A nonlinear mixed effect model to study albumin-mediated drug transport into endothelial cells in vitro.
Bonnefois G (1), Payen L (2), Guitton J (3), Freyer G (1), Rouvet I (4), Zabot MT (4), Tod M (1,5)
(1)EA3738 Ciblage Thérapeutique en Oncologie, Faculté de Médecine Lyon-Sud, Oullins France; (2)Institut des Sciences Pharmaceutiques et Biologiques, Lyon France; (3)Centre Hospitalier Lyon-Sud, Laboratoire de biochimie-toxicologie, Hospices Civils de Lyon, F-69495, Pierre Bénite, France; (4)Centre de biotechnologie cellulaire, Centre de Biologie et de Pathologie Est - CHU Lyon, 69677 Bron, France; (5) Hospices Civils de Lyon, France
Objectives: Albumin penetrates into endothelial cells after binding to albondin, and is recycled to the surface by FcRn [1]. The objective was to study the impact of albumin-mediated drug transport on drug intracellular concentration in vitro. Sorafenib, an antiangiogenic drug with high affinity for albumin, was used as a probe.
Methods: Human Pulmonary Artery Endothelial Cells were used for cell culture. Cell cultures were exposed to different concentrations of sorafenib (0, 0.0064, 0.021, 0.064, 0.64 µM) and albumin (0, 14.5, 43.5, 145, 725 µM) in triplicate. Sorafenib intracellular concentration was measured by LC-MSMS. A mixed-effect regression model was fitted to the apparent intracellular sorafenib concentrations (NONMEM 7). Two compartments were considered: the medium (A) and the cells (B). Albumin active transport in each direction was assumed to be either nonlinear or linear. In the linear case, the transports were characterized by their clearance in each direction. Sorafenib was assumed to cross the cell membrane by passive diffusion, and by active transport of the albumin-sorafenib complex, with the same rate as albumin. In each compartment, sorafenib was allowed to bind reversibly to albumin. The dissociation constant of this equilibrium was assumed to be the same in both compartments. The unknown factor of dilution of the intracellular content was also estimated. An explicit solution of the steady-state model was derived.
Results: Direct evidence of albumin penetration into cells was obtained by FITC-albumin. The full nonlinear and the reduced linear models could be fitted to the data, but there was no evidence of a better fit with the full model (p = 0.10). The distribution of NPDEs for the linear model was random and showed no trend with increasing concentrations of either albumin or sorafenib. Removing the active transport from the final model (i.e., fixing CLAB = 0) resulted in a significantly worse fit (p < 0.0001). The ratio of albumin concentration A/B was equal to 7.
Conclusions: Albumin increases sorafenib penetration either by enhancing diffusion or active transport. Simulations show that the intracellular total concentration of sorafenib would be much lower if albumin was not actively transported. In case of rapid drug elimination from the cells, this active transport may enhance intracellular free drug concentration. The model equation may be applied to other drugs, binding proteins and cell types.
References:
[1] Jonghan Kim, William L Hayton, John M Robinson, and Clark L Anderson. Kinetics of fcrn-mediated recycling of igg and albumin in human : pathophysiology and therapeutic implications using a simplied mechanism-based model. Clin Immunol, 122(2) :146155, Feb 2007.
