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IPEC-J2 rMdr1a, a New Cell Line with Functional Expression of Rat P-glycoprotein Encoded by Rat Mdr1a for Drug
Lasse Saaby1, Josefine Trasborg2, Mikkel A Rasmussen1
1Bioneer A/S, Kogle Alle 2, DK-2970 Hørsholm, Denmark.
Abstract:
The efflux pump P-glycoprotein (P-gp) affects drug distribution after absorption in humans and animals. P-gp is encoded by the multidrug resistance gene (MDR1) gene in humans, while rodents (the most common preclinical animal model) express the two isoforms Mdr1a and Mdr1b. Differences in substrate selectivity has also been reported. Our aim was to generate an in vitro cell model with tight barrier properties, expressing functional rat Mdr1a P-gp, as an in vitro tool for investigating species differences. The IPEC-J2 cell line forms extremely tight monolayers and was transfected with a plasmid carrying the rat Mdr1a gene sequence. Expression and P-gp localization at the apical membrane was demonstrated with Western blots and immunocytochemistry. Function of P-gp was shown through digoxin transport experiments in the presence and absence of the P-gp inhibitor zosuquidar. Bidirectional transport experiments across monolayers of the IPEC-J2 rMDR1a cell line and the IPEC-J2 MDR1 cell line, expressing human P-gp, showed comparable magnitude of transport in both the absorptive and efflux direction. We conclude that the newly established IPEC-J2 rMdr1a cell line, in combination with our previously established cell line IPEC-J2 MDR1, has the potential to be a strong in vitro tool to compare P-gp substrate profiles of rat and human P-gp.
Insights
Researchers developed a new rat P-glycoprotein (P-gp) cell model for comparing drug transport differences between rats and humans. This in vitro tool aids in understanding species-specific drug distribution and potential drug interactions.
Area of Science:
- Pharmacology
- Cell Biology
- Drug Metabolism
Background:
- P-glycoprotein (P-gp) is an efflux pump influencing drug distribution in humans and animals.
- Rodents express Mdr1a and Mdr1b isoforms, differing in substrate selectivity from human P-gp (encoded by MDR1).
- Investigating species-specific P-gp activity is crucial for preclinical drug development.
Purpose of the Study:
- To create an in vitro cell model with tight barrier properties expressing functional rat Mdr1a P-gp.
- To establish a tool for investigating species differences in P-gp-mediated drug transport.
- To compare P-gp substrate profiles between rat and human P-gp.
Main Methods:
- Transfection of the IPEC-J2 cell line with the rat Mdr1a gene.
- Confirmation of P-gp expression and apical localization via Western blot and immunocytochemistry.
- Assessment of P-gp function using digoxin transport assays with and without zosuquidar.
Main Results:
- The IPEC-J2 rMdr1a cell line exhibited tight monolayer properties.
- Rat Mdr1a P-gp expression and localization were confirmed at the apical membrane.
- Functional P-gp activity was demonstrated through digoxin transport studies.
- Comparable transport magnitudes were observed in both absorptive and efflux directions between rat and human P-gp expressing cell lines.
Conclusions:
- The IPEC-J2 rMdr1a cell line provides a functional in vitro model of rat P-gp.
- This model, alongside the IPEC-J2 MDR1 (human P-gp) line, enables direct comparison of rat and human P-gp substrate profiles.
- The established cell lines offer a valuable tool for studying species differences in drug disposition and P-gp interactions.
