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  • Role of multidrug resistance-associated protein 4 in the basolateral efflux of hepatically derived enalaprilat.

Role of multidrug resistance-associated protein 4 in the basolateral efflux of hepatically derived enalaprilat.

Drug metabolism and disposition: the biological fate of chemicals (2014-06-25)
Brian C Ferslew, Kathleen Köck, Arlene S Bridges, Kim L R Brouwer
ABSTRACT

Hepatic uptake and efflux transporters govern the systemic and hepatic exposure of many drugs and metabolites. Enalapril is a pharmacologically inactive prodrug of enalaprilat. Following oral administration, enalapril is converted to enalaprilat in hepatocytes and undergoes translocation into the systemic circulation to exert its pharmacologic effect by inhibiting angiotensin-converting enzyme. Although the transport proteins governing hepatic uptake of enalapril and the biliary excretion of enalapril and enalaprilat are well established, it remains unknown how hepatically derived enalaprilat translocates across the basolateral membrane into the systemic circulation. In this study, the role of ATP-binding cassette transporters in the hepatic basolateral efflux of enalaprilat was investigated using membrane vesicles. ATP-dependent uptake of enalaprilat into vesicles expressing multidrug resistance-associated protein (MRP) 4 was significantly greater (∼3.8-fold) than in control vesicles. In contrast, enalaprilat was not transported to a significant extent by MRP3, and enalapril was not transported by either MRP3 or MRP4. The functional importance of MRP4 in the basolateral excretion of derived enalaprilat was evaluated using a novel basolateral efflux protocol developed in human sandwich-cultured hepatocytes. Under normal culture conditions, the mean intrinsic basolateral efflux clearance (CLint ,basolateral) of enalaprilat was 0.026 ± 0.012 µl/min; enalaprilat CLint,basolateral was significantly reduced to 0.009 ± 0.009 µl/min by pretreatment with the pan-MRP inhibitor MK-571. Results suggest that hepatically derived enalaprilat is excreted across the hepatic basolateral membrane by MRP4. Changes in MRP4-mediated basolateral efflux may alter the systemic concentrations of this active metabolite, and potentially the efficacy of enalapril.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Enalaprilat dihydrate, ≥98% (HPLC)
Supelco
Enalaprilat, analytical standard
USP
Enalaprilat, United States Pharmacopeia (USP) Reference Standard
Enalaprilat dihydrate, European Pharmacopoeia (EP) Reference Standard
Enalapril maleate, European Pharmacopoeia (EP) Reference Standard
Sigma-Aldrich
Ethylenediaminetetraacetic acid, SAJ special grade, ≥99.0%
Sigma-Aldrich
Ethylenediaminetetraacetic acid disodium salt solution, BioUltra, for molecular biology, pH 8.0, ~0.5 M in H2O
Sigma-Aldrich
Ethylenediaminetetraacetic acid, BioUltra, ≥99.0% (KT)
Sigma-Aldrich
Ethylenediaminetetraacetic acid, anhydrous, crystalline, BioReagent, suitable for cell culture
Sigma-Aldrich
Ethylenediaminetetraacetic acid, purified grade, ≥98.5%, powder
Sigma-Aldrich
Ethylenediaminetetraacetic acid, anhydrous, BioUltra, ≥99% (titration)
Sigma-Aldrich
Ethylenediaminetetraacetic acid, ACS reagent, 99.4-100.6%, powder
Sigma-Aldrich
Ethylenediaminetetraacetic acid solution, 0.02% in DPBS (0.5 mM), sterile-filtered, BioReagent, suitable for cell culture
Supelco
Enalapril Maleate, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
Ethylenediaminetetraacetic acid, 99.995% trace metals basis
Sigma-Aldrich
Ethylenediaminetetraacetic acid, ≥98.0% (KT)
USP
Enalapril maleate, United States Pharmacopeia (USP) Reference Standard