IN VITRO–IN VIVO SCALE-UP OF DRUG TRANSPORT ACTIVITIES - Drug Transporters: Molecular Characterization and Role in Drug Disposition - page 567

a species difference in the substrate specificities of common transporters such as
MRP2, or the expression of multiple transporters with substrate specificities that differ
from each other. Shilling et al. have demonstrated that the interspecies variation of
the uptake clearance depends on the substrate by using CMVs prepared from rats,
dogs, monkeys, and humans, implying the species difference of the expression and
function of each efflux transporter. 31 Some specific inhibitors of efflux transporters
have been proposed. For example, Ko143 preferentially inhibits BCRP-mediated
transport, 32 while PSC833 and LY335979 inhibit the MDR1-mediated transport more
potently than transport via other efflux transporters. 33 , 34 By evaluating the effect of
transporter-specific inhibitors on the ATP-dependent transport of test compounds in
human CMVs, the relative contribution of each transporter to the biliary excretion
can be assessed.
Recently, LeCluyse et al. have demonstrated that a collagen-sandwich culture
enabled the hepatocytes to form a bile canalicular “pocket” between the adjacent
cells, 35 and depletion of Ca 2 + from the incubation medium disrupted the bile canali-
culi rapidly. 36 The advantage of this culture configuration is that the polarity and
expression level of uptake and efflux transporters are fully retained for several days,
unlike normal culture on rigid collagen, and the biliary excretion of compounds
can be evaluated in intact cell systems by differential cumulative uptake in mono-
layers preincubated in Ca 2 + -containing buffer and Ca 2 + -free buffer. 37 , 38 Liu et al.
have found that the in vitro biliary clearance of five compounds (inulin, salicylate,
methotrexate, [D-pen 2 , 5 ]enkephalin, and taurocholate) in rat hepatocytes calculated
from the amount excreted into a bile canalicular pocket divided by the area under
the incubation medium concentration-time profile correlated well with their in vivo
intrinsic biliary clearance, suggesting that this system is useful for prediction of the
in vivo biliary excretion of many compounds 37 (Figure 19.4). Tian et al. succeeded in
applying siRNAs targeted to Mrp2 and Mrp3 in sandwich-cultured rat hepatocytes. 39

150

100

50

0

0 25 50 75

In Vitro Biliary Cl (mL/min . kg)

FIGURE 19.4. Relationship between in vivo intrinsic biliary clearance and in vitro esti-
mated biliary clearance calculated from the result in 96-hour sandwich-cultured hepatocytes for
model substrates, inulin (open square), salicylate (closed diamond), methotrexate (open circle),
[D-pen2,5]enkephalin (closed triangle) and taurocholate (closed circle). The dashed line is the
fit of a linear regression equation to the data ( r 2

= 0 . 99). [From ref. 37, with the kind permission
of the American Society for Pharmacology and Experimental Therapeutics.]