Isomerization of (Z,Z) to (E,E)1-Bromo-2,5-bis-(3-hydroxycarbonyl-4-hydroxy)styrylbenzene in Strong Base: Probes for Amyloid Plaques in the Brain

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Abstract
In developing probes for detecting β-amyloid (Aβ) plaques in the brain of Alzheimer's disease (AD), we have synthesized 1-bromo-2,5-bis-(3-hydroxycarbonyl-4-hydroxy)styrylbenzene (5, BSB). Due to the presence of two double bonds, formation of four different isomers is possible. Four isomers, E , E - 5, E , Z -5, Z , E - 5, and Z , Z -5, were prepared. Surprisingly, all showed strong fluorescent labeling of Aβ plaques in the brain of postmortem brain sections of patients with confirmed AD. In vitro binding assay also showed that all four isomers of BSB (E , E - 5, E , Z -5, Z , E - 5, and Z , Z -5) displayed a similar high binding affinity inhibiting the binding of [125I]E , E - 6, 1-iodo-2,5-bis-(3-hydroxycarbonyl-4-methoxy)styrylbenzene (IMSB) to Aβ1-40 aggregates. The inhibition constants (Ki) of E , E - 5, E , Z -5, Z , E - 5, and Z , Z -5 were 0.11 ± 0.01, 0.19 ± 0.03, 0.27 ± 0.06, and 0.13 ± 0.02 nM, respectively. Due to the fact that geometric stability of these styrylbenzenes is unknown, and the conversion of Z , Z -5 to E , E - 5 may occur automatically in the binding or labeling assaying conditions, we have investigated the kinetics of conversion of Z , Z -5 to E , E - 5 by NMR in D2O/NaOD at elevated temperatures (70, 95, and 115 °C). The activation energy was determined to be 14.15 kcal/mol. The results strongly suggest that the isomeric conversion at room temperature in aqueous buffer solution is unlikely. All of the styrylbenzene isomers clearly showed potential as useful tools for studying Aβ aggregates in the brain. The data suggest that, despite the rigidity of this series of styrylbenzenes, the binding sites on Aβ aggregates may have certain flexibility and the binding pockets could be adaptable for binding to other smaller ligands. Such information could be exploited to develop new ligands for detecting amyloid plaques in AD.