The Mechanism Of A Novel Acetylcholinesterase Inhibitor Bis(9)-(-)-Meptazinol Improving The Learining And Memory Function Of APP/PS1 Mice

Alzheimer’s disease（AD）is a progressive neurodegenerative disease characterized by cognitive impairment,which is the most common form of dementia in elderly patients.In recent years,the number of AD patients is growing rapidly,placing a heavy economic burden on their families and the society.The pathogenosis of AD is complicated,involving many influential factors.Senile plaques（SP）,neurofibrillary tangles（NFT）,and degenerative changes in cholinergic neurons are the prominent pathological features of AD.At present,the most commonly used drugs for the treatment of AD are acetylcholinesterase inhibitors（ACh EIs）.However,ACh EIs can only alleviate the cognitive symptoms of AD,but can not reverse its pathological processesof AD.Bis（9）-（-）-meptazinol（B9M）is a novel dual-binding ACh E inhibitor,developed based on the design strategy of Multi-Target-Directed Ligand（MTDL）.B9M could bind to the catalytic anionic site and peripheral anionic site of ACh E simultaneously,and inhibit ACh E activity and ACh E-induced Aβaccumulation effectively.In this study,APP/PS1 double transgenic mice were chosen as AD mouse model.Behavioral tests including nest-building,novel object recognition and Morris water maze were preformed to determine the effects of B9M on the cognitive impairment of APP/PS1 mice.The ACh E activity,Aβlevels and neuroinflammation in the brain of AD mice was detected by immunohistochemical staining and ELISA to explore the disease-modifying properties of B9M.The LPS-induced BV2 cells were used to further confirm the effects of B9M on neuroinflammation.Moreover,the tissue distribution and excretion of B9M in rats and mice was determined by a LC-MS/MS method previously established to improve understanding of the distribution and metabolism of B9M in brain and body.The results showed that the subcutaneous administration of B9M for 4 weeks（0.2μg/kg/day,0.6μg/kg/day,and 2μg/kg/day）could significantly improve nesting scores,novel object discrimination index and spatial learning and memory in APP/PS1 mice.B9M treatment could effectively inhibit ACh E activity,decrease the number of Aβplaques and the levels of insoluble and soluble Aβ₄₀ and Aβ₄₂,and reduce the activation of microglial and astrocyte and the expression of inflammatory factors including IL-1β,IL-6 and TNF-αin the cortex and hippocampus of APP/PS1 mice as well.The results on cell inflammation model further confirmed the anti-inflammatory effect of B9M.Pharmacokinetics results indicated B9M could distribute widely in various tissues in rats.The unchanged drug of B9M excreted in urine and feces was less than 3%,which suggested that a big amount of B9M was metabolized in the body and excreted out of the body in the form of metabolite.In conclusion,the results in this study suggested that the novel dual-binding ACh E inhibitor,B9M,could ameliorate the learning and memory deficits and pathological features of APP/PS1 double transgenic mice.B9M could be widely distributed and rapidly excreted after subcutaneous.B9M is a promising candidate for the treatment of AD.Since the distribution of B9M in brain is very limited and the metabolism of B9M is very fast and thorough,so more attention should be paid on the pharmacological effect of the metabolites of B9M in future study.