8.4 Reaction Phenotyping
It is extremely important to gain an early understanding of the enzymes
predicted to be important for metabolic clearance of an NCE so that some level
of predictions can be made as to the potential for drug–drug interactions and
polymorphic clearance in humans can be made. It should be noted that the true
contribution of individual enzymes to the clearance of a compound will in most
cases be difficult to predict until the human ADME study is complete and the
contribution of metabolic clearance to overall clearance of the drug has been
determined.
The major goal of phenotyping at the discovery stage is to make predictions
regarding the potential for the dependence of a single enzyme for a major
fraction (usually defined as>20%) of the clearance of a NCE. The type of
information that can reasonably generated at this stage is the contribution of
individual CYP enzymes to the intrinsic clearance in human liver microsomes
using well-established experimental techniques. Thein vitroinformation can be
augmented with in vivo information in animal models establishing the
contribution of oxidative biotransformation to the overall clearance, but this
preclinical information cannot be directly extrapolated to humans and does not
add to the prediction of contribution of individual CYP enzymes. The
estimation of fractional clearance by transferase enzymes producing direct
conjugates, esterases, or other drug metabolizing enzymes remains problematic
as does the prediction of transporters to the direct clearance via biliary or
urinary excretion. This information is certainly important for the design of
early clinical drug–drug interaction studies but in some cases may be used in
the discovery stage for decision-making purposes.
8.5 Assessment of Potential Toxicology of Metabolites
8.5.1 Reactive Metabolite StudiesIn vitro
The metabolism of drugs to reactive intermediates followed by covalent
binding to cellular components is hypothesized to be the basis for the acute or
idiosyncratic toxicities caused by some drugs (Guengerich, 2001; Ju and
Uetrecht, 2002; Kaplowitz, 2005; Liebler and Guengerich, 2005; Park et al.,
2005; Walgren et al., 2005). The testing of new drug candidates for their
potential to form reactive metabolites has received much recent attention
(Baillie, 2006; Guengerich, 2005; Uetrecht, 2003;). Reactive intermediates are
most commonly thought to arise through the generation of high energy
intermediates during the oxidation of drugs by CYP enzymes (Amacher, 2006;
Guengerich, 2003, 2006). Examples of these intermediates are epoxides,
oxirenes, arene oxides, and quinoid species. Myeloperoxidase is another human
oxidative enzyme that is known to catalyze the formation of reactive
intermediates. Reactive esters formed by the conjugation of carboxylic acids
246 DRUG METABOLISM RESEARCH