Safety, tolerability, and pharmacokinetics of TG-1000, a new molecular entity against influenza virus: first-in-human study.

Background: The cap-snatching mechanism of influenza virus mRNA transcription is strongly suppressed by TG-1000, a prodrug rapidly metabolized into TG-0527, is a potent cap-dependent nucleic acid endonuclease inhibitor. Herein, we aimed to assess the safety, tolerability, and pharmacokinetics of TG-1000 in healthy participants and the effect of food on the pharmacokinetics and safety of TG-1000. Method: The study was divided into 2 parts: Part A [Single Ascending-Dose (SAD) study, 10-160 mg] and Part B [Food-Effect (FE) study, 40 mg] were launched sequentially. The study included 66 participants for both investigations. We administered different TG-1000 capsules or placebo doses per the study protocol and collected blood samples for pharmacokinetic assessments at specific times. In plasma, TG-1000 and its active metabolite TG-0527 were assayed, and PK parameters were determined. Results: In SAD, the increase in AUC was less than the proportional increase in dose over the 20-160 mg dose range, while the increase in Cmax was proportional to the increase in dose. In the 10-160 mg dose range, T1/2, λz and Tmax of TG-0527 were dose-independent; and T1/2 and Tmax were within 33.8-39.4 h and 3.02-6 h, respectively. In FE, the AUC0-inf, AUC0-last, and Cmax of TG-0527 decreased by approximately 17.52%, 18.76%, and 41.35%, respectively, and the Tmax delay was around 1.50 h. No serious adverse events occurred during the studies. Conclusion: Overall, TG-1000 was well tolerated and exhibited an acceptable safety and PK profile, supporting further clinical investigation of TG-1000 for the treatment of influenza.

Pharmacokinetics of Nifedipine-Sustained Release Tablets in Healthy Subjects After a Single Oral Administration: Bioequivalence Analysis and Food Effects.

We compared newly developed delayed-release oral tablets (test) of 30-mg nifedipine (NFP) with its marketed counterpart (30 mg; reference) in healthy adult Chinese volunteers to assess the former's bioequivalence. This was a randomized, open-label, four-period, crossover trial study including fasting and fed trials. The participants were randomly administered test or reference formulations (1:1 ratio) throughout each period, with a 7-day washout period. In the next session, they were administered the alternate products. Liquid chromatography-tandem mass spectrometry and WinNonlin software were used to evaluate the bioequivalence of the maximum plasma concentration (Cmax ) of NFP and the area under the concentration-time curve (AUC). In total, 46 and 48 people participated in the fasting and postprandial trials. In both groups, the 90% confidence intervals of geometric mean ratios of Cmax , AUC from time zero to time t, and AUC from time zero to infinity were in the equivalence range (80%-125%). When NFP was administered concomitantly with a high-fat meal, time to maximum concentration was approximately twofold earlier, absorption was approximately 4.8% less, and Cmax exhibited a slight change relative to those under fasting conditions. Moreover, no serious adverse events were recorded in the participants. The present findings confirm the bioequivalence of test and reference formulations of NFP tablets under fasting and postprandial conditions.

Pharmacokinetics of Amoxicillin and Clavulanate Potassium for Suspension (200 mg/28.5 mg) in Healthy Subjects: Sample Add Stabilizer Study and Food Effects.

The present study compares the pharmacokinetics of amoxicillin and clavulanate potassium suspension (200 mg/28.5 mg) during fasting and postprandial conditions, and the sample adds a stabilizer study. Two randomized, crossover trials were conducted in an open-label, single-center study (a fasting trial and a postprandial trial). In each part of the study, the subjects were randomly assigned to receive either test or reference products (200 mg/28.5 mg) in a 1:1:1 ratio, followed by the alternative products after a 7-day washout period. Plasma amoxicillin and clavulanic acid concentrations were analyzed by liquid chromatography-tandem mass spectrometry. WinNonlin software was used to evaluate the pharmacokinetic parameters (noncompartmental model). The formulations were considered bioequivalent if the geometric means of area under the plasma concentration-time curve (AUC) and maximum plasma concentration (Cmax ) of amoxicillin and clavulanic acid were within the predetermined bioequivalence range established by average bioequivalence (ABE) or reference-scaled ABE. Tolerability was assessed throughout the study. The postprandial trial and the fasting study each had 12 volunteers. Under fasting and postprandial conditions, the 90%CI for the ratio of geometric means of amoxicillin of Cmax , AUC from time 0 to the last measurable concentration, and AUC from time 0 to infinity were within the ABE acceptance limits (80%-125%); the geometric means of clavulanic acid of Cmax (critbound, -0.03; point estimate, 1.07) were within the reference-scaled ABE acceptance limits, and the AUC from time 0 to the last measurable concentration and AUC from time 0 to infinity were within the ABE acceptance limits (80%-125%). Time to maximum concentration of amoxicillin was delayed 1.0 hour with high-fat meals compared to fasting conditions. Meantime, high-fat meals decreased the exposure of clavulanic acid by nearly 40%. No serious adverse events were found among the subjects. The bioequivalence of test and reference amoxicillin and clavulanate potassium for suspension was validated in this study under fasting and postprandial conditions.

Pharmacokinetics of Esomeprazole Magnesium After Single Oral Doses in Healthy Subjects: Bioequivalence Study and Food Effects.

This study was designed to evaluate the bioequivalence of the newly developed delayed-release oral suspension (test) 40 mg esomeprazole magnesium compared to its marketed counterpart (40 mg; reference) in healthy adult Chinese subjects. We conducted randomized, open-label, two-period, single-dose, two-way crossover trials over a 7-day washout period, comprising a fasting trial and a fed trial. The subjects were administered the test or reference products in a 1:1 ratio at random throughout each period. Then, in the next session, they received the alternate products. Liquid chromatography-tandem mass spectrometry and WinNonlin software were used to assess the bioequivalence of esomeprazole peak plasma concentration (Cmax ) and area under the concentration-time curve (AUC). Overall, 33 subjects participated in the fasting trial and 42 subjects participated in the fed trial. Under both situations, the 90% confidence interval for the ratio of geometric means of Cmax , AUC0-t , and AUC0-∞ were within equivalence ranges (80%-125%). In these trials, no severe adverse events or protocol violations were observed. Moreover, when esomeprazole was administered while fed, the tmax was delayed, and both Cmax and AUC were reduced. The results of this research suggest that the test and reference formulations were bioequivalent under fasting and fed states.

Hypoglycemic flavonoids from Selaginella tamariscina (P.Beauv.) Spring.

Six flavonoids, namely, three undescribed biflavonoids, one undescribed 8-aryl flavonoid, and two known compounds, were isolated from Selaginella tamariscina (P.Beauv.) Spring. The structures and absolute configurations of those undescribed compounds were established by NMR spectroscopy data, HRESIMS analyses and electronic circular dichroism (ECD) analyses. In addition, all the isolates were evaluated for their hypoglycemic activity in HepG2 cells. Involvenflavone H, I, and J significantly increased glucose consumption in both normal and insulin-resistant HepG2 cells. Interestingly, these three compounds can effectively upregulate the protein expression of glucokinase (GCK) and adenylate cyclases (ADCYs). These results suggested that involvenflavone H, I, and J (especially involvenflavone J) may have potent hypoglycemic activity, which also provided promising molecular targets for the treatment of diabetes.

Trichocladabiflavone A, a chalcone-flavonone type biflavonoid from Selaginella trichoclada Alsto.

A chalcone-flavonone type biflavonoid, trichocladabiflavone A (1), along with eight known biflavonoids (2-9) were isolated from the 70% EtOH extract of Selaginella trichoclada. Their structures were elucidated by extensive spectroscopic analyses. Compound 1 was the first chalcone-flavonone type biflavonoid reported in the genus Selaginella. Moreover, compound 1 exhibited moderate cytotoxicity against DU145, MCF-7 and PC3 human cancer cell lines.

Pharmacokinetics of Flunarizine Hydrochloride After Single Oral Doses in Healthy Subjects: Bioequivalence Study and Food Effects.

We designed a study to compare the newly developed 5-mg flunarizine hydrochloride capsules (test) to that of its marketed counterpart (5-mg; reference) among healthy adult Chinese volunteers. We performed an open-label, single-center study that consisted of 2 randomized, crossover trials, including a fasting trial and a fed trial. In each part of the study, the subjects were randomly assigned to either receive the test or reference products (5-mg flunarizine) in a 1:1 ratio. Subjects then received the alternative products, following a 14-day washout period. Concentrations of plasma flunarizine were analyzed using liquid chromatography-tandem mass spectrometry. Pharmacokinetic parameters (noncompartmental model) were evaluated using the WinNonlin software. The analysis of variance and Food and Drug Administration bioequivalence statistical criterion of 90% confidence interval for 80% to 125% range (set at P ≤ .05) of geometric means ratios of test: reference product for peak plasma concentration, area under the plasma concentration-time curve (AUC) from time 0 to time t, and AUC from time 0 to infinity were determined. Tolerability was evaluated during the entire study period. Overall, 23 volunteers completed the fasting study, while 40 volunteers completed the fed study. The test formulation was found to be bioequivalent to the marketed formulation, as the 90% confidence interval for the ratio of geometric means of peak plasma concentration (fasting: 87.61%-101.67%; fed: 87.38%-104.06%), AUC from time 0 to time t (fasting: 89.44%-99.92%; fed: 92.65%-98.28%), and AUC from time 0 to infinity (fasting: 95.02%-104.33%; fed: 90.41%-96.96%) were within equivalence limits (80-125%) under both the fasting and fed conditions. When flunarizine was given alongside high-fat meals, time to maximum concentration was delayed ≈3.5 hours compared to fasting conditions. Meantime, high-fat meals increased its exposure by nearly 50%. Furthermore, there were no serious adverse events found among the subjects. This study confirmed that test and reference flunarizine hydrochloride capsules were bioequivalent under fasting and postprandial conditions.

Cytotoxic effects of the biflavonoids isolated from Selaginella trichoclada on MCF-7 cells and its potential mechanism.

A new biflavonoid, (2''S)-6''-methyl-2'',3''-dihydroochnaflavone (1), along with two known ochnaflavones (2, 3), four known amentoflavones (4-7) and two known robustaflavones (8, 9) were obtained from the 70% EtOH extract of Selaginella trichoclada. The chemical structures of isolated compounds were elucidated by extensive spectroscopic analyses. Overall, compounds 1-9 displayed moderate cytotoxic effects against human breast cancer MCF-7 cell lines. Among them, compounds 2 and 8 exhibited relatively strong cytotoxic effects against MCF-7 cells with an IC50 value of 7.7 and 6.9 µΜ, respectively. The results of RNA-sequencing and KEGG functional enrichment analysis showed that 8 could induce ferroptosis in MCF-7 cells by down-regulating the expression of ferroptosis-related genes including ACSL4, NOXO1, NOXA1, ACSL5, STEAP3, LPCAT3, ATG7 and TP53. Then 8 could inhibit the expression of ACSL4 proteins through molecule docking analysis, which showed a strong interaction of - 11.89 Kcal/mol binding energy. Those results indicate that 8 could be chemotherapy agents to fight drug resistance in breast cancer by down-regulating the expression level of ACSL4 proteins via ferroptosis, which needs to be further certified in vitro.

Method development and validation of sildenafil, N-desmethylsildenafil and N1,N4-desmethylsildenafil by LC-MS/MS and its application.

This study aimed to develop an LC-MS/MS method for determining sildenafil and its metabolites N-desmethylsildenafil and N1,N4-desmethylsildenafil in human plasma and applying it to a pharmacokinetic study of sildenafil in healthy volunteers. Sildenafil-d8 was used as the internal standard. Plasma samples were pretreated via protein precipitation with acetonitrile. The extractives were then separated on an ACQUITY UPLC BEH C18 (50-mm × 2.1-mm, 1.7-µm) column using gradient elution. The aqueous and organic mobile phases were ammonium formate 2 mM supplemented with 0.1% formic acid in water and acetonitrile, respectively, and the flow rate was 0.3 mL/min. An electrospray ionization source was applied, and multiple reaction monitoring was operated in the positive mode with selective channels at m/z 475.30 â†’ 100.10, 461.20 â†’ 283.30, 483.30 â†’ 108.10, and 449.00 â†’ 283.00 for sildenafil, sildenafil-d8, N-desmethylsildenafil, and N1,N4-desmethylsildenafil, respectively. The linear calibration curves of sildenafil and its metabolites spanned 1.0-1000 ng/mL. The lower limit of quantification was 1.0 ng/mL. The extractive recovery of analytes from the biological matrix was more than 90% and the matrix effect complied with relevant provisions. The intra- and inter-day precisions of sildenafil and its metabolite were <10%. The intra- and inter-day accuracy of sildenafil, N-desmethylsildenafil, and N1,N4-desmethylsildenafil was more than 99%. The method is highly sensitive and selective, and it was successfully applied to the bioequivalence studies of 100-mg sildenafil citrate tablets in 40 healthy Chinese volunteers.

Identification of a new natural biflavonoids against breast cancer cells induced ferroptosis via the mitochondrial pathway.

Breast cancer is one of the major malignant tumors in females, and currently, recurrence and metastasis are the main obstacles preventing effective breast cancer treatment. Biflavonoids of secondary metabolites from plants are excellent anticancer agents to fight sensitive and resistant breast cancer cell lines. In this study, six C-3'-C-6″ biflavonoids, including one new robustaflavone A (1, RF-A) and five known robustaflavone derivatives (2-6), were isolated from Selaginella trichoclada for the first time. We aimed to evaluate the inhibitory effects of compounds 1-6 against human breast cancer MCF-7 cells. Among the six compounds, RF-A showed the strongest activity, decreasing cell viability with an IC50 value of 11.89 µΜ. Furthermore, RF-A strikingly induced MCF-7 nonapoptotic cell death through ferroptosis by enhancing the expression of VDAC2 channels and reducing the expression of Nedd4 E3 ubiquitin ligase, leading to lipid peroxidation and ROS production. The results suggested that RF-A has potential as a novel breast cancer treatment through its regulation of the mitochondrial VDAC2 and Nedd4 pathways.

Bioequivalence and Evaluation Parameters Based on the Pharmacodynamics of Miglitol in Healthy Volunteers.

The aim of this study was to explore the bioequivalence of miglitol based on pharmacodynamic properties. The study was performed as a single-dose, randomized, open-label, 3-period, 3-way crossover trial over a 7-day washout period. Forty-eight subjects were randomly assigned into 3 groups: (1) miglitol test formulation/sucrose coadministration, (2) miglitol reference formulation/sucrose coadministration, and (3) sucrose administration alone. Serum glucose concentrations were measured by the hexokinase detection method. The peak serum glucose concentration (Cmax ) and the area under the serum glucose concentration-time curve through 4 hours (AUC0-4h ) were used as the main pharmacodynamic parameters to evaluate bioequivalence. The 90% confidence intervals for the geometric mean ratios of Cmax and AUC0-4h were 94.81%-101.07% and 98.82%-100.72%, respectively, which were all within the bioequivalence range of 80.00%-125.00%. The test and reference formulations of miglitol were pharmacodynamically bioequivalent during the trial.

Two new biflavanoids from Selaginella trichoclada Alsto.

Two new robustaflavones, (±)-trichocladabiflavone A (1) and uncinatabiflavone E (2), along with seven known biflavanoids (3-9) were isolated from the 70% EtOH extract of Selaginella trichoclada. Their structures and absolute configurations were established by extensive spectroscopic and circular dichroism (CD) analyses. Compound 1 was resoluted into optically pure enantiomers (+)-1 and (-)-1 by chiral-phase HPLC. Moreover, compounds 1 and 2 exhibited moderate cytotoxicity against A549 and HepG2 human cancer cell lines.

Method for evaluating the human bioequivalence of acarbose based on pharmacodynamic parameters.

OBJECTIVE: To explore a method for evaluating the bioequivalence of acarbose based on pharmacodynamic parameters using a single-dose, randomized-sequence, three-way crossover study of acarbose test (T) and reference (R) formulations. METHODS: Baseline-adjusted, pre-dose value deduction, and direct comparison methods were used to evaluate the geometric T/R ratios and 90% confidence intervals (CIs) of the ln-transformed pharmacodynamic parameters to identify the most suitable evaluation system. Twelve participants were randomly divided into three groups to receive treatment in the following sequences: TRR, RTR, and RRT, each including a 7-day washout period between treatment periods. The serum glucose concentration (baseline) was determined. Pharmacodynamic parameters, including the maximum reduction in serum glucose concentrations (ΔCSG,max) and difference of the AUC of glucose between before and after acarbose exposure (ΔAUEC), were tested. RESULTS: Using the direct comparison method, the geometric mean ratios of CSG,max, AUEC(0-2h), and AUEC(0-4h) were 94.13%, 97.82% and 99.76%, respectively. The 90% CIs of the geometric T/R ratios for CSG,max, AUEC(0-2h), and AUEC(0-4h) all fell between 80% and 125%. Conversely, ΔCSG,max and ΔAUEC(0-4h) were less reliable measures of acarbose bioequivalence. CONCLUSIONS: Pre-dose value deduction and direct comparison methods can be initially considered suitable for assessing acarbose bioequivalence.

Seladelicatulasine A-G, C27 steroidal glycosides with cholinesterase inhibitory activities from Selaginella delicatula.

Seven undescribed C27 steroidal glycosides, Seladelicatulasine A-G, including six cholestanol glycosides and one spirostanol glycoside, were isolated from Selaginella delicatula. Their structures were elucidated by 1D/2D NMR spectra and HRESIMS analyses. The absolute configurations of the sugars were determined by enzymatic hydrolysis and GC/MS analyses. These cholestanol glycosides were isolated from the family Selaginellaceae for the first time. Seladelicatulasine F is characterized as a rare B-5,6-secosteroid. In addition, all the compounds were evaluated for their inhibitory activities against cholinesterase (AChE/BChE) and monoamine oxidase (MAO-A/MAO-B). These steroidal glycosides displayed selective inhibition activities on cholinesterase. Seladelicatulasine A, B and E inhibited the AChE activity with IC50 values of 0.31, 0.09, and 0.04 µM, respectively. Seladelicatulasine A and F showed the strongest inhibition activity on BChE with IC50 values of 0.37 and 0.65 µM, respectively.

Palhinosides A-H: Flavone Glucosidic Truxinate Esters with Neuroprotective Activities from Palhinhaea cernua.

Palhinosides A-H (1-8), new flavone glucosidic truxinate esters, including ß-truxinate and µ-truxinate forms, were isolated from Palhinhaea cernua. Their structures were elucidated by extensive spectroscopic methods and chemical analyses. The flavone glucoside cyclodimers possess a unique cyclobutane ring in their carbon scaffolds. Compounds 2-7 represent three pairs of stereoisomers (2/3, 4/5, 6/7). The protective effects of 1-8 against the damage of HT-22 cells induced by l-glutamate were evaluated, and compounds 4 and 5 showed better neuroprotective effects than the positive control, Trolox.

Validation of the ADVIA Centaur® XP system for the determination of insulin and its application.

In this study, a direct chemiluminescent immunoassay for the determination of human serum insulin levels using the ADVIA Centaur® XP system was validated. Dilution recovery, linearity, precision, sensitivity, between analyzer variation, reference interval and stability were analyzed. The linear range of the insulin assay was from 0.64 to 277.27 mU/L. Intra- and inter-assay coefficients of variation were 3.67-7.96% and 4.66-8.69%, respectively. The lower and upper limits of quantification were 0.61 mU/L and 8872.64 mU/L, respectively. In terms of between analyzer variation, our study showed comparable results with a good correlation of r2 = 0.9934. The human serum insulin reference interval was in the range of 3.0-25.0 mU/L. Serum insulin can be kept for 7 days between 2-8 °C and 18-26 °C, and the corresponding results for -20 °C and -70 °C were 1 month and 6 months are reported. We proved that this insulin assay was robust and the analytical performance met the requirements. We successfully applied this insulin assay to a bioequivalence study of miglitol in 48 healthy Chinese subjects. The miglitol bioequivalence study was evaluated based on pharmacokinetic and pharmacodynamic parameter endpoints. The results demonstrated that the test formulation and the reference formulation were bioequivalent.

Pictalignans D-F, three new neolignan derivatives from Selaginella picta.

Three new neolignan derivatives (1-3), together with three known isolariciresinol derivatives (4-6) were isolated from Selaginella picta. Their structures were elucidated by spectroscopic methods (1D/2D NMR, HRESIMS and CD). All isolated compounds were assayed on the neuroprotective activity against the injury of HT-22 cells induced by L-Glutamate in vitro. All compounds displayed potent protective effect on HT-22 cells.

Two new anthraquinone derivatives and one new triarylbenzophenone analog from Selaginella tamariscina.

Two new anthraquinone derivatives, selaginones A (1) and B (2), and one new triarylbenzophenone analog, selagibenzophenone B (3), were isolated from Selaginella tamariscina (Beauv.) Spring. Their structures were established by 1D-, 2D-NMR and HR-ESI-MS data. Compounds 1 and 2 represent the uncommon examples of aryl substituted anthraquinone derivatives. Especially, compound 2 is a unique anthranone with exceptional structural feature, in which a p-hydroxyphenyl moiety is attached to the C-10 position. Compound 3 is the second naturally occurring triarylbenzophenone and showed moderate activity against SMCC-7721 and MHCC97-H cell lines with IC50 values of 39.8, 51.5 µM respectively.

Implementation of a reference-scaled average bioequivalence approach for highly variable generic drug products of atorvastatin in Chinese subjects
.

OBJECTIVE: The purpose of this study was to evaluate the bioequivalence of two formulations of atorvastatin using the reference-scaled average bioequivalence (RSABE) method and to study the pharmacokinetics of atorvastatin in healthy Chinese subjects under fed conditions. MATERIALS AND METHODS: A single-dose, randomized, open-label, four-way crossover study was conducted in healthy Chinese subjects after informed consent was obtained. Healthy subjects were randomly assigned to receive 20 mg of either the test or reference formulation, following a 7-day washout period. The formulations were considered bioequivalent if 90% confidence intervals (CIs) for the ln-transformed ratios and ratio of geometric means (GMR) of AUC and Cmax of atorvastatin were within the bioequivalence range (80 - 125%). Plasma atorvastatin, ortho-hydroxy atorvastatin and para-hydroxy atorvastatin concentrations were analyzed by liquid chromatography-tandem mass spectrometry. Tolerability was assessed during the entire study period. RESULTS: ANOVA indicated that the period, sequence, and formulation had no significant effect on the pharmacokinetic parameters (p < 0.05). The test formulation was bioequivalent to the marketed formulation as the 90% CIs for natural log-transformed ratios of atorvastatin of Cmax (88.45 - 103.57%), AUC0-t (98.08 - 104.89%) and AUC0-∞ (98.15 - 104.87%) were within equivalence limits (80 - 125%). No serious adverse events were found among the subjects. CONCLUSION: The RSABE approach was successful in evaluating the bioequivalence of these two formulations. This study confirmed that test and reference atorvastatin calcium tablets were bioequivalent under fed condition.

Anti-cholinesterase activities of constituents isolated from Lycopodiastrum casuarinoides.

Phytochemical investigation of the ethyl acetate extract of Lycopodiastrum casuarinoides (Spring) Holub (Lycopodiaceae) led to the isolation of nine compounds, including two new serratene triterpenoids, serrat-14-en-3α,21α-diol (1), 26-nor-8-oxo-21-one-α-onocerin (6), one new abietane diterpenoid, lycocasuarinone A (7), one new sesquiterpene acid, 7, 9-diene-1,4-epoxy-2-hydroxy-10-carboxylic acid (8) and one new chromone derivative, 5,7-dihydroxy-2-methyl esterchromone (9), together with four known serratene triterpenoids (2-5). Abietane diterpenoid (7) and sesquiterpene acid (8) from Lycopodiastrum casuarinoides are reported for the first time. Their structures and stereochemistry were unambiguously elucidated by spectroscopic analysis and comparison with known ones. All the compounds were tested for acetylcholinesterase (AChE) and butyrocholinesterase (BuChE) inhibitory activities. Bioactivity assays revealed that compound 6 exhibited the most potent AChE inhibitory effect.