Species differences in sinusoidal and canalicular efflux transport of mycophenolic acid 7-O-glucuronide in sandwich-cultured hepatocytes

Abstract

Metabolism and sinusoidal/canalicular efflux of mycophenolic acid (MPA) was investigated using sandwich-cultured hepatocytes (SCHs). After applying MPA to SCHs from humans, wild-type rats, and multidrug resistance-associated protein (Mrp) 2-deficient rats, the MPA metabolites 7-O-glucuronide (MPAG) and acyl glucuronide (AcMPAG) were detected in the intracellular compartment of the SCHs. Sinusoidal efflux of MPAG was detected in all SCH preparations including Mrp2-deficient rat SCHs, whereas canalicular efflux of MPAG was observed in wild-type rat and human SCHs but not in Mrp2-deficient rat SCHs. The ratio of canalicular efflux to net (canalicular plus sinusoidal) efflux was 37 ± 8% in wild-type rat SCHs, while the ratio in human SCHs was significantly lower (20 ± 2%, P < 0.05), indicating species differences in the direction of hepatic MPAG transport. This 20% ratio in human SCHs corresponds to a high sinusoidal MPAG efflux (80%) that can in part account for the urine-dominated recovery of MPAG in humans. Both sinusoidal and canalicular MPAG efflux in rat SCHs shows a good correspondence to urinary and biliary recovery of MPAG after MPA dosing. The sinusoidal efflux of AcMPAG in human SCHs was detected from one out of three donors, suggesting donor-to-donor variation. In conclusion, this study demonstrates the predictive value of SCHs for elucidating the interplay of metabolism and efflux transport, in addition to demonstrating a species difference between rat and human in sinusoidal and canalicular efflux of MPAG.

Keywords: Biliary excretion; glucuronidation; hepatocytes; mycophenolic acid; sinusoidal efflux; transporter.

Figure 1

Typical time course of MPAG, AcMPAG, and MPA efflux to buffer after applying MPA to rat, Mrp2 KO rat, and human SCH. Closed circles (●) and open squares (□) represent mass in Plus (+) buffer (Mass (+)) and mass in Minus (−) buffer (Mass (−)), respectively. Mass (+) can represent sinusoidal efflux while Mass (−) can represent net efflux (sinusoidal plus canalicular efflux). Data are expressed as mean ± SD (n = 3). *P < 0.05, **P < 0.01, and ***P < 0.001, respectively.

Figure 2

Relative sinusoidal/canalicular efflux of MPAG among rat, Mrp2 KO rat, and human SCH. Given observations of statistical significance in rat and human SCHs between Mass (+) and Mass (−) (n = 3), analysis to determine relative sinusoidal/canalicular efflux was conducted. The results of three independent SCH preparations are expressed as closed circles (●). Mean ± SD lines are also plotted. ^aIn Mrp2 KO rat SCHs, statistic significance was not observed between Mass (+) and Mass (−) (n = 3) in two independent SCH preparations. *P < 0.05.