Rapid assessment of P-glycoprotein inhibition and induction in vitro.
OBJECTIVE
Using rhodamine123 (RH123) cell exclusion. 17 clinically used compounds were screened for their inhibitory effect on P-glycoprotein (P-gp), which was compared with the drugs' inhibitory activity against CYP3A4. The same assay was used to study induction of P-gp activity.


METHODS
P-gp inhibition was assessed using RH123 accumulation into LS180V cells as well as Rh123 transport across Caco-2 mono-layers. Inhibition of CYP3A4 was determined in human liver microsomes using triazolam-4-hydroxylation. Induction of P-gp expression and activity was measured using western blot analysis and RH123 accumulation into LS180V cells, respectively.


RESULTS
The observed inhibition of RH123 cell exclusion ranged from little or no effect (digoxin, indinavir, fexofenadine) up to a nearly 10-fold increase in RH 123 accumulation (ivermectin, terfenadine). No correlation between P-gp and CYP3A4 inhibition was observed. The rank order in P-gp inhibitory potency for terfenadine, verapamil, ritonavir. and indomethacin was identical in both LS180V and Caco-2 models. Ritonavir and St. John's wort extract showed a concentration-dependent P-gp induction, with good correlation between western blot analysis and RH123 accumulation.


CONCLUSIONS
The RH123 accumulation assay in LS180V cells can be used as a valuable screening tool to study both inhibition and induction of P-gp activity and expression. This assay has the potential to predict P-gp-mediated alterations in intestinal absorption of drugs.
Perloff
Michael D
MD
Department of Pharmacology and Experimental Therapeutics, Tufts University School of Medicine, 136 Harrison Avenue, Boston, Massachusetts 02111, USA.
Störmer
Elke
E
von Moltke
Lisa L
LL
Greenblatt
David J
DJ
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D015153:Blotting, Western; D002460:Cell Line; D051544:Cytochrome P-450 CYP3A; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005456:Fluorescent Dyes; D006801:Humans; D066298:In Vitro Techniques; D008856:Microscopy, Fluorescence; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D020112:Rhodamine 123
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D020112:Rhodamine 123; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human