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

Molecular modeling techniques were used to derive a predictive model for substrates of cytochrome P450 2D6, an isozyme known to metabolize only compounds with one or more basic nitrogen atoms. Sixteen substrates, 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 site and basic nitrogen atom were fitted into one model by postulating an interaction of the basic nitrogen atom with a negatively charged carboxylate group on the protein. This acidic residue anchors and neutralizes the positively charged basic nitrogen atom of the substrates. In case of "5-angstrom substrates" this interaction probably occurs with the carboxylic oxygen atom nearest to the oxidation site, whereas in the case of "7-angstrom substrates" this interaction takes place at the other oxygen atom. Furthermore, all substrates exhibit a coplanar conformation near the oxidation site and have negative molecular electrostatic 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 studied so that this region of the active site can accommodate a variety of N-substituents. Therefore, the substrate specificity of P450 2D6 most likely is determined by the distance between oxidation site and basic nitrogen atom, by steric constraints near the oxidation site, and by the degree of complementarity between the MEPs of substrate and protein in the planar region adjacent to the oxidation site. The predictive value of the model was evaluated by investigating the P450 2D6 mediated metabolism of four new compounds comprising at least 14 oxidative metabolic routes. According to our model, 4 of the metabolic routes were predicted to be mediated by P450 2D6, whereas 10 were not. The involvement of P450 2D6 in these 14 metabolic reactions was investigated in man in vivo and/or in vitro. From these experimental results it appeared that 3 of the 4 predicted metabolic routes were mediated by P450 2D6 and 11 were not, closely matching the predictions from the model. Thus, 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 "debrisoquine/sparteine" type of polymorphism in a very early stage of the development of drugs.