Lisa L. von Moltke

Saint John's wort: An in vitro analysis of P-glycoprotein induction due to extended exposure

Chronic use of Saint Johns wort (SJW) has been shown to lower the bioavailability for a variety of co‐administered drugs including indinavir, cyclosporin, and digoxin. Decreases in intestinal absorption through induction of the multidrug resistance transporter, P‐glycoprotein (P‐gp), may explain decreased bioavailability. The present study characterized the response of P‐gp to chronic and acute exposure of SJW and hypericin (HYP, a presumed active moiety within SJW) in an in vitro system. Experiments were performed with 3 to 300 μg ml−1 of methanol‐extracted SJW and 0.03 to 3 μM HYP, representing low to high estimates of intestinal concentrations. In induction experiments, LS‐180 intestinal carcinoma cells were exposed for 3 days to SJW, HYP, vehicle or a positive control (ritonavir). P‐gp was quantified using Western blot analysis. P‐gp expression was strongly induced by SJW (400% increase at 300 μg ml−1) and by HYP (700% at 3 μM) in a dose‐dependent fashion. Cells chronically treated with SJW had decreased accumulation of rhodamine 123, a P‐gp substrate, that was reversed with acute verapamil, a P‐gp inhibitor. Fluorescence microscopy of intact cells validated these findings. In Caco‐2 cell monolayers, SJW and HYP caused moderate inhibition of P‐gp‐attributed transport at the maximum concentrations tested. SJW and HYP significantly induced P‐gp expression at low, clinically relevant concentrations. Similar effects occurring in vivo may explain the decreased bioavailability of P‐gp substrate drugs when co‐administered with SJW.

Differential modulation of P-glycoprotein expression and activity by non-nucleoside HIV-1 reverse transcriptase inhibitors in cell culture.

AbstractPurpose. This study investigated the effects of the non-nucleoside HIV-1 reverse transcriptase inhibitors (NNRTI) nevirapine (NVR), efavirenz (EFV), and delavirdine (DLV) on P-glycoprotein (P-gp) activity and expression to anticipate P-gp related drug-drug interactions associated with combination therapy. Methods. NNRTIs were evaluated as P-gp substrates by measuring differential transport across Caco-2 cell monolayers. Inhibition of P-gp mediated rhodamine123 (Rh123) transport in Caco-2 cells was used to assess P-gp inhibition by NNRTIs. Induction of P-gp expression and activity in LS180V cells following 3-day exposure to NNRTIs was measured by western blot analysis and cellular Rh123 uptake, respectively. Results. The NNRTIs showed no differential transport between the basolateral to apical and apical to basolateral direction. NNRTI transport in either direction was not affected by the P-gp inhibitor verapamil. DLV inhibited Rh123 transport, causing a reduction to 15% of control at 100 μM (IC50 = 30 μM). NVR caused a concentration-dependent induction of P-gp expression in LS180V cells resulting in a 3.5-fold increase in immunoreactive P-gp at 100 μM NVR. Induction attributable to EFV and DLV was quantitatively smaller. NVR significantly reduced cellular uptake of Rh123 into LS180V cells, indicating increased drug efflux due to induced P-gp activity; effects of EFV and DLV were smaller. Acute DLV treatment of LS180V cells previously induced with NVR or ritonavir did not reverse the decreased Rh123 cell accumulation. Conclusions. NNRTIs show differential effects on P-gp activity and expression in vitro. Clinical studies are required to elucidate the clinical importance of potential drug interactions.

Fexofenadine Transport in Caco‐2 Cells: Inhibition with Verapamil and Ritonavir

This study investigated fexofenadine (FXD) transport and the inhibition of FXD transport in Caco‐2 cell monolayer transwells, usingrhodamine 123 (RH123) transport as a positive control. FXD transport from the basolateral (B) to apical (A) compartment was fivefold higher than A to B transport. FXD transport was linear with respect to time (up to 270 min) and concentration (up to 300 μm). Similar results were seen with the positive control RH123. Ritonavir (100 μM) and verapamil (100 μm) reduced transport of FXD and RH123 by more than 80%, whereas transport was not inhibited by 100 m indomethacin or 2 mM probenecid. This suggests predominantly P‐glycoprotein (P‐gp)‐mediated transport as opposed to transport by multidrug resistance protein. In concentration‐response experiments, FXD transport was inhibited by verapamil and ritonavir with IC50 values of 6.5 μm and 5.4 μm, respectively. Results from this in vitro study demonstrate differential transport of FXD across Caco‐2 cell monolayers and inhibition of FXD transport by established P‐gp inhibitors. The findings support the use of FXD as an index or probe compound to reflect P‐gp activity in vivo.

P‐glycoprotein Interactions of Nefazodone and Trazodone in Cell Culture

This study investigated the effects of nefazodone (NFZ) and trazodone (TZD) on P‐glycoprotein (P‐gp) activity and expression in cell culture. NFZ and TZD showed no differential transport between the basolateral to apical and apical to basolateral direction across Caco‐2 cell monolayers. Transport in either direction was not affected by verapamil. NFZ was a potent inhibitor (IC50 = 4.7 M) of rhodamine123 (Rh123)B to A transport across Caco‐2 cell monolayers, while TZD had minimal effect. Following 72‐hour exposure of LS180V cells to NFZ and TZD (10 M), a twofold increase in immunoreactive P‐gp was observed. Rh123 accumulation into these cells was reduced to 65% and 74% of control by NFZ and TZD (10 M), respectively. It was concluded that differential rates of transport of NFZ and TZD in Caco‐2 cells were not evident. However, NFZ is an inhibitor of P‐gp activity at clinically relevant in vivo concentrations and may have the potential to increase bioavailability of coadministered compounds that are substrates for transport. Concentrations of NFZ and TZD achieved in the intestine after chronic oral dosing may induce P‐gp expression and reduce absorption of coadministered drugs.