Evidence for the presence of three distinct binding sites for the thioflavin T class of Alzheimer's disease PET imaging agents on β-amyloid peptide fibrils

Abstract

Imaging the progression of Alzheimer's disease would greatly facilitate the discovery of therapeutics, and a wide range of ligands are currently under development for the detection of β-amyloid peptide (Aβ)-containing plaques by using positron emission tomography. Here we report an in-depth characterization of the binding of seven previously described ligands to in vitro generated Aβ-(1-40) polymers. All of the compounds were derived from the benzothiazole compound thioflavin T and include 2-[4′-(methylamino) phenyl]benzothiazole and 2-(4′-dimethylamino-)phenyl-imidazo[1,2-a]- pyridine derivatives, 2-[4′-(dimethylamino)phenyl]-6-iodobenzothiazole and 2-[4′-(4″-methylpiperazin-1-yl)phenyl]-6-iodobenzothiazole, and a benzofuran compound (5-bromo-2-(4-dimethylaminophenyl)benzofuran). By using a range of fluorescent and radioligand binding assays, we find that these compounds display a more complex binding pattern than described previously and are consistent with three classes of binding sites on the Aβ fibrils. All of the compounds bound with very high affinity (low nM Kd) to a low capacity site (BS3) (1 ligand-binding site per ∼300 Aβ-(1-40) monomers) consistent with the previously recognized binding site for these compounds on the fibrils. However, the compounds also bound with high affinity (Kd ∼100 nM) to either one of two additional binding sites on the Aβ-(1-40) polymer. The properties of these sites, BS1 and BS2, suggest they are adjacent or partially overlapping and have a higher capacity than BS3, occurring every ∼35 or every ∼4 monomers of Aβ-(1-40)-peptide, respectively. Compounds appear to display selectivity for BS2 based on the presence of a halogen substitution (2-[4′-(dimethylamino)phenyl]-6- iodobenzothiazole, 2-[4′-(4″-methylpiperazin-1-yl)phenyl]-6- iodobenzothiazole, and 5-bromo-2-(4-dimethylaminophenyl)benzofuran) on their aromatic ring system. The presence of additional ligand-binding sites presents potential new targets for ligand development and may allow a more complete modeling of the current positron emission tomography data. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.