Myricetin: a naturally occurring regulator of metal-induced amyloid-β aggregation and neurotoxicity.

One of the most severe and incurable forms of neurodegeneration, Alzheimer’s disease (AD), is characterized in the brain by the accumulation of aggregated amyloid-b (Ab) peptides. In the diseased brain, elevated concentrations of metals, such as Fe, Cu, and Zn, are found in Ab plaques. It has been proposed that metal ions, such as Cu and Zn, can bind to Ab ; this causes enhanced peptide aggregation, and in the case of redox active metal ions (e.g. , Cu), the generation of reactive oxygen species (ROS) leading to oxidative stress and neuronal death. While peptide aggregation and oxidative stress have been implicated in AD progression, the role of metal ions associated with Ab species in the development of this disease remains unclear. To clarify the function of metal ions in Ab-related pathological events, small molecule-based tools that contain bifunctionality for probing both metal ions and Ab have been sought. Several small molecules have been fashioned according to a rational structure-based design strategy to target metal-associated Ab species (metal–Ab species) and to interrogate metal-induced Ab aggregation and neurotoxicity. 9–14] Due to the range of possible conformations of metal–Ab that could be involved in AD neuropathogenesis, discovery of novel structural frameworks that can target these species might advance progress for this design strategy. One tactic to identify new classes of basic structural scaffolds is through screening of naturally occurring compounds, such as flavonoids. Flavonoids are a class of polyphenolic compounds that are abundant in natural products, such as berries, fruits, and vegetables, and have been investigated as potential therapeutic agents in human diseases including cancer, cardiovascular disease, and AD. These naturally occurring compounds have been shown independently to chelate metal ions and to interact with Ab, suggesting their potential bifunctionality toward metal–Ab species. One of the flavonoid compounds, myricetin (Scheme 1), previously demonstrated an anti-amyloidogenic effect through its reversible binding to fibrillar Ab but not to the monomeric species. In the case of its metal binding property, prior studies have shown that myricetin has multiple potential sites for metal chelation including positions between the 4-oxo and the 3or 5-OH groups (Scheme 1) that can form complexes with a binding stoichiometry of 1:1 or 1:2, metal/myricetin. Despite the known interactions of myricetin or other members of the flavonoid family with metal ions and Ab, their influence on metal-induced Ab aggregation pathways and neurotoxicity has not been investigated. Herein, we report that myricetin, exhibiting bifunctionality (metal chelation and Ab interaction), was capable of modulating Cuand Zn-induced Ab aggregation and neurotoxicity in vitro and in human neuroblastoma cells. To the best of our knowledge, this is the first example to establish the reactivity of the flavonoid myricetin toward metal– Ab species. Before conducting reactivity studies of myricetin with metal–Ab species, its metal binding properties were confirmed by UV/Vis experiments. In agreement with the previous reports, a bathochromic shift of the optical band was observed upon addition of one equivalent of CuCl2 to myricetin (25 mm) in HEPES (20 mm), pH 7.4, NaCl (150 mm ; ca. 377 to 450 nm, Figure S1 in the Supporting Information). Similarly, a bathochromic shift in the optical spectrum (ca. 377 to 400 nm, Figure S1 in the Supporting Information) was visible when one equivalent of Zn was introduced to myricetin. Moving forward, to determine if metal chelation by myricetin was possible in the presence of Ab, one equivalent of myricetin was added to a preincubated solution of Ab (25 mm) and CuCl2 or ZnCl2 (25 mm). New absorption bands that resembled metal-incubated myricetin in Ab-free solutions appeared (Figure S1 in the Supporting Information). These results suggest that myricetin could compete with Ab for binding to metal ions, with possible implications for reactivity in metal-induced Ab events (vide infra). The reactivity of myricetin toward in vitro metal-induced Ab aggregation was probed according to previously reported procedures (Figures 1 and 2). Generally, if a small molecule can inhibit formation of or promote the disassembly of Ab aggregates, more soluble, smaller-sized Ab species will be produced. The relative MW distribution of these Ab species can be Scheme 1. Chemical structure of myricetin (3,5,7-trihydroxy-2-(3,4,5-trihydroxyphenyl)-4H-1-benzopyran-4-one). Potential donor atoms for metal chelation are highlighted in bold.