Mass Balance and Metabolic Pathways of Eliapixant, a P2X3 Receptor Antagonist, in Healthy Male Volunteers.

BACKGROUND: Overactive adenosine triphosphate signaling via P2X3 homotrimeric receptors is implicated in multiple conditions. To fully understand the metabolism and elimination pathways of eliapixant, a study was conducted to assess the pharmacokinetics, mass balance, and routes of excretion of a single oral dose of the selective P2X3 receptor antagonist eliapixant, in addition to an in vitro characterization. METHODS: In this single-center open-label non-randomized non-placebo-controlled phase I study, healthy male subjects (n = 6) received a single dose of 50 mg eliapixant blended with 3.7 MBq [14C]eliapixant as a PEG 400-based oral solution. Total radioactivity and metabolites excreted in urine and feces, and pharmacokinetics of total radioactivity, eliapixant, and metabolites in plasma were assessed via liquid scintillation counting and high-performance liquid chromatography-based methods coupled to radiometric and mass spectrometric detection. Metabolite profiles of eliapixant in human in vitro systems and metabolizing enzymes were also investigated. RESULTS: After administration as an oral solution, eliapixant was rapidly absorbed, reaching maximum plasma concentrations within 2 h. Eliapixant was eliminated from plasma with a mean terminal half-life of 48.3 h. Unchanged eliapixant was the predominant component in plasma (72.6% of total radioactivity area under the curve). The remaining percentage of drug-related components in plasma probably represented the sum of many metabolites, detected in trace amounts. Mean recovery of total radioactivity was 97.9% of the administered dose (94.3-99.4%) within 14 days, with 86.3% (84.8-88.1%) excreted via feces and 11.6% (9.5-13.1%) via urine. Excretion of parent drug was minimal in feces (0.7% of dose) and urine (≈ 0.5%). In feces, metabolites formed by oxidation represented > 90% of excreted total radioactivity. The metabolites detected in the in vitro experiments were similar to those identified in vivo. CONCLUSION: Complete recovery of administered eliapixant-related radioactivity was observed in healthy male subjects with predominant excretion via feces. Eliapixant was almost exclusively cleared by oxidative biotransformation (> 90% of dose), with major involvement of cytochrome P450 3A4. Excretion of parent drug was of minor importance (~ 1% of dose). CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov: NCT04487431 (registered 27 July 2020)/EudraCT number: 2020-000519-54 (registered 3 February 2020), NCT02817100 (registered 26 June 2016), NCT03310645 (registered 16 October 2017).

Eliapixant is a drug that acts on structures in the body called P2X3 receptors that are involved in several conditions, including chronic cough, overactive bladder, and endometriosis-related pain. When evaluating a new drug, it is important to know how it is being removed from the body by natural mechanisms. We performed a study in which six healthy male volunteers took a single dose of eliapixant, and we investigated what happened to the drug after it was taken. We measured the amount of eliapixant in the volunteers' blood, urine, and feces, and also measured the compounds formed when eliapixant was broken down naturally by the body ("metabolites"). We also used human cells in the laboratory to investigate how the different metabolites of eliapixant are formed. Almost three-quarters of eliapixant in the blood had not been broken down at all, while the remaining one-quarter had been converted into many different metabolites. A total of 2 weeks after taking eliapixant, almost all of it had been converted to metabolites and eliminated from the body (mostly in feces, but also a small amount in urine). The most important organ for breaking down eliapixant is the liver. The information from this study will help doctors determine whether eliapixant is likely to interfere with other drugs taken simultaneously, and whether patients with liver or kidney problems might take longer than healthy people to remove it from their bodies.

Metabolism and Disposition of the Novel Oral Factor XIa Inhibitor Asundexian in Rats and in Humans.

BACKGROUND AND OBJECTIVES: Current anticoagulants pose an increased risk of bleeding. The development of drugs targeting factor XIa, like asundexian, may provide a safer treatment option. A human mass­balance study was conducted to gain a deeper understanding of the absorption, distribution, metabolism, excretion, and potential for drug-drug interaction of asundexian. Additionally, an overview of the biotransformation and clearance pathways for asundexian in humans and bile-duct cannulated (BDC) rats in vivo, as well as in vitro in hepatocytes of both species, is reported. METHODS: The mass balance, biotransformation, and excretion pathways of asundexian were investigated in six healthy volunteers (single oral dose of 25 mg [14C]asundexian) and in BDC rats (intravenous [14C]asundexian 1 mg/kg). RESULTS: Overall recovery of radioactivity was 101% for humans (samples collected up to 14 days after dosing), and 97.9% for BDC rats (samples collected in the 24 h after dosing). Radioactivity was mainly excreted into feces in humans (80.3%) and into bile/feces in BDC rats (> 94%). The predominant clearance pathways in humans were amide hydrolysis to metabolite M1 (47%) and non-labeled M9 with subsequent N-acetylation to M10; oxidative biotransformation was a minor pathway (13%). In rats, hydrolysis of the terminal amide to M2 was the predominant pathway. In human plasma, asundexian accounted for 61.0% of total drug-related area under the plasma concentration-time curve (AUC); M10 was the major metabolite (16.4% of the total drug-related AUC). Excretion of unmetabolized drug was a significant clearance pathway in both species (human, ~ 37%; BDC rat, ~ 24%). The near-complete bioavailability of asundexian suggests negligible limitations on absorption and first-pass metabolism. Comparison with radiochromatograms from incubations with human or rat hepatocytes indicated consistency across species and a good overall in vitro/in vivo correlation. CONCLUSIONS: Similar to preclinical experiments, total asundexian-derived radioactivity is cleared quantitatively predominantly via feces. Excretion occurs mainly via amide hydrolysis and as the unchanged drug.

Structural and Mechanistic Investigation of the Unusual Metabolism of Nifurtimox.

The oral antiparasitic drug nifurtimox has been used to treat Chagas disease for more than 50 years. Historical studies determined that very little nifurtimox is excreted unchanged, but contemporaneous preclinical studies of radiolabeled nifurtimox found almost all of the radiolabel was rapidly excreted, suggesting that metabolism is extensive. Attempts to study nifurtimox metabolism have had limited success, yet this knowledge is fundamental to characterizing the pharmacokinetics and pharmacodynamics of the drug. We conducted in vitro studies using hepatic and renal sources with 14C-labeled nifurtimox as substrate and obtained samples of urine, plasma, and feces from rats administered 2.5 mg/kg [14C]-nifurtimox, and samples of human urine and plasma from phase 1 clinical studies in which participants received a single dose of 120 mg nifurtimox. Analysis of metabolites was done by high-performance liquid chromatography (HPLC)-high-resolution mass spectrometry (HRMS) and HRMS/MS with offline liquid scintillation counting of radiolabeled samples. Surprisingly, only traces of a few metabolites were identified from in vitro incubations with hepatocytes and subcellular fractions, but more than 30 metabolites were identified in rat urine, mostly with atypical mass changes. We developed an HRMS scouting method for the analysis of human samples based on the sulfur atom in nifurtimox and the natural abundance of 34S, as well as a characteristic tandem mass spectrometry (MS/MS) fragmentation of nifurtimox and metabolites. Fragmentation patterns on HRMS/MS were used to propose structures for 18 metabolites (22 including stereoisomers), and based on these structures, the six most abundant products were synthesized and the structures of the synthetic forms were confirmed by HRMS and two-dimensional nuclear magnetic resonance (2D NMR). Overall, we determined that the metabolism of nifurtimox is almost certainly not mediated by typical hepatic and renal drug-metabolizing enzymes, and instead is rapidly metabolized mainly by reduction or nucleophilic attack, with some evidence of oxidation. Knowledge of the most abundant metabolites of nifurtimox affords the possibility of future studies to investigate levels of exposure and possible drug-drug interactions.

Pharmacokinetics, excretion, distribution, and metabolism of 60-kDa polyethylene glycol used in BAY 94-9027 in rats and its value for human prediction.

The covalent binding of proteins with polyethylene glycol (PEG) molecules is a valuable tool to extend the half-life of many biotherapeutics, including factor VIII (FVIII) products to treat patients with haemophilia A. Although PEG has low toxicity, accumulation of large PEG molecules (>20-30 kDa) with long-term exposure is a potential concern. Thus, it is important to determine whether sufficient excretion processes exist for PEG molecules used in biotherapeutics. BAY 94-9027 is an extended-half-life FVIII product modified through addition of a 60-kDa (branched: dual 30-kDa) PEG molecule. BAY 1025662 is the 60-kDa PEG moiety used for PEGylation of BAY 94-9027. This study investigated the pharmacokinetic (PK) properties, distribution, and excretion of BAY 1025662 in rats in order to predict estimated 60-kDa PEG PK properties in patients. Plasma concentrations in male rats after a single 11-mg/kg intravenous dose of BAY 1025662 (approximating the cumulative PEG-60 exposure in patients during 30 years of BAY 94-9027 treatment) decreased with an initial half-life of 119 h (5 days) in the interval of 114-336 h post administration. Single-dose mass balance studies using radiolabeled BAY 1025662 ([prop-14C]BAY 1025662) showed that 30.4% of radioactivity was excreted within 1 week and 79.1% by Day 168 (primarily in urine). The terminal half-life of radioactivity elimination was approximately 24 days in blood and plasma and was 31-68 days in the majority of other organs up to Day 168. Elimination was nearly complete at the end of the experiment on Day 168; only ~4% of residual radioactivity was present in the animal body. There was no irreversible binding of radioactivity to any tissues and no penetration of the blood-brain barrier. Based on these results, very low steady-state concentrations of 60-kDa PEG were predicted in patients treated with BAY 94-9027, and the validity of these predictions was supported by clinical studies in which almost all 179 patients receiving BAY 94-9027 for prophylaxis had undetectable PEG in plasma for up to >5 years; those with detectable PEG levels demonstrated concentrations within the predicted range. These combined preclinical and clinical observations suggest that excretion processes are in place for high-molecular-weight PEGs such as the PEG-60 moiety used in BAY 94-9027.

Enzymatic Late-Stage Oxidation of Lead Compounds with Solubilizing Biomimetic Docking/Protecting groups.

Late-stage functionalization of lead compounds is of high interest in drug discovery since it offers an easy access to metabolites and derivatives of a lead compound without the need to redesign an often long multistep synthesis. Owing to their high degree of chemoselectivity, biocatalytic transformations, enzymatic oxidations in particular, are potentially very powerful because they could allow the synthesis of less lipophilic derivatives of a lead compound. In the majority of cases, enzymatic oxidations have been used in an empirical way as their regioselectivity is difficult to predict. In this publication, the concept of using docking/protecting groups in a biomimetic fashion was investigated, which could help steer the regioselectivity of a P450BM3 -mediated oxidation. A novel set of docking/protecting groups was designed that can be cleaved under very mild conditions and address the often problematic aqueous solubility of the substrates. Vabicaserin was used as tool compound containing typical groups such as basic, aliphatic, and aromatic moieties. The results were rationalized with the help of in silico docking and molecular dynamic studies.

Emerging technologies for metabolite generation and structural diversification.

Multiple technologies have emerged for structural diversification and efficient production of metabolites of drug molecules. These include expanded use of enzymatic and bioorganic transformations that mimic biological systems, biomimetic catalysis and electrochemical techniques. As this field continues to mature the breadth of transformations is growing beyond simple oxidative processes due in part to parallel development of more efficient catalytic methods for functionalization of unactivated scaffolds. These technologies allow for efficient structural diversification of both aromatic and aliphatic substrates in many cases via single step reactions without the use of protecting groups.

A planar chiral [2.2]paracyclophane derived N-heterocyclic stannylene.

The reaction of pseudo-ortho-4,12-N,N'-diphenyldiamino-[2.2]paracyclophane ((±)-3) with Sn[N(SiMe(3))(2)](2) results in the formation of the monomeric planar chiral N-heterocyclic stannylene (±)-4, featuring a unique [2.2]paracyclophane backbone, which has been characterized by an X-ray diffraction study.

N-heterocyclic carbene-catalyzed hydroacylation of unactivated double bonds.

An intramolecular N-heterocyclic carbene (NHC)-catalyzed hydroacylation of unactivated double bonds is reported. Systematic variation of the catalyst structure revealed an N-mesitylthiazolylidene annulated with a seven-membered ring to be especially reactive. This NHC enables a unique C-C bond-forming reaction to afford substituted chroman-4-ones in moderate to excellent yields, even ones containing all-carbon quaternary centers.

Palladium-catalyzed intramolecular direct arylation of benzoic acids by tandem decarboxylation/C-H activation.

A novel protocol for the palladium-catalyzed intramolecular direct arylation of benzoic acids is reported, which combines decarboxylation and C-H activation into one efficient process. With the optimized catalyst system of palladium trifluoroacetate, silver carbonate, and the proper solvent mixture (5% DMSO/dioxane), dibenzofuran derivatives are formed in excellent yield and selectivity.

Anionic Indole N-carbamoyl N --> C translocation. A directed remote metalation route to 2-Aryl- and 2-heteroarylindoles. synthesis of benz[a]carbazoles and Indeno[1,2-b]indoles.

A new LDA-induced anionic N-C carbamoyl migration of 2-arylindoles (7) is reported. Treatment of N-carbamoylindoles 10 and 13, readily available by direct and ipso-borodesilylative Suzuki-Miyaura cross-coupling routes from 8 and 12, respectively, provides a general route to functionalized 2-arylindoles 11 and 14, respectively (Tables 1 and 2). The reaction has been applied to the synthesis of benzo[ a]carbazoles 16 and indeno[1,2- b]indoles 18, and its intramolecularity has been established by a crossover experiment (Scheme 4).