Abstract
Generating accurate in vitro data is crucial for in vitro to in vivo extrapolation and pharmacokinetic predictions. The use of HEK293 cells overexpressing OATP1B1 and OATP1B3 in protein-free buffer and 100% human plasma incubations was explored for the uptake of four OATP substrates, pravastatin, rosuvastatin, repaglinide and pitavastatin. Differences were observed for each parameter (Km,u, Vmax, CLint, and Pdif,u) obtained from the buffer and human plasma incubations in both cells, and in general, the fold differences increased as plasma protein binding increased. The fold change in Km,u values ranged from 2.03-1020, and the fold change in Vmax values ranged from 1.22-97.4. As a result, the CLint values generated in the plasma incubations were 1.66-51.9 fold higher than the values generated in protein-free buffer in both cells. The unbound passive diffusion was also consistently higher in the human plasma incubations for all four compounds, with a fold difference range of 2.06-38.4. These shifts in the presence and absence of human plasma suggest that plasma proteins may play a role in both the active uptake and passive diffusion processes. The results also support the idea of a transporter-induced protein-binding shift, where high protein binding may not limit the uptake of compounds that have high affinity for transporters. The addition of plasma to incubations leading to higher CLint values for transporter substrates helps mitigate the underprediction commonly noted with in vitro to in vivo extrapolation.
SIGNIFICANCE STATEMENT The current investigation brought new perspective on how to mitigate the underprediction commonly noted with in vitro to in vivo extrapolation for OATP substrates. It also supports the idea of a transporter-induced protein-binding shift, where high protein binding may not limit the uptake of compounds that have high affinity for transporters.
- in vitro-in vivo prediction (IVIVE)
- liver/hepatic
- protein binding
- Uptake transporters (OATP, OAT, OCT, PEPT, MCT, NTCP, ASBT, etc.)
- The American Society for Pharmacology and Experimental Therapeutics