Understanding the determinants of selectivity in drug metabolism through modeling of dextromethorphan oxidation by cytochrome P450

Abstract

Cytochrome P450 enzymes play key roles in the metabolism of the majority of drugs. Improved models for prediction of likely metabolites will contribute to drug development. In this work, two possible metabolic routes (aromatic carbon oxidation and O-demethylation) of dextromethorphan are compared using molecular dynamics (MD) simulations and density functional theory (DFT). The DFT results on a small active site model suggest that both reactions might occur competitively. Docking and MD studies of dextromethorphan in the active site of P450 2D6 show that the dextromethorphan is located close to heme oxygen in a geometry apparently consistent with competitive metabolism. In contrast, calculations of the reaction path in a large protein model [using a hybrid quantum mechanical-molecular mechanics (QM/MM) method] show a very strong preference for O-demethylation, in accordance with experimental results. The aromatic carbon oxidation reaction is predicted to have a high activation energy, due to the active site preventing formation of a favorable transition-state structure. Hence, the QM/MM calculations demonstrate a crucial role of many active site residues in determining reactivity of dextromethorphan in P450 2D6. Beyond substrate binding orientation and reactivity of Compound I, successful metabolite predictions must take into account the detailed mechanism of oxidation in the protein. These results demonstrate the potential of QM/MM methods to investigate specificity in drug metabolism.C ytochrome P450 enzymes (P450s) form one of the most powerful defense mechanisms of living organisms: They protect against xenobiotics via an oxidative pathway, which in most instances leads to a less harmful and more soluble product with faster excretion. The same mechanism is involved in the metabolism of most drugs and alters the pharmacological activity of many drugs. As a consequence, P450-mediated transformations of drug candidates are of crucial importance in the pharmaceutical industry. The roles of P450s are manifold. Oxidation by P450s can lead to toxic products, but, on the other hand, local activation of, e.g., anticancer prodrugs by P450s to lethal intracellular toxins at the site of the tumor is an important strategy (1). Drug compounds can induce P450 expression but can also inhibit them in various ways (2-4). The metabolic clearance of most drugs depends on P450s, and they have been implicated in a large number of drug-drug interactions (5, 6), which can result in fatalities (7,8). Interactions of drug candidates with P450s must be taken into account during the drug discovery process if the expensive and time-consuming development of active compounds with hidden toxic effects is to be avoided.It is increasingly recognized that methods capable of predicting P450 oxidative activity for a given substrate, and also the predominant site(s) of metabolism, could contribute significantly to drug development. There are many aspects to this problem, including isoform selectivity (i.e., which P450 isoform will contribute mos...

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