Effect of food and polymorphisms in SLCO2B1, CYP3A4 and UGT1A4 on pharmacokinetics of abiraterone and its metabolites in Chinese volunteers.

AIMS: Abiraterone acetate, a prodrug of abiraterone (ABI), provides an efficient therapeutic option for metastatic castration-resistant prostate cancer patients. ABI undergoes extensive metabolism in vivo and is transformed into active metabolites Δ4 -abiraterone and 3-keto-5α-abiraterone as well as inactive metabolites abiraterone sulfate and abiraterone N-oxide sulfate. We aimed to examine the effect of polymorphisms in SLCO2B1, CYP3A4 and UGT1A4 on the pharmacokinetics of ABI and its metabolites. METHODS: In this study, 81 healthy Chinese subjects were enrolled and divided into 2 groups for fasted (n = 45) and fed (n = 36) studies. Plasma samples were collected after administering a 250 mg abiraterone acetate tablet followed by liquid chromatography-tandem mass spectrometry analysis. Genotyping was performed on a MassARRAY system. The association between SLCO2B1, CYP3A4, UGT1A4 genotype and pharmacokinetic parameters of ABI and its metabolites was assessed. RESULTS: Food effect study demonstrated high fat meal remarkedly increased systemic exposure of ABI and its metabolites. The geometric mean ratio and 90% confidence interval of area under the plasma concentration-time curve from time 0 to the time of the last quantifiable concentration (AUC0-t ) and maximum plasma concentration (Cmax ) of ABI in fed state vs. fasted state were 351.64% (286.86%-431.04%) and 478.45% (390.01%-586.94%), respectively, while the corresponding results were ranging from 145.11% to 269.42% and 150.10% to 478.45% for AUC0-t and Cmax of ABI metabolites in fed state vs. fasted state, respectively. The SLCO2B1 rs1077858 had a significant influence on AUC0-t and Cmax , while 7 other SLCO2B1 variants prolonged half-life of ABI under both fasted and fed conditions. As for ABI metabolites, the systemic exposure of Δ4 -abiraterone, abiraterone sulfate and abiraterone N-oxide sulfate as well as the elimination of 3-keto-5α-abiraterone were significantly affected by SLCO2B1 polymorphisms. Polymorphisms in CYP3A4 and UGT1A4 did not significantly affect pharmacokinetics of ABI and its metabolites. CONCLUSION: Polymorphisms in SLCO2B1 were significantly related to the pharmacokinetic variability of ABI and its metabolites under both fasted and fed conditions.

Molecular mechanisms of hepatoprotective effect of tectorigenin against ANIT-induced cholestatic liver injury: Role of FXR and Nrf2 pathways.

Cholestatic liver injury is caused by toxic action or allergic reaction, resulting in abnormality of bile formation and excretion. Few effective therapies have become available for the treatment of cholestasis. Herein, we found that tectorigenin (TG), a natural isoflavone, showed definite protective effects on alpha-naphthylisothiocyanate (ANIT)-induced cholestatic liver injury, significantly reversing the abnormality of plasma alanine/aspartate aminotransferase, total/direct bilirubin and alkaline phosphatase, as well as hepatic reactive oxygen species, catalase and superoxide dismutase. Importantly, the targeted metabolomic determination found that BA homeostasis could be well maintained in TG-treated cholestatic mice, especially the levels of glycocholic acid, tauromuricholic acid, taurocholic acid, taurolithocholic acid, tauroursodeoxycholic acid and taurodeoxycholic acid. Overall, primary/secondary and amidated/unamidated bile acid (BA) levels were significantly altered upon ANIT stimulation but could be restored by TG intervention to certain extents. In addition, TG boosted the expression of farnesoid x receptor (FXR), which in turn upregulated multidrug resistance protein 2 (MRP2) and bile salt export pump (BSEP) to accelerate the excretion of BA. Meanwhile, TG enhanced the expression of Nrf2 and its upstream genes PI3K/Akt and downstream target genes HO-1, NQO1, GCLC and GCLM to strengthen the antioxidant capacity. Taken together, TG plays a vital role in maintaining BA homeostasis and ameliorating cholestatic liver injury through regulating FXR-mediated BA efflux and Nrf2-mediated antioxidative pathways.

A topical thermosensitive hydrogel system with cyclosporine A PEG-PCL micelles alleviates ulcerative colitis induced by TNBS in mice.

Ulcerative colitis (UC) is an idiopathic, chronic, relapsing disease. In most cases, only the distal colon is affected, and the colonic stasis or fast colonic transit through the inflamed colon usually results in reduced exposure of the distal inflamed colon. Although the immunosuppressant cyclosporine A (CsA) has been used in patients with severe colitis who do not respond to corticosteroids, the clinical application of CsA remains limited due to the systemic toxicities and insufficient accumulation at the site of action for the intravenous and oral routes. In this study, we loaded CsA into the amphipathic poly(ethylene glycol)-poly(ε-caprolactone) (PEG-PCL) micelles and then embedded them in hydrogels consisting of chitosan, poloxamer 188, and poloxamer 407 to construct a thermosensitive and mucoadhesive hydrogel drug delivery system (PLCP). The PLCP presented a high drug-loading capacity and showed a stable and rapid gelation rate after rectal administration into the body. Compared to CsA-loaded micelles and Sandimmun (Neoral®), the developed thermosensitive gel exhibited prolonged retention on the inflamed colon, as seen from in vitro adhesion and in vivo distribution experiments. It also fast mitigated colitis symptoms in TNBS-treated mice by regulating the expression levels of proinflammatory cytokines (TNF-α, IL-1ß, COX-2, and iNOS2), anti-inflammatory cytokines (IL-10, Nrf2, NQO1, and HO-1), and other relevant biochemical factors. Our results suggested that CsA-loaded micelle thermal hydrogel system could be a promising strategy by enhancing the retention in the diseased colon and promoting the relief and recovery of UC.

The important role of glycerophospholipid metabolism in the protective effects of polyphenol-enriched Tartary buckwheat extract against alcoholic liver disease.

Alcoholic liver disease (ALD) is a mounting public health problem with significant medical, economic and social burdens. Tartary buckwheat (F. tataricum (L.) Gaertn, bitter buckwheat) is a kind of healthy and nutritious food, which has been demonstrated to protect against ALD, but the underlying mechanism has not been fully studied. Herein, we aimed to elucidate the beneficial effects of Tartary buckwheat extract (mainly composed of polyphenols including rutin, quercetin, kaempferol and kaempferol-3-O-rutinoside) in terms of lipid metabolism with the aid of lipidomic analysis. In our study, we employed C57BL/6J mice and a Lieber-DeCarli alcohol liquid diet to construct an ALD model and found that Tartary buckwheat extract was able to prevent ALD-induced histopathological lesions, liver injury and abnormal plasma lipid levels. These beneficial effects might be attributed to the regulation of energy metabolism-related genes (SIRT1, LKB1 and AMPK), lipid synthesis-related genes (ACC, SREBP1c and HMGR) and lipid oxidation-related genes (PPARα, CPT1 and CPT2). In addition, lipidomic profiling and KEGG pathway analysis showed that glycerophospholipid metabolism contributed the most to elucidating the regulatory mechanism of Tartary buckwheat extract. In specific, chronic ethanol intake reduced the level of phosphatidylcholines (PC) and increased the level of phosphatidylethanolamines (PE) in the liver, resulting in a decrease in the PC/PE ratio, which could be all significantly restored by Tartary buckwheat extract intervention, indicating that the Tartary buckwheat extract might regulate PC/PE homeostasis to exert its lipid-lowering effect. Overall, we demonstrated that Tartary buckwheat extract could prevent ALD by modulating hepatic glycerophospholipid metabolism, providing the theoretical basis for its further exploitation as a medical plant or nutritional food.

Enteric Polymer-Based Amorphous Solid Dispersions Enhance Oral Absorption of the Weakly Basic Drug Nintedanib via Stabilization of Supersaturation.

The pH−induced crystallization of weakly basic drugs in the small intestine limits oral bioavailability. In this study, we investigated the solubilization and inhibitory effects on nintedanib in the presence of enteric polymers (HPMCAS LG, HPMCAS MG, Eudragit L100 55, and Eudragit L100). These polymers provided maintenance of supersaturation by increasing the solubility of nintedanib in PBS 6.8 in a concentration-dependent manner, and the improved ranking was as follows: Eudragit L100 > Eudragit L100 55 > HPMCAS MG > HPMCAS LG. After being formulated into amorphous solid dispersions (ASDs) by a solvent evaporation method, the drug exhibited an amorphous state. The pH shift dissolution results of polymer-ASDs demonstrated that four polymers could effectively maintain the drug supersaturation even at the lowest ratio of nintedanib and polymer (1:1, w/w). Eudragit L100−ASD could provide both acid resistance and the favorable mitigation of crystallization in GIF. In comparison to the coarse drug, the relative bioavailability of Eudragit L100−ASD was 245% after oral administration in rats, and Tmax was markedly delayed from 2.8 ± 0.4 h to 5.3 ± 2.7 h. Our findings indicate that enteric ASDs are an effective strategy to increase the intestinal absorption of nintedanib by improving physiologically generated supersaturation and subsequent crystallization.

Hepatotoxicity of the Major Anthraquinones Derived From Polygoni Multiflori Radix Based on Bile Acid Homeostasis.

Polygoni Multiflori Radix (PMR), the dried root of Polygonum Multiflorum Thunb., has been widely used as traditional Chinese medicines in clinical practice for centuries. However, the frequently reported hepatotoxic adverse effects hindered its safe use in clinical practice. This study aims to explore the hepatotoxic effect of PMR extract and the major PMR derived anthraquinones including emodin, chrysophanol, and physcion in mice and the underlying mechanisms based on bile acid homeostasis. After consecutively treating the ICR mice with PMR extract or individual anthraquinones for 14 or 28 days, the liver function was evaluated by measuring serum enzymes levels and liver histological examination. The compositions of bile acids (BAs) in the bile, liver, and plasma were measured by LC-MS/MS, followed by Principal Component Analysis (PCA) and Partial Least Squares Discriminate Analysis (PLS-DA). Additionally, gene and protein expressions of BA efflux transporters, bile salt export pump (Bsep) and multidrug resistance-associated protein 2 (Mrp2), were examined to investigate the underlying mechanisms. After 14-day administration, mild inflammatory cell infiltration in the liver was observed in the physcion- and PMR-treated groups, while it was found in all the treated groups after 28-day treatment. Physcion and PMR extract induced hepatic BA accumulation after 14-day treatment, but such accumulation was attenuated after 28-day treatment. Based on the PLS-DA results, physcion- and PMR-treated groups were partially overlapping and both groups showed a clear separation with the control group in the mouse liver. The expression of Bsep and Mrp2 in the physcion- and PMR-treated mouse liver was decreased after 14-day treatment, while the downregulation was abrogated after 28-day treatment. Our study, for the first time, demonstrated that both PMR extract and tested anthraquinones could alter the disposition of either the total or individual BAs in the mouse bile, liver, and plasma via regulating the BA efflux transporters and induce liver injury, which provide a theoretical basis for the quality control and safe use of PMR in practice.

Simultaneous determination of abiraterone and its five metabolites in human plasma by LC-MS/MS: Application to pharmacokinetic study in healthy Chinese subjects.

In this study, a rapid, simple and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated to simultaneously quantify abiraterone (ABI), a widely used anti-metastatic castration-resistant prostate cancer drug, and its metabolites comprising Δ4-abiraterone (D4A), 3-keto-5α-abiraterone (5αA), abiraterone N-oxide (A-NO), abiraterone sulfate (A-Sul) and abiraterone N-oxide sulfate (A-NO-Sul) in human plasma. The analytes were extracted by protein precipitation with acetonitrile and ideal chromatographic separation was achieved on ACE-C18 column (2.1 × 50 mm, 5 µm) using a gradient elution. Triple Quad™ 6500+ mass spectrometer equipped with an electrospray ionization (ESI) source was used and the multiple reaction mode (MRM) was performed. In terms of method validation, good linearity was observed in preassigned validated concentration range for each analyte of interest. Both intra- and inter-batch accuracy was within the range of 87.6-113.8% for all analytes, while intra- and inter-batch precision was below 14.0%. Additionally, both low matrix effects and high recovery were obtained. All analytes remained stable in human plasma at room temperature for 4 h, on wet ice for 8 h, at - 80 °C for 42 d, over three freeze-thaw cycles and under auto-sampler temperature (4 °C) for 48 h post sample preparation. Subsequently, the validated LC-MS/MS method was applied for pharmacokinetic study in healthy Chinese volunteers following an oral dose of 250 mg abiraterone acetate tablet under fasted conditions. Our study for the first time reported the pharmacokinetic parameters of the ABI metabolites in Chinese subjects.

Au3+ -Functionalized UiO-67 Metal-Organic Framework Nanoparticles: O2•- and •OH Generating Nanozymes and Their Antibacterial Functions.

The synthesis and characterization of Au3+ -modified UiO-67 metal-organic framework nanoparticles, Au3+ -NMOFs, are described. The Au3+ -NMOFs reveal dual oxidase-like and peroxidase-like activities and act as an active catalyst for the catalyzed generation of O2•- under aerobic conditions or •OH in the presence of H2 O2 . The two reactive oxygen species (ROS) agents O2•- and •OH are cooperatively formed by Au3+ -NMOFs under aerobic conditions, and in the presence of H2 O2. The Au3+ -NMOFs are applied as an effective catalyst for the generation ROS agents for antibacterial and wound healing applications. Effective antibacterial cell death and inhibition of cell proliferation of Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) bacterial colonies are demonstrated in the presence of the Au3+ -NMOFs. In addition, in vivo experiments demonstrate effective wound healing of mice wounds infected by S. aureus, treated by the Au3+ -NMOFs.

mPEG2k-PCLx Polymeric Micelles Influence Pharmacokinetics and Hypoglycemic Efficacy of Metformin through Inhibition of Organic Cation Transporters in Rats.

Increasing evidence has shown that nanocarriers have effects on several efflux drug transporters. To date, little is known about whether influx transporters are also modulated. Herein, we investigated the impact of amphiphilic polymer micelles on the uptake function of organic cation transporters (OCTs) and the influence on the pharmacokinetics and pharmacodynamics of metformin, a well-characterized substrate of OCTs. Five types of polymeric micelles (mPEG2k-PCL2k, mPEG2k-PCL3.5k, mPEG2k-PCL5k, mPEG2k-PCL7.5k, and mPEG2k-PCL10k) were prepared to evaluate the inhibition of hOCT1-3-overexpressing Madin-Darby canine kidney cells. The mPEG2k-PCLx micelles played an inhibitory role above the critical micelle concentration. The inhibitory potency could be ranked as mPEG2k-PCL2k > mPEG2k-PCL3.5k > mPEG2k-PCL5k > mPEG2k-PCL7.5k > mPEG2k-PCL10k, which negatively declined with the increase of molecular weight of the hydrophobic segment. The inhibitory effects of polymeric micelles on the hOCT1 isoform were the most pronounced, with the lowest IC50 values ranging from 0.106 to 0.280 mg/mL. The mPEG2k-PCL2k micelles distinctly increased the plasma concentration of metformin and significantly decreased Vss by 35.6% (p < 0.05) after seven consecutive treatments in rats, which was interrelated with the restrained metformin distribution in the liver and kidney. The uptake inhibition of micelles on hepatic and renal rOcts also diminished the glucose-lowering effect of metformin and fasting insulin levels in the oral glucose tolerance test. Consistent with the inhibitory effects, the mRNA and protein levels of rOct1 and rOct2 were decreased in the liver, kidney, and small intestine. The present study demonstrated that mPEG2k-PCLx micelles could inhibit the transport function of OCTs, indicating a potential risk of drug-drug interactions during concomitant medication of nanomedicine with organic cationic drugs.

Enhanced therapeutic efficacy of a novel colon-specific nanosystem loading emodin on DSS-induced experimental colitis.

BACKGROUND: Ulcerative colitis (UC) is an intricate enteric disease with a rising incidence that is closely related to mucosa-barrier destruction, gut dysbacteriosis, and immune disorders. Emodin (1,3,8-trihydroxy-6-methyl-9,10-anthraquinone, EMO) is a natural anthraquinone derivative that occurs in many Polygonaceae plants. Its multiple pharmacological effects, including antioxidant, immune-suppressive, and anti-bacteria activities, make it a promising treatment option for UC. However, its poor solubility, extensive absorption, and metabolism in the upper gastrointestinal tract may compromise its anti-colitis effects. PURPOSE: EMO was loaded in a colon-targeted delivery system using multifunctional biomedical materials and the enhanced anti-colitis effect involving mucosa reconstruction was investigated in this study. METHODS: EMO-loaded Poly (DL-lactide-co-glycolide)/EudragitⓇ S100/montmorillonite nanoparticles (EMO/PSM NPs) were prepared by a versatile single-step assembly approach. The colon-specific release behavior was characterized in vitro and in vivo, and the anti-colitis effect was evaluated in dextran sulfate sodium (DSS)-induced acute colitis in mice by weight loss, disease activity index (DAI) score, colon length, histological changes, and colitis biomarkers. The integrity of the intestinal mucosal barrier was evaluated through transwell co-culture model in vitro and serum zonulin-related tight junctions and mucin2 (MUC2) in vivo. RESULTS: EMO/PSM NPs with a desirable hydrodynamic diameter (~ 235 nm) and negative zeta potential (~ -31 mV) could prevent the premature drug release (< 4% in the first 6 h in vitro) in the upper gastrointestinal tract (GIT) and boost retention in the lower GIT and inflamed colon mucosa in vivo. Compared to free EMO-treatment of different doses in UC mice, the NPs could enhance the remedial efficacy of EMO in DAI decline, histological remission, and regulation of colitis indicators, such as myeloperoxidase (MPO), nitric oxide (NO), and glutathione (GSH). The inflammatory factors including induced nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), TNF-α, and IL-1ß were suppressed by EMO/PSM NPs at both mRNA and protein levels. The obtained NPs could also promote the regeneration of the mucosal barrier via reduced fluorescein isothiocyanate (FITC)-dextran leakage in the transwell co-culture model and decreased serum zonulin levels, which was demonstrated to be associated with the upregulated tight junctions (TJs)-related proteins (claudin-2, occludin, and zo-1) and MUC2 at mRNA level. Moreover, the NPs could contribute to attenuating the liver injury caused by free EMO under excessive immune inflammation. CONCLUSION: Our results demonstrated that EMO/PSM NPs could specifically release EMO in the diseased colon, and effectively enhance the anti-colitis effects of EMO related to intestinal barrier improvement. It can be considered as a novel potential alternative for oral colon-targeted UC therapy by increasing therapeutic efficacy and reducing side-effects.

Erratum to: Kinetic Characterization of Tyrosinase-catalyzed Oxidation of Four Polyphenols.

The article "Kinetic Characterization of Tyrosinase-catalyzed Oxidation of Four Polyphenols", written by Wan-yu LIU, Congming ZOU, Jian-hua HU, Zi-jun XU, Lu-qin SI, Jun-jun LIU, Jian-geng HUANG, was originally published electronically on the publisher's internet portal on May 2020 without open access. With the author(s)' decision to opt for Open Choice, the copyright of the article is changed to © The Author(s) 2020 and the article is forthwith distributed under a Creative Commons Attribution 4.0 International License ( https://creativecommons.org/licenses/by/4.0/ ), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.The original article has been corrected.

Kinetic Characterization of Tyrosinase-catalyzed Oxidation of Four Polyphenols.

Phenolic compounds such as chlorogenic acid, cryptochlorogenic acid, neochlorogenic acid and caffeic acid are widely distributed in fruits, vegetables and traditional Chinese medicines with a wide range of biological activities. Tyrosinase plays a critical role in the food industry, but recent studies have proposed unexplored aspects of clinical application. Tyrosinase-catalyzed oxidation of four polyphenols as well as its underlying mechanism remains unclear. In the current work, we investigated the kinetic properties of tyrosinase-catalyzed oxidation of the four polyphenols of interest. To measure the unstable o-quinone products, an analytical method using 3-methyl-2-benzothiazolinone hydrazone (MBTH) was established. The optimal incubation time, buffer pH, temperature and enzyme concentration for the enzyme activity in the presence of each polyphenol of interest were investigated. Under the final optimized conditions, the kinetics and substrate specificity of four polyphenols were examined. Kinetic data showed that tyrosinase had the greatest substrate affnity to chlorogenic acid compared with its isomers and caffeic acid. The catalytic effciency with chlorogenic acid was 8- to 15-fold higher than that with the other 3 polyphenols. Molecular docking study demonstrated that the tight binding of chlorogenic acid at the peripheral site should be the major reason for the specifcity to chlorogenic acid. In light of this, the rational design of high-affnity inhibitors against tyrosinase may focus on the binding of both the Cu site and peripheral site. This study will supply a basis for the selection of phenolic acids in food industry and health care.

Development, validation and comparison of three LC-MS/MS methods for determination of endogenous striatal oleoyl ethanolamine in mice.

Obesity has become a severe public health problem worldwide. An endogenous fatty acid ethanolamine oleoyl ethanolamine (OEA) is reported to be capable of reducing body weight and food intake by increasing striatal extracellular dopamine concentration. However, association between obesity and striatal OEA level remains unknown. As such, it is necessary to develop a sensitive and reliable method to quantitate OEA concentration in striatum. Because true endogenous analytes free blank matrix is not available, surrogate analyte, surrogate matrix and background subtraction methods are often employed for the analysis of endogenous compounds. In this study, three liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods were developed and validated for the determination of OEA concentration in mouse brain homogenate. Interestingly, stability results found that OEA-d4 degraded in brain homogenate under room temperature, while OEA level remarkably increased with time. Since lowering temperature could observably decelerate the endogenous transformation of OEA, sample collection and preparation were carried out under ice-bath condition. Hexane: isopropanol (9:1, v/v) was employed as an extractant for liquid-liquid extraction. After method validation, three methods were applied to quantify OEA in striatum homogenate from C57B6/L mice following normal and high fat diet feeding for 4 months. Results from three methods all showed the striatal OEA level in obesity group was significantly higher than control group and obesity-resist group, which indicated that obesity might be associated with elevated striatal OEA level.

Pharmacokinetics and Novel Metabolite Identification of Tartary Buckwheat Extracts in Beagle Dogs Following Co-Administration with Ethanol.

Alcoholic liver disease (ALD) has become a critical global public health issue worldwide. Tartary buckwheat extracts exhibit potential therapeutic effects against ALD due to its antioxidant and anti-inflammatory activities. However, in vivo pharmacokinetics and metabolite identification of tartary buckwheat extracts have not been clearly elucidated. Accordingly, the current manuscript aimed to investigate pharmacokinetics and to identify novel metabolites in beagle dogs following oral co-administration of tartary buckwheat extracts and ethanol. To support pharmacokinetic study, a simple LC-MS/MS method was developed and validated for simultaneous determination of quercetin and kaempferol in beagle dog plasma. The conjugated forms of both analytes were hydrolyzed by ß-glucuronidase and sulfatase followed by liquid-liquid extraction using methyl tert-butyl ether. In addition, another effective approach was established using advanced ultrafast liquid chromatography coupled with a Q-Exactive hybrid quadrupole orbitrap high resolution mass spectrometer to identify the metabolites in beagle dog biological samples including urine, feces, and plasma. The pharmacokinetic study demonstrated that the absolute oral bioavailability for quercetin and kaempferol was determined to be 4.6% and 1.6%, respectively. Oral bioavailability of quercetin and kaempferol was limited in dogs probably due to poor absorption, significant first pass effect, and biliary elimination, etc. Using high resolution mass spectrometric analysis, a total of nine novel metabolites were identified for the first time and metabolic pathways included methylation, glucuronidation, and sulfation. In vivo pharmacokinetics and metabolite identification results provided preclinical support of co-administration of tartary buckwheat extracts and ethanol in humans.

A PepT1 mediated medicinal nano-system for targeted delivery of cyclosporine A to alleviate acute severe ulcerative colitis.

To effectively alleviate acute severe ulcerative colitis (ASUC), we developed a colon-specific delivery system-PLGA-KPV/MMT/CS multifunctional medicinal nanoparticles loaded with cyclosporine A (CyA). The lysine-proline-valine (KPV) tripeptide, which possesses anti-inflammatory properties and high affinity to peptide transporter 1 (PepT1), can target therapy-related cells (colonic epithelial cells and macrophages) via overexpression of PepT1. Montmorillonite (MMT)/chitosan (CS) coating can reduce CyA leakage in the upper gastrointestinal tract (GIT) and enhance nanoparticle adhesion to the inflamed colon. The bio-distribution demonstrated that nanoparticles can specifically accumulate in the inflamed tissues and can be retained for up to 36 h. After being treated with the CyA-PLGA-KPV/MMT/CS nanoparticles (PKMCN), the mice with DSS-induced ulcerative colitis exhibited significant improvements in body weight, colon length, and disease activity index. Moreover, biochemistry and immunohistochemical analysis showed that the PKMCN treatment group performed as well as the healthy group. Intriguingly, PKMCN without CyA also presented marked therapeutic effects. Our results suggested that PKMCN could be a promising drug delivery system for ASUC therapy by targeting inflamed cells, prolonging curative time, and mitigating colitis.

The sub-chronic impact of mPEG2k-PCLx polymeric nanocarriers on cytochrome P450 enzymes after intravenous administration in rats.

Recent studies indicated obvious impacts of nano-carriers on cytochrome P450 enzymes (CYP450s) in vitro, but the effects in vivo are still unknown. In the present research, mPEG2k-PCLx micelles with different length of hydrophobic block (2000-10,000 Da) were intravenously administrated into rats for 14 days to evaluate the sub-chronic influences in the metabolic function of hepatic CYP450s. Although CYP1A1/B2 was susceptible to mPEG2k-PCLx micelles compared with other CYP isoenzymes, induction was mainly observed and varied with micelle type, administration dose, and CYP isoform. Interestingly, mPEG2k-PCL3.5k micelles at 5 mg/kg increased the activity of CYP1A2, CYP2B1, CYP2C6, CYP2C11, and CYP3A1/2 while mPEG2k-PCL5k micelles only induced the latter three enzymes at 75 mg/kg. The mRNA expression of corresponding CYPs was mostly up-regulated by mPEG2k-PCL3.5k micelles whilst less effect in protein level except for CYP3A1/2. Moreover, mPEG2k-PCL3.5k micelles could affect the pharmacokinetic properties of phenacetin (CYP1A2), tolbutamide (CYP2C6), omeprazole (CYP2C11), and midazolam (CYP3A1/2) with a decrease of 19.6% in Cmax, 20.5% in AUC0-t, 31.6% in AUC0-t, and 40.1% in Cmax at 5 mg/kg, respectively (P < 0.05 or P < 0.01). These results unveiled nano-DDS might be involved in nanocarrier-drug interaction by intervening in the activity of CYP450s.

Simultaneous Quantitation of a Novel α1/ß1-Blocker TJ0711 and Its Two Metabolites in Dog Plasma Using LC-MS/MS and Its Application to a Pharmacokinetic Study after Intravenous Infusion.

TJ0711∙HCl, which is a novel α1/ß1 adrenoceptor blocking agent with a ratio of 1:1 for α1/ß1, is designed to treat and prevent perioperative hypertension. M1 and M3 were identified as important metabolites in vitro for either antihypertension activity or the major metabolite production. In order to obtain a pharmacokinetic profile of both TJ0711 and its metabolites, a rapid, selective, and reliable LC-MS/MS method was developed and validated for simultaneous determination of TJ0711 and two metabolites in beagle dog plasma via efficiently separating two interferential metabolites M16 and M4 from M1 and M3, respectively. Chromatographic separation was achieved on a Waters CORTECS C18⁺ column (2.1 × 100 mm, 2.7 µm). The mass spectrometric detection was carried out in positive ion MRM mode with ESI⁺ source. Protein precipitation was used in sample preparation and provided good recovery without a matrix effect. Good linearity was observed at the ranges of 0.5⁻100 ng/mL for TJ0711 and M3, 0.1⁻20 ng/mL for M1. Additional validation results were within the acceptance limits followed U.S. FDA guidelines for bioanalytical method validation. This method was successfully applied to an intravenous infusion pharmacokinetic study of TJ0711 at dosing rates of 3, 6, and 12 µg/kg/min in anesthetized beagle dogs for the first time. TJ0711 and its two metabolites exhibited effective proportionality in the dosage of 3 to 12 µg/kg/min. Neither TJ0711 nor its metabolites showed significant differences in pharmacokinetic parameters such as t1/2, CL, and Vss among three dose groups.

In vitro effect of mPEG2k-PCLx micelles on rat liver cytochrome P450 enzymes.

The present study was aimed to evaluate the effects of amphiphilic copolymer micelles on six major hepatic cytochrome P450 (CYP) isoforms. A series of mPEG2k-PCLx polymeric micelles (mPEG2k-PCL2k, mPEG2k-PCL3.5k, mPEG2k-PCL5k and mPEG2k-PCL10k) ranging from 20 to 100 nm were prepared to investigate the inhibitory or inductive activities by in vitro incubations of rat liver microsomes and primary rat hepatocytes. Inhibition of these polymeric micelles on CYP1A2, CYP2B1, CYP2C6, CYP2C11, CYP2D2 and CYP3A1/2 isoenzymes were observed above their critical micelle concentrations (>10 µg·mL-1) and in a concentration-dependent manner. The mPEG2k-PCL2k micelles showed the strongest inhibition of CYP1A2, followed by CYP2C11. The micelles with lower molecular weight PCL segment exhibited more potent inhibitory potential. Induction on CYP1A2, CYP2B1 and CYP3A1/2 activity (2.1-7.2-fold, 1.5-2.4-fold and 1.3-3.0-fold, respectively) were detected at all tested concentrations (0.1-1000 µg·mL-1 or 0.1-100 µg·mL-1). Accordingly, most of the mRNA levels were upregulated. As demonstrated in ex vivo fluorescence imaging results, the mPEG2k-PCLx micelles mainly accumulated in the liver after intravenous administration. In conclusion, mPEG2k-PCLx micelles can interfere with the normal metabolic function of CYP450s in vitro, indicating polymeric micelles as promising drug nano-carriers might cause micelle-drug interaction and the in vivo interaction deserves further investigation.

Oral Bioavailability of Kinsenoside in Beagle Dogs Measured by LC-MS/MS: Improvement of Ex Vivo Stability of a Lactone-Containing Compound.

Kinsenoside (KD), an active compound isolated from Anoectochilus roxburghii, has demonstrated multiple pharmacological activities including hepatoprotection, antihyperliposis, antihyperglycemia, and antiosteoporosis. To the best of our knowledge, there are no available data concerning its preclinical pharmacokinetics and bioavailability in beagle dogs. To support preclinical pharmacokinetic and bioavailability study, a reliable LC-MS/MS method was developed for KD concentration measurements in beagle dog plasma. The chromatographic separation was achieved on a Waters Atlantis® Hilic Silica column with an optimum mobile phase consisting of 5 mM ammonium acetate in water (pH 3.0 adjusted with acetic acid) and acetonitrile at a flow rate of 0.2 mL/min. Mass spectrometric analyses were carried out by monitoring multiple reaction monitoring transitions at m/z 265.2→102.9 for KD and m/z 174.0→128.0 for l-phenyl-d5-alanine-2,3,3-d3 (IS). The stability of KD in beagle dog whole blood and plasma was systematically evaluated. Lowering the temperature played a more critical role in stabilizing KD than decreasing the pH and adding esterase inhibitors, indicating that the major reason for instability of KD was probably due to chemical hydrolysis rather than esterase-mediated degradation. The currently developed method was validated and applied to a pharmacokinetic and bioavailability study of KD in beagle dogs following oral administration at a dose of 3 mg/kg. The absolute oral bioavailability for KD was determined to be 27.6%. Compared with typical glycosides, KD has a better bioavailability and is suitable for developing an oral dosage form.

Simultaneous LC-MS/MS bioanalysis of etoposide and paclitaxel in mouse tissues and plasma after oral administration of self-microemulsifying drug-delivery systems.

In this study, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated to simultaneously determine the anticancer drugs etoposide and paclitaxel in mouse plasma and tissues including liver, kidney, lung, heart, spleen and brain. The analytes were extracted from the matrices of interest by liquid-liquid extraction using methyl tert-butyl ether-dichloromethane (1:1, v/v). Chromatographic separation was achieved on an Ultimate XB-C18 column (100 × 2.1 mm, 3 µm) at 40°C and the total run time was 4 min under a gradient elution. Ionization was conducted using electrospray ionization in the positive mode. Stable isotope etoposide-d3 and docetaxel were used as the internal standards. The lower limit of quantitation (LLOQ) of etoposide was 1 ng/g tissue for all tissues and 0.5 ng/mL for plasma. The LLOQ of paclitaxel was 0.4 ng/g tissue and 0.2 ng/mL for all tissues and plasma, respectively. The coefficients of correlation for all of the analytes in the tissues and plasma were >0.99. Both intra- and inter-day accuracy and precision were satisfactory. This method was successfully applied to measure plasma and tissue drug concentrations in mice treated with etoposide and paclitaxel-loaded self-microemulsifying drug-delivery systems.