Application of a new MDCKII-MDR1 cell model to measure the extent of drug distribution in vitro at equilibrium for prediction of in vivo unbound brain-to-plasma drug distribution

Intro: Reliable estimates of drug uptake from blood to brain parenchyma are crucial in CNS drug discovery and development. While in vivo K p,uu,brain estimates are the gold standard for investigating brain drug disposition, animal usage is a limitation to high throughput application. This study investigates an in vitro model using P-gp expressing MDCKII-MDR1 cells for predicting in vivo brain drug penetration. Methods: In vitro equilibrium distribution studies were conducted in apical and basolateral solutions with high protein content to estimate K p,brain and K p,uu,brain values. The correlation between in vitro and in vivo K p,brain values for a set of compounds was examined. Results: We observed a good correlation between in vitro and in vivo K p,brain values (R 2 = 0.69, Slope: 1.6), indicating that the in vitro model could predict in vivo drug brain penetration. The ‘unilateral (Uni-L)’ in vitro setup correctly classified 5 out of 5 unrestricted compounds and 3 out of 5 restricted compounds. Possible reasons for the observed disparities for some compounds have been discussed, such as difference in transport areas between in vitro and in vivo settings and effect of pH changes. Conclusion: The in vitro assay setup developed in this study holds promise for predicting in vivo drug brain penetration in CNS drug discovery. The correlation between in vitro and in vivo K p,brain values, underscores that the model may have potential for early-stage screening. With minor refinements, this in vitro approach could reduce the reliance on in vivo experiments, accelerating the pace of CNS drug discovery and promoting a more ethical research approach.