A family of huprine-tacrine heterodimers has been developed to simultaneously block the active and peripheral sites of acetylcholinesterase (AChE). Their dual site binding for AChE, supported by kinetic and molecular modeling studies, results in a highly potent inhibition of the catalytic activity of human AChE and, more importantly, in the in vitro neutralization of the pathological chaperoning effect of AChE toward the aggregation of both the β-amyloid peptide (Aβ) and a prion peptide with a key role in the aggregation of the prion protein. Huprine-tacrine heterodimers take on added value in that they display a potent in vitro inhibitory activity toward human butyrylcholinesterase, self-induced Aβ aggregation, and β-secretase. Finally, they are able to cross the blood-brain barrier, as predicted in an artificial membrane model assay and demonstrated in ex vivo experiments with OF1 mice, reaching their multiple biological targets in the central nervous system. Overall, these compounds are promising lead compounds for the treatment of Alzheimer's and prion diseases.

A new family of dual binding site acetylcholinesterase (AChE) inhibitors has been designed, synthesized, and tested for their ability to inhibit AChE, butyrylcholinesterase (BChE), AChE-induced and self-induced β-amyloid (Aβ) aggregation and β-secretase (BACE-1), and to cross the blood-brain barrier. The new heterodimers consist of a unit of racemic or enantiopure huprine Y or X and a donepezil-related 5,6-dimethoxy-2-[(4-piperidinyl)methyl]indane moiety as the active site and peripheral site to mid-gorge-interacting moieties, respectively, connected through a short oligomethylene linker. Molecular dynamics simulations and kinetics studies support the dual site binding to AChE. The new heterodimers are potent inhibitors of human AChE and moderately potent inhibitors of human BChE, AChE-induced and self-induced Aβ aggregation, and BACE-1, and are predicted to be able to enter the central nervous system (CNS), thus constituting promising multitarget anti-Alzheimer drug candidates with the potential to modify the natural course of this disease.

Background: Different studies have established that cholinergic neurodegeneration could be a major pathological feature of Alzheimer’s disease (AD). Thus, enhancement of the central cholinergic neurotransmission has been regarded as one of the most promising strategies for the symptomatic treatment of AD, mainly by means of reversible acetylcholinesterase inhibitors (AChEIs). The cognitive-enhancing properties of both huprine X, a new AChEI, and the structurally related huperzine A, as well as their effects on the regulation of several neurochemical processes related to AD have been studied in triple transgenic mice (3xTg-AD). Methods: Seven-month-old homozygous 3xTg-AD male mice, which received chronic intraperitoneal treatment with either saline, huprine X (0.12 µmol·kg^–1) or huperzine A (0.8 µmol·kg^–1) were subjected to a battery of behavioural tests after 3 weeks of treatment and thereafter the brains were dissected to study the neurochemical effects induced by the two AChEIs. Results: Treatments with huprine X and huperzine A improved learning and memory in the Morris water maze and some indicators of emotionality without inducing important adverse effects. Moreover, huprine X and huperzine A activate protein kinase C/mitogen-activated protein kinase pathway signalling, α-secretases (ADAM 10 and TACE) and increase the fraction of phospho-glycogen synthase kinase 3-β. Conclusion: Results obtained herein using a sample of 3xTg-AD animals strongly suggest that the treatment with the two AChEIs not only improves the cognitive performance of the animals but also induces some neurochemical changes that could contribute to the beneficial effects observed.

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Background: Several studies implicate acetylcholinesterase (AChE) in the pathogenesis of Alzheimer’s disease (AD), raising the question of whether inhibitors of AChE also might act in a disease-modifying manner. Huprine X (HX), a reversible AChE inhibitor hybrid of tacrine and huperzine A, has shown to affect the amyloidogenic process in vitro. In this study, the aim was to investigate whether HX could affect the AD-related neuropathology in vivoin two mouse models. Methods:Tg2576 (K670M/N671L) (APPswe) and 3xTg-AD (K670M/N671L, PS1M146V, tauP301L) mice were treated with HX (0.12 µmol/kg, i.p., 21 days) or saline at 6–7 months. Human β-amyloid (Aβ) was measured by ELISA, synaptophysin by Western blot and α7 neuronal nicotinic acetylcholine receptors (nAChRs) were analyzed by [¹²⁵I]α-bungarotoxin autoradiography. Results: Treatment with HX reduced insoluble Aβ1–40 (about 40%) in the hippocampus of 3xTg-AD mice, while showing no effect in APPswe mice. Additionally, HX markedly increased cortical synaptophysin levels (about 140%) and decreased (about 30%) the levels of α7 nAChRs in the caudate nucleus of 3xTg-AD mice, while increasing (about 10%) hippocampal α7 nAChRs in APPswe mice. Conclusion: The two mouse models react differently to HX treatment, possibly due to their differences in brain neuropathology. The modulation of Aβ and synaptophysin by HX in 3xTg-AD mice might be due to its suggested interaction with the peripheral anionic site on AChE, and/or via cholinergic mechanisms involving activation of cholinergic receptors. Our results provide further evidence that drugs targeting AChE affect some of the fundamental processes that contribute to neurodegeneration, but whether HX might act in a disease-modifying manner in AD patients remains to be proven.