Design, Synthesis And Evaluation Of BACE1 Inhibitors For The Treatment Of Alzheimer's Disease

Alzheimer's disease(AD),also called senile dementia,is the most common a progressive neurodegenerative disease.It is characterized by the presence of numerous extracellular amyloid plaques in the brain and of intracellular neurofibrillary tangles(NFTs).With the increase of human life span and the ageing of the population,this disease has brought a huge economic burden to both family and society.Alzheimer's is a degenerative and terminal disease for which there is currently no known cure.Four medications are currently approved to treat the cognitive manifestations of AD by regulatory agencies.Three are acetylcholinesterase inhibitors and the other is memantine,an NMDA receptor antagonist.No drug has an indication for delaying or halting the progression of the disease.Amyloid peptides(Aβ),which contains 40-42 amino acids,is the main component of senile plaques that lead to neuronal toxicity and cell death.Accumulation of Aβ, known as the amyloid hypothesis,is considered to play a central role in the pathology and subsequent cognitive decline of AD.Aβis produced fromβ-amyloid precursor protein(βAPP)by the sequential cleavage of theβ-andγ-secretase.Because the initial proteolytic cleavage of APP byβ-secretase(β-amyloid cleaving enzyme-1,BACE1)is considered to be the rate-limiting step in the processing of APP to Aβ,BACE1 has become an attractive therapeutic target for the treatment and prevention of AD.Many of BACE1 inhibitors have been discovered since BACE1 was identified in the late of 1999.However,most of them derived from transition-state analogues,such as statine,homostatine and hydroxyethylamine,are peptidomimetics.These inhibitors are usually to have low stability and poor oral bioavailability,which most likely precludes their full development as AD drugs.In addition,because of the localization of BACE1 in the central nervous system,inhibitors should be required to have sufficient lipophilicity and low-molecular weight in order to permeate the blood brain barrier.Based on the crystal structure of BACE1 and the critical interactive characteristics of the OM99-2 with BACE1,a novel virtual library of BACE1 inhibitors was constructed by using CombiLibMaker Model of Sybyl 6.91 program package and virtual screened by molecular docking(Dock 4.0 program)against active site of BACE1.The first target compounds were chosen to synthesize according to their score of virtual screening.Six research stages were included in the whole research work.Combining the structure-activity relationship(SAR)of the BACE1 inhibitory activity of the target compounds of every research stage and computer-assisted drug design,seven series of 114 nonpeptidomimetic small molecular compounds were designed and synthesized.The structure of these compounds had been confirmed by ¹H-NMR and MS.These target compounds were synthesized by seven different synthetic routes.The reaction conditions of every synthetic procedure were carefully investigated.In the course of synthesizing these compounds,the method of selectively hydrolyzing one of bi-esterfunctions in the diethyl 2,2-dimethyl-1,3-dioxolane -4,5-dicarboxylate was optimized.This method is suitable to the hydrolyzing others 1,3-dioxolane -4,5-dicarboxylate derivatives.Cleaving the acetal functions in 1,3-dioxolane-4,5-dicarboxamide detivatives,we discovered that the relationship between the reactive activities and the size of the group conjugating to the carboxyl is that the larger the group,the more difficult the reaction.Epoxide derivatives,the key intermediate of the control compound 2,were synthesized by the redesigned and implemented stereoselectivity synthetic method.Although this method exhibits lower chiral selectivity comparable to that of the report in literatures,the attractiveness of this methodology stems from its simplicity,convenience,hight yield,and suitable to amplify preparation.Also this method avoids low-temperature,expensive lithium reagent,anhydrous and no oxygen conditions.Various HEA derivatives were conveniently obtained from this intermediate.These target compounds were detected in vitro for inhibition against BACE1 by using a fluorescence resonance energy transfer(FRET)assay.It has been shown that 52 compounds had high inhibitory effects against BACE1 with inhibitiory rate of more than 50%at the concentration of 100μM,the most potent compound L80407 having an IC₅₀value of 178nM.In addition,six compounds with remarkable BACE1 inhibitory activity in vitro were evaluated by enzyme-linked immunospecific assay (ELISA),which was used to measure inhibitory activity by determining contents of Aβ42 in HEK293 cells.Although the results indicated that compounds LL70705 and L80407 among them displayed low inhibitory activity with inhibitiory rate of 23.68 and 23.49%,respectively,at the concentration of 50nM,it suggests that these inhibitors possess excellent cell permeability. The SAR data gleaned from the BACE1 inhibitory activity of the target compounds were investigated.The SAR results and the binding model of active compounds to BACE1 by docking could obtain a foundation and guide molecular design of BACE 1 inhibitor in the future research.