Studies On Substrate Selectivity And Molecular Design Of Several Drug Metabolizing Enzymes

Covalent compounds have gained significant attention from pharmacologists and have been widely applied in chemical biology and medicinal chemistry in recent years due to their unique mode of action with the targets and their advantages of high biological activity,long residence time,low dose of drug administration and low ratio of off-target.However,covalent fragments may also possess collateral drug binding with other endogenous substances such as glutathione and DNA in the body,thereby causing unnecessary toxic and side effects.In addition,compounds may covalently bind to metabolic enzymes in vivo for biotransformation and metabolism,and their metabolic selectivity is governed by various factors.Therefore,it is of great significance to explore the factors that affect the reactivity and metabolic selectivity of covalent fragments and to introduce the common methods for evaluating the reactivity and selectivity,which can guide the optimization and modification of preclinical compounds.This dissertation mainly focuses on the metabolic selectivity mediated by metabolic enzymes in drug design and the effect of the intrinsic reactivity of compounds on metabolic selectivity.At the same time,according to the binding characteristics and intrinsic reactivity of the ligand,several strategies are combined to modify and optimize the molecular structure.In the first chapter,the factors affecting the covalent reactivity and selectivity of compounds and the corresponding drug design methods are systematically introduced.In the second chapter,the possible metabolic mechanism of cyanogroup in the gliptin analogues catalyzed by dipeptidyl peptidse 4(DPP-4)and the reasons for its metabolic selectivity are revealed by molecular simulation.DPP-4 is an important drug target of type ? diabetes mellitus,and its inhibitors own the advantages of increasing glucose tolerance,lowering blood glucose level,being effective and well tolerated with no adverse effects such as hypoglycemia and weight gain.It has been reported that DPP-4 can selectively metabolize peptide-mimic inhibitors containing cyanogroup,while the factors that affect the metabolic profile are still unknown.Combining with experiments,we applied molecular dynamics simulation and quantum chemistry calculation to explore the metabolic selectivity of cyanopyrrolidines mediated by DPP-4.The metabolism of cyanopyrrolidines by DPP-4 invloved multiple steps,in which two water molecules also participated.We observed that the influence of solvent environments within the catalytic site was not the reason for metabolic selectivity,but the reaction energy of water molecules nucleophilic attack on imine intermediates during the metabolic reaction process was.Additionally,both the calculated results and the experimental data revealed that the cyanogroup was first metabolized into an amide intermediate product,and then the compound was finally hydrolyzed into a carboxyl product according to the mechanism of serine proteolytic enzyme.This work shed light on understanding the metabolic mechanism of cyanopyrrolidines catalyzed by DPP-4,and further revealed the metabolic preference of DPP-4 as a potential drug metabolic enzyme.Cytochrome P450 1As(CYP1As)are one subfamily of the important drug metabolic enzymes and play a crucial role in the metabolism of endogenous and exogenous substances.Among them,CYP1A1 has been reported to involve a variety of tumor-related gene mutation and tumor formation,which has attracted extensive attention in recent years.In the third chapter,combining molecular docking and quantum chemistry calculations with the experimental results,we optimized the known substrate of CYP1A1 and yielded novel fluorescent substrate of CYP1A1 with fluorescence changes before and after metabolism based on the fluorescent mechanism of intramolecular charge transfer.We illustrated the key residues in CYP1 As affecting the binding affinity of compounds with targets,and the electron effects influencing the metabolic selectivity of compounds were also explored.Besides,the electron excitation properties and the contribution and influence of fragments on the electron excitation were also clarified.In summary,a new type of CYP1A1 fluorescent molecular probe was discovered in this study,which might have great prospects for the biological imaging of endogenous CYP1A1 in living cells and the further study on the biological functions of CYP1A1 in complex biological systems.Aldehyde oxidase(AOX)is a drug metabolizing molydbo-flavoenzyme that has gained increasing attention because of contribution to the biotransformation in phase I metabolism of xenobiotics.Unfortunately,the intra-and inter-species variations in AOX activity and lack of reliable and predictive animal models make evaluation of AOX-catalyzed metabolism prone to be misleading.In the forth study,we developed an improved computational model integrating both atom-level and molecule-level features to predict whether a drug-like molecule is a potential human AOX(hAOX)substrate and to identify the corresponding sites of metabolism.Additionally,we combined the proposed computational strategy and in vitro experiments for evaluating the metabolic property of a series of epigenetic-related drug candidates still in early stage of development.In summary,this study provides an improved strategy to evaluate the liability of molecules toward hAOX and offers useful information for accelerating the drug design and optimization stage.