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The quest in finding an everlasting panacea to the pernicious impact of sickle cell disease (SCD) in the society hit a turn of success since the recent discovery of a small molecule reversible covalent inhibitor, Voxelotor. A drug that primarily promotes the stability of oxygenated hemoglobin and inhibit the polymerization of HbS by enhancing hemoglobin's affinity for oxygen has opened a new frontier in drug discovery and development. Despite eminent efforts made to reproduce small molecules with better therapeutic targets, none has been successful. To this end, we employed the use of structure-based computational techniques with emphasis on the electrophilic warhead group of Voxelotor to harness novel covalent binders that could elicit better therapeutic response against HbS. The PubChem database and DataWarrior software were used to design random molecules using Voxelotor's electrophilic functionality. Following the compilation of these chemical entities, a high-throughput covalent docking-based virtual screening campaign was conducted which revealed three (Compound_166, Compound_2301, and Compound_2335) putative druglike candidates with higher baseline energy value compared to the standard drug. Subsequently, in silico ADMET profiling was carried out to evaluate their pharmacokinetics and pharmacodynamics properties, and their stability was evaluated for 1 μs (1 μs) using molecular dynamics simulation. Finally, to prioritize these compounds for further development in drug discovery, MM/PBSA calculations was employed to evaluate their molecular interactions and solvation energy within the HbS protein. Despite the admirable druglike and stability properties of these compounds, further experimental validations are required to establish their preclinical relevance for drug development., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Sickle cell disease (SCD) is a devastating, multisystemic disorder that affects millions of people worldwide. The earliest clinical manifestations of SCD can affect infants as young as 6 months of age, and pediatric patients are at risk for acute and life-threatening complications. Early intervention with treatments that target the underlying pathophysiological mechanism of SCD, sickle hemoglobin (HbS) polymerization, are expected to slow disease progression and circumvent disease-associated morbidity and mortality., Procedure: The HOPE-KIDS 1 trial (NCT02850406) is an ongoing four-part, phase 2a, open-label, single- and multiple-dose study to evaluate the pharmacokinetics, efficacy, and safety of voxelotor-a first-in-class HbS polymerization inhibitor-in patients aged 6 months to 17 years with SCD. Initial findings from a cohort of 45 patients aged 4 to 11 years who received voxelotor treatment for up to 48 weeks are reported., Results: Hemoglobin (Hb) response, defined as a >1.0 g/dl increase from baseline, was achieved at week 24 by 47% (n = 16/34) of patients with Hb measurements at baseline and week 24. At week 24, 35% (n = 12/34) and 21% (n = 7/34) of patients had a >1.5 g/dl increase and a >2.0 g/dl increase from baseline in Hb concentration, respectively. Concurrent improvements in hemolytic markers were observed. Voxelotor was well tolerated in this young cohort, with no newly emerging safety signals., Conclusions: Based on its mechanism as an HbS polymerization inhibitor, voxelotor improves Hb levels and markers of hemolysis and has the potential to mitigate SCD-related complications; these results support its use in patients aged ≥4 years., (© 2022 The Authors. Pediatric Blood & Cancer published by Wiley Periodicals LLC.)

Oxbryta (voxelotor) is a small-molecule inhibitor of sickle hemoglobin (Hb) polymerization approved for patients with sickle cell disease (SCD) aged greater than or equal to 12 years at a dose of 1500 mg once daily (q.d.). Voxelotor binds preferentially to Hb, and voxelotor partitioning into red blood cells is an effective predictor of Hb occupancy. The objectives of these analyses were to develop a population pharmacokinetic (PopPK) model for voxelotor in both plasma and whole blood in adults and adolescents to support the dose selection for optimal target engagement and to identify covariates that have a significant effect on voxelotor pharmacokinetics (PK) in plasma and whole blood. An integrated plasma and whole blood PopPK model with two compartments, first-order absorption and elimination, and a site-of-action effect compartment adequately described the concentration-time profiles of voxelotor in plasma and whole blood in patients treated up to 72 weeks. Covariates with significant effects on voxelotor PK included baseline blood volume on apparent volume of the central compartment and time-varying hematocrit and dose on whole blood partitioning, indicating that clinical markers of voxelotor effect can, in turn, influence its PK. Furthermore, the model confirmed that voxelotor PK in plasma and whole blood is linear with dose and time and comparable for adults and adolescents. No clinically important covariate effects on voxelotor PK that warranted dose adjustment were identified in this analysis. Overall, the PopPK analyses contributed significantly to the voxelotor label and support 1500 mg q.d. as the therapeutic dose in adults and adolescents with SCD., (© 2022 Global Blood Therapeutics, Inc. CPT: Pharmacometrics & Systems Pharmacology published by Wiley Periodicals LLC on behalf of American Society for Clinical Pharmacology and Therapeutics.)

Two open-label studies assessed the safety, tolerability, and pharmacokinetics of Oxbryta (voxelotor) in subjects with hepatic or renal impairment. Eight subjects with severe renal impairment (estimated glomerular filtration rate <30 mL/min/1.73 m 2 ) and 8 healthy age-, sex-, and body mass index-matched controls were administered a single oral dose of voxelotor 900 mg. Seven patients with mild (Child-Pugh A), moderate (Child-Pugh B), and severe (Child-Pugh C) hepatic impairment and healthy age-, sex-, and body mass index-matched controls (7:7:7:7) were administered a single oral dose of voxelotor 1500 mg, except those with severe hepatic impairment (600 mg). There was no apparent effect of renal function on the excretion of voxelotor based on comparable half-life values between subjects with severe renal impairment and healthy matched controls. Mean area under the concentration-time curve from time 0 to infinity (AUC 0-inf ) values were lower by approximately 50% (plasma) and 25% (whole blood) in subjects with severe renal impairment compared with controls. Accordingly, dose adjustment is not required in patients with severe renal impairment. Voxelotor plasma and whole-blood exposures were slightly increased in subjects with mild and moderate hepatic impairment. Mean AUC 0-inf values were approximately 9% to 18% higher compared with those of healthy matched controls. Dose adjustment is therefore not required in patients with mild or moderate hepatic impairment. Voxelotor mean AUC 0-inf values were approximately 90% higher in subjects with severe hepatic impairment. A lower voxelotor dose (1000 mg) is recommended for patients with severe hepatic impairment. Voxelotor was well tolerated in all treatment groups., (© 2020 The Authors. The Journal of Clinical Pharmacology published by Wiley Periodicals LLC on behalf of American College of Clinical Pharmacology.)

A simple, sensitive, and rapid UHPLC-MS/MS method was developed for the simultaneous determination of veratraldehyde and its metabolite veratric acid in rat plasma. Cinnamaldehyde was used as an internal standard (IS) and the one-step protein precipitation method with 0.2% formic acid in acetonitrile (mobile phase B) was used for the sample extraction. Reversed C18 column (YMC-Triart C18 column, 50 mm × 2.0 mm, 1.9 µm) was used for chromatographic separation and was maintained at 30 °C. The total run time was 4.5 min and the electrospray ionization in positive mode was used with the transition m/z 167.07 → 139.00 for veratraldehyde, m/z 183.07 → 139.00 for veratric acid, and m/z 133.00 → 55.00 for IS. The developed method exhibited good linearity ( r 2   ≥  0.9977), and the lower limits of quantification ranged from 3 to 10 ng/mL for the two analytes. Intra-day precision and accuracy parameters met the criteria (within ±15%) during the validation. The bioanalytical method was applied for the determination of veratraldehyde and veratric acid in rat plasma after oral and percutaneous administration of 300 and 600 mg/kg veratraldehyde. Using the analytical methods established in this study, we can confirm the absorption and metabolism of veratraldehyde in rats for various routes.

Escherichia coli and Staphylococcus aureus are important agents of urinary tract infections that can often evolve to severe infections. The rise of antibiotic-resistant strains has driven the search for novel therapies to replace the use or act as adjuvants of antibiotics. In this context, plant-derived compounds have been widely investigated. Cuminaldehyde is suggested as the major antimicrobial compound of the cumin seed essential oil. However, this effect is not fully understood. Herein, we investigated the in silico and in vitro activities of cuminaldehyde, as well as its ability to potentiate ciprofloxacin effects against S. aureus and E. coli. In silico analyses were performed by using different computational tools. The PASS online and SwissADME programmes were used for the prediction of biological activities and oral bioavailability of cuminaldehyde. For analysis of the possible toxic effects and the theoretical pharmacokinetic parameters of the compound, the Osiris, SwissADME and PROTOX programmes were used. Estimations of cuminaldehyde gastrointestinal absorption, blood brain barrier permeability and skin permeation by using SwissADME; and drug likeness and score by using Osiris, were also evaluated The in vitro antimicrobial effects of cuminaldehyde were determined by using microdilution, biofilm formation and time-kill assays. In silico analysis indicated that cuminaldehyde may act as an antimicrobial and as a membrane permeability enhancer. It was suggested to be highly absorbable by the gastrointestinal tract and likely to cross the blood brain barrier. Also, irritative and harmful effects were predicted for cuminaldehyde if swallowed at its LD50. Good oral bioavailability and drug score were also found for this compound. Cuminaldehyde presented antimicrobial and anti-biofilm effects against S. aureus and E. coli.. When co-incubated with ciprofloxacin, it enhanced the antibiotic antimicrobial and anti-biofilm actions. We suggest that cuminaldehyde may be useful as an adjuvant therapy to ciprofloxacin in S. aureus and E. coli-induced infections., Competing Interests: The authors have declared that no competing interests exist.

A novel cyclodextrin (CD)-based controlled release system was developed in the small intestine to control the rate of drug release, on the premise of enteric-coated tablets. The system was designed based on the enzymes exogenous β-cyclodextrin glycosyltransferase (β-CGTase) and endogenous maltase-glucoamylase (MG), wherein MG is secreted in the small intestine and substituted by a congenerous amyloglucosidase (AG). The vanillin-/curcumin-β-CD complexes were prepared and detected by Fourier transform infrared (FT-IR), thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC), and host CD degradation was measured based on the glucose yield. The combination of β-CGTase and AG was also functional in the CD complex system. The variations in the concentrations of added β-CGTase, with AG constantly in excess, could effectively alter the rate of host CD degradation and guest release by monitoring glucose production and color disappearance, thus, demonstrating that guest release in the CD complex system could be precisely controlled by changing the amount of β-CGTase used. Thus, the in vitro simulation of the system indicated that a novel controlled release system, based on endogenous MG, could be established in the small intestine. The CD-based controlled release system can be potentially applied in drug delivery and absorption in the small intestine.

Objectives: Cuminaldehyde self-emulsified nanoemulsion (CuA-SEN) was prepared and optimised to improve its oral bioavailability and antihepatotoxicity., Methods: Cuminaldehyde self-emulsified nanoemulsion was developed through the self-nanoemulsification method using Box-Behnken Design (BBD) tool while appropriate physicochemical indices were evaluated. The optimised CuA-SEN was characterised via droplet size (DS), morphology, polydispersity index (PDI), zeta potential (ZP), entrapment efficiency, in-vitro release, and pharmacokinetic studies while its antihepatotoxicity was evaluated., Key Findings: Cuminaldehyde self-emulsified nanoemulsion with acceptable characteristics (mean DS-48.83 ± 1.06 nm; PDI-0.232 ± 0.140; ZP-29.92 ± 1.66 mV; EE-91.51 ± 0.44%; and drug-loading capacity (DL)-9.77 ± 0.75%) was formulated. In-vitro drug release of CuA-SEN significantly increased with an oral relative bioavailability of 171.02%. Oral administration of CuA-SEN to CCl 4 -induced hepatotoxicity mice markedly increased the levels of superoxide dismutase, glutathione and catalase in serum. Also, CuA-SEN reduced the levels of tumour necrosis factor-alpha and interleukin-6 in both serum and liver tissues while aspartate aminotransferase, alanine aminotransferase and malonaldehyde levels were significantly decreased., Conclusions: These findings showed that the improved bioavailability of cuminaldehyde via SEN provided an effective approach for enhancing antioxidation, anti-inflammation and antihepatotoxicity of the drug., (© 2019 Royal Pharmaceutical Society.)

Aims: Voxelotor (previously GBT440) is a haemoglobin (Hb) modulator that increases Hb-oxygen affinity, thereby reducing Hb polymerization and sickling of red blood cells (RBCs), being developed as a once-daily oral drug to treat sickle cell disease (SCD). This first-in-human study evaluated the safety, tolerability, pharmacokinetics and pharmacodynamics of voxelotor in healthy volunteers and SCD patients., Methods: A total of 40 healthy volunteers (100, 400, 1000, 2000 or 2800 mg) and 8 SCD patients (1000 mg) were randomly assigned to a single dose of voxelotor once daily (n = 6 per group) or placebo (n = 2 per group). Twenty-four healthy volunteers received multiple doses of voxelotor once daily for 15 days (300, 600 or 900 mg, n = 6 per group) or placebo (n = 2 per group)., Results: Voxelotor was well tolerated and exhibited a linear pharmacokinetic profile and a half-life ranging from 61 ± 7 h to 85 ± 7 h. High partitioning into the RBC compartment provides evidence of highly specific binding to Hb. Voxelotor exhibited a concentration-dependent left-shift of oxygen equilibrium curves. Percent Hb modification following 900 mg voxelotor for 15 days was 38 ± 9%. Terminal half-life of voxelotor in SCD patients (50 ± 3 h) was shorter than in healthy volunteers. Evaluation of erythropoietin, exercise testing, and haematologic parameters were consistent with normal oxygen delivery during both rest and exercise., Conclusion: This first-in-human study demonstrates voxelotor was well tolerated in SCD patients and healthy volunteers and established proof of mechanism on increasing Hb-oxygen affinity., (© 2019 Global Blood Therapeutics. British Journal of Clinical Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Background: Danshen is a first-line traditional Chinese medicine derived from Salvia miltiorrhiza Bunge consisting mainly of tanshinone IIA, tanshinol, protocatechuic aldehyde, and salvianolic acid B, it is widely used to treat cardiovascular diseases based on the synergistic effect of its multiple active components. Recent studies have indicated that the overall effect of traditional Chinese medicine is closely related to the in vivo coexistence of a variety of active components., Hypothesis: The prolongation of the coexistence of the four active components in Danshen in vivo by regulating their pharmacokinetic processes may contribute to better efficiency., Methods/study Designs: Individual sustained-release pellets of the four main active components in Danshen were respectively prepared according to the optimised formulations developed in our previous studies to modulate their in vivo processes, in which the desired release profiles of each kind of sustained-release pellets for formulation optimisation were calculated based on the point-area deconvolution and circadian rhythm of variant angina. The four kinds of sustained-release pellets were filled into capsules on the basis of the original weight ratio of the four active components in purified Salvia miltiorrhiza extract for further in vitro release and pharmacokinetic and pharmacodynamic investigations., Results: The release behaviours of the combined Danshen capsules composed of the four kinds of sustained-release pellets were evaluated in three media with different pH levels (pH 1.2, 6.8, and pure water). The release profiles of each kind of sustained-release pellets in pH 6.8 PBS and pH 1.2 HCl were similar to the release profile of those in pure water (similarity factors f 2  > 50). Pharmacokinetic studies revealed that the four kinds of sustained-release pellets in the combined Danshen capsules possessed the same T max and similar and extended MRT. Moreover, pharmacodynamic studies indicated that the combined Danshen capsules had much better anti-angina effects than commercial Danshen capsules according to comprehensive evaluations via electrocardiogram, serum index (CK-MB, cTn-I, ET, and NO), myocardial oxidative damage, and myocardial pathologic biopsy., Conclusion: Sustained-release preparations can markedly prolong the in vivo coexistence of multiple components in Danshen to enhance their overall effects, which provides a potent strategy for developing the combination therapy of traditional Chinese medicine., (Copyright © 2018. Published by Elsevier GmbH.)

New treatments directly targeting polymerization of sickle hemoglobin (HbS), the proximate event in the pathophysiology of sickle cell disease (SCD), are needed to address the severe morbidity and early mortality associated with the disease. Voxelotor (GBT440) is a first-in-class oral therapy specifically developed to treat SCD by modulating the affinity of hemoglobin (Hb) for oxygen, thus inhibiting HbS polymerization and downstream adverse effects of hemolytic anemia and vaso-occlusion. GBT440-001 was a phase 1/2 randomized, double-blind, placebo-controlled, single and multiple ascending dose study of voxelotor in adult healthy volunteers and patients with SCD, followed by a single-arm, open-label extension study. This report describes results of voxelotor (500-1000 mg per day) in patients with sickle cell anemia. The study evaluated the safety, tolerability, pharmacokinetic, and pharmacodynamic properties of voxelotor and established proof of concept by improving clinical measures of anemia, hemolysis, and sickling. Thirty-eight patients with SCD received 28 days of voxelotor 500, 700, or 1000 mg per day or placebo; 16 patients received 90 days of voxelotor 700 or 900 mg per day or placebo. Four patients from the 90-day cohort were subsequently enrolled in an extension study and treated with voxelotor 900 mg per day for 6 months. All patients who received multiple doses of voxelotor for ≥28 days experienced hematologic improvements including increased Hb and reduction in hemolysis and percentage of sickled red cells, supporting the potential of voxelotor to serve as a disease-modifying therapy for SCD. Voxelotor was well tolerated with no treatment-related serious adverse events and no evidence of tissue hypoxia. These trials were registered at www.clinicaltrials.gov as #NCT02285088 and #NCT03041909., (© 2019 by The American Society of Hematology.)

Infections represent one of the most frequent causes of arthroplasty revision. Thus, design of new antimicrobial scaffolds to reduce implant rejections, bone infections and associated medical costs is highly desired. In recent years, essential oil components (EOCs) have merged as compounds with significant antimicrobial activity that can be attached to specific surfaces to enhance and prolong their antimicrobial effect. Herein calcium phosphate CaP regenerative materials have been coated with a vanillin derivative to combine its original bone regeneration properties with antimicrobial action of EOCs. Materials in form of microparticles and blocks were prepared and fully characterized. Clonogenic viability tests demonstrated that low concentrations of material (10 mg·mL -1 ) resulted effective to kill 100% of E. coli DH5α bacteria. Additionally, vanillin containing scaffolds did not display any toxic effect over cells, yet they preserve the ability to express alkaline phosphatase (ALPL), collagen type 1, chain α1 (COL1A1) and bone gamma-carboxyglutamic acid-containing protein or osteocalcin (BGLAP), which are genes typically expressed by osteoblasts. These results demonstrate that commercially available scaffolds can be functionalized with EOCs, achieving antimicrobial activity and open up a new approach for the treatment and prevention of infection. STATEMENT OF SIGNIFICANCE: During the last years, the interest in bone regenerative materials with antibiotic properties has increased, since prosthesis infection is one of the most usual complications in implant surgery. In this work, we report a hybrid system composed by a calcium phosphate material (powders and scaffolds) functionalized with the derivative of an essential oil component (EOC). Our purpose was to provide the calcium phosphate material with antimicrobial activity without harming its bone regenerative capability. The obtained results were encouraging, which opens up the possibility of developing new modified materials for the prevention and treatment of bone infection., (Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

The three analytes of the Traditional Chinese Medicine ZibuPiyin Recipe (ZBPYR), namely, liquiritin, protocatechuic aldehyde and rosmarinic acid, may synergistically play an important role in regulating memory and learning. However, the pharmacokinetic behaviors of these compounds after their co-administration remain unclear. To this end, a selective and sensitive ultra-performance liquid chromatography-tandem mass spectrometry method was developed and validated in rat plasma for the study of these three major bioactive ingredients in ZBPYR. The analytes in the plasma samples were separated on a Shiseido Capcell core C18 column using bendrofluazide as an internal standard, with a gradient mobile phase system of acetonitrile-water containing 0.1% formic acid. Electrospray ionization in the negative-ion mode and multiple reaction monitoring were used to identify and quantify the three analytes. All of the calibration curves showed good linearity (r > 0.992) over the concentration range, with a lower limit of quantification of 5 ng/mL. The precision of the analytical method was evaluated by intra- and inter-day assays, and the percentage of relative standard deviation (SD) was within 15%. Satisfactory extraction efficiency (between 83.4 and 99.4%) and matrix effects (76.4-107.4) were obtained by liquid-liquid extraction. The pharmacokinetic results showed that the three bioactive ingredients were rapidly absorbed and had a short terminal half-life in rats after oral administration of ZibuPiyin recipe. This UPLC-MS-MS study method used in this study may be useful for assessing the pharmacokinetic characteristics of various compounds, which would be helpful in determining their clinical potential., (© The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

A sensitive, specific and accurate liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous determination of seven constituents of the Zaoren Anshen prescription (ZAP) in rat plasma after oral administration of the ZAP: spinosin, salvianic acid A, 6'''-feruloylspinosin, protocatechualdehyde, salvianolic acid B, schisandrin and deoxyschisandrin. The plasma samples and the internal standard (IS) sulfamethoxazole were extracted using acetonitrile. Chromatographic separation was performed with an Agilent HC-C 18 column using a gradient elution profile and a mobile phase consisting of 0.01% formic acid in water (A) and acetonitrile (B). The analytes were quantified simultaneously in a single run using an ion trap mass spectrometer operated in the multiple reaction monitoring mode and electrospray ion-source polarity in the positive and negative modes. The calibration curves for spinosin, salvianic acid A, 6'''-feruloylspinosin, protocatechualdehyde, salvianolic acid B, schisandrin and deoxyschisandrin were linear over the concentration ranges of 2.90-1160, 2.50-1000, 1.80-720, 0.65-260, 2.50-1000, 8.00-1600 and 1.30-520 ng/mL, respectively. The intra- and inter-day precisions in terms of relative standard deviation were <18.9%, and the accuracies in terms of relative error were within ±14.2%. Consequently, the proposed method was successfully applied to the pharmacokinetic analysis of these seven major active compounds in rats administered ZAP. These results will facilitate research aiming to predict the effectiveness of the optimal dose of ZAP and might be beneficial for the therapeutic use of ZAP in the clinical setting., (Copyright © 2017 John Wiley & Sons, Ltd.)

Background: Protocatechuic aldehyde (PAL) and hydroxysafflor yellow A (HSYA) are 2 effective ingredients of Danhong Injection, which is extensively used for the clinical treatment of cardio-cerebrovascular diseases. This study aims to investigate the pharmacokinetic differences between single and combined medication of PAL and HSYA and analyze the interaction of the above effective components in hyperlipidemia rats., Methods: Thirty male SD rats were randomly divided into the control group (n = 6) and the model group (n = 24). The hyperlipidemia model was established by feeding with superfatted forage. The successful model rats were then randomly divided into the PAL group (16 mg/kg), the HSYA group (10 mg/kg), and the combination group (16 mg/kg + 10 mg/kg). Administration through tail-vein, and orbital blood was sampled at different time points. The mass concentration of PAL and HSYA was determined by high performance liquid chromatography (HPLC-DAD). Analysis of pharmacokinetic parameters was conducted by using DAS 3.2.6 software and SPSS 19.0 statistical analysis software., Results: According to the parameters of statistical moment of non-compartmental model, there was a significant difference in plasma clearance (CL) between the PAL group and the drug combination group (p < 0.01), as well as in the area under the first moment of the plasma concentration-time curve and the elimination half-life (t1/2) between the HSYA group and the drug combination group (p < 0.01) but no obvious differences about the blood concentration time curve area, the average dwell time (MRT), and the peak concentration (Cmax; p > 0.05)., Conclusion: The combined medication of PAL and HSYA could increase the plasma CL significantly and have a great influence on the absorption of HSYA in rats with hyperlipidemia., (© 2018 S. Karger AG, Basel.)

Adaptation to hypoxia requires compensatory mechanisms that affect O 2 transport and utilization. Decreased hemoglobin (Hb) O 2 affinity is considered part of the physiological adaptive process to chronic hypoxia. However, this study explores the hypothesis that increased Hb O 2 affinity can complement acute physiological responses to hypoxia by increasing O 2 uptake and delivery compared with normal Hb O 2 affinity during acute severe hypoxia. To test this hypothesis, Hb O 2 affinity in mice was increased by oral administration of 2-hydroxy-6-{[(2 S )-1-(pyridine-3-carbonyl)piperidin-2yl] methoxy}benzaldehyde (GBT1118; 70 or 140 mg/kg). Systemic and microcirculatory hemodynamics and oxygenation parameters were studied during hypoxia in awake-instrumented mice. GBT1118 increased Hb O 2 affinity and decreased the Po 2 at which 50% of Hb is saturated with O 2 (P 50 ) from 43 ± 1.1 to 18.3 ± 0.9 mmHg (70 mg/kg) and 7.7 ± 0.2 mmHg (140 mg/kg). In a dose-dependent fashion, GBT1118 increased arterial O 2 saturation by 16% (70 mg/kg) and 40% (140 mg/kg) relative to the control group during 5% O 2 hypoxia. In addition, a GBT1118-induced increase in Hb O 2 affinity reduced hypoxia-induced hypotension compared with the control group. Moreover, microvascular blood flow was higher during hypoxia in GBT1118-treated groups than the control group. The increased O 2 saturation and improved blood flow in GBT1118-treated groups preserved higher interstitial tissue Po 2 than in the control group during 5% O 2 hypoxia. In conclusion, increased Hb O 2 affinity enhanced physiological tolerance to hypoxia, as evidenced by improved hemodynamics and tissue oxygenation. Therefore, pharmacologically induced increases in Hb O 2 affinity become a potential therapeutic approach to improve tissue oxygenation in pulmonary diseases characterized by severe hypoxemia. NEW & NOTEWORTHY This study establishes that pharmacological modification of hemoglobin O 2 affinity can be a promising and novel therapeutic strategy for the treatment of hypoxic hypoxia and paves the way for the clinical development of molecules that prevent hypoxemia., (Copyright © 2017 the American Physiological Society.)

Darapladib, a lipoprotein-associated phospholipase A2 (Lp-PLA2) inhibitor, failed to demonstrate efficacy for the primary endpoints in two large phase III cardiovascular outcomes trials, one in stable coronary heart disease patients (STABILITY) and one in acute coronary syndrome (SOLID-TIMI 52). No major safety signals were observed but tolerability issues of diarrhea and odor were common (up to 13%). We hypothesized that genetic variants associated with Lp-PLA2 activity may influence efficacy and tolerability and therefore performed a comprehensive pharmacogenetic analysis of both trials. We genotyped patients within the STABILITY and SOLID-TIMI 52 trials who provided a DNA sample and consent (n = 13,577 and 10,404 respectively, representing 86% and 82% of the trial participants) using genome-wide arrays with exome content and performed imputation using a 1000 Genomes reference panel. We investigated baseline and change from baseline in Lp-PLA2 activity, two efficacy endpoints (major coronary events and myocardial infarction) as well as tolerability parameters at genome-wide and candidate gene level using a meta-analytic approach. We replicated associations of published loci on baseline Lp-PLA2 activity (APOE, CELSR2, LPA, PLA2G7, LDLR and SCARB1) and identified three novel loci (TOMM5, FRMD5 and LPL) using the GWAS-significance threshold P≤5E-08. Review of the PLA2G7 gene (encoding Lp-PLA2) within these datasets identified V279F null allele carriers as well as three other rare exonic null alleles within various ethnic groups, however none of these variants nor any other loci associated with Lp-PLA2 activity at baseline were associated with any of the drug response endpoints. The analysis of darapladib efficacy endpoints, despite low power, identified six low frequency loci with main genotype effect (though with borderline imputation scores) and one common locus (minor allele frequency 0.24) with genotype by treatment interaction effect passing the GWAS-significance threshold. This locus conferred risk in placebo subjects, hazard ratio (HR) 1.22 with 95% confidence interval (CI) 1.11-1.33, but was protective in darapladib subjects, HR 0.79 (95% CI 0.71-0.88). No major loci for tolerability were found. Thus, genetic analysis confirmed and extended the influence of lipoprotein loci on Lp-PLA2 levels, identified some novel null alleles in the PLA2G7 gene, and only identified one potentially efficacious subgroup within these two large clinical trials.

To study the pharmacokinetics-pharmacodynamics correlation of protocatechuic aldehyde and hydroxysafflor yellow A alone or their combination use in rats with hyperlipidemia. In this study, the hyperlipidemia model was established by intravenous injection of protocatechuic aldehyde (20 mg•kg⁻¹) and hydroxysafflor yellow A (12 mg•kg⁻¹). The HPLC-DAD method was applied to determine the plasma concentration of protocatechuic aldehyde and hydroxysafflor yellow A at different time points and draw the drug effect-time curve. Meanwhile, the platelet activating factors (PAF) and plasma a granule membrane protein (GMP-140) contents were determined at different time points to draw the time-effect curve. Then DAS 3.2.6 software was used to process the data, analyze their correlation, and compare the difference of pharmacokinetics and pharmacodynamics of protocatechuic aldehyde and hydroxysafflor yellow A in hyperlipidemia rats after alone or their combined application, so as to evaluate the effect of protocatechuic aldehyde and hydroxysafflor yellow A on hyperlipidemia rats. According to the result, the pharmacokinetics and pharmacodynamics process of protocatechuic aldehyde and hydroxysafflor yellow A in hyperlipidemia rats after alone or their combination were consistent to the three-compartment model. In model group, the plasma PAF and GMP-140 were significantly increased, and the PAF and GMP-140 in vivo contents were decreased in a certain time after treatment. The effects of protocatechuic aldehyde and hydroxysafflor yellow A against the pharmacodynamic action may be related with their level in vivo, and their plasma concentration was positively related to the PAF and GMP-140 contents. The pharmacodynamic indexes were better after the combined use of protocatechuic aldehyde and hydroxysafflor yellow A, with certain influence on each other in hyperlipidemia rats; at the same time, it also reflected the rationality of protocatechuic aldehyde and hydroxysafflor yellow A combined application., Competing Interests: The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose., (Copyright© by the Chinese Pharmaceutical Association.)

Scope: Silicon (Si) is one of the most abundant trace elements in the body. Although pharmacokinetics data described its absorption from the diet and its body excretion, the mechanisms involved in the uptake and transport of Si across the gut wall have not been established., Methods and Results: Caco-2 cells were used as a well-accepted in vitro model of the human intestinal epithelium to investigate the transport, across the intestinal barrier in both the absorption and excretion directions, of Si supplied as orthosilicic acid stabilized by vanillin complex (OSA-VC). The transport of this species was found proportional to the initial concentration and to the duration of incubation, with absorption and excretion mean rates similar to those of Lucifer yellow, a marker of paracellular diffusion, and increasing in the presence of EGTA, a chelator of divalents cations including calcium. A cellular accumulation of Si, polarized from the apical side of cells, was furthermore detected., Conclusion: These results provide evidence that Si, ingested as a food supplement containing OSA-VC, crosses the intestinal mucosa by passive diffusion via the paracellular pathway through the intercellular tight junctions and accumulates intracellularly, probably by an uptake mechanism of facilitated diffusion. This study can help to further understand the kinetic of absorption of Si., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The present study was aimed to develope a proliposomal formulation to decrease the hepatic first-pass metabolism of protocatechualdehyde (PD), followed by pellet coating to modify the drug release for angina chronotherapy. PD proliposomes were prepared by depositing PD-phospholipid complex on mannitol powders to improve the drug encapsulation. Afterwards, the PD proliposomes were prepared into pellet cores via extrusion-spheronization using 10% κ-carrageenan as pelletization aid prior to the development of PD sustained-release pellets (PD-SRPs). Eudragit® NE 30D was chosen as coating material and the desired drug release profile of PD-SRPs was calculated for formulation optimization by deconvolution based on the circadian rhythm of variant angina. A high similarity factor (f2=85.72) was achieved when the coating weight was 30% and the sustained release behavior also prevented the destruction of liposomes by gastric fluids. Pharmacokinetic studies revealed a basically consistent trend between the actual and the predicted plasma concentration-time curve with absolute percent errors (%PE) of concentrations <10% in 2-12h. Meanwhile, a relative bioavailability of 200% was achieved compared with pure PD. Therefore, the development of proliposomes-based PD-SRPs was an effective strategy to provide both improved oral bioavailability and desired drug plasma concentration-time course for angina chronotherapy., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Salvianolic acids, the well-known active components in Salvia miltiorrhiza, have been shown to possess markedly pharmacological activities. However, due to the complex in vivo course after administration, the pharmacologically active forms are still poorly understood. In present study, we evaluated the stability of eight major salvianolic acids from Danshen extract under different chemical and physiological conditions. We also quantitatively explained the absorption, metabolism and excretion of these salvianolic acids in rats after gastric-administration, which was carried out by simultaneously determining the amounts of salvianolic acids and their metabolites in the rat gastrointestinal contents, gastrointestinal mucosa, plasma, bile and urine. We found that: 1) protocatechuic aldehyde (PAL) was much stable whether in acidic environment (pH4.0) or in alkaline environment (pH8.0), while other salvianolic acids were stable in acidic environment and instable in alkaline environment; 2) PAL, salvianoli acid A (SAA) and salvianolic acid B (SAB) were instable whether in rat stomach or in small intestine, while other salvianolic acids were stable in rat stomach and instable in small intestine; 3) after gastric-administration, except PAL and Danshensu (DSS), other phenolic acids would be metabolized into DSS and caffeic acid (CA) in the rat gastrointestinal tract before absorption, and only free and glucuronidated PAL, CA and DSS were detected in rat plasma, bile and urine. In conclusion, it was the free and glucuronidated PAL, CA and DSS rather than the prototypes of other salvianolic acids that were present in plasma with considerable concentrations after gastric-administration., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Although exertional dyspnea and worsening hypoxia are hallmark clinical features of idiopathic pulmonary fibrosis (IPF), no drug currently available could treat them. GBT1118 is a novel orally bioavailable small molecule that binds to hemoglobin and produces a concentration-dependent left shift of the oxygen-hemoglobin dissociation curve with subsequent increase in hemoglobin-oxygen affinity and arterial oxygen loading. To assess whether pharmacological modification of hemoglobin-oxygen affinity could ameliorate hypoxemia associated with lung fibrosis, we evaluated GBT1118 in a bleomycin-induced mouse model of hypoxemia and fibrosis. After pulmonary fibrosis and hypoxemia were induced, GBT1118 was administered for eight consecutive days. Hypoxemia was determined by monitoring arterial oxygen saturation, while the severity of pulmonary fibrosis was assessed by histopathological evaluation and determination of collagen and leukocyte levels in bronchoalveolar lavage fluid. We found that hemoglobin modification by GBT1118 had strong antihypoxemic therapeutic effects with improved arterial oxygen saturation to near normal level. Moreover, GBT1118 treatment significantly attenuated bleomycin-induced lung fibrosis, collagen accumulation, body weight loss, and leukocyte infiltration. This study is the first to suggest the beneficial effects of hemoglobin modification in fibrotic lungs and offers a promising and novel therapeutic strategy for the treatment of hypoxemia associated with chronic fibrotic lung disorders in human, including IPF., (© 2016 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.)

In order to provide scientific basis for clinical selection of drugs, to compare and analyze the effective constitutes and the intestinal absorption in vivo in rats of the compound salvia tablets and compound salvia dropping pills (taken as the representatives). Determine the contents of tanshinol, protocatechuic aldehyde, salvianolic acid B and tanshinone II A, cryptotanshinone, ginseng saponin Rg1 and Rb1 in the compound salvia tablets and compound salvia dropping pills by High Performance Liquid Chromatography (HPLC). The intestinal absorption condition of the tanshinol, protocatechuic aldehyde, salvianolic acid B of the compound salvia tablets and compound salvia dropping pills in rats were detected by intestinal perfusion experiment. Only the intake of protocatechuic aldehyde in the compound salvia tablets was higher than in the compound dropping pills, the intake of the other 6 effective constitutes were all lower than in the compound dropping pills. The intestinal absorption of protocatechuic aldehyde was rather complete, while the intestinal absorption of tanshinol and salvianolic acid B were not significant. The duodenum was the main absorption region of these three components. The absorption of protocatechuic aldehyde was different in different regions of the intestines. Each intake of the effective constitutes in the tablets and dropping pills were significantly different, and the rat intestinal absorption of part of the components were different.

A very simple and selective high-performance liquid chromatography electrospray ionization tandem mass spectrometry (LC-MS-MS) method was developed for simultaneous determination and pharmacokinetic study of protocatechuic aldehyde (PAL) and its active metabolite protocatechuic acid (PCA). The method involves a simple liquid-liquid extraction with ethyl acetate. The separation was performed on a Hypersil GOLD C18 column (2.1 × 150 mm, 3.0 µm; particle, Thermo, USA) with isocratic elution using a mobile phase consisted of methanol and water (containing 0.1% formic acid) at a flow rate of 0.2 mL/min. The detection of target compounds was done by using low-energy collision dissociation tandem mass spectrometry (CID-MS-MS) using the selective reaction monitoring scan mode. The method was linear for all analytes over the investigated range for all correlation coefficients greater than 0.9950. The lower limits of quantification were 2.0 ng/mL for PAL and PCA. The intra- and interday precisions (relative standard deviation, RSD %) were <6.84 and 5.54%, and the accuracy (relative error, RE %) was between -2.85 and 0.74% (n = 6). The developed method was applied to study the pharmacokinetics of PAL and its major active metabolite PCA in rat plasma after oral and intravenous administration of PAL., (© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background and Objectives: Darapladib is a lipoprotein-associated phospholipase A2 (Lp-PLA2) inhibitor. This study evaluated the pharmacokinetics, pharmacodynamics and safety of darapladib in healthy Chinese subjects., Methods: Twenty-four subjects received darapladib 160 mg orally, approximately 1 hour after a standard breakfast, as a single dose and once daily for 28 days. Non-compartmental methods were used to determine the single and multiple dose pharmacokinetics of darapladib and its metabolite SB-553253. Repeat dose Lp-PLA2 activity and safety were evaluated., Results: Systemic exposure (AUC(0-T), Cmax geometric mean (CVb%)) of darapladib was higher after multiple-dosing (519 ng.h/mL (33.3%), 34.4 ng/mL (49.9%)) compared to single-dose administration (153 ng.h/mL (69.0%), 17.9 ng/mL (55.2%). The steady-state accumulation ratio was less than unity (Rs = 0.80), indicating time-dependent pharmacokinetics of darapladib. Darapladib steady-state was reached by Day 14 of once daily dosing. Systemic exposure to SB-553253 was lower than darapladib with median (SB-553253: darapladib) ratios for AUC(0-τ) of 0.0786 for single dose and 0.0532 for multiple dose administration. On Day 28, pre-dose and maximum inhibition of Lp-PLA2 activity was approximately 70% and 75% relative to the baseline value, respectively and was dependent of darapladib concentration. The most common adverse events (≥ 21% subjects) were abnormal faeces, abnormal urine odour, diarrhoea and nasopharyngitis., Conclusion: Darapladib 160 mg single and repeat doses were profiled in healthy Chinese subjects. Single dose systemic exposure to darapladib in healthy Chinese subjects was consistent with that observed previously in Western subjects whereas steady-state systemic exposure was approximately 65% higher in Chinese than Western subjects. The Lp-PLA2 activity and adverse event profile were similar in healthy Chinese and previous reports in Western subjects. Ethnic-specific dose adjustment of darapladib is not considered necessary for the Chinese population., Trial Registration: ClinicalTrials.gov NCT02000804.

Aim: Darapladib is a potent and reversible orally active inhibitor of lipoprotein-associated phospholipase A2 (Lp-PLA2 ). The aim of the study was to assess the effects of severe renal impairment on the pharmacokinetics and safety/tolerability of darapladib compared with normal renal function., Methods: This was an open label, parallel group study of darapladib following 10 day once daily 160 mg oral dosing in subjects with normal (n = 8) and severe renal impairment (estimated glomerular filtration rate <30 ml min(-1) 1.73 m(-2) , n = 8). Plasma concentrations of total and unbound darapladib as well as total darapladib metabolites were determined in samples obtained over 24 h on day 10., Results: Plasma concentrations of total and unbound darapladib as well as all three metabolites were higher in subjects with severe renal impairment. Area under the plasma concentration vs. time curve between time zero and 24 h (AUC(0,24 h) and maximum plasma concentration (Cmax ) of total darapladib in severely renally impaired subjects were 52% and 59% higher than those in the matched healthy subjects, respectively. Similar results were found with the darapladib metabolites. Darapladib was highly plasma protein bound with 0.047% and 0.034% unbound circulating in plasma in severely renally impaired and healthy subjects, respectively. Unbound plasma darapladib exposures were more than two-fold higher in severely renally impaired subjects than in healthy controls. Adverse events (AE) were reported in 38% of healthy subjects and 75% of severely renally impaired subjects, most of which were mild or moderate in intensity., Conclusions: The results of this study showed that darapladib exposure was increased in subjects with severe renal impairment compared with healthy controls. However, darapladib was generally well tolerated in both groups., (© 2015 The British Pharmacological Society.)

A rapid and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed for the simultaneous determination of the four major active ingredients, danshensu, protocatechuic aldehyde, rosmarinic acid, and ligustrazine, in the traditional Chinese medicine Shenxiong glucose injection in rat plasma. Acidified and alkalized plasma samples were extracted using ethyl acetate, and separated on a Waters C18 column (2.1mm×50mm, 1.7μm) by using a gradient mobile phase system of acetonitrile-water containing 0.1% formic acid and luteoloside as an internal standard. Electrospray ionization in the positive-ion mode and multiple reaction monitoring were used to identify and quantitate the active components. All calibration curves showed good linearity (r>0.994) over the concentration range, with a lower limit of quantification (LLOQ) between 0.02 and 0.21μg/mL. The precision of the in vivo study was evaluated by intra- and inter-day assays, and the percentage of relative standard deviation was within 15%. Moreover, satisfactory extraction efficiency was obtained (between 83.94 and 117.81%) by liquid-liquid extraction. The validated method was successfully applied in a pharmacokinetic study in rats after intravenous administration of Shenxiong glucose injection. The results showed that the four bioactive ingredients in Shenxiong glucose injection have linear pharmacokinetic properties in rats after intravenous injection within the administered dose range and partially different ones compared to single ingredient., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Purpose: To investigate the potential of lipoprotein-associated phospholipase A2 inhibition as a novel mechanism to reduce edema and improve vision in center-involved diabetic macular edema (DME)., Design: Prospective, multicenter, randomized, double-masked, placebo-controlled phase IIa study., Participants: Fifty-four center-involved DME patients randomized 2:1 to receive darapladib (n = 36) or placebo (n = 18)., Methods: Darapladib 160 mg or placebo monotherapy was administered orally once daily for 3 months, and patients were followed up monthly for 4 months., Main Outcome Measures: Mean change from baseline in best-corrected visual acuity (BCVA) and the center subfield and center point of the study eye at month 3 as determined by spectral-domain optical coherence tomography., Results: Five patients in the study received intravitreal anti-vascular endothelial growth factor rescue therapy before the day 90 assessment, 2 of 36 (6%) in the darapladib arm and 3 of 18 (17%) in the placebo arm. Administration of 160 mg darapladib for 3 months resulted in statistically significant mean improvements, from baseline to month 3, in BCVA of 4.1 Early Treatment Diabetic Retinopathy Study (ETDRS) letters (95% confidence interval [CI], 2.3-5.8) and of 57 μm in central subfield thickness (95% CI, -84 to -30) in the study eyes. An increase in BCVA of 1.7 ETDRS letters (95% CI, -1.0 to 4.4) and a decrease in center subfield thickness of 34 μm (95% CI, -75 to 6.8) for the placebo group were not significant. No ocular severe adverse events (SAEs) or SAEs considered related to darapladib were reported. One SAE of myocardial infarction, not considered related to darapladib, was reported, and 1 SAE of severe diarrhea was reported in a placebo patient, subsequently withdrawn from the study. Study eye ocular adverse events (AEs) and nonocular AEs were similar between treatment groups., Conclusions: Once-daily oral darapladib administered for 3 months demonstrated modest improvements in vision and macular edema that warrant additional investigation of this novel lipoprotein-associated phospholipase A2 inhibitory mechanism for the treatment of DME., (Copyright © 2015 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

A simple, sensitive and reliable gradient elution high performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI-MS) method was developed for quantifying helicid in dog plasma. The limit of detection (LOD) and the lower limit of quantitation (LLOQ) were 0.3 and 1 ng/mL, respectively. This method was validated for selectivity, linearity, accuracy and precision, extraction recoveries, matrix effects, carry-over, cross-talk, dilution integrity, stability and incurred sample reanalysis (ISR). Bioavailability and pharmacokinetic parameters of helicid in beagle dogs were researched from a two period crossover design study. After intravenous administration (i.v.), helicid had a mean (± SD) AUC0-∞ of 12062.06 ± 2482.69 ng/mL h and terminal half-life (t1/2 z) of 2.91 ± 1.37 h, while Cmax was 35613.23 ± 8157.18 ng/mL. Following intragastric gavage administration (i.g.), AUC0-∞ was 7589.16 ± 1797.20 ng/mL h along with a longer t1/2 z of 4.10 ± 4.35 h. Cmax was researched at 0.58 ± 0.20 h. The absolute bioavailability (F) of helicid was 15.74 ± 1.87%., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Purpose: DMS612 is a dimethane sulfonate analog with bifunctional alkylating activity and preferential cytotoxicity to human renal cell carcinoma (RCC) in the NCI-60 cell panel. This first-in-human phase I study aimed to determine dose-limiting toxicity (DLT), maximum tolerated dose (MTD), pharmacokinetics (PK), and pharmacodynamics (PD) of DMS612 administered by 10-minute intravenous infusion on days 1, 8, and 15 of an every-28-day schedule., Experimental Design: Patients with advanced solid malignancies were eligible. Enrollment followed a 3+3 design. PKs of DMS612 and metabolites were assessed by mass spectroscopy and PD by γ-H2AX immunofluorescence., Results: A total of 31 patients, including those with colorectal (11), RCC (4), cervical (2), and urothelial (1) cancers, were enrolled. Six dose levels were studied, from 1.5 mg/m(2) to 12 mg/m(2). DLTs of grade 4 neutropenia and prolonged grade 3 thrombocytopenia were observed at 12 mg/m(2). The MTD was determined to be 9 mg/m(2) with a single DLT of grade 4 thrombocytopenia in 1 of 12 patients. Two patients had a confirmed partial response at the 9 mg/m(2) dose level, in renal (1) and cervical (1) cancer. DMS612 was rapidly converted into active metabolites. γ-H2AX immunofluorescence revealed dose-dependent DNA damage in both peripheral blood lymphocytes and scalp hairs., Conclusions: The MTD of DMS12 on days 1, 8, and 15 every 28 days was 9 mg/m(2). DMS612 appears to be an alkylating agent with unique tissue specificities. Dose-dependent PD signals and two partial responses at the MTD support further evaluation of DMS612 in phase II trials., (©2014 American Association for Cancer Research.)