Cell-to-Medium Concentration Ratio Overshoot in the Uptake of Statins by Human Hepatocytes in Suspension, but Not in Monolayer: Kinetic Analysis Suggesting a Partial Loss of Functional OATP1Bs.

Suspended human hepatocytes (SHH) have long been used in assessing hepatic drug uptake, while plated human hepatocytes in short-term monolayer culture (PHH) have gained use in recent years. This study aimed to cross-evaluate SHH and PHH in measuring the hepatic uptake mediated by organic anion transporting polypeptide 1Bs (OATP1Bs). We compared the time courses of cell-to-medium (C/M) concentration ratios and initial uptake clearance values of the OATP1B substrates (pitavastatin, rosuvastatin, cerivastatin, pravastatin, dehydropravastatin, and SC-62807) between SHH and PHH. For all compounds except cerivastatin, the C/M ratios in SHH displayed an apparent overshoot (an initial increase followed by a decrease) during the 180-min uptake experiment, but not in PHH. Based on the literature evidence suggesting the possible internalization of OATP1Bs in primary hepatocytes, separate experiments measured the drug uptake after varying lengths of pre-incubation in the drug-free medium. The initial uptake clearances of pitavastatin and rosuvastatin declined in SHH beyond an apparent threshold time of 20-min drug-free pre-incubation, but not in PHH. Kinetic modeling quantitatively captured the decline in the active uptake clearance in SHH, and more than half of the active uptake clearances of pitavastatin and rosuvastatin were prone to loss during the 180-min uptake experiment. These results suggested a partial, time-delayed loss of the functional OATP1Bs in SHH upon prolonged incubation. Our results indicate that PHH is more appropriate for experiments where a prolonged incubation is required, such as estimation of unbound hepatocyte-to-medium concentration ratio (Kp,uu) at the steady-state.

Investigation of the impact of substrate selection on in vitro organic anion transporting polypeptide 1B1 inhibition profiles for the prediction of drug-drug interactions.

The risk assessment of organic anion transporting polypeptide (OATP) 1B1-mediated drug-drug interactions (DDIs) is an indispensable part of drug development. We previously reported that in vitro inhibitory potencies of several inhibitors on OATP1B1 depend on the substrates when prototypical substrates, estradiol-17ß-glucuronide (E2G), estrone-3-sulfate, and sulfobromophthalein were used as test substrates. The purpose of this study was to comprehensively investigate this substrate-dependent inhibition of OATP1B1 using clinically relevant OATP1B1 inhibitors and substrate drugs. Effects of cyclosporine A (CsA), rifampin, and gemfibrozil on OATP1B1-mediated uptake of 12 substrate drugs were examined in OATP1B1-expressing human embryonic kidney 293 cells. The Ki values (µM) for CsA varied from 0.0771 to 0.486 (6.3-fold), for rifampin from 0.358 to 1.23 (3.4-fold), and for gemfibrozil from 9.65 to 252 (26-fold). Except for the inhibition of torasemide uptake by CsA and that of nateglinide uptake by gemfibrozil, the Ki values were within 2.8-fold of those obtained using E2G as a substrate. Preincubation potentiated the inhibitory effect of CsA on OATP1B1 with similar magnitude regardless of the substrates. R values calculated based on a static model showed some variation depending on the Ki values determined with various substrates, and such variability could have an impact on the DDI predictions particularly for a weak-to-moderate inhibitor (gemfibrozil). OATP1B1 substrate drugs except for torasemide and nateglinide, or E2G as a surrogate, is recommended as an in vitro probe in the inhibition experiments, which will help mitigate the risk of false-negative DDI predictions potentially caused by substrate-dependent Ki variation.

Evaluation of Oatp and Mrp2 activities in hepatobiliary excretion using newly developed positron emission tomography tracer [11C]dehydropravastatin in rats.

We developed a pravastatin derivative, sodium (3R,5R)-3,5-dihydroxy-7-((1S,2S,6S,8S)-6-hydroxy-2-methyl-8-((1-[(11)C]-(E)-2-methyl-but-2-enoyl)oxy)-1,2,6,7,8,8a-hexahydronaphthalen-1-yl)heptanoate ([(11)C]DPV), as a positron emission tomography (PET) probe for noninvasive measurement of hepatobiliary transport, and conducted pharmacokinetic analysis in rats as a feasibility study for future clinical study. Transport activities of DPV in freshly isolated rat hepatocytes and rodent multidrug resistance-associated protein 2 (rMrp2; human, MRP2)-expressing membrane vesicles were similar to those of pravastatin. Rifampicin diminished the uptake of DPV and pravastatin by the hepatocytes, with similar inhibition potency. [(11)C]DPV underwent biotransformation to produce at least two metabolites in rat, but metabolism of [(11)C]DPV occurred negligibly in human hepatocytes during a 90-minute incubation. After intravenous injection, [(11)C]DPV was mainly distributed to the liver and kidneys, where the tissue uptake clearances (CLuptake,liver and CLuptake,kidney) were blood-flow-limited (73.6 ± 4.8 and 24.6 ± 0.6 ml/min per kilogram, respectively). Systemic elimination of [(11)C]DPV was delayed in rifampicin-treated rat and an Mrp2-deficient mutant rat, Eisai hyperbilirubinemic mutant rat (EHBR). Rifampicin treatment decreased both CLuptake,liver and CLuptake,kidney of [(11)C]DPV by 30% (P < 0.05), whereas these parameters were unchanged in EHBR. Meanwhile, the canalicular efflux clearance (CLint,bile) of [(11)C]DPV, which was 12.2 ± 1.5 ml/min per kilogram in the control rat, decreased by 60% and 89% in rifampicin-treated rat and EHBR (P < 0.05), respectively. These results indicate that [(11)C]DPV is taken up into the liver by organic anion-transporting polypeptides (rodent, Oatps; human, OATP) and excreted into bile by Mrp2 in rat, and that rifampicin may inhibit Mrp2 as well as Oatps, and consequently increase systemic exposure of [(11)C]DPV. PET using [(11)C]DPV is feasible for studies prior to the future clinical investigation of OATP and MRP2 functionality, especially for personalized medicine.

Laparoscopic surgery for the ascending colon cancer associated with malrotation of the midgut.

Malrotation of the midgut is a congenital anomaly of the gastrointestinal tract that usually presents in neonates. Moreover, synchronous colon cancer has rarely been reported. In the present article, we report a preliminary experience with laparoscopic approach for intestinal malrotation with early colon cancer in a 68-year-old woman who presented with bloody stools. Colonoscopy revealed a lateral spreading tumor of the ascending colon. An air-barium contrast enema showed that the entire colon lay within the left hemiabdomen. A computed tomography revealed the superior mesenteric vein rotation sign. At surgery, a condition of malrotation of the midgut was observed: the third and the fourth part of the duodenum descended vertically without Treitz's ligament, and the small bowel and colon were located in the right and left side of the abdominal cavity, respectively. We mobilized the terminal ileum and the right colon with laparoscopic approach. A 3-cm abdominal incision was made via the umbilicus. Right colectomy with lymph node dissection was achieved following extracorporealization. Pathological examination revealed well-differentiated tubular adenocarcinoma without nodal involvement. The patient had an uneventful postoperative course. Laparoscopic surgery for colon cancer associated with malrotation of the midgut is feasible and a promising method because of its less invasiveness and its adaptability to the malrotation without extending the skin incision.

Microdosing clinical study: pharmacokinetic, pharmacogenomic (SLCO2B1), and interaction (grapefruit juice) profiles of celiprolol following the oral microdose and therapeutic dose.

The authors evaluated the contribution of the SLCO2B1 polymorphism to the pharmacokinetics of celiprolol at a microdose (MD) and therapeutic dose (TD) and compared pharmacokinetic proportionality between the 2 dose forms in 30 SLCO2B1 genotype-matched healthy volunteers. Three drugs (celiprolol, fexofenadine, and atenolol) were orally administered as a cassette dosing following the MD (totally 97.5 µg) and then a TD (100 mg) of celiprolol, with and without grapefruit juice. The mean AUC(0-24) of celiprolol was lower in SLCO2B1*3/*3 individuals (775 ng·h/mL) than in *1/*3 (1097 ng·h/mL) and *1/*1 (1547 ng·h/mL) individuals following the TD, and this was confirmed in population pharmacokinetic analysis with statistical significances; however, SLCO2B1 genotype-dependent differences disappeared following the MD. Dose-normalized AUC of celiprolol at the MD was much lower than that at the TD, explained by the saturation of the efflux transporter. Thus, the effect of SLCO2B1 polymorphism on the AUC of celiprolol clearly observed only at the TD may be due to the saturation of the efflux transport systems.