variability in pharmacokinetics observed for isoniazid. Recombinant forms
of these enzymes are commercially available, and human liver cytosol serves
as another source. Acetyl coA is the required cofactor for this enzyme
family.
15.4.3 Sulfation Reaction Phenotyping
Sulfation, using 3^0 -phosphoadenosine-5^0 -phosphosulphate (PAPS) as cofactor,
is rarely a primary clearance mechanism for drugs relative to glucuronidated
drugs or those cleared by cytochrome P450 enzymes. Exceptions include
minoxidil, acetaminophen at therapeutic doses, and ethinylestradiol.
Recombinant enzymes are commercially available for sulfotransferase pheno-
typing (Schneider and Glatt, 2004), but selective inhibitors have yet to be
identified.
15.5 Transporter Phenotyping
Transporters have the potential to contribute to clearance. Strictly speaking,
a reaction is not taking place, but since the consequence of transporter
action may be the same as with a drug metabolizing enzyme (removal of
unchanged drug from the plasma), inclusion in this chapter is warranted. A
good example of the developing field of transporter phenotyping is that for
hepatic uptake of pitivastatin, which modeled the‘‘relative activity factor’’
approach previously taken for cytochrome P450 enzymes (Hirano et al.,
2004). The kinetics of pitivastatin uptake was monitored in cells transfected
with the hepatic uptake transporters OATP1B1 and OATP1B3. The hepatic
uptake of pitivastatin was also modeled using the ‘‘relative abundance’’
approach with protein expression levels of OATP1B1 and OATP1B3
measured in transfectants and human hepatocytes. Both methods predicted
approximately 90 and 10% contributions of OATP1B1 and OATP1B3 to the
hepatic uptake of pitivastatin in human liver, respectively (Hirano et al.,
2004). However the overall hepatic uptake in hepatocytes using the ‘‘relative
abundance’’ approach overestimated the observed hepatic clearance. This
over estimation may be due to (1) differences in the recovery of the
transporter protein in samples for Western Blot analysis for transfectants
and hepatocytes or (2) the total amount of protein in the whole-cell crude
membrane may not be reflective of the functional transporter on the cell
surface. The pitivastatin example provides a significant advance over the
previously applied approach, where compounds were screened in cells
transfected with various recombinant transporters. Although this formerly
applied approach provided assessment of potential contribution of
individual transporters, it could not offer an assessment of relative
contributions, unlike the relative activity factor approach (Hirano et al.,
2004).
488 REACTION PHENOTYPING