Re-expression of epigenetically silenced PTPRR by histone acetylation sensitizes RAS-mutant lung adenocarcinoma to SHP2 inhibition.

Silenced protein tyrosine phosphatase receptor type R (PTPRR) participates in mitogen-activated protein kinase (MAPK) signaling cascades during the genesis and development of tumors. Rat sarcoma virus (Ras) genes are frequently mutated in lung adenocarcinoma, thereby resulting in hyperactivation of downstream MAPK signaling. However, the molecular mechanism manipulating the regulation and function of PTPRR in RAS-mutant lung adenocarcinoma is not known. Patient records collected from the Cancer Genome Atlas and Gene Expression Omnibus showed that silenced PTPRR was positively correlated with the prognosis. Exogenous expression of PTPRR suppressed the proliferation and migration of lung cancer cells. PTPRR expression and Src homology 2 containing protein tyrosine phosphatase 2 (SHP2) inhibition acted synergistically to control ERK1/2 phosphorylation in RAS-driven lung cancer cells. Chromatin immunoprecipitation assay revealed that HDAC inhibition induced enriched histone acetylation in the promoter region of PTPRR and recovered PTPRR transcription. The combination of the HDAC inhibitor SAHA and SHP2 inhibitor SHP099 suppressed the progression of lung cancer markedly in vitro and in vivo. Therefore, we revealed the epigenetic silencing mechanism of PTPRR and demonstrated that combination therapy targeting HDAC and SHP2 might represent a novel strategy to treat RAS-mutant lung cancer.

ACAT1 deficiency in myeloid cells promotes glioblastoma progression by enhancing the accumulation of myeloid-derived suppressor cells.

Glioblastoma (GBM) is a highly aggressive and lethal brain tumor with an immunosuppressive tumor microenvironment (TME). In this environment, myeloid cells, such as myeloid-derived suppressor cells (MDSCs), play a pivotal role in suppressing antitumor immunity. Lipometabolism is closely related to the function of myeloid cells. Here, our study reports that acetyl-CoA acetyltransferase 1 (ACAT1), the key enzyme of fatty acid oxidation (FAO) and ketogenesis, is significantly downregulated in the MDSCs infiltrated in GBM patients. To investigate the effects of ACAT1 on myeloid cells, we generated mice with myeloid-specific (LyzM-cre) depletion of ACAT1. The results show that these mice exhibited a remarkable accumulation of MDSCs and increased tumor progression both ectopically and orthotopically. The mechanism behind this effect is elevated secretion of C-X-C motif ligand 1 (CXCL1) of macrophages (Mφ). Overall, our findings demonstrate that ACAT1 could serve as a promising drug target for GBM by regulating the function of MDSCs in the TME.

Chlorogenic Acid Induced Neuroblastoma Cells Differentiation via the ACAT1-TPK1-PDH Pathway.

BACKGROUND: Chlorogenic acid (CHA) has been shown to have substantial biological activities, including anti-inflammatory, antioxidant, and antitumor effects. However, the pharmacological role of CHA in neuroblastoma has not yet been assessed. Neuroblastoma is a type of cancer that develops in undifferentiated sympathetic ganglion cells. This study aims to assess the antitumor activity of CHA against neuroblastoma and reveal its mechanism of action in cell differentiation. METHODS: Be(2)-M17 and SH-SY5Y neuroblastoma cells were used to confirm the differentiation phenotype. Subcutaneous and orthotopic xenograft mouse models were also used to evaluate the antitumor activity of CHA. Seahorse assays and metabolomic analyses were further performed to investigate the roles of CHA and its target ACAT1 in mitochondrial metabolism. RESULTS: CHA induced the differentiation of Be(2)-M17 and SH-SY5Y neuroblastoma cells in vivo and in vitro. The knockdown of mitochondrial ACAT1, which was inhibited by CHA, also resulted in differentiation characteristics in vivo and in vitro. A metabolomic analysis revealed that thiamine metabolism was involved in the differentiation of neuroblastoma cells. CONCLUSIONS: These results provide evidence that CHA shows good antitumor activity against neuroblastoma via the induction of differentiation, by which the ACAT1-TPK1-PDH pathway is involved. CHA is a potential drug candidate for neuroblastoma therapy.

S-72, a Novel Orally Available Tubulin Inhibitor, Overcomes Paclitaxel Resistance via Inactivation of the STING Pathway in Breast Cancer.

Microtubule-targeting agents are widely used as active anticancer drugs. However, drug resistance always emerges after their long-term use, especially in the case of paclitaxel, which is the cornerstone of all subtypes of breast cancer treatment. Hence, the development of novel agents to overcome this resistance is vital. This study reports on a novel, potent, and orally bioavailable tubulin inhibitor called S-72 and evaluated its preclinical efficacy in combating paclitaxel resistance in breast cancer and the molecular mechanisms behind it. We found that S-72 suppresses the proliferation, invasion and migration of paclitaxel-resistant breast cancer cells in vitro and displays desirable antitumor activities against xenografts in vivo. As a characterized tubulin inhibitor, S-72 typically inhibits tubulin polymerization and further triggers mitosis-phase cell cycle arrest and cell apoptosis, in addition to suppressing STAT3 signaling. Further studies showed that STING signaling is involved in paclitaxel resistance, and S-72 blocks STING activation in paclitaxel-resistant breast cancer cells. This effect further restores multipolar spindle formation and causes deadly chromosomal instability in cells. Our study offers a promising novel microtubule-destabilizing agent for paclitaxel-resistant breast cancer treatment as well as a potential strategy that can be used to improve paclitaxel sensitivity.

SHMT2 Promotes Gastric Cancer Development through Regulation of HIF1α/VEGF/STAT3 Signaling.

The metabolic enzymes involved in one-carbon metabolism are closely associated with tumor progression and could be potential targets for cancer therapy. Recent studies showed that serine hydroxymethyltransferase 2 (SHMT2), a crucial enzyme in the one-carbon metabolic pathway, plays a key role in tumor proliferation and development. However, the precise role and function of SHMT2 in gastric cancer (GC) remain poorly understood. In this study, we presented evidence that SHMT2 was necessary for hypoxia-inducible factor-1α (HIF1α) stability and contributed to GC cells' hypoxic adaptation. The analysis of datasets retrieved from The Cancer Genome Atlas and the experimentation with human cell lines revealed a marked increase in SHMT2 expression in GC. The SHMT2 knockdown in MGC803, SGC7901, and HGC27 cell lines inhibited cell proliferation, colony formation, invasion, and migration. Notably, SHMT2 depletion disrupted redox homeostasis and caused glycolytic function loss in GC cells under hypoxic circumstances. Mechanistically, we discovered SHMT2 modulated HIF1α stability, which acted as a master regulator of hypoxia-inducible genes under hypoxic conditions. This, in turn, regulated the downstream VEGF and STAT3 pathways. The in vivo xenograft experiments showed that SHMT2 knockdown markedly reduced GC growth. Our results elucidate the novel function of SHMT2 in stabilizing HIF1α under hypoxic conditions, thus providing a potential therapeutic strategy for GC treatment.

Variable p53/Nrf2 crosstalk contributes to triptolide-induced hepatotoxic process.

This study was to investigate the potential mechanism of triptolide-induced hepatotoxicity. We found a novel and variable role of p53/Nrf2 crosstalk in triptolide-induced hepatotoxic process. Low doses of triptolide led to adaptive stress response without obvious toxicity, while high levels of triptolide caused severe adversity. Correspondingly, at the lower levels of triptolide treatment, nuclear translocation of Nrf2 as well as its downstream efflux transporters multidrug resistance proteins and bile salt export pump expressions were significantly enhanced, so did p53 pathways that also increased; at a toxic concentration, total and nuclear accumulations of Nrf2 decreased, while p53 showed an obvious nuclear translocation. Further studies showed the cross-regulation between p53 and Nrf2 after different concentrations of triptolide treatment. Under mild stress conditions, Nrf2 induced p53 highly expression to maintain the pro-survival outcome, while p53 showed no obvious effect on Nrf2 expression and transcriptional activity. Under high stress conditions, the remaining Nrf2 as well as the largely induced p53 mutually inhibited each other, leading to a hepatotoxic result. Nrf2 and p53 could physically and dynamically interact. Low levels of triptolide enhanced the interaction between Nrf2 and p53. Reversely, p53/Nrf2 complex dissociated at high levels of triptolide treatment. Altogether, variable p53/Nrf2 crosstalk contributes to triptolide-induced self-protection and hepatotoxicity, modulation of which may be a potential strategy for triptolide-induced hepatotoxicity intervention.

Liquiritin alleviates alpha-naphthylisothiocyanate-induced intrahepatic cholestasis through the Sirt1/FXR/Nrf2 pathway.

Liquiritin (LQ) is an important monomer active component in flavonoids of licorice. The objective of this study was to evaluate the hepatoprotective effects of LQ in cholestatic mice. LQ (40 or 80 mg/kg) was intragastrically administered to mice once daily for 6 days, and mice were treated intragastrically with a single dosage of ANIT (75 mg/kg) on the 5th day. On the 7th day, mice were sacrificed to collect blood and livers. The mRNA and protein levels were determined by qRT-PCR and western blot assay. We also conducted systematical assessments of miRNAs expression profiles in the liver. LQ ameliorated ANIT-induced cholestatic liver injury, as evidenced by reduced serum biochemical markers and attenuated pathological changes in liver. Pretreatment of LQ reduced the increase of malondialdehyde, TNF-α, and IL-1ß induced by ANIT. Moreover, ANIT suppressed the expression of Sirt1 and FXR in liver tissue, which was weakened in the LQ pre-treatment group. LQ enhanced the nuclear expression of Nrf2, which was increased in the ANIT group. LQ also increased the mRNA expressions of bile acid transporters Bsep, Ntcp, Mrp3, and Mrp4. Furthermore, a miRNA deep sequencing analysis revealed that LQ had a global regulatory effect on the hepatic miRNA expression. Kyoto Encyclopedia of Genes and Genomes functional enrichment analysis showed that the differentially expressed miRNAs were mainly related to metabolic pathways, endocytosis, and MAPK signaling pathway. Collectively, LQ attenuated hepatotoxicity and cholestasis by regulating the expression of Sirt1/FXR/Nrf2 and the bile acid transporters, indicating that LQ might be an effective approach for cholestatic liver diseases.

Glycyrrhetinic Acid Protects α-Naphthylisothiocyanate- Induced Cholestasis Through Regulating Transporters, Inflammation and Apoptosis.

Glycyrrhetinic acid (GA), the active metabolic product of Glycyrrhizin (GL) that is the main ingredient of licorice, was reported to protect against α-naphthylisothiocyanate (ANIT)- induced cholestasis. However, its protective mechanism remains unclear. In our work, the cholestatic liver injury in mice was caused by ANIT and GA was used for the treatment. We assessed cholestatic liver injury specific indexes, histopathological changes, bile acid transporters, inflammation and apoptosis. The results of liver biochemical index and histopathological examination showed that GA markedly attenuated ANIT-induced liver injury. Mechanism research suggested that GA could activate the expression of farnesoid x receptor (FXR) and its downstream bile acids transporters Na+/taurocholate co-transporting polypeptide (NTCP), bile salt export pump (BSEP) and multidrug resistance-associated protein 2 (MRP2), as well as the nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream proteins MRP3, MRP4. These transporters play a vital role in mediating bile acid homeostasis in hepatocytes. Moreover, GA could significantly inhibit the ANIT-induced activation of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) inflammatory pathway and the increase of tumor necrosis factor-α (TNF-α) concentration in serum. Also, GA protected against ANIT-induced mitochondrial apoptosis by regulating the expression of Bcl-2, Bax, cleaved caspase 3 and cleaved caspase 9. In conclusion, GA alleviates the hepatotoxicity caused by ANIT by regulating bile acids transporters, inflammation and apoptosis, which suggests that GA may be a potential therapeutic agent for cholestasis.

Role of MicroRNA-155 in Triptolide-induced hepatotoxicity via the Nrf2-Dependent pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Triptolide (TP), the main bioactive and toxic ingredient of Tripterygium wilfordii Hook F, causes severe toxicity, particularly for hepatotoxicity. However, the underlying mechanisms for its hepatotoxicity are not entirely clear. AIM OF THE STUDY: The purpose of the study was to explore the role of miR-155, a microRNA closely related to various liver injuries and a regulator of the nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant pathway, in TP-induced liver injury in vitro and in vivo. MATERIALS AND METHODS: First, in vitro L02 cells were treated with different concentrations of TP. The protein levels of Nrf2 and its downstream genes Heme oxygenase1 (HO-1) were determined by Western blot. The mRNA expression of miR-155, Nrf2, NAD(P)H: quinone oxidoreductase 1 (NQO1) and HO-1 were measured using qRT-PCR. And we transfected miR-155 inhibitor and miminc before TP treatment to determine the mRNA and/or protein levels of miR-155, Nrf2 and HO-1. Then, we further confirmed the interaction between miR-155 and Nrf2 pathway in TP-induced hepatic injury in BALB/C mice. The degree of liver injury was determined by HE staining and serum biochemical. The mRNA expression of miR-155 was examined with qRT-PCR and Nrf2 and HO-1 gene expression in liver were evaluated by immunohistochemistry and/or Western blot. RESULTS: The results showed that TP significantly induced the expression of miR-155 both in L02 cells and in rodents liver tissue, and the inhibition of miR-155 could mitigate the hepatic damages caused by TP. Further experiments demonstrated that the inhibition of miR-155 reversed the down-regulation of Nrf2 and HO-1 by TP, while the miR-155 mimic enhanced the effects of TP. Animal experiments also showed that the inhibition of miR-155 by miR-155 antagomir reversed the decrease of Nrf2 induced by TP administration. CONCLUSIONS: These results indicated that miR-155 played an important role in TP-induced hepatotoxicity by regulating the Nrf2 signaling pathway.

Celastrol slows the progression of early diabetic nephropathy in rats via the PI3K/AKT pathway.

BACKGROUND: Diabetic nephropathy serves as one of the most regular microvascular complications of diabetes mellitus and is the main factor that causes end-stage renal disease and incident mortality. As the beneficial effect and minute adverse influence of Celastrol on the renal system requires further elucidation, the renoprotective function of Celastrol in early diabetic nephropathy was investigated. METHODS: In high-fat and high-glucose diet/streptozotocin-induced diabetic rats which is the early diabetic nephropathy model, ALT, AST, 24 h urinary protein, blood urea nitrogen, and serum creatinine content were observed. Periodic acid-Schiff staining, enzyme-linked immunosorbent assay, immunohistochemical analysis, reverse transcription-polymerase chain reaction, and western blot analysis were used to explore the renoprotective effect of Celastrol to diabetic nephropathy rats and the underlying mechanism. RESULTS: High dose of Celastrol (1.5 mg/kg/d) not only improved the kidney function of diabetic nephropathy (DN) rats, and decreased the blood glucose and 24 h urinary albumin, but also increased the expression of LC3II and nephrin, and downregulated the expression of PI3K, p-AKT, and the mRNA level of NF-κB and mTOR. CONCLUSION: Celastrol functions as a potential therapeutic substance, acting via the PI3K/AKT pathway to attenuate renal injury, inhibit glomerular basement membrane thickening, and achieve podocyte homeostasis in diabetic nephropathy.

Nrf2-dependent antioxidant response mediated the protective effect of tanshinone IIA on doxorubicin-induced cardiotoxicity.

Doxorubicin (DOX), a potent and widely used anticancer agent, can give rise to severe cardiotoxicity that limits its clinical use by inducing oxidative stress. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is the central regulator of cellular responses to electrophilic/oxidative stress, which serves a critical role in maintenance of normal cardiac function. Tanshinone IIA (Tan IIA) has previously been reported to protect against DOX-induced cardiotoxicity. The aim of the present study was to elucidate whether Nrf2 signaling serves a role in the underlying mechanism. In the animal model, DOX induced acute cardiotoxicity, whereas Tan IIA pretreatment reduced the activity of myocardial enzymes, and increased activity of the antioxidant enzymes superoxide dismutase, catalase and glutathione (GSH). Furthermore, Tan IIA pretreatment (3-10 µM) significantly increased the cell viability and markedly restored morphological changes in DOX-injured H9c2 cells, decreased the generation of reactive oxygen species, and increased the level of intracellular GSH. Additionally, Tan IIA pretreatment also induced the nuclear accumulation of Nrf2 and its downstream genes heme oxygenase-1, NAD(P)H dehydrogenase (quinone) 1, and glutamate-cysteine ligase catalytic subunit in both the mice cardiac tissues and H9c2 cells. Nrf2 knockdown by small interfering RNA downregulated Tan IIA-induced Nrf2 activation and reversed the effect of Tan IIA on the DOX-induced inhibition of cell viability. These results suggest that the Nrf2-dependent antioxidant response mediates the protective effect of Tan IIA on DOX-induced cardiotoxicity.

Broad targeting of triptolide to resistance and sensitization for cancer therapy.

Cancer cell resistance to current anticancer therapeutics as well as the side effects are still obstacles to successful cancer therapy. Hence, the development of novel anticancer agents or therapeutics is of vital significance, and especially rational adjuvant therapies containing low-cost natural products with multiple targets have attracted great interests. Triptolide, the main biocomponent of Tripterygium wilfordii Hook F, is restricted in clinical applications mainly due to its severe systemic toxicities, although it has shown strong antitumor activities in preclinical studies. Mounting evidence suggests that triptolide at low doses as an adjuvant therapeutic agent circumvents resistance to current anticancer therapies, enhances the anticancer effectiveness, and relieves toxicities of both triptolide and anticancer therapies. Furthermore, several unique antitumor targets of triptolide make it superior to other therapeutics. The molecular mechanisms of triptolide-induced anti-resistance and sensitization effects include changes in ATP-binding cassette transporters, induction of apoptosis pathways, increase in tumor suppressors and decrease in oncogenic factors, and interactions with the RNA polymerase II complex; targeting cancer stem cells and tumor-microenvironment-mediated resistance are also involved. Besides, some synthetic derivatives and novel delivery systems of triptolide are also developed to enhance the water-solubility and reduce the toxicity, which will also be discussed.

Mechanisms of Triptolide-Induced Hepatotoxicity and Protective Effect of Combined Use of Isoliquiritigenin: Possible Roles of Nrf2 and Hepatic Transporters.

Triptolide (TP), the main bioactive component of Tripterygium wilfordii Hook F, can cause severe hepatotoxicity. Isoliquiritigenin (ISL) has been reported to be able to protect against TP-induced liver injury, but the mechanisms are not fully elucidated. This study aims to explore the role of nuclear transcription factor E2-related factor 2 (Nrf2) and hepatic transporters in TP-induced hepatotoxicity and the reversal protective effect of ISL. TP treatment caused both cytotoxicity in L02 hepatocytes and acute liver injury in mice. Particularly, TP led to the disorder of bile acid (BA) profiles in mice livers. Combined treatment of TP with ISL effectively alleviated TP-induced hepatotoxicity. Furthermore, ISL pretreatment enhanced Nrf2 expressions and nuclear accumulations and its downstream NAD(P)H: quinine oxidoreductase 1 (NQO1) expression. Expressions of hepatic P-gp, MRP2, MRP4, bile salt export pump, and OATP2 were also induced. In addition, in vitro transport assays identified that neither was TP exported by MRP2, OATP1B1, or OATP1B3, nor did TP influence the transport activities of P-gp or MRP2. All these results indicate that ISL may reduce the hepatic oxidative stress and hepatic accumulations of both endogenous BAs and exogenous TP as well as its metabolites by enhancing the expressions of Nrf2, NQO1, and hepatic influx and efflux transporters. Effects of TP on hepatic transporters are mainly at the transcriptional levels, and changes of hepatic BA profiles are very important in the mechanisms of TP-induced hepatotoxicity.

Simple and rapid UPLC-MS/MS method for quantification of entecavir in human plasma: Application to a bioequivalence study
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A simple and fast ultra-performance liquid chromatography-tandem mass spectrometry method was developed and validated to determine entecavir in human plasma with the stable isotopically labeled internal standard entecavir-13C215N. Samples (100 µL each) were pretreated by protein precipitation with methanol, and then separated on an ACQUITY UPLC BEH C18 analytical column (2.1 × 50 mm, 1.7 µm) with a simple isocratic elution. The detection was operated by a positive ionization electrospray mass spectrometry in multiple reaction monitoring mode. The method had a short chromatographic run time of 2 minutes, and obtained sharp peaks of entecavir and the internal standard. Good linearity was found within 0.1 - 20 ng/mL. The intra- and inter-day precision and accuracy met the acceptance criteria, and no matrix effect was observed. This method was successfully applied in a bioequivalence study of two kinds of entecavir tablets in healthy Chinese volunteers. And the results showed that no significant differences were found between the test and reference preparations in pharmacokinetic parameters (p > 0.05) by ANOVA. The 90% confidence intervals for the geometric mean ratios (test/reference) of Cmax, AUC0-tlast, and AUC0-∞ fell within the bioequivalence acceptance criteria (80 - 125%). No significant difference was found in tmax between the two preparations. The two one-sided t-tests showed that these two products were bioequivalent.
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Genotyping as a Key Element of Sample Size Optimization in Bioequivalence of Risperidone Tablets.

BACKGROUND AND OBJECTIVES: Risperidone is a derivative of benzisoxazole and is widely used for schizophrenia and other psychiatric illnesses in both adults and children. Previous studies have confirmed that it is a highly variable drug (within-subject variability ≥ 30%). To reduce the large sample size required for bioequivalence researches on highly variable drugs, a role for genotyping in the design of the bioequivalence study was employed. METHODS: A randomized, open-label, two-period crossover study was adopted: 20 subjects with specific genotypes carrying cytochrome P450 (CYP) 2D6*10 were randomized to two groups to receive a single oral dose of trial formulation or reference formulation with a 2-week washout period. Blood concentrations of risperidone (parent drug) and 9-hydroxy risperidone (active metabolite) were measured by high-performance liquid chromatography-tandem mass spectrometry. RESULTS: Eighteen out of the 20 subjects completed the study (two did not finish the test in the second period). The pharmacokinetic parameters of AUClast, AUC∞ and Cmax for the 18 subjects after a single oral dose of the trial or reference preparation were 216.1 ± 88.7 and 220.5 ± 96.8 ng·h/mL; 221.6 ± 93.1 and 226.4 ± 103.5 ng·h/mL; 36.7 ± 10.3 and 36.0 ± 10.2 ng/mL, respectively. The CVw of risperidone in natural logarithm-transformed Cmax was 22.4 and 25.38% for 9-hydroxy risperidone. CONCLUSIONS: The test formulation met the Food and Drug Administration guidelines and regulation criteria for bioequivalence. By controlling the genotype, it could actually help reduce the CVw, which may be a feasible method to decrease the sample size for the bioequivalence study of highly variable drugs.

The Ethanol Extract of Licorice (Glycyrrhiza uralensis) Protects against Triptolide-Induced Oxidative Stress through Activation of Nrf2.

To investigate the potential role of nuclear factor erythroid 2-related factor 2 (Nrf2) in licorice ethanol extract (LEE) against triptolide- (TP-) induced hepatotoxicity, HepG2 cells were exposed to LEE (30, 60, and 90 mg·L-1) for 12 h and then treated with TP (50 nM) for 24 h. Besides, an acute liver injury model was established in ICR mice by a single dose of TP (1.0 mg·kg-1, i.p.). Relevant oxidant and antioxidant mediators were analyzed. TP led to an obvious oxidative stress as evidenced by increasing levels of ROS and decreasing GSH contents in HepG2 cells. In vitro results were likely to hold true in in vivo experiments. LEE protected against TP-induced oxidative stress in both in vitro and in vivo conditions. Furthermore, the decreased level of Nrf2 in the TP-treated group was observed. The mRNA levels of downstream genes decreased as well in ICR mice liver, whereas they increased in HepG2 cells. In contrast, LEE pretreatment significantly increased the level of Nrf2 and its downstream genes. LEE protects against TP-induced oxidative stress partly via the activation of Nrf2 pathway.

Danshen modulates Nrf2-mediated signaling pathway in cisplatin-induced renal injury.

Danshen, an efficacious agent for cardiovascular diseases, has been found to play an essential role in kidney injury. In the present study, the effect of Danshen on cisplatin-induced renal dysfunction was investigated in a mouse model. Danshen was administered to mice at a dose of 3 g/kg 4 days before and 3 days after cisplatin treatment. A single intraperitoneal injection of 20 mg/kg cisplatin was used to induce nephrotoxicity. The mice were sacrificed 72 h after cisplatin intoxication. Biochemical parameters including serum creatinine and blood urea nitrogen were analyzed. Histopathological changes of kidney tissues were detected using HE staining. Antioxidant enzymes (GSH-Px and SOD) and peroxidative product (MDA) were detected. Protein expressions of Nrf2 and its target genes including HO-1 and NQO1 were measured by Western blotting. The results showed that pretreatment with Danshen significantly reduced serum creatinine and blood urea nitrogen in the cisplatin-treated mice. Histopathological examination showed that Danshen mitigated the renal damage induced by cisplatin. Moreover, Danshen restored the activities of antioxidant enzymes (GSH-Px and SOD) and normalized the MDA contents in renal tissues. Western blotting revealed that Danshen enhanced the expressions of Nrf2 and its target genes in cisplatin-exposed mice. It was suggested that Danshen protects against the cisplatin-induced renal impairment in the mice, which is potentially associated with the upregulation of Nrf2-mediated signaling pathway.

Pharmacokinetics and relative bioavailability of a generic amisulpride tablet in healthy Chinese volunteers
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OBJECTIVE: To assess and compare the pharmacokinetic properties and bioavailability of a newly developed formulation of amisulpride with those of a conventional formulation in healthy Chinese volunteers under fasting state. MATERIALS AND METHODS: A single-dose, two-sequence crossover study was designed. 20 healthy subjects (14 males and 6 females) were randomized into two groups. A single oral dose of amisulpride (200 mg) was given after an overnight fast of 12 hours. Blood samples were taken at scheduled time spots and separated by a washout period of 14 days. Plasma concentration of amisulpride was measured by high-performance liquid chromatography-fluorescence detector (HPLC-FLD) method. RESULTS: The pharmacokinetic parameters of AUC0-tlast, AUC0-∞, and Cmax for the 20 subjects after a single oral dose of the trial preparation or the reference preparation were 4,767.2 and 4,856.3 ng×h×mL-1; 4,891.7 and 5,043.2 ng×h×mL-1; 584.7 and 586.3 ng×mL-1, respectively. The relative bioavailability was 98.9 ± 14.5%. No significant difference was found among the main pharmacokinetic parameters in the two preparations by ANOVA. The 90% confidence intervals for the geometric mean ratios (test/reference) of Cmax and AUC0-tlast were 90.7 - 109.1% and 92.5 - 103.6%, respectively, meeting the predetermined criteria (80 - 125%) for bioequivalence. No serious adverse events were reported. CONCLUSION: The study demonstrated that the two preparations met the regulatory criteria for bioequivalence and both formulations were well tolerated.
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Glycyrrhetinic Acid Accelerates the Clearance of Triptolide through P-gp In Vitro.

Triptolide (TP) is an active ingredient isolated from Tripterygium wilfordii Hook. f. (TWHF), which is a traditional herbal medicine widely used for the treatment of rheumatoid arthritis and autoimmune disease in the clinic. However, its adverse reactions of hepatotoxicity and nephrotoxicity have been frequently reported which limited its clinical application. The aim of this study was to investigate the mechanism of glycyrrhetinic acid (GA) effecting on the elimination of TP in HK-2 cells and the role of the efflux transporters of P-gp and multidrug resistance-associated proteins (MRPs) in this process. An ultra performance liquid chromatography-electrospray ionization-mass spectrometry (UPLC-ESI-MS) analytical method was established to determine the intracellular concentration of TP. In order to study the role of efflux transporters of P-gp and MRPs in GA impacting on the accumulation of TP, the inhibitors of efflux transporters (P-gp: verapamil; MRPs: MK571) were used in this study. The results showed that GA could enhance the elimination of TP and reduce the TP accumulation in HK-2 cells. Verapamil and MK571 could increase the intracellular concentration of TP; in addition, GA co-incubation with verapamil significantly increased the TP cellular concentration compared with the control group. In conclusion, GA could reduce the accumulation of TP in HK-2 cells, which was related to P-gp. This is probably one of the mechanisms that TP combined with GA to detoxify its toxicity. Copyright © 2017 John Wiley & Sons, Ltd.

Pharmacokinetics and relative bioavailability of two allopurinol tablets in healthy Chinese volunteers
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OBJECTIVE: To evaluate the pharmacokinetics and relative bioavailability of two allopurinol tablets in healthy Chinese volunteers. METHODS: A single-center, randomized, cross-over, two-period study design was conducted in healthy male subjects who were identified as not carrying the HLA-B*58:01 allele. Under fasting conditions, a single oral dose of 300 mg test or reference tablets was given with a 1-week washout period. The blood samples were collected for up to 12 hours after the administration and the plasma concentrations of allopurinol were determined by high performance liquid chromatography. Subject interviews and physical examinations were done over regular intervals to monitor the adverse events. RESULTS: 18 subjects were enrolled in the study, and none dropped out. The main pharmacokinetic parameters of allopurinol test and reference preparations were as follows: AUC0-tlast was 6,725.1 ± 1,390.0 ng×h×mL-1 and 6,425.6 ± 1,257.6 ng×h×mL-1; AUC0-∞ was 7,069.1 ± 1,503.2 ng×h×mL-1 and 6,750.6 ± 1,347.7 ng×h×mL-1; tmax was 1.3 ± 0.8 hours and 1.3 ± 0.8 hours; Cmax was 2,203.7 ± 557.4 ng×mL-1 and 2,310.8 ± 662.8 ng×mL-1; and T1/2 was 2.0 ± 1.6 hours and 1.7 ± 0.7 hours. The relative bioavailability was 105.1 ± 12.6%. The 90% confidence intervals for the geometric mean ratios (test/reference) of Cmax, AUC0-tlast, and AUC0-∞ of both preparations fell within the bioequivalence acceptance criteria (80 - 125%). No adverse events were found or reported during the study. CONCLUSION: The test allopurinol preparations and the reference preparations are bioequivalent and both are well tolerated.
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