Simulation-based assessment of the P-glycoprotein expression-activity relationship shows a drug and system dependency

In vitro-in vivo extrapolation (IVIVE) of P-glycoprotein (P-gp) transporter activity bears two important assumptions: (1) P-gp expression (i.e., concentration) is linearly related to P-gp activity; and (2) this relation is drug independent. However, conflicting experimental results have been obtained about this relationship. This study therefore aimed to theoretically explore the P-gp expression-activity relationship (EAR). A P-gp membrane kinetic binding model was used to explore the P-gp EAR, and how this is impacted by different drugs as well as drug dose and initial P-gp expression. The model included passive permeability and drug interaction with P-gp described through the association (kon), dissociation (koff), and efflux rate (ke) constants. Simulations were first performed for 7 P-gp substrates, assuming an initial P-gp expression of 1000 µM and drug concentrations of 1 µM. Subsequently, the P-gp concentration was varied (2-300%) to derive the EAR. Next, to determine the impact of drug-specific and system-dependent properties on the P-gp EAR, virtual drugs varying in koff and ke were simulated, simultaneously considering different initial P-gp expressions and drug concentrations. Our simulations show that the P-gp EAR is not always linear, with the EAR showing both linear and non-linear behaviour depending on the drug. The koff/ke ratio of a drug was found to be an important determinant of the P-gp EAR, which shifted towards non-linearity for lower koff/ke ratios. Additionally, the P-gp EAR was more likely to be non-linear for higher ratios of the initial P-gp expression to drug concentration. In conclusion, this study shows that P-gp expression is not always proportional to P-gp activity, thereby explaining experimental contradictions about the EAR. This implies that using P-gp expression as single biomarker for P-gp activity needs to be reconsidered.