In Vitro Stimulation of Multidrug Resistance-Associated Protein 2 Function Is Not Reproduced In Vivo in Rats.

Ravindranath Gilibili,Vishwanath Kurawattimath,Hong Shen,T. Mariappan,Sagnik Chatterjee,Bokka Murali,Yurong Lai

doi:10.3390/pharmaceutics10030125

Abstract

Previously we reported that coproporphyrin-I (CP-I) is an optimal probe substrate for multidrug resistance-associated protein 2 (MRP2), and stimulation of MRP2-mediated transport is probe substrate-dependent. In the present investigation, we assessed if the in vitro stimulation is physiologically relevant. Similar to human MRP2 transport, CP-I was transported by rat Mrp2 in a typical Michaelis-Menten kinetics with apparent Km and Vmax values of 15 ± 6 µM and 161 ± 20 pmol/min/mg protein, respectively. In vivo Mrp2 functions were monitored by biliary and renal secretion of CP-I and its isomer CP-III, in bile-duct cannulated rats before and after treatment with mitoxantrone, progesterone, and verapamil. These compounds stimulated Mrp2-mediated CP-I transport in vitro. No significant increase in biliary or renal clearances, as well as in the cumulative amount of CP-I or CP-III eliminated in bile, were detected following treatment with the in vitro stimulators, indicating an in vitro to in vivo disconnect. In presence of 10 µM bilirubin, the in vitro stimulation was suppressed. We concluded that the in vitro stimulation of CP-I transport mediated by Mrp2 is not translatable in vivo, and proposed that the presence of endogenous compounds such as bilirubin in the liver may contribute to the in vitro to in vivo disconnect.

Highlights

It is becoming increasingly evident that multidrug resistance-associated protein 2 (MRP2/Mrp2, denoted by gene ABCC2/Abcc2) plays an important role in disposition and elimination to regulate pharmacokinetics and pharmacodynamic of xenobiotics [1,2]
We have previously shown that the stimulators used in the study of Heredi-Szabo et al, benzbromarone and indomethacin, are inhibitors of MRP2, when CP-I is used as a probe substrate [10]
We first showed that CP-I follows a Michaelis-menten kinetics in rat Mrp2 vesicles

Summary

Introduction

It is becoming increasingly evident that multidrug resistance-associated protein 2 (MRP2/Mrp, denoted by gene ABCC2/Abcc2) plays an important role in disposition and elimination to regulate pharmacokinetics and pharmacodynamic of xenobiotics [1,2]. MRP2 facilitates the efflux of glucuronide and glutathione conjugates of endobiotics, such as bilirubin conjugates and dianionic bile acids [4], as well as few anionic xenobiotics, such as methotrexate [5]. The inhibition of MRP2 activity is important for drug hepatic disposition and elimination, and can cause potential drug-drug interactions (DDI) and hepatotoxicity [6]. A two-fold increase in methotrexate plasma levels due to MRP2 polymorphism has been reported [7]. Changes in MRP2 activity in vivo is usually associated with profound changes in liver exposure, and minor/no effect on plasma exposure, and it usually remains undetected [8]. It was shown that atorvastatin liver concentration increases 1.64-fold, in the presence of metformin, an Mrp inhibitor, without affecting the plasma exposure in Sprague Dawley (SD) rats [9]

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