Studies On Synthesis And Biological Evaluation Of Multifunctional PARP-1 And Cholinesterase Inhibitors As Anti-alzheimer's Disease Agents

ObjectiveIn order to search for new anti-AD drug based on both PARP-1 and cholinesterase inhibition,21 novel compounds incorporating 4-benzylpyridazinone and benzylpiperidine moiety were designed and synthesized using Olaparib and donepezil as lead compounds.Their inhibitory effects on PARP-1,AChE and BuChE in vitro were tested.The compound with the best biological activity was screened out,and then its effect on cognitive function and exercise capacity of AD mice induced by A?_25-35 was evaluated.Preliminary prediction of the interaction mode of the most potent compound with the targets was carried out by computer-assisted drug design.Method1.Design and synthesis of target compounds:The main pharmcophore of Olaparib was unchanged,and the cyclopropionyl piperazine group was replaced with the pharmacophore benzylpiperazine of donepezil to obtain 21 noval target compounds.2-Formoxyl benzoic acid was used as a start material,after cyclization,oxidation,reduction,2-fluoro-5-[?4-oxo-3,4-dihydropyridazin-1-yl?methyl]benzoic acid was generated;The substituted benzyl bromide is used as a raw material,it reacted with tert-butyloxo-aminopiperidine,tert-butyloxo-aminomethylpiperidineand hydroxypiperidine to obtain an amino group-containing substituted benzyl piperidine,after deprotecting the t-butoxycarbonyl group,the amidation reaction with 2-fluoro-5-[?4-oxo-3,4-dihydropyridazin-1-yl?methyl]benzoic acid was carried out to give the desired products.2.Biological activities evaluations of the target compounds:?1?The inhibitory activities of target compounds against AChE and BuChE were evaluated by Ellman method and the inhibitory activity of the target compounds against PARP-1 enzyme was detected by HT homologous PARP fluorescence inhibition kit?Trevigen,Cat#4690-096-K?.?2?The proliferation inhibition activity of all target compounds on MDA-MB-436 was evaluated by MTT assay.?3?Mice were injected with aggregated A?_25-35 into lateral ventricles,the effect of drug on behavior of AD mice was observed after administration of compound 34.3.Molecular docking:AutoDock Vina was used to explore its molecular recognition mechanism and binding mode between compounds and proteins?AChE,BuChE and PARP-1?.Result1.In this study,21 target compounds were designed and synthesized.The structures of derivatives were confirmed by ¹H-NMR,¹³C-NMR,IR and MS.2.The protease inhibition assay screened several compounds that inhibited the activity of AChE,BuChE and PARP-1 simultaneously.The IC₅₀ of 34 for AChE,BuChE and PARP-1 were 13.481±2.154?M,1.630±0.523?M and 8.18±2.81nM,respectively.Behavioral experiments were carried out to test the effects of this compound on cognitive function and exercise capacity of AD mice.It was found that after continuous administration of compound 34 for two weeks can improve cognitive function in AD mice to some extent,compared with AD model group.3.Molecular docking studies showed that compound 34 could form hydrogen bonds with Gly863 and Ser904 of PARP-1.At the same time,compound 34 can be wholly placed in the active catalytic domain of BuChE,which may be the reason for its better inhibitory activity against butyrylcholinesterase than other compounds.ConclusionIn this study,21 novel compounds were designed and synthesized.Several compounds which can simultaneously inhibit AChE,BuChE and PARP-1 were screened out by protease inhibition experiments,of which compound 34 is better than oters.Animalbehavioralexperimentsshowedthatcompound34can improve cognitive function in AD mice to some extent after intragastric administration at a dose of 10 mg/kg/d for 2 weeks.