Absorption, Distribution, Metabolism, and Excretion of [14C]-Volixibat in Healthy Men: Phase 1 Open-Label Study.

BACKGROUND AND OBJECTIVES: Volixibat is a potent inhibitor of the apical sodium-dependent bile acid transporter in development for the treatment of nonalcoholic steatohepatitis. This phase 1, open-label study investigated the absorption, distribution, metabolism, and excretion of [14C]-volixibat in heathy men. METHODS: Eligible men (n = 8) aged 18-50 years (body mass index 18.0-30.0 kg/m2; weight >50 kg) received a single oral dose of [14C]-volixibat 50 mg containing ~5.95 µCi radioactivity. The primary objectives were to assess the pharmacokinetics of [14C]-volixibat and to determine the total radioactivity in whole blood, plasma, urine, and feces at pre-selected time points over 6 days. The secondary objectives were to characterize metabolites and to assess the safety and tolerability. RESULTS: Low concentrations of volixibat (range 0-0.179 ng/mL) were detected in plasma up to 8 h following administration; the pharmacokinetic parameters could not be calculated. No radioactivity was observed in plasma or whole blood. The percentage (mean ± standard deviation) of total radioactivity in urine was 0.01 ± 0.007%. The vast majority (92.3 ± 5.25%) of volixibat was recovered in feces (69.2 ± 33.1% within 24 h). Unchanged volixibat was the only radioactive component detected in feces. Adverse events were mild in severity and mostly gastrointestinal. Changes in laboratory values were not clinically meaningful. CONCLUSIONS: Following oral administration, [14C]-volixibat was excreted unchanged from the parent compound almost exclusively via fecal excretion, indicating that the drug is minimally absorbed. Consistent with other studies, adverse events were primarily gastrointestinal in nature. ClinicalTrials.gov identifier NCT02571192.

Highly sensitive liquid chromatography-tandem mass spectrometry method for quantification of a new bone anabolic agent, TAK-778, in human serum.

A liquid chromatography-tandem mass spectrometric (LC-MS-MS) method for the highly sensitive determination of a new bone-anabolic agent, TAK-778 in human serum was developed. The internal standard (I.S.) used was deuterated TAK-778. TAK-778 and I.S. were extracted from serum samples with diethyl ether at neutral pH. A turbo ion spray interface was used as the ion source of LC-MS-MS, and the analysis was performed in the selected reaction monitoring mode. The lower limit of quantification was 0.02 ng/ml when 0.4 ml of serum was used, and the standard curve was linear in the range of 0.02-10 ng/ml. The method was precise; the intra- and inter-day precision of the method was not more than 17.9%. The accuracy of the method was good with the deviations between added and calculated concentration of TAK-778 being typically within 9.0%.

Application of 1-alkylamines to a liquid chromatographic/turbo ionspray tandem mass spectrometric method for quantifying metabolites of a new bone anabolic agent, TAK-778, in human serum.

We investigated the application of alkylamines, as additives to the mobile phase, to a quantification method for the metabolites, M-III and M-IV, of TAK-778, which is a new bone anabolic agent, in human serum using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Prior to setting up the analytical method, we found that 1-alkylamines co-existing with M-III and M-IV in the turbo ionsprayed solution formed 1-alkylammonium adduct molecules of these metabolites during the ionization process, and the abundance of the adduct ions was considerably higher than that of protonated molecules ([M + H](+)s) of these metabolites. Based on these findings, we investigated a variety of 1-alkylamines and their spiked concentrations in the mobile phase for LC/MS/MS analysis to obtain higher sensitivities for the quantification of these metabolites. After these examinations, we found that 1-hexylamine at a final concentration of 0.05 mmol l(-1) was the most suitable additive for the mobile phase, and set the selected reaction monitoring (SRM) ions for the 1-hexylammonium adduct molecule and [M + H](+), allowing about a fivefold gain in the SRM chromatographic peak compared with that without 1-hexylamine. The adduct ion was considered to be formed by interaction between the amino group of 1-hexylamine and the phosphoryl group of M-III and M-IV. The internal standard (I.S.) used was deuterated M-III for each metabolite. The analytes and I.S. were extracted with diethyl ether from serum samples at neutral pH and injected into the LC/MS/MS system with a turbo ionspray interface. The limit of quantification for both analytes was 0.5 ng ml(-1) when 0.1 ml of serum was used, and the calibration curves were linear in the range 0.5-100 ng ml(-1). The method was precise; the intra- and inter-day precisions of the method were not more than 5.6%. The accuracy of the method was good, with deviations between added and calculated concentrations of M-III and M-IV being typically within 16.6%. This method provided reliable pharmacokinetic data for M-III and M-IV after the intramuscular administration of TAK-778 sustained-release formulation in humans.

Sustained-release microcapsules of a bone formation stimulant, TAK-778, for local injection into a fracture site.

The benzothiepin derivative (2R,4S)-(-)-N-(4-diethoxyphosphorylmethylphenyl)-1,2,4,5-tetrahydro-4-methyl-7,8-methylenedioxy-5-oxo-3-benzothiepin-2-carboxamide (TAK-778) is a potent bone formation stimulant. A sustained release formulation was prepared by encapsulating the drug into biodegradable microcapsules for local application to fracture repair in rats. The microcapsules consisted of TAK-778 (10% w/w) and a biodegradable polymer, copoly (d,l-lactic/glycolic acid), with a copolymer ratio of 85:15 (mol/mol) and an average molecular weight of 15 k. The TAK-778 amount at the injection site progressively diminished for 4 weeks after administration, and the serum level of TAK-778 was sustained over the same period. The local concentration of TAK-778 after administration of the microcapsules was simulated by a two-compartment open model. In the model, a first-order release rate constant and a transfer rate constant were obtained from the release profile of the microcapsules and the serum level of TAK-778 after administration of the TAK-778 solution, respectively. Localization at the injection site was examined by radiography using microcapsules in which iodoform was encapsulated as a contrast agent. The microcapsules formed a clot at the injection site, and their spread was narrowly restricted. The local concentration was calculated to be maintained within the range 10(-3)-10(-6) M over 4 weeks on the assumption that the dose and spread volume were 5 mg of TAK-778/site and 3 mL, respectively.

A one-step enzyme-linked immunosorbent assay for a novel osteoblast differentiation-promoting compound, TAK-778 in serum.

TAK-778, ((2R,4S)-(-)-N-[(4-diethoxyphosphorylmethyl)phenyl]- 1,2,4,5-tetrahydro-4-methyl-7,8-methylenedioxy-5-oxo-3-benzothiepin-2- carboxamide), is a novel osteoblast differentiation-promoting compound, and its sustained-release formulation is expected to be clinically used for the enhancement of fracture healing. To date, TAK-778 levels in serum have been measured using conventional reverse-phase HPLC with inferior sensitivity and time-consuming procedures. We have produced polyclonal antibodies against TAK-778 by using one of its derivatives coupled with a carrier protein, and developed a one-step ELISA for the determination of TAK-778 in serum. The antibodies had minimal cross-reactivities to the biologically inactive metabolites including the oxidized-form (0.36%) and the cleavaged-form (0.00083%). The competitive ELISA was accomplished within three hours with a detection limit of 0.5 ng/ml serum, and the coefficients of variations for samples ranging from 1 ng/ml to 80 ng/ml were 1.1-4.4% in an intra-assay and 3.1-9.6% in an inter-assay. The present ELISA is so rapid, sensitive and selective for TAK-778 that it could be conveniently used in a clinical field for the determination of many serum specimens.

[Densitometric determination of dithiadenoxide in blood]. / Densitometrické stanovení dithiadenoxidu v krevní plazme.

The present paper describes a method of dithiadenoxide determination in blood plasma. Dithiadenoxide was reduced to dithiaden with zinc dust in acidified plasma. The solution was alkalified and dithiaden extracted into n-hexane and isolated on a thin layer of silica gel using the system chloroform--n-heptane-methanol (60:22:18). After spraying the plate with a solution of m-nitrophenyldiazoniumfluoroborate in acid medium, dithiaden stains were evaluated densitometrically in the absorption maximum of the reaction product at 565 nm (RF = 0.37). Detection limit of dithiadenoxide in the proposed method is 0.004 microgram in an applied amount.

High-performance liquid chromatographic assay of a new non-steroidal anti-inflammatory agent, 2-(10,11-dihydro-10-oxodibenzo[b,f]thiepin-2-yl)propionic acid, in human plasma and urine.

A high-performance liquid chromatographic method for the quantitation of a new anti-inflammatory agent, 2-(10,11-dihydro-10-oxodibenzo[b,f]thiepin-2-yl)propionic acid (CN-100; I), has been developed. The assay consists in extracting samples containing I and mefenamic acid, the internal standard, under acidic conditions and analysis by reversed-phase chromatography using ultraviolet detection at 330 nm. Preliminary plasma concentration-time and cumulative urinary excretion profiles from a healthy subject following oral administration of the tablet formulation are presented. This method is simple, sensitive and reproducible and is applicable to studies of the pharmacokinetic behaviour of I in humans.