(Mdr1) or Mrp2 mutations have been identified in mice (Lankas et al., 1997;
Kwei et al., 1999), rats (Buechler et al., 1996; Elferink et al., 1989; Kurisu et al.,
1991), or dogs (Paul et al., 1987). It is a useful tactic to use multiple gene
knockout animals together to phenotype the impact of transporters on the
pharmacokinetics of a given drug. By using Bcrp1(-/-) mice and EHBR rats,
Hirano et al. (2005) demonstrated that biliary clearance of pitavastatin was
accounted for to a large extent by Bcrp in mice and to a small extent by Mrp2
in rats. These findings differed for pravastatin, whose biliary clearance is
largely mediated by Mrp2. A similar strategy has been applied for investigating
the transporters involved in the transport of sulfasalazine. By comparing the
PK profiles of sulfasalazine after IV and PO administration to wild type,
Bcrp1(-/-) and Mdr1(-/-) mice, it was shown that Bcrp1, but not P-gp, was the
important determinant for the oral bioavailability and the elimination of
sulfasalazine in mice and that sulfasalazine can be potentially utilized as a
specificin vivoprobe for Bcrp1 (Zaher et al., 2006).
Caution needs to be taken when using these genetic animal models to
elucidate transport mechanisms since deletion of one transporter can often
cause alteration in the expression of other transporters or enzymes as well as
change the physiology of the knockout animal. For example, the protein
expression and activity of CYP3A2, 1A2, 2B1, and 2C11 in liver microsomes
prepared from EHBR were lower than those in SD rats (Ohmori et al., 1991).
The protein and mRNA expression levels of Mrp3 were significantly increased
in the liver and kidney of EHBR compared to wild type SD rats. On the
contrary, the protein and mRNA expressions of Oatp1 and Oatp2 were
significantly decreased in the liver of EHBR.
6.6.3.2 Transporter Chemically Knockout Models Mice or rats treated with a
transporter inhibitor can provide useful information on the role of the
transporter in absorption, elimination, and tissue distribution of a drug. The
chemically knockout animal model is commonly used for the evaluation of the
role of efflux transporters. The specificity of an inhibitor is critical when using
chemically knockout animal models to evaluate the role of a specific
transporter in the PK and toxicity of a drug. Whereas selective inhibitors are
now available for P-gp and BCRP, no selective inhibitors exist for the MRP
family of transporters. GF120918 (an inhibitor for P-gp and BCRP (Jonker
et al., 2000)), LY335979 (a selective P-gp inhibitor (Shepard et al., 2003)), and
Ko143 (a selective BCRP inhibitor (Allen et al., 2002)) can be safely used in
rats or mice to chemically knockout the corresponding efflux transporter(s).
However, no specific uptake transporter inhibitor for use inin vivostudies has
been identified.
The combination of chemical and genetic knockout mice have been used to
phenotype the role of efflux transporters in the absorption, disposition, or
elimination of drugs. For example, etoposide has been demonstrated to be a
P-gp and BCRP substrate using the cellular transport assay in P-gp or
Bcrp- transfected cell lines. However, treatment of P-gp-deficient mice with
METHODS TO EVALUATE TRANSPORTER SUBSTRATE 183