Mihwa Kwon

Enhanced Oral Bioavailability of Morin Administered in Mixed Micelle Formulation with PluronicF127 and Tween80 in Rats

To overcome the low oral bioavailability of morin, a mixed micelle formulation with pharmaceutical excipients that facilitate solubilization and modulate P-glycoprotein (P-gp) was developed and evaluated in vitro and in vivo rats. Morin-loaded mixed micelle formulation with a morin-PluronicF127-Tween80 ratio of 1 : 10 : 0.02 (w/w/w) was prepared by a thin-film hydration method. The solubility, size distribution, drug encapsulation efficiency, and percent drug loading of the formulation were characterized. Subsequently, in vivo pharmacokinetic parameters of morin loaded in a PluronicF127 and Tween80 mixed-micelle formulation were investigated in rats. Absolute bioavailability of morin was dramatically increased by the oral administration of morin-loaded PluronicF127 and Tween80 mixed micelle from 0.4% to 11.2% without changing the systemic clearance and half-life. In Caco-2 cells, absorption permeability of morin from the novel formulation was increased 3.6-fold compared with that of morin alone. P-gp inhibition by cyclosporine A (CsA) increased absorptive permeability of morin 2.4-fold but decreased the efflux of morin by 52%, which was consistent with increased plasma concentration of morin in the pretreatment of CsA in rats. The morin formulation inhibited P-gp transport activity by 83.1% at 100 µM as morin concentration. Moreover, morin formulation increased paracellular permeability of Lucifer yellow by 1.6-1.8 fold. In conclusion, enhanced oral bioavailability of morin from morin-loaded PluronicF127 and Tween80 mixed micelle formulation can be attributed to increased intestinal permeation of morin, which was mediated at least by P-gp inhibition and enhanced paracellular route.

Organic anion transporter 3- and organic anion transporting polypeptides 1B1- and 1B3-mediated transport of catalposide

We investigated the in vitro transport characteristics of catalposide in HEK293 cells overexpressing organic anion transporter 1 (OAT1), OAT3, organic anion transporting polypeptide 1B1 (OATP1B1), OATP1B3, organic cation transporter 1 (OCT1), OCT2, P-glycoprotein (P-gp), and breast cancer resistance protein (BCRP). The transport mechanism of catalposide was investigated in HEK293 and LLC-PK1 cells overexpressing the relevant transporters. The uptake of catalposide was 319-, 13.6-, and 9.3-fold greater in HEK293 cells overexpressing OAT3, OATP1B1, and OATP1B3 transporters, respectively, than in HEK293 control cells. The increased uptake of catalposide via the OAT3, OATP1B1, and OATP1B3 transporters was decreased to basal levels in the presence of representative inhibitors such as probenecid, furosemide, and cimetidine (for OAT3) and cyclosporin A, gemfibrozil, and rifampin (for OATP1B1 and OATP1B3). The concentration-dependent OAT3-mediated uptake of catalposide revealed the following kinetic parameters: Michaelis constant (Km) =41.5 μM, maximum uptake rate (Vmax) =46.2 pmol/minute, and intrinsic clearance (CLint) =1.11 μL/minute. OATP1B1- and OATP1B3-mediated catalposide uptake also showed concentration dependency, with low CLint values of 0.035 and 0.034 μL/minute, respectively. However, the OCT1, OCT2, OAT1, P-gp, and BCRP transporters were apparently not involved in the uptake of catalposide into cells. In addition, catalposide inhibited the transport activities of OAT3, OATP1B1, and OATP1B3 with half-maximal inhibitory concentration values of 83, 200, and 235 μM, respectively. However, catalposide did not significantly inhibit the transport activities of OCT1, OCT2, OAT1, P-gp, or BCRP. In conclusion, OAT3, OATP1B1, and OATP1B3 are major transporters that may regulate the pharmacokinetic properties and may cause herb–drug interactions of catalposide, although their clinical relevance awaits further evaluation.

Transport characteristics and transporter‐based drug–drug interactions of TM‐25659, a novel TAZ modulator

The in vitro metabolic stability and transport mechanism of TM‐25659, a novel TAZ modulator, was investigated in human hepatocytes and human liver microsomes (HLMs) based on the preferred hepatobiliary elimination in rats. In addition, the in vitro transport mechanism and transporter‐mediated drug–drug interactions were evaluated using oocytes and MDCKII cells overexpressing clinically important drug transporters. After a 1 h incubation in HLMs, 92.9 ± 9.5% and 95.5 ± 11.6% of the initial TM‐25659 remained in the presence of NADPH and UDPGA, respectively. Uptake of TM‐25659 readily accumulated in human hepatocytes at 37 ºC (i.e. 6.7‐fold greater than that at 4 ºC), in which drug transporters such as OATP1B1 and OATP1B3 were involved. TM‐25659 had a significantly greater basal to apical transport rate (5.9‐fold) than apical to basal transport rate in the Caco‐2 cell monolayer, suggesting the involvement of an efflux transport system. Further studies using inhibitors of efflux transporters and overexpressing cells revealed that MRP2 was involved in the transport of TM‐25659. These results, taken together, suggested that TM‐25659 can be actively influxed into hepatocytes and undergo biliary excretion without substantial metabolism. Additionally, TM‐25659 inhibited the transport activities of OATP1B1 and OATP1B3 with IC50 values of 36.3 and 25.9 μm, respectively. TM‐25659 (100 μm) increased the accumulation of the probe substrate by 160% and 213%, respectively, through the inhibition of efflux function of P‐gp and MRP2. In conclusion, OATP1B1, OATP1B3, P‐gp and MRP2 might be major transporters responsible for the pharmacokinetics and drug–drug interaction of TM‐25659, although their contribution to in vivo pharmacokinetics needs to be further investigated. Copyright

Pharmacokinetics of Jaspine B and Enhancement of Intestinal Absorption of Jaspine B in the Presence of Bile Acid in Rats

We aimed to investigate the pharmacokinetics and the underlying mechanisms of the intestinal absorption, distribution, metabolism, and excretion of Jaspine B in rats. The oral bioavailability of Jaspine B was 6.2%, but it decreased to 1.6% in bile-depleted rats and increased to 41.2% (normal) and 23.5% (bile-depleted) with taurocholate supplementation (60 mg/kg). Consistent with the increased absorption in the presence of bile salts, rat intestinal permeability of Jaspine B also increased in the presence of 10 mM taurocholate or 20% bile. Further studies demonstrated that the enhanced intestinal permeability with bile salts was due to increased lipophilicity and decreased membrane integrity. Jaspine B was designated as a highly tissue-distributed compound, because it showed large tissue to plasma ratios in the brain, kidney, heart, and spleen. Moreover, the recovery of Jaspine B from the feces and urine after an intravenous administration was about 6.3%, suggesting a substantial metabolism of Jaspine B. Consistent with this observation, 80% of the administered Jaspine B was degraded after 1 h incubation with rat liver microsomes. In conclusion, the facilitated intestinal permeability in the presence of bile salts could significantly increase the bioavailability of Jaspine B and could lead to the development of oral formulations of Jaspine B with bile salts. Moreover, the highly distributed features of Jaspine B in the brain, kidney, heart, and spleen should be carefully considered in the therapeutic effect and toxicity of this compound.

Differential Cytotoxic Effects of Jaspine B in Various Cancer Cells

Jaspine B is an anhydrophytosphingosine that is isolated from a marine sponge. Because of its structural similarity to sphingosine, it shows anti-cancer effects in human carcinomas. Therefore, this study aims to investigate its anti-proliferative effect on various cancer cells and to correlate its association with the intracellular accumulation of Jaspine B in relevant cancer cells. The anti-proliferative effect of Jaspine B in various cancer cells was determined by a cell viability test, and the intracellular concentration of Jaspine B in relevant cancer cells was determined using mass spectrometry coupled with liquid chromatography. The correlation coefficient and p value between the cytotoxicity and the cell accumulation of Jaspine B were determined using SPSS 16.1. The cytotoxicity of Jaspine B varied depending on the type of cancer cell when compared the EC 50 values of Jaspine B. Breast and melanoma cancer cells were susceptible to Jaspine B, whereas renal carcinoma cells were resistant. The intracellular concentrations of Jaspine B had a reciprocal correlation with the EC 50 values in the same cells (r = 0.838). The results suggested that the anti-proliferative effect of Jaspine B was associated with the cellular accumulation of this compound. However, Jaspine B was not a substrate for P-glycoprotein and breast cancer resistance protein, as major efflux pumps caused multidrug resistance. The maintenance of a high intracellular concentration is crucial for the cytotoxic effect of Jaspine B; however, efflux pumps may not be a controlling factor for Jaspine B-related resistance in cancer cells.

Ursodeoxycholate Restores Biliary Excretion of Methotrexate in Rats with Ethinyl Estradiol Induced-Cholestasis by Restoring Canalicular Mrp2 Expression

The in vivo relevance of ursodeoxycholate (UDCA) treatment (100 mg/kg/day, per oral tid for 5 days before cholestasis induction followed by the same dosing for 5 days) on hepatic function was investigated in rats with 17α-ethinylestradiol (EE, 10 mg/kg, subcutaneous for 5 days)-induced experimental cholestasis. The bile flow rate and the expression level of hepatic multidrug resistance-associated protein 2 (Mrp 2) that were decreased in cholestasis were restored after UDCA treatment. Consistent with this, the biliary excretion clearance (CLexc,bile) of a representative Mrp2 substrate—methotrexate (MTX)—was decreased in cholestatic rats but was restored after UDCA treatment. Consequently, the plasma concentrations of MTX, which were increased by cholestasis, were decreased to control levels by UDCA treatment. Thus, the restoration of CLexc,bile appears to be associated with the increase in Mrp2 expression on the canalicular membrane by UDCA treatment followed by Mrp2-mediated biliary excretion of MTX. On the other hand, the hepatic uptake clearance (CLup,liver) of MTX was unchanged by cholestasis or UDCA treatment, suggestive of the absence of any association between the uptake process and the overall biliary excretion of MTX. Since UDCA has been known to induce the expression of canalicular MRP2 in humans, UDCA treatment might be effective in humans to maintain or accelerate the hepatobiliary elimination of xenobiotics or metabolic conjugates that are MRP2 substrates.