (AD) senile plaques prompted us to assess if AChE could trigger the PrP peptides aggregation as well. Consequently, the efficacy of AChE on the PrP peptide spanning-residues 106-126 aggregation containing a coumarin fluorescence probe (coumarin-PrP 106-126) was studied. Kinetics of coumarin-PrP 106-126 aggregation showed a significant increase of maximum size of aggregates (MSA), which was dependent on AChE concentration. AChE-PrP 106-126 aggregates showed the tinctorial and optical amyloid properties as determined by polarized light and electronic microscopy analysis. A remarkable inhibition of MSA was obtained with propidium iodide, suggesting that AChE triggers PrP 106-126 and Abeta aggregation through a similar mechanism. Huprines (AChE inhibitors) also significantly decreased MSA induced by AChE as well, unveiling the potential interest for some AChE inhibitors as a novel class of potential anti-prion drugs.     

Heterologous amyloid seeding: revisiting the role of acetylcholinesterase in Alzheimer's disease

Neurodegenerative diseases associated with abnormal protein folding and ordered aggregation require an initial trigger which may be infectious, inherited, post-inflammatory or idiopathic. Proteolytic cleavage to generate vulnerable precursors, such as amyloid-beta peptide (Abeta) production via beta and gamma secretases in Alzheimer's Disease (AD), is one such trigger, but the proteolytic removal of these fragments is also aetiologically important. The levels of Abeta in the central nervous system are regulated by several catabolic proteases, including insulysin (IDE) and neprilysin (NEP). The known association of human acetylcholinesterase (hAChE) with pathological aggregates in AD together with its ability to increase Abeta fibrilization prompted us to search for proteolytic triggers that could enhance this process. The hAChE C-terminal domain (T40, AChE(575-614)) is an exposed amphiphilic alpha-helix involved in enzyme oligomerisation, but it also contains a conformational switch region (CSR) with high propensity for conversion to non-native (hidden) beta-strand, a property associated with amyloidogenicity. A synthetic peptide (AChE(586-599)) encompassing the CSR region shares homology with Abeta and forms beta-sheet amyloid fibrils. We investigated the influence of IDE and NEP proteolysis on the formation and degradation of relevant hAChE beta-sheet species. By combining reverse-phase HPLC and mass spectrometry, we established that the enzyme digestion profiles on T40 versus AChE(586-599), or versus Abeta, differed. Moreover, IDE digestion of T40 triggered the conformational switch from alpha- to beta-structures, resulting in surfactant CSR species that self-assembled into amyloid fibril precursors (oligomers). Crucially, these CSR species significantly increased Abeta fibril formation both by seeding the energetically unfavorable formation of amyloid nuclei and by enhancing the rate of amyloid elongation. Hence, these results may offer an explanation for observations that implicate hAChE in the extent of Abeta deposition in the brain. Furthermore, this process of heterologous amyloid seeding by a proteolytic fragment from another protein may represent a previously underestimated pathological trigger, implying that the abundance of the major amyloidogenic species (Abeta in AD, for example) may not be the only important factor in neurodegeneration.     

Inhibitors of amyloid beta-protein aggregation mediated by GM1-containing raft-like membranes

The aggregation (fibril formation) of amyloid beta-protein (Abeta) is considered to be a crucial step in the etiology of Alzheimer's disease (AD). The inhibition of Abeta aggregation and/or decomposition of fibrils formed in aqueous solution by small compounds have been studied extensively for the prevention and treatment of AD. However, recent studies suggest that Abeta aggregation also occurs in lipid rafts mediated by a cluster of monosialoganglioside GM1. This study examined the effects of representative compounds on Abeta aggregation and fibril destabilization in the presence of GM1-containing raft-like liposomes. Among the compounds tested, nordihydroguaiaretic acid (NDGA), rifampicin (RIF), tannic acid (TA), and quercetin (QUE) showed strong fibrillization inhibitory activity. NDGA and RIF inhibited the binding of Abeta to GM1 liposomes by competitively binding to the membranes and/or direct interaction with Abeta in solution, thus at least partly preventing fibrils from forming. Coincubation of Abeta with NDGA, RIF, and QUE in the presence of GM1 liposomes resulted in elongate particles, whereas the presence of TA yielded protofibrillar structures. TA and RIF also destabilized fibrils. The most potent NDGA prevented Abeta-induced toxicity in PC12 cells by inhibiting Abeta accumulation. Furthermore, a comparison of the inhibitory effects of various compounds between aqueous-phase and GM1-mediated aggregation of Abeta suggested that the two aggregation processes are not identical.