In-vitro characterization of coding variants with predicted functional implications in the efflux transporter...

Area of Science:

• Pharmacogenomics
• Molecular Biology
• Drug Metabolism

Background:

• Multidrug resistance-associated protein 4 (MRP4) is a key efflux transporter involved in drug disposition and toxicity.
• Genetic variations in MRP4 can alter its function, potentially leading to adverse drug reactions.
• Understanding MRP4 variant function is crucial for personalized medicine and predicting drug efficacy.

Purpose of the Study:

• To experimentally characterize the functional impact of common MRP4 coding variants.
• To investigate the effects of these variants on transporter activity and protein localization.
• To assess the substrate-specific effects and potential mechanisms underlying altered MRP4 function.

Main Methods:

• Selected ten common MRP4 coding variants in Europeans, focusing on nine predicted to be functional.
• Assessed protein localization and activity using HEK293T cells over-expressing wildtype or variant MRP4.
• Quantified intracellular accumulation of taurocholic acid (TCA) and estradiol 17-β-glucuronide (E217βG) as prototypic substrates.

Main Results:

• V458M and T1142M variants exhibited reduced activity for both E217βG and TCA.
• L18I, G187W, K293E, and R531Q variants showed substrate-dependent increases in activity.
• V458M displayed reduced cell surface expression, while G187W showed a trend for reduced surface expression.

Conclusions:

• Six of nine predicted functional MRP4 variants demonstrated moderately altered activity with varying mechanisms.
• Altered activity may stem from changes in protein surface expression or intrinsic transporter function.
• These findings highlight the clinical importance of MRP4 variants and the need for further cell-based studies with multiple substrates.