The Roles of Neutrophil-Derived Myeloperoxidase (MPO) in Diseases: The New Progress.

Myeloperoxidase (MPO) is a heme-containing peroxidase, mainly expressed in neutrophils and, to a lesser extent, in monocytes. MPO is known to have a broad bactericidal ability via catalyzing the reaction of Cl- with H2O2 to produce a strong oxidant, hypochlorous acid (HOCl). However, the overproduction of MPO-derived oxidants has drawn attention to its detrimental role, especially in diseases characterized by acute or chronic inflammation. Broadly speaking, MPO and its derived oxidants are involved in the pathological processes of diseases mainly through the oxidation of biomolecules, which promotes inflammation and oxidative stress. Meanwhile, some researchers found that MPO deficiency or using MPO inhibitors could attenuate inflammation and tissue injuries. Taken together, MPO might be a promising target for both prognostic and therapeutic interventions. Therefore, understanding the role of MPO in the progress of various diseases is of great value. This review provides a comprehensive analysis of the diverse roles of MPO in the progression of several diseases, including cardiovascular diseases (CVDs), neurodegenerative diseases, cancers, renal diseases, and lung diseases (including COVID-19). This information serves as a valuable reference for subsequent mechanistic research and drug development.

The anti-hyperlipidemic effects of Poria cocos (Schw.) Wolf extract: Modulating cholesterol homeostasis in hepatocytes via PPARα pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Poria cocos (Schw.) Wolf (Polyporaceae, P.cocos), which is born on the pine root, has a history of more than two thousand years of medicine in China. P.cocos was first recorded in the Shennong's Herbal Classic, studies have proved its lipid-lowering effect. AIM OF STUDY: The aim of study was to investigate the underlying mechanism of P.cocos extract on hyperlipidemia. MATERIALS AND METHODS: Male Sprague-Dawley (SD) rats aged 9-12 weeks were intraperitoneally (IP) injected with Triton-WR 1339 to establish an acute hyperlipidemia model. At 0 h and 20 h after the model was established, low and high doses of P.cocos extract or simvastatin were given twice. After 48 h, the rats were sacrificed, and liver and serum samples were collected for analysis. The cell model was constructed by treating L02 cells with 1% fat emulsion-10% FBS-RPMI 1640 medium for 48 h. At the same time, low and high doses of P.cocos extract and simvastatin were administered. Oil red O staining was used to evaluate the lipid accumulation in the cells, and H&E staining was used to evaluate the liver lesions of rats. Real-time quantitative PCR and western blotting were used to detect the expressions of lipid metabolism-related genes. RESULTS: P.cocos extract relieved lipid accumulation in vitro and alleviated hyperlipidemia in vivo. Both gene and protein expressions of peroxisome proliferator-activated receptor α (PPARα) were shown to be up-regulated by P.cocos extract. Additionally, P.cocos extract down-regulated the expressions of fatty acid synthesis-related genes sterol regulatory element-binding protein-1 (SREBP-1), Acetyl-CoA Carboxylase 1 (ACC1) and fatty acid synthase (FAS), while up-regulated the expressions of cholesterol metabolism-related genes liver X receptor-α (LXRα), ATP-binding cassette transporter A1 (ABCA1), cholesterol 7alpha-hydroxylase (CYP7A1) and low density lipoprotein receptor (LDLR), which were reversed by the treatment with the PPARα inhibitor GW6471. CONCLUSION: P.cocos extract ameliorates hyperlipidemia and lipid accumulation by regulating cholesterol homeostasis in hepatocytes through PPARα pathway. This study provides evidence that supplementation with P.cocos extract could be a potential strategy for the treatment of hyperlipidemia.

Development and validation of a chiral UPLC-MS/MS method for quantifying S-Oxiracetam and R-Oxiracetam in human plasma, urine and feces: Application to a phase-I clinical pharmacokinetic study.

A chiral UPLC-MS/MS method was developed and validated to determine oxiracetam enantiomers in human plasma, urine, and feces. The R-Oxiracetam and S-Oxiracetam were quantified using a CHIRALPAK ®AD3 column at 25 â„ƒ, and the resolution was greater than 3.2. The S-Oxiracetam is the eutomer that isresponsible for the treatment of various brain damage. Isocratic elution was conducted at a flow rate of 0.9 mL/min for 6 min using the mixture of methanol and acetonitrile (methanol:acetonitrile, 15:85) containing 0.3‰ formic acid. The methods showed linearity at the range of 0.5-100 µg/mL for each oxiracetam enantiomer. A comprehensive validation process was carried out, covering aspects including linearity, selectivity, carryover, accuracy, precision, interferences, matrix effect, recovery, dilution integrity and stability in matrix and solution. The validated methods were successfully applied to quantifying R-Oxiracetam and S-Oxiracetam in human plasma, urine, and feces of 12 healthy subjects treated with either a single dose of 2 g S-Oxiracetam injection or 4 g Oxiracetam injection in a phase-I clinical trial. There was no significant difference for plasma pharmacokinetic parameters of S-Oxiracetam between the two regimens (P＞0.05). The S-Oxiracetam and Oxiracetam were primarily eliminated through urine in their original form, with cumulative excretion rates of 92.16% and 85.92%, respectively, within 24 h after administration. Enantiomers interconversion was not observed in the plasma, urine, or feces. The results of this study suggest that replacing 4 g Oxiracetam injection with 2 g S-Oxiracetam injection could offer clinical benefits by lowering the dosage and mitigating potential risks, based on the pharmacokinetic characteristics.

Overexpression of Glutathione S-Transferases in Human Diseases: Drug Targets and Therapeutic Implications.

Glutathione S-transferases (GSTs) are a major class of phase II metabolic enzymes. Besides their essential role in detoxification, GSTs also exert diverse biological activities in the occurrence and development of various diseases. In the past few decades, much research interest has been paid to exploring the mechanisms of GST overexpression in tumor drug resistance. Correspondingly, many GST inhibitors have been developed and applied, solely or in combination with chemotherapeutic drugs, for the treatment of multi-drug resistant tumors. Moreover, novel roles of GSTs in other diseases, such as pulmonary fibrosis and neurodegenerative diseases, have been recognized in recent years, although the exact regulatory mechanisms remain to be elucidated. This review, firstly summarizes the roles of GSTs and their overexpression in the above-mentioned diseases with emphasis on the modulation of cell signaling pathways and protein functions. Secondly, specific GST inhibitors currently in pre-clinical development and in clinical stages are inventoried. Lastly, applications of GST inhibitors in targeting cell signaling pathways and intracellular biological processes are discussed, and the potential for disease treatment is prospected. Taken together, this review is expected to provide new insights into the interconnection between GST overexpression and human diseases, which may assist future drug discovery targeting GSTs.

Treatment effects of laparoscopy versus laparotomy on heterotopic pregnancy after in vitro fertilization and embryo transfer.

OBJECTIVE: To compare the treatment effects of laparoscopy versus laparotomy on heterotopic pregnancy (HP) after in vitro fertilization-embryo transfer (IVF-ET). METHODS: The retrospective case-control study enrolled 109 patients diagnosed with HP after IVF-ET treatment in our hospital from January 2009 to March 2020. All patients received surgical treatment by either laparoscopy or laparotomy. Data for general characteristics, diagnostic features, surgical parameters, as well as perinatal and neonatal outcomes were collected. RESULTS: Sixty-two patients received laparoscopy and 47 received laparotomy. Significantly lower percentage of large hemoperitoneum (P = 0.001), shorter surgery duration (P < 0.001), less intraoperative blood loss (P = 0.001), higher rates of general anesthesia (P < 0.001), and lower cesarean section rates for singletons (P = 0.003) were found in the laparoscopy group. The perinatal and neonatal outcomes were comparable between the two groups. When interstitial pregnancy was considered alone, the surgical blood loss was significantly reduced in the laparoscopy group (P = 0.021), but there was no significant difference in hemoperitoneum, surgery duration, or perinatal and neonatal outcomes in singletons. CONCLUSION: Both laparoscopy and laparotomy are effective surgical treatments for HP after IVF-ET. Laparoscopy is minimally invasive but laparotomy can be an alternative in emergency situations.

Mechanistic Study of Icaritin-Induced Inactivation of Cytochrome P450 2C9.

Icaritin (ICT) is a prenylflavonoid derivative that has been approved by National Medical Products Administration for the treatment of hepatocellular carcinoma. This study aims to evaluate the potential inhibitory effect of ICT against cytochrome P450 (CYP) enzymes and to elucidate the inactivation mechanisms. Results showed that ICT inactivated CYP2C9 in a time-, concentration-, and NADPH-dependent manner with Ki = 1.896 µM, Kinact = 0.02298 minutes-1, and Kinact/Ki = 12 minutes-1 mM-1, whereas the activities of other CYP isozymes was minimally affected. Additionally, the presence of CYP2C9 competitive inhibitor, sulfaphenazole, superoxide dismutase/catalase system, and GSH all protected CYP2C9 from ICT-induced activity loss. Moreover, the activity loss was neither recovered by washing the ICT-CYP2C9 preincubation mixture nor the addition of potassium ferricyanide. These results, collectively, implied the underlying inactivation mechanism involved the covalent binding of ICT to the apoprotein and/or the prosthetic heme of CYP2C9. Furthermore, an ICT-quinone methide (QM)-derived GSH adduct was identified, and human glutathione S-transferases (GST) isozymes GSTA1-1, GSTM1-1, and GSTP1-1 were shown to be substantially involved in the detoxification of ICT-QM. Interestingly, our systematic molecular modeling work predicted that ICT-QM was covalently bound to C216, a cysteine residue located in the F-G loop downstream of substrate recognition site (SRS) 2 in CYP2C9. The sequential molecular dynamics simulation confirmed the binding to C216 induced a conformational change in the active catalytic center of CYP2C9. Lastly, the potential risks of clinical drug-drug interactions triggered by ICT as a perpetrator were extrapolated. In summary, this work confirmed that ICT was an inactivator of CYP2C9. SIGNIFICANCE STATEMENT: This study is the first to report the time-dependent inhibition of CYP2C9 by icaritin (ICT) and the intrinsic molecular mechanism behind it. Experimental data indicated that the inactivation was via irreversible covalent binding of ICT-quinone methide to CYP2C9, while molecular modeling analysis provided additional evidence by predicting C216 as the key binding site which influenced the structural confirmation of CYP2C9's catalytic center. These findings suggest the potential of drug-drug interactions when ICT is co-administered with CYP2C9 substrates clinically.

Pharmacokinetic study of Strongylocentrotus nudus egg polysaccharide in rats and beagles using a 3H-labeling method.

Strongylocentrotus nudus egg polysaccharide (SEP) extracted from sea urchins has potential anticancer activity. However, little is known about its pharmacokinetic properties. To investigate the pharmacokinetics of SEP, it was radiolabeled with tritium. Furthermore, a sensitive, selective, and rapid liquid scintillation counter (LSC) method for quantifying 3H-SEP in biological matrix was validated. The lower quantification limit of the method was 4 Bq. The relative standard deviations (RSDs) of the intra- and inter-day precision were <3.0% and <3.9%, respectively. 3H-SEP was successfully applied to investigate the pharmacokinetics of SEP after intravenous administration of 20, 40, and 80 mg/kg (40 µCi/kg) in rats and 5, 10, and 20 mg/kg (6 µCi/kg) in beagles. The AUC(0-t) of SEP at three different doses was 487.81 ± 39.99 mg/L*h, 1,003.10 ± 95.94 mg/L*h, and 2,188.84 ± 137.73 mg/L*h in rats and 144.12 ± 3.78 mg/L*h, 322.62 ± 28.03 mg/L*h, and 754.17 ± 37.79 mg/L*h in beagles. The terminal elimination half-life (t1/2) of SEP was longer in beagles (204.29 ± 139.34 h) than in rats (35.48 ± 6.04 h). The concentration of SEP in plasma declined rapidly in both rats and beagles. All the study results provide detailed pharmacokinetic profiles of SEP in two kinds of animals, which will be helpful for further development.

Development and validation of an LC-MS/MS method for simultaneous determination of SH-1028, an irreversible third-generation EGFR TKI, and two of its metabolites in human plasma: application in clinical pharmacokinetics.

SH-1028 is a novel, potent, and highly selective epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) developed for the treatment of T790M Mutation-positive non-small cell lung cancer (NSCLC). The objective was to develop an LC-MS/MS method for the simultaneous determination of SH-1028 and its metabolites, Imp2 and Imp3, in human plasma.The plasma samples were extracted through protein precipitation with acetonitrile on wet ice conditions. A rapid, sensitive, and specific method was developed and successfully applied to evaluate the pharmacokinetic (PK) properties of SH-1028 in patients with advanced NSCLC following single and multiple doses of SH-1028 (60 mg).After single-dose administration, the Cmax of SH-1028, Imp2, and Imp3 was 11.2, 50.2, and 7.99 ng/mL, respectively. The mean AUC0-24 h was 138, 602, and 76.7 h*ng/mL, respectively. And the terminal half-life time was 19.9, 14.4, and 26.1 h, respectively. After multiple-dose administration, SH-1028 exhibited a slight accumulation, with a mean accumulation ratio (RAUC) of 2.00.The study assessed the PK properties of SH-1028 following single and multiple doses in patients with advanced NSCLC and would provide meaningful information for the further development of SH-1028.

Glutathione S-Transferases Mediate In Vitro and In Vivo Inactivation of Genipin: Implications for an Underlying Detoxification Mechanism.

Genipin (GP), the reactive metabolite of geniposide (GE), is responsible for GE-induced hepatotoxicity. As a potential detoxification pathway, the inactivation of GP by glutathione S-transferases (GSTs) has not yet been characterized. In this study, the thiol-GSH conjugates of GP, M532-1 and M532-2 were first identified and the catalytic activities of GSTs were investigated both in vitro and in vivo. GSTA1-1 and GSTA4-4 showed high activity in the formation of both thiol-GSH conjugates, whereas GSTA4-4 specifically catalyzed M532-2 formation in vitro. The active GST isoforms protect against alkylation of N-acetylcysteine (NAC), a classic model nucleophile. GST inhibition attenuated M532-1 formation in rat bile, confirming the in vivo catalytic role of GSTs. In conclusion, this study demonstrated the inactivation of GP by GSTs and implied that interindividual variability of GSTs may be a risk factor for susceptibility to GE-induced hepatotoxicity.

Subacute oral toxicology and toxicokinetics of pterostilbene, a novel Top1/Tdp1 inhibiting anti-tumor reagent.

This study evaluated the subacute toxicity and toxicokinetics of a potential anti-cancer drug candidate, pterostilbene, in rats. Animals were orally administered at two repeated doses of 200 and 500 mg/kg for 28 days. No mortality was observed during the 28 days of continuous administration of pterostilbene. Body weight and food consumption in each group increased steadily, while no significant difference was found. Liver weight in the 500 mg/kg female, but not male group increased with mild cytoplasmic vacuoles observed in histopathological study. Toxicokinetics was assessed by measuring plasma concentrations of pterostilbene on the first and 28th day of administration using UPLC-MS/MS. Toxicokinetic parameters showed that AUC0-t significantly increased in all animals, while the increase in females was greater than males. System exposure of pterostilbene appeared to be linear within the administrated dose range. In conclusion, our findings suggested a minimal subacute toxicity profile of pterostilbene, which could strongly support further development of this compound as a novel anti-cancer agent.

Phase I Metabolism of Pterostilbene, a Dietary Resveratrol Derivative: Metabolite Identification, Species Differences, Isozyme Contribution, and Further Bioactivation.

Pterostilbene (PTE), a dietary derivative of resveratrol, displayed pleiotropic health-promoting activities. This study aimed to explore the metabolic profiles and species differences of the phase I metabolism of PTE and to investigate subsequent detoxification after PTE bioactivation. PTE was found to be biotransformed to two pharmacologically active metabolites, pinostilbene and 3'-hydroxypterostilbene, in vivo and in vitro with substantial species differences. Human CYP1A2 was proved to be mainly responsible for the demethylation and 3'-hydroxylation of PTE, with its contribution to a demethylation of 94.5% and to a 3'-hydroxylation of 97.9%. An in vitro glutathione trapping experiment revealed the presence of an ortho-quinone intermediate formed by further oxidation of 3'-hydroxypterostilbene. Human glutathione S-transferase isoforms A2, T1, and A1 inactivated the ortho-quinone intermediate by catalyzing glutathione conjugation, implicating a potential protective pathway against PTE bioactivation-derived toxicity. Overall, this study provided a comprehensive view of PTE phase I metabolism and facilitated its further development as a promising nutraceutical.

Effects of ginkgo leaf tablet on the pharmacokinetics of rosiglitazone in rats and its potential mechanism.

CONTEXT: Ginkgo leaf tablet (GLT), a traditional Chinese herbal formula, is often combined with rosiglitazone (ROS) for type 2 diabetes mellitus treatment. However, the drug-drug interaction between GLT and ROS remains unknown. OBJECTIVE: To investigate the effects of GLT on the pharmacokinetics of ROS and its potential mechanism. MATERIALS AND METHODS: The pharmacokinetics of 10 mg/kg ROS with 100/200 mg/kg GLT as single-dose and 10-day multiple-dose administration were investigated in Sprague-Dawley rats. In vitro, the effects of GLT on the activity of CYP2C8 and CYP2C9 were determined in recombinant human yeast microsomes and rat liver microsomes with probe substrates. RESULTS: The t1/2 of ROS increased from 2.14 ± 0.38 (control) to 2.79 ± 0.37 (100 mg/kg) and 3.26 ± 1.08 h (200 mg/kg) in the single-dose GLT administration. The AUC0-t (139.69 ± 45.46 vs. 84.58 ± 39.87 vs. 66.60 ± 15.90 h·µg/mL) and t1/2 (2.75 ± 0.70 vs. 1.99 ± 0.44 vs. 1.68 ± 0.35 h) decreased significantly after multiple-dose GLT treatment. The IC50 values of quercetin, kaempferol, and isorhamnetin, GLT main constituents, were 9.32, 7.67, and 11.90 µmol/L for CYP2C8, and 27.31, 7.57, and 4.59 µmol/L for CYP2C9. The multiple-dose GLT increased rat CYP2C8 activity by 44% and 88%, respectively. DISCUSSION AND CONCLUSIONS: The metabolism of ROS is attenuated in the single dose of GLT by inhibiting CYP2C8 and CYP2C9 activity, and accelerated after the multiple-dose GLT treatment via inducing CYP2C8 activity in rats, indicating that the clinical dose of ROS should be adjusted when co-administrated with GLT.

The Effects of Immediate vs Gradual Reduction in Nicotine Content of Cigarettes on Smoking Behavior: An Ecological Momentary Assessment Study.

Background: In recent years, much research has examined the effects of various interventions and treatments for smoking cessation. The results suggest that interventions targeting changes of nicotine content can help smokers reduce tobacco use or quit smoking. A number of clinical studies show that smokers who received an immediate reduction in nicotine content to very low levels have significantly greater reductions in the number of cigarettes smoked and toxic substance exposure compared to those with gradual reductions. However, from the perspective of smoking craving, whether the immediate and gradual reduction in nicotine content reduce smoking by reducing cravings needs further investigation. Methods: 74 eligible Participants were randomly allocated to one of the two experimental conditions: (1) immediate reduction to 0.1 mg of nicotine per cigarette (n = 40); (2) gradual reduction from 1.0 (0.8 g ~ 1.2 mg) to 0.1 mg of nicotine per cigarette (n = 34). All participants completed 1-week baseline period during which they smoked their usual cigarette, followed by 16-week of interventions. The primary outcomes included cigarette cravings and number of cigarettes smoked per day (CPD); secondary outcomes included the number of cigarette-free day and emotional states. Results: Among the 52 participants [51 (98.1%) men; mean (SD) age, 33.44 (6.71) years; mean (SD) CPD, 16.83 (9.94)] who completed the trial, significantly lower cravings for cigarettes were observed in the immediate (n = 25) vs. gradual nicotine reduction group (n = 27) in the morning (t = -2.072, p = 0.039) and after dinner (t = -2.056, p = 0.041). Compared with the baseline daily smoking, the number of cigarettes smoked per day was significantly reduced at the beginning of week 12 in the immediate nicotine reduction group (p = 0.001) and at week 16 in the gradual nicotine reduction group (p < 0.001). The number of participants with any cigarette-free day was not significantly different between the groups (p = 0.198). The number of cigarette-free days was significantly more in the immediate vs. gradual nicotine reduction group (p = 0.027). Conclusions: The significantly lower cravings were observed in the immediate vs. gradual nicotine reduction group, and led to faster reduction in the number of CPD, and a significant increase in the number of cigarette-free days. These findings add to the evidence base for reduced nicotine content in cigarettes. Clinical Trial Registration: ClinicalTrials.gov, identifier: ChiCTR2100048216.

Metabolites identification and species comparison of Oroxylin A, an anti-cancer flavonoid, in vitro and in vivo by HPLC-Q-TOF-MS/MS.

Oroxylin A, the main component of Scutellaria baicalensis Georigi has been widely studied due to its well-known pharmacological effects. According to previous studies, Oroxylin A with low bioavailability was converted into glucuronidation and sulphonated metabolites, which had high exposure in plasma and generated certain activities. It is necessary to study the metabolites and metabolic pathways of Oroxylin A.This study aimed to explore the metabolites of Oroxylin A in liver microsomes, primary hepatocyte incubation samples of five different species (human, monkey, dog, mouse, rat), and in bile, urine and faeces of rats.It would provide a systematic description of metabolic pathway of Oroxylin A. Also, a method of high-performance liquid chromatography combined with quadrupole time-of-flight mass spectrometry detector (HPLC-Q-TOF-MS/MS) for identification of each metabolite in various biological matrices was developed.This experiment illustrated that phase II metabolites were the main form of Oroxylin A in vitro and in excretion of rats, accompanied with a small amount of phase I metabolites.Furthermore, there were obvious species differences among the metabolism in vitro, especially in phase II. Monkeys and rats may be more suitable for preclinical research than dogs and mice as non-rodent or rodent species.

TNF-α-dependent neuronal necroptosis regulated in Alzheimer's disease by coordination of RIPK1-p62 complex with autophagic UVRAG.

Background: Neuronal death is a major hallmark of Alzheimer's disease (AD). Necroptosis, as a programmed necrotic process, is activated in AD. However, what signals and factors initiate necroptosis in AD is largely unknown. Methods: We examined the expression levels of critical molecules in necroptotic signaling pathway by immunohistochemistry (IHC) staining and immunoblotting using brain tissues from AD patients and AD mouse models of APP/PS1 and 5×FAD. We performed brain stereotaxic injection with recombinant TNF-α, anti-TNFR1 neutralizing antibody or AAV-mediated gene expression and knockdown in APP/PS1 mice. For in vitro studies, we used TNF-α combined with zVAD-fmk and Smac mimetic to establish neuronal necroptosis models and utilized pharmacological or molecular biological approaches to study the signaling pathways. Results: We find that activated neuronal necroptosis is dependent on upstream TNF-α/TNFR1 signaling in both neuronal cell cultures and AD mouse models. Upon TNF-α stimulation, accumulated p62 recruits RIPK1 and induces its self-oligomerization, and activates downstream RIPK1/RIPK3/MLKL cascade, leading to neuronal necroptosis. Ectopic accumulation of p62 is caused by impaired autophagy flux, which is mediated by UVRAG downregulation during the TNF-α-promoted necroptosis. Notably, UVRAG overexpression inhibits neuronal necroptosis in cell and mouse models of AD. Conclusions: We identify a finely controlled regulation of neuronal necroptosis in AD by coordinated TNF-α signaling, RIPK1/3 activity and autophagy machinery. Strategies that could fine-tune necroptosis and autophagy may bring in promising therapeutics for AD.

Effect of Pterostilbene, a Natural Derivative of Resveratrol, in the Treatment of Colorectal Cancer through Top1/Tdp1-Mediated DNA Repair Pathway.

Topoisomerase 1 (Top1) inhibitor is an effective anticancer drug, but several factors limit its clinical application such as drug inactivation, tyrosyl-DNA phosphodiesterase 1 (Tdp1)-mediated tumor drug resistance, and its toxicity. Our previous study identified pterostilbene (PTE) and resveratrol (RE) to suppress these two proteins by binding to their active center. PTE and RE could inhibit the proliferation of various colorectal cancer cells, induce cell apoptosis, and make cell cycle stay in G2/M phase in vitro. PTE and RE could decrease Top1 and Tdp1 contents and mRNA expression in wild-type, constructed Tdp1 overexpressing CL187, Top1- or Tdp1- silenced CL187 cell lines. PTE exhibited excellent antitumor activity in subcutaneous CL187 transplantation model (TGI = 79.14 ± 2.85%, 200 mg/kg, i.p.) and orthotopic transplantation model (TGI = 76.57 ± 6.34%, 100 mg/kg, i.p.; TGI = 72.79 ± 4.06%, 500 mg/kg, i.g.) without significant toxicity. PTE had no significant inhibitory effect on non-tumor cell proliferation in vitro and would not induce damage to liver, kidney, and other major organs. Overall, PTE and RE can inhibit the activity of Top1 enzyme and inhibit the DNA damage repair pathway mediated by Top1/Tdp1, and can effectively inhibit colorectal cancer development with low toxicity, thus they have great potential to be developed into a new generation of anti-tumor drugs.

Identification of Triterpene Acids in Poria cocos Extract as Bile Acid Uptake Transporter Inhibitors.

Literature reports that Poria cocos reduces blood lipid levels; however, the underlying mechanism remains unclear. Blood lipid levels are closely related to the enterohepatic circulation of bile acids, where uptake transporters playing a significant role. P. cocos extract is commonly used in traditional prescriptions and food supplements in China. We investigated the effects of P. cocos and its five triterpene acids on bile acid uptake transporters, including intestinal apical sodium-dependent bile acid transporter (ASBT) and hepatic sodium/taurocholate cotransporting polypeptide (NTCP). Triterpene acids were fingerprinted by high-performance liquid chromatography-TripleTOF and quantified by ultraperformance liquid chromatography/tandem mass spectrometry. The inhibitory effect of P. cocos and its five major representative triterpene acids on ASBT and NTCP was investigated by in vitro assays using Xenopus oocytes expressing ASBT and NTCP. P. cocos extract exhibited significant inhibitory effects with half-maximum inhibition constants of 5.89 µg/ml and 14.6 µg/ml for NTCP and ASBT, respectively. Among five triterpene acids, poricoic acid A, poricoic acid B, and polyporenic acid C significantly inhibited NTCP function. Poricoic acid A, poricoic acid B, and dehydrotumulosic acid significantly inhibited ASBT function. The representative triterpene acid, poricoic acid A, was identified as a competitive inhibitor of NTCP with an inhibitory constant of 63.4 ± 18.7 µM. In conclusion, our results indicate that both P. cocos extract and its major triterpenes are competitive inhibitors of ASBT and NTCP. Accordingly, it was suggested that competitive inhibition of these bile acid transporters is one of the underlying mechanisms for the hypolipidemic effect of P. cocos. SIGNIFICANCE STATEMENT: Poria cocos, a commonly used Chinese herbal medicine and food supplement, demonstrates significantly inhibitory effects on the function of apical sodium-dependent bile acid transporter and sodium/taurocholate cotransporting polypeptide. P. cocos has potential to reduce the blood lipid through inhibition of these uptake transporters in enterohepatic circulation of bile acid.

Influences of ABC transporter and CYP3A4/5 genetic polymorphisms on the pharmacokinetics of lenvatinib in Chinese healthy subjects.

PURPOSE: To investigate whether the CYP3A4/5 and ABC transporter genetic polymorphisms could affect the pharmacokinetics of lenvatinib in Chinese healthy subjects. METHODS: Thirty-two healthy Chinese volunteers were enrolled and took oral administration of 8 mg lenvatinib. Plasma concentration of lenvatinib was determined by UPLC-MS/MS, the CYP3A4*1G, CYP3A5*3, ABCB1 (3435 C>T, 1236 C>T, 2677 G>T/A), ABCG2 (421 C>A, 34 G>A), and ABCC2-24 C>T genotypes were determined by SnapShot Technique. RESULTS: In ABCB1 3435T carriers (n = 19), AUC0-120h (815.7 (701.9-923.9) ng·h/mL) and AUC0-∞ (843.3 (722.2-977.7) ng·h/mL) were significantly higher than ABCB1 3435CC homozygous subjects (n = 13, 575.3 (513.7-756.9) ng·h/mL and 590.0 (540.5-782.0) ng·h/mL, respectively); on the contrary, the clearance (CL/F) of ABCB1 3435T carriers was significantly lower (9.5 (8.2-11.1) L/h vs. 13.6 (10.4-14.8) L/h). And the Cmax in CYP3A4*1G/*1G allele carrier subjects was higher than *1 carrier (73.4 ng/mL vs. 53.5 (46.1-60.6) ng/mL), but did not reach the level of significantly statistical difference. Genetic polymorphisms of ABCC2, ABCG2, and CYP3A5 could not influence pharmacokinetic parameters of lenvatinib. CONCLUSIONS: This work presented an evidence that the ABCB1 3435 C>T polymorphism could significantly affect the exposure and clearance of lenvatinib. These findings may explain the reasons for the huge inter-individual differences in lenvatinib, and should contribute to clinical individualized treatment.

A rapid HPLC-MS/MS method for determining busulfan in hemolytic samples from children with hematopoietic stem cell transplantation.

A rapid and sensitive method for the quantitative detection of busulfan (BU) in children's hemolytic samples by HPLC-tandem mass spectrometry (MS/MS) was established. In this study, the sample preparation procedure involved a one-step protein precipitation with acetonitrile (ACN) solution, and the HPLC-MS/MS method used Hypersil GOLD C18 . The mobile phase consisted of 10 mM ammonium acetate solution (containing 0.1% formic acid) and ACN with a flow rate of 0.4 mL/min. Multiple reaction monitoring modes were used for quantitative analysis and the ion pairs of BU and BU-d8 were m/z 263.9 → 150.9 and 272.0 → 159.0, respectively. BU had a good linearity in the range of 0.01-10 µg mL-1 . The intra- and inter-day relative error was between -7.21% and 8.26%, and the coefficient of variation was less than 12.64%. The average extraction recovery rate in plasma samples was 99.76% ± 6.53%, and the matrix in normal plasma and hemolyzed plasma had no significant effect on the detection results. Normal and hemolytic samples could maintain good stability at 4, 25 and -40°C. As a result, this method is particularly suitable for determining BU in hemolytic samples from children with hematopoietic stem cell transplantation (HSCT), and this study provides the methodological basis for further research on the pharmacokinetics of BU in children with HSCT.

A systematic metabolic pathway identification of Common Gardenia Fruit (Gardeniae Fructus) in mouse bile, plasma, urine and feces by HPLC-Q-TOF-MS/MS.

Gardeniae Fructus was a traditional Chinese medicine (TCM) containing various biological ingredients including iridoids and crocetins, monocyclic monoterpenes, organic acids, and flavonoids. However, few systematic identification studies of the bioactive components in vivo have been reported. Herein, the ingredients and metabolites of Gardeniae Fructus were investigated using high-performance liquid chromatography coupled with high-sensitivity Q-TOF mass spectrometry. A total of 45 prototype compounds in Gardeniae Fructus extract were tentatively identified. After oral administration, 69 of prototypes and metabolites were identified from mice bile, plasma, urine, and feces, in which, 31 compounds were prototypes, and 38 chemicals were metabolites. The in vivo biotransformation pathways of these metabolites were also proposed including phase I (hydrolysis, hydrogenation, oxidation, loss of O, and ketone formation, decarboxylation) and phase II reactions (glycine, cysteine, glutathione, and glutamine, and sulfate conjugation, and glucuronidation). For the first time, our results had revealed systematic metabolic profiles of ingredients in Gardeniae Fructus extract in vivo of mice and replenished novel knowledge into the explanation of effective material and/or toxicological basis of Gardeniae Fructus which deserves further investigation.