Study On The Drug Interaction Of Octahydroaminoacridine Succinate In Vitro

Alzheimer's Disease(AD)is a common central nervous system disease.Statistically,there are approximately 46 million people worldwide suffer from AD.By 2050,the number is expected to 130 million,AD is the fourth leading cause of death in the elderly after heart disease,tumors and cerebrovascular disease.Octahydroaminoacridine succinate is a self-dependent drug for the treatment of AD,which is a kind of new acetyl cholinesterase inhibitor,it is very different from other acetyl cholinesterase inhibitor drugs,literature indicated that octahydroaminoacridine succinate could slow down alzheimer's patients beta amyloid effectively,octahydroaminoacridine succinate had good effectiveness in the prevention and treatment of AD,cerebral hemorrhage,stroke sequela,and teenagers lack of concentration syndrome.Octahydroaminoacridine succinate is a novel compound.Currently,the metabolic study of octahydroaminoacridine succinate is mainly focused on the preclinical pharmacokinetic study in vivo,no literature was available on the mechanism of of drugdrug Interation(DDI).In the drug development process,if DDI was not found in time,side effects would occur in the clinical tral,which will lead to the failure of drug development eventually.Thus,the research on DDI plays a crucial role in the clinical medication.Therefore,the octahydroaminoacridine succinate was chosen as the test subjects.Firstly,An LC-MS/MS(Liquid chromatography-tandem mass spectrometry,LC-MS/MS)method for quantitation of octahydroaminoacridine succinate was established,and research DDI in the metabolic process of octahydroaminoacridine succinate through the vitro study,the main research content as follows:(1)Establishment of LC-MS/MS quantitative analysis method for octahydroaminoacridine succinate in liver microsomesBased on LC-MS/MS technology,the quantitative analysis method for octahydroaminoacridine succinate in liver microsomes was established for the first time,then the precision and accuracy,linear range,residue,extraction recovery and the matrix effect was investigated,the results showed that the method could quantify octahydroaminoacridine succinate accurately,and the method could be applied to the DDI study of octahydroaminoacridine succinate in vitro.(2)Study on the metabolic stability of octahydroaminoacridine succinateThe incubation system of octahydroaminoacridine succinate in human liver microsomes was established and the metabolic stability of octahydroaminoacridine succinate was studied.The results showed that octahydroaminoacridine succinate was linearly eliminated in human liver microsomes within 0-180 min.The elimination rate constant and half-life of amber octadecridine in liver microsomes were k= 0.0026min-1,t1/2 = 266.5 min.Which showed the metabolism of octahydroaminoacridine succinate was stable.(3)Identification of the octahydroaminoacridine succinate CYP450 metabolic pathwayBased on liver microsomal incubation system,there were 7 kinds of main CYP450 enzymes(CYP1A2,CYP2B6,CYP2C8,CYP2C9,CYP2C19,CYP2D6,and CYP3A4)were chosen as the research object,and octahydroaminoacridine succinate CYP450 enzymes metabolic pathways was identified by selective chemical inhibition method and genetic recombination anthropogenic CYP450 enzyme hydrolysis,verification method of selective inhibitors of experimental results showed that the CYP3A4 and CYP2C8 played an important role in the metabolism of octahydroaminoacridine succinate by chemical inhibitor method.However,genetic recombination anthropogenic CYP450 enzymatic method showed that octahydroaminoacridine succinate did not metabolize obviously in CYP3A4 and CYP2C8 pure enzyme incubation system,inferring chemical inhibitor method and gene recombination anthropogenic CYP450 enzymatic method,we assumed that the metabolism of octahydroaminoacridine succinate was completed by several kinds of CYP450 enzymes,a single subtype CYP450 enzyme could not metabolize octahydroaminoacridine succinate.(4)Inhibitory effects of octahydroaminoacridine succinate on CYP450 enzymesThe "Cocktail" mixed probe substrate incubation system was established to investigate the inhibitory effect of amber hydroaminoacrine on 7 CYP450 enzyme subtypes(CYP1A2,CYP2B6,CYP2C8,CYP2C9,CYP2C19,CYP2D6 and CYP3A4)for the first time.The results showed that there was no activity of inhibition for 7 major CYP450 subtypes in the octahydroaminoacridine succinate concentration from 0.005 ?M to 0.5 ?M.Half-inhibitory concentration(IC50)of octahydroaminoacridine succinate revealed that were all greater than 50 ?M,indicating that the octahydroaminoacridine succinate had no inhibitory effects on seven CYP450 enzyme.In summary,in view of the octahydroaminoacridine succinate DDI in the fields of research,this project established an LC-MS/MS analysis method for the quantitation of octahydroaminoacridine succinate,then identified the CYP450 metabolic pathway of octahydroaminoacridine succinate by genetic recombination anthropogenic CYP450 enzymatic method and chemical inhibitors method respectively.And established a "Cocktail" mixed probe substrates incubation system ti study the inhibitory effects of octahydroaminoacridine succinate on CYP450 enzymes,providing an important reference for safety and effectiveness of octahydroaminoacridine succinate DDI in the clinical application of evaluation.