Molecular Simulation Studies On Drug Metabolism And Plasma Protein Binding

When drugs are administered to living organisms, they undergo a series of events such as absorption, distribution, metabolism and excretion (ADME).The cytochromes P450 (CYP) are the major-drug metabolizing enzymes in human, and play an important role in metabolizing and detoxicating various exogenous and endogenous compounds. Currently, approximately 70% of drugs on the market are metabolized through P450s. Therefore, research into P450s has been quite attractive due to its importance on drug metabolism and detoxication. On the other hand, most drugs undergo reversible binding to plasma proteins. Only the free fraction of a drug is available to diffuse from the vascular system thereby enabling interaction with therapeutic targets. The extent of drug binding to plasma proteins has a significant impact on its pharmacodynamic and pharmacokinetic properties. Therefore, understanding of the plasma protein binding behavior of a drug is quite essential both in drug discovery and clinical medication safety.However, the situation in drug metabolism and plasma protein binding becomes complicated, since:1.Cooperative behavior of many P450s has been related to the clinically important phenomenon of drug-drug interactions; 2.The polymorphism of P450s results in inter-individual differences in drug metabolism, which can be a risk factor for adverse drug effects; 3. Stereoselective plasma protein binding may result in pharmacodynamic and pharmacokinetic differences between enantiomers, thereby affecting the therapeutic effect or producing side-effects. Therefore, understanding the underlying mechanisms of those problems is quite essential in developing safe and effective drugs. Besides, genetic polymorphism of P450s has an important role in susceptibility to cancer.With the progress of computer hardware, as well as the development of theoretical knowledge, molecular simulation methods such as molecular docking, molecular dynamics and free energy calculation have been useful in exploring protein-ligand interactions. In the present study, molecular simulations were conducted to give some insights in cooperative behavior of P450s, polymorphism of P450s, and stereoselective plasma protein binding. In the first part of our work, CYP3A4 and testosterone were chosen as an example to explore the cooperative behavior. In the second part, CYP2A13*2,*5,*6,*8 and *9 were studied for its altered function. In the last part, propafenone and mexiletine were selected to study the stereoselective binding with AGP.1. Molecular simulations of testosterone in the active site of CYP 3A4:an insight into the cooperativityTo gain some insights into the cooperativity, molecular simulations were carried out for CYP3A4 with one and two testosterone molecules in their putative binding sites. Results showed that the effector testosterone can stabilize the active testosterone in an alternative conformation conductive to oxidation through hydrophobic interactions. Several residues were suggested to have important roles in cooperativity using energy decomposition, which was in accordance with previous mutation studies by other researchers. Besides, conformational changes were also observed and may associate in the cooperativity.2. Study on the altered functions of CYP2A13*2,*5,*6,*8 and *9 using molecular simulations Molecular simulations were carried out for CYP2A13*1,*2,*5,*6,*8 and *9. The substitutions in variants could induce conformational deviations and may lead to changes of the following aspects:substrate egress abilities, substrate recognition region and interaction with the reductase. We also investigated the enzymatic activity of CYP2A13 variants on coumarin. According to the computational results, none of the mutations directly altered the binding modes and binding energies of coumarin in the active site, indicating they should have similar Km values between wild type and each variant, which is in accordance with previous experimental results from Chen’s group. Besides, minor changes in distance between Fe of heme and reactive site in coumarin were also observed, which may explain the differences in the Vmax values upon residue substitutions.3. Molecular simulations on enantioselective binding of chiral drugs with a 1-AGPMolecular simulations were conducted to illuminate the stereoselective binding mechanisms of propafenone and mexiletine enantiomers with AGP at an atom level. The calculated binding energy difference trends between propafenone and mexiletine or between their enantiomers were correlated well with previous experimental results. The stereoselective binding was mainly attributed to hydrophobic interactions. The different binding modes were found between enantiomers, and the responsible residues were also analyzed, with the notable contribution of Arg90 and Ser89 both in propafenone and mexiletine.