A PREDICTIVE MODEL FOR SUBSTRATES OF CYTOCHROME P450-DEBRISOQUINE (2D6)

Molecular modeling techniques were used to derive a predictive model f or substrates of cytochrome P450 2D6, an isozyme known to metabolize o nly compounds with one or more basic nitrogen atoms. Sixteen substrate s, accounting for 23 metabolic reactions, with a distance of either 5 angstrom ("5-angstrom substrates", e.g., debrisoquine) or 7 angstrom ( "7-angstrom substrates", e.g., dextromethorphan) between oxidation sit e and basic nitrogen atom were fitted into one model by postulating an interaction of the basic nitrogen atom with a negatively charged carb oxylate group on the protein. This acidic residue anchors and neutrali zes the positively charged basic nitrogen atom of the substrates. In c ase of "5-angstrom substrates" this interaction probably occurs with t he carboxylic oxygen atom nearest to the oxidation site, whereas in th e case of "7-angstrom substrates" this interaction takes place at the other oxygen atom. Furthermore, all substrates exhibit a coplanar conf ormation near the oxidation site and have negative molecular electrost atic potentials (MEPs) in a part of this planar domain approximately 3 angstrom away from the oxidation site. No common features were found in the neighbourhood of the basic nitrogen atom of the substrates stud ied so that this region of the active site can accommodate a variety o f N-substituents. Therefore, the substrate specificity of P450 2D6 mos t likely is determined by the distance between oxidation site and basi c nitrogen atom, by steric constraints near the oxidation site, and by the degree of complementarity between the MEPs of substrate and prote in in the planar region adjacent to the oxidation site. The predictive value of the model was evaluated by investigating the P450 2D6 mediat ed metabolism of four new compounds comprising at least 14 oxidative m etabolic routes. According to our model, 4 of the metabolic routes wer e predicted to be mediated by P450 2D6, whereas 10 were not. The invol vement of P450 2D6 in these 14 metabolic reactions was investigated in man in vivo and/or in vitro. From these experimental results it appea red that 3 of the 4 predicted metabolic routes were mediated by P450 2 D6 and 11 were not, closely matching the predictions from the model. T hus, the computer-assisted predictions seem to correlate well with the experimental results, and hence the presented model may be useful in identifying metabolic pathways that might be subject to the "debrisoqu ine/sparteine" type of polymorphism in a very early stage of the devel opment of drugs.