Alzheimer's disease (AD) onset and progression are associated with the dysregulation of multiple and complex physiological processes and a successful therapeutic approach should therefore address more than one target. Two new chemical entities, the easily accessible heterocyclic scaffolds 1,3-diphenylbenzo [e][1,2,4ltriazin-7(1H)-one (benzotriazinone I) and 2-phenyl-6H-[1,2,4]triazino[5,6,1-Acarbazol-6-one (triazafluoranthenone II), were explored for their multitarget-directed inhibition of beta-amyloid (A beta) fibrillization and acetyl- (AChE) and/or butyryl- (BChE) cholinesterase, three valuable targets for AD therapy. Introduction of appropriate amine substituents at positions 6 and 5 on scaffold I and II, respectively, allowed the preparation of a series of compounds that were tested as A beta(1-40) aggregation and cholinesterase inhibitors. Potent inhibitors of A beta self-aggregation were discovered and among them benzotriazinone 7 exhibited an outstanding IC50 equal to 0.37 mu M. Compounds bearing a basic amine linked to the heterocyclic scaffold through a linear alkyl chain of varying length also afforded good ChE inhibitors. In particular, benzotriazinone 24 and triazafluoranthenone 38 were endowed with an interesting multiple activity, the former displaying IC50 values of 1.4, 1.5 and 1.9 mu M on A beta aggregation and AChE and BChE inhibition, respectively, and the latter showing IC50 values of 1.4 and an outstanding 0.025 mu M in the A beta aggregation and BChE inhibition, respectively. Benzotriazinone 24 and triazafluoranthenone 29, selected owing to their suitable aqueous solubility and A beta aggregation inhibition, were submitted to a time course kinetic assay followed with thioflavin T (ThT) spectrofluorimetry, circular dichroism (CD) and transmission electron microscopy (TEM). Experimental data indicated that 24 acted at a low concentration ratio (10 mu M 24 vs. 50 mu M A beta), stabilizing the unstructured A beta peptide and inhibiting fibrillogenesis, and that 29 also acted as fibrillization inhibitor, but likely enhancing and stabilizing the beta-sheet arrangement of A beta to yield protofibrillar species as detected by TEM. (C) 2012 Elsevier Masson SAS. All rights reserved.
