Alzheimer's disease (AD) is an age-related disorder that threatens to become an epidemic as the world population ages. Neurotoxic oligomers of A␤42 are believed to be the main cause of AD; therefore, disruption of A␤ oligomerization is a promising approach for developing therapeutics for AD. Formation of A␤42 oligomers is mediated by intermolecular interactions in which the C terminus plays a central role. We hypothesized that peptides derived from the C terminus of A␤42 may get incorporated into oligomers of A␤42, disrupt their structure, and thereby inhibit their toxicity. We tested this hypothesis using A␤ fragments with the general formula A␤(x؊42) (x ‫؍‬ 28 -39). A cell viability screen identified A␤(31-42) as the most potent inhibitor. In addition, the shortest peptide, A␤(39 -42), also had high activity. Both A␤(31-42) and A␤(39 -42) inhibited A␤-induced cell death and rescued disruption of synaptic activity by A␤42 oligomers at micromolar concentrations. Biophysical characterization indicated that the action of these peptides likely involved stabilization of A␤42 in nontoxic oligomers. Computer simulations suggested a mechanism by which the fragments coassembled with A␤42 to form heterooligomers. Thus, A␤(31-42) and A␤(39 -42) are leads for obtaining mechanism-based drugs for treatment of AD using a systematic structure-activity approach.Alzheimer's disease ͉ amyloid ␤-protein ͉ inhibitor design A lzheimer's disease (AD) is the predominant cause of dementia and one of the leading causes of death among elderly people. It is estimated that there are currently Ϸ27 million people suffering from AD worldwide (1). Because the world population is aging rapidly, if no cure is found in the near future AD will become an epidemic (2).The amyloid cascade hypothesis proposed that amyloid ␤-protein (A␤) fibrils-an aggregated form of A␤ found in amyloid plaques in the brains of patients with AD-were the neurotoxic agents causing AD (3). However, in recent years, multiple lines of evidence have led to a revision of this view, and today the primary toxins causing AD are believed to be early-forming A␤ oligomers rather than A␤ fibrils (4, 5). This paradigm shift suggests that efforts toward development of therapeutic agents targeting A␤ assembly should be directed at A␤ oligomers rather than fibrils. In particular, genetic, physiologic, and biochemical data indicate that oligomers of the 42-aa form of A␤, A␤42, are most strongly linked to the etiology of AD (6-9) and therefore are a particularly attractive target for inhibitor design.Several groups have reported small-molecule inhibitors of A␤ oligomerization (10-13). The importance of understanding the mechanism of inhibition recently has been highlighted (14) after findings that many small-molecule inhibitors of fibrillogenesis may act nonspecifically, likely making them unsuitable for treating amyloid-related disorders (15). In addition, inhibition of fibril formation may actually lead to stabilization of toxic oligomers (16). Interestingly, when oligomers are stabiliz...

No abstract

A key event in Alzheimer’s disease (AD) is age-dependent, brain accumulation of amyloid β-protein (Aβ) leading to Aβ self-association into neurotoxic oligomers. Previously, we showed that Aβ oligomerization and neurotoxicity could be inhibited by C-terminal fragments (CTFs) of Aβ42. Because these CTFs are highly hydrophobic, we asked if they themselves aggregated and if so, what parameters regulated their aggregation. To answer these questions, we investigated the dependence of CTF aqueous solubility, aggregation kinetics and morphology on peptide length and sequence, and the correlation between these characteristics and inhibition of Aβ42-induced toxicity. We found that CTFs up to 8-residues long were soluble at concentrations >100 µM and had a low propensity to aggregate. Longer CTFs were soluble at ~1–80 µM and most, but not all, readily formed β-sheet-rich fibrils. Comparison to Aβ40-derived CTFs showed that the C-terminal dipeptide I41-A42 strongly promoted aggregation. Aggregation propensity correlated with previously reported tendency to form β-hairpin conformation but not with inhibition of Aβ42-induced neurotoxicity. The data enhance our understanding of the physical characteristics that affect CTF activity and advance our ability to design, synthesize, and test future generations of inhibitors.

Oligomeric forms of amyloid β-protein (Aβ) are key neurotoxins in Alzheimer's disease (AD). Previously, we found that C-terminal fragments (CTFs) of Aβ42 interfered with assembly of fulllength Aβ42 and inhibited Aβ42-induced toxicity. To decipher the mechanism(s) by which CTFs affect Aβ42 assembly and neurotoxicity, here, we investigated the interaction between Aβ42 and CTFs using photo-induced cross-linking and dynamic light scattering. The results demonstrate that distinct parameters control CTF inhibition of Aβ42 assembly and Aβ42-induced toxicity. Inhibition of Aβ42-induced toxicity was found to correlate with stabilization of oligomers with hydrodynamic radius (R H ) = 8-12 nm and attenuation of formation of oligomers with R H = 20-60 nm. In contrast, inhibition of Aβ42 paranucleus formation correlated with CTF solubility and the degree to which CTFs formed amyloid fibrils themselves but did not correlate with inhibition of Aβ42-induced toxicity. Our findings provide an important insight into the mechanisms by which different CTFs inhibit the toxic effect of Aβ42 and suggest that stabilization of non-toxic Aβ42 oligomers is a promising strategy for designing inhibitors of Aβ42 neurotoxicity. KeywordsAmyloid β-protein; aggregation; neurotoxicity; inhibitor; dynamic light scattering Alzheimer's Disease (AD) is the most common neurodegenerative disease, affecting over 35 million people worldwide (1). Abundant evidence suggests that oligomeric forms of amyloid * To whom correspondence should be addressed: Department of Neurology, David Geffen School of Medicine, University of California at Los Angeles, Neuroscience Research Building 1, Room 451, 635 Charles E. Young Drive South, Los Angeles, CA 90095-7334, USA. gbitan@mednet.ucla.edu. Tel.: +1-310-206-2082; Fax.: +1-310-206-1700 The sequences of such inhibitors were based on random selection (9), selfrecognition of the central hydrophobic cluster (CHC) of Aβ (10-13), or structural modifications of sequences from the CHC or C-terminal regions (14-17). As the hypothesis of the cause of AD shifted from Aβ fibril formation and deposition to oligomeric Aβ, the design strategy for peptide inhibitors for treatment of AD was adjusted to target Aβ oligomerization. In view of the important role of the C-terminus in Aβ42 assembly and toxicity, we hypothesized that the C-terminal fragments (CTFs) of Aβ42 might disrupt Aβ42 assembly and inhibit its neurotoxicity. In a previous study, we confirmed this hypothesis and found that Aβ42 CTFs, Recently, we reported a systematic characterization of biophysical properties of all the CTFs in the original series (19), to which we added for additional structural insight two Aβ40 CTFs, Aβ(34-40) and , and a fragment derived from the putative folding nucleus of Aβ, Aβ(21-30) (20). We found that most of Aβ42 CTFs longer than 8 residues readily formed β-sheet-rich fibrils, whereas the shorter CTFs did not. The two Aβ40 CTFs were substantially less prone to aggregation than their Aβ42 CTF counterparts (19). Surprisingly, Aβ(30-40)...

Amyloid β-protein (Aβ) is central to the pathology of Alzheimer’s disease (AD). Of the two predominant Aβ alloforms, Aβ1–40 and Aβ1–42, the latter forms more toxic oligomers. C-terminal fragments (CTFs) of Aβ were recently shown to inhibit Aβ1–42 toxicity in vitro. Here we studied Aβ1–42 assembly in the presence of three effective CTF inhibitors and an ineffective fragment, Aβ21–30. Using a discrete molecular dynamics approach that recently was shown to capture key differences between Aβ1–40 and Aβ1–42 oligomerization, we compared Aβ1–42 oligomer formation in the absence and presence of CTFs or Aβ21–30 and identified structural elements of Aβ1–42 that correlated with Aβ1–42 toxicity. CTFs co-assembled with Aβ1–42 into large heterooligomers containing multiple Aβ1–42 and inhibitor fragments. In contrast, Aβ21–30 co-assembled with Aβ1–42 into heterooligomers containing mostly a single Aβ1–42 and multiple Aβ21–30 fragments. The CTFs, but not Aβ21–30, decreased the β-strand propensity of Aβ1–42 in a concentration-dependent manner. CTFs and Aβ21–30 had a high binding propensity to the hydrophobic regions of Aβ1–42 but only CTFs were found to bind the Aβ1–42 region A2-F4. Consequently, only CTFs but not Aβ21–30 reduced the solvent accessibility of Aβ1–42 in the region D1-R5. The reduced solvent accessibility of Aβ1–42 in the presence of CTFs was comparable to the solvent accessibility of Aβ1–40 oligomers formed in the absence of Aβ fragments. These findings suggest that the region D1-R5, which was more exposed to the solvent in Aβ1–42 than in Aβ1–40 oligomers, is involved in mediating Aβ1–42 oligomer neurotoxicity.