PhD Scientific Days 2025

Pharmaceutical Sciences and Health Technologies I.

Investigating in vitro species differences and prediction of CNS penetration using MDR1 overexpressing cells

Name of the presenter

Sóskuti Emőke

Institute/workplace of the presenter

Charles River Laboratories Hungary Kft.

Authors

Emőke Sóskuti^1,2, Nóra Szilvásy¹, Csilla Temesszentandrási-Ambrus¹, György Várady³, Zsuzsanna Gáborik¹

1: Charles River Laboratories Hungary Kft.
2: Doctoral School of Semmelweis University, Molecular Medicine Division
3: HUN-REN Hungarian Research Network

Text of the abstract

Determining blood-brain barrier penetration of CNS targeting drugs early in drug discovery is challenging. Efforts to implement the 3R initiative, and the need to reveal species differences call for more predictive models.
Here, we created endogenous Abcb1KO-MDCKII assays, where overexpression of the MDR1 from human, mouse, rat or cynomolgus monkey are comparable. Efflux ratios (ER) were determined, and found to be comparably similar across cell lines, suggesting limited species specificities. In vivo Kp,uu,brain data from literature and ERs determined here were used to predict BBB penetration classification.
Using a cutoff value of 0.3 for Kp,uu,brain and 2 for ERs, the best sensitivity was achieved with the hMDR1 cells, which has the largest transporter abundance, therefore its resolution for weaker substrates is highest. For quantitative prediction different published models were used. The best correlation between in vivo and predicted Kp,uu,brain was achieved with mathematically parametrized calculation using scaling factors. Inhibitory potency of known MDR1 interactors using digoxin or quinidine was also tested. Inhibitory potency of many drugs is highly substrate dependent; here, generally lower IC50s with quinidine were found. In some cases, IC50 difference between orthologs was resolved by transporter abundance correction, but this correlation is dependent on substrate-inhibitor interaction, suggesting dependence on the mode of inhibition.
In summary, this is the first report of systematic comparison of inhibitor and substrate interactions of multiple compounds across MDR1 orthologs. Species differences in DDI potential can be investigated, but transporter abundance and mode of inhibition need to be considered. Our in vitro assays can be integrated into drug discovery screening programs to predict BBB penetration, also, to reveal differences in transporter susceptibility between preclinical and human, improving the translatability of in vivo preclinical data.