Research

13 pages, 1754 KiB

Open AccessArticle

In Vitro Metabolism Study of Seongsanamide A in Human Liver Microsomes Using Non-Targeted Metabolomics and Feature-Based Molecular Networking

by Zhexue Wu, Geum Jin Kim, So-Young Park, Jong Cheol Shon, Kwang-Hyeon Liu and Hyukjae Choi

Pharmaceutics 2021, 13(7), 1031; https://doi.org/10.3390/pharmaceutics13071031 - 07 Jul 2021

Cited by 1 | Viewed by 2518

Abstract

Seongsanamide A is a bicyclic peptide with an isodityrosine residue discovered in Bacillus safensis KCTC 12796BP which exhibits anti-allergic activity in vitro and in vivo without significant cytotoxicity. The purpose of this study was to elucidate the in vitro metabolic pathway and potential [...] Read more.

Seongsanamide A is a bicyclic peptide with an isodityrosine residue discovered in Bacillus safensis KCTC 12796BP which exhibits anti-allergic activity in vitro and in vivo without significant cytotoxicity. The purpose of this study was to elucidate the in vitro metabolic pathway and potential for drug interactions of seongsanamide A in human liver microsomes using non-targeted metabolomics and feature-based molecular networking (FBMN) techniques. We identified four metabolites, and their structures were elucidated by interpretation of high-resolution tandem mass spectra. The primary metabolic pathway associated with seongsanamide A metabolism was hydroxylation and oxidative hydrolysis. A reaction phenotyping study was also performed using recombinant cytochrome P450 isoforms. CYP3A4 and CYP3A5 were identified as the major metabolic enzymes responsible for metabolite formation. Seongsanamide A did not inhibit the cytochrome P450 isoforms commonly involved in drug metabolism (IC₅₀ > 10 µM). These results will contribute to further understanding the metabolism and drug interaction potential of various bicyclic peptides. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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19 pages, 3958 KiB

Open AccessArticle

In Vitro Metabolism of Donepezil in Liver Microsomes Using Non-Targeted Metabolomics

by Sin-Eun Kim, Hyung-Ju Seo, Yeojin Jeong, Gyung-Min Lee, Seung-Bae Ji, So-Young Park, Zhexue Wu, Sangkyu Lee, Sunghwan Kim and Kwang-Hyeon Liu

Pharmaceutics 2021, 13(7), 936; https://doi.org/10.3390/pharmaceutics13070936 - 23 Jun 2021

Cited by 5 | Viewed by 2896

Abstract

Donepezil is a reversible acetylcholinesterase inhibitor that is currently the most commonly prescribed drug for the treatment of Alzheimer’s disease. In general, donepezil is known as a safe and well-tolerated drug, and it was not associated with liver abnormalities in several clinical trials. [...] Read more.

Donepezil is a reversible acetylcholinesterase inhibitor that is currently the most commonly prescribed drug for the treatment of Alzheimer’s disease. In general, donepezil is known as a safe and well-tolerated drug, and it was not associated with liver abnormalities in several clinical trials. However, rare cases of drug-related liver toxicity have been reported since it has become commercially available. Few studies have investigated the metabolic profile of donepezil, and the mechanism of liver damage caused by donepezil has not been elucidated. In this study, the in vitro metabolism of donepezil was investigated using liquid chromatography–tandem mass spectrometry based on a non-targeted metabolomics approach. To identify metabolites, the data were subjected to multivariate data analysis and molecular networking. A total of 21 donepezil metabolites (17 in human liver microsomes, 21 in mice liver microsomes, and 17 in rat liver microsomes) were detected including 14 newly identified metabolites. One potential reactive metabolite was identified in rat liver microsomal incubation samples. Metabolites were formed through four major metabolic pathways: (1) O-demethylation, (2) hydroxylation, (3) N-oxidation, and (4) N-debenzylation. This study indicates that a non-targeted metabolomics approach combined with molecular networking is a reliable tool to identify and detect unknown drug metabolites. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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16 pages, 3250 KiB

Open AccessArticle

Laser Capture Microdissection Coupled Capillary Immunoassay to Study the Expression of PCK-2 on Spatially-Resolved Islets of Rat Langerhans

by Shashank Pandey, Zdenek Tuma, Tereza Smrhova, Miroslava Cedikova, Tereza Macanova and Magdalena Chottova Dvorakova

Pharmaceutics 2021, 13(6), 883; https://doi.org/10.3390/pharmaceutics13060883 - 15 Jun 2021

Cited by 1 | Viewed by 1912

Abstract

The platform for precise proteomic profiling of targeted cell populations from heterogeneous tissue sections is developed. We demonstrate a seamless and systematic integration of LCM with an automated cap-IA for the handling of a very small-sized dissected tissues section from the kidney, liver [...] Read more.

The platform for precise proteomic profiling of targeted cell populations from heterogeneous tissue sections is developed. We demonstrate a seamless and systematic integration of LCM with an automated cap-IA for the handling of a very small-sized dissected tissues section from the kidney, liver and pancreatic Langerhans islet of rats. Our analysis reveals that the lowest LCM section area ≥ 0.125 mm² with 10 µm thickness can be optimized for the detection of proteins through LCM-cap-IA integration. We detect signals ranging from a highly-abundant protein, β-actin, to a low-abundance protein, LC-3AB, using 0.125 mm² LCM section from rat kidney, but, so far, a relatively large section is required for good quality of results. This integration is applicable for a highly-sensitive and accurate assessment of microdissected tissue sections to decipher hidden proteomic information of pure targeted cells. To validate this integration, PCK2 protein expression is studied within Langerhans islets of normal and diabetic rats. Our results show significant overexpression of PCK2 in Langerhans islets of rats with long-term diabetes. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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15 pages, 4527 KiB

Open AccessArticle

Nontargeted Metabolomics by High-Resolution Mass Spectrometry to Study the In Vitro Metabolism of a Dual Inverse Agonist of Estrogen-Related Receptors β and γ, DN203368

by Sin-Eun Kim, Seung-Bae Ji, Euihyeon Kim, Minseon Jeong, Jina Kim, Gyung-Min Lee, Hyung-Ju Seo, Subin Bae, Yeojin Jeong, Sangkyu Lee, Sunghwan Kim, Taeho Lee, Sung Jin Cho and Kwang-Hyeon Liu

Pharmaceutics 2021, 13(6), 776; https://doi.org/10.3390/pharmaceutics13060776 - 31 May 2021

Cited by 1 | Viewed by 2421

Abstract

DN203368 ((E)-3-[1-(4-[4-isopropylpiperazine-1-yl]phenyl) 3-methyl-2-phenylbut-1-en-1-yl] phenol) is a 4-hydroxy tamoxifen analog that is a dual inverse agonist of estrogen-related receptor β/γ (ERRβ/γ). ERRγ is an orphan nuclear receptor that plays an important role in development and homeostasis and holds potential as a novel [...] Read more.

DN203368 ((E)-3-[1-(4-[4-isopropylpiperazine-1-yl]phenyl) 3-methyl-2-phenylbut-1-en-1-yl] phenol) is a 4-hydroxy tamoxifen analog that is a dual inverse agonist of estrogen-related receptor β/γ (ERRβ/γ). ERRγ is an orphan nuclear receptor that plays an important role in development and homeostasis and holds potential as a novel therapeutic target in metabolic diseases such as diabetes mellitus, obesity, and cancer. ERRβ is also one of the orphan nuclear receptors critical for many biological processes, such as development. We investigated the in vitro metabolism of DN203368 by conventional and metabolomic approaches using high-resolution mass spectrometry. The compound (100 μM) was incubated with rat and human liver microsomes in the presence of NADPH. In the metabolomic approach, the m/z value and retention time information obtained from the sample and heat-inactivated control group were statistically evaluated using principal component analysis and orthogonal partial least-squares discriminant analysis. Significant features responsible for group separation were then identified using tandem mass spectra. Seven metabolites of DN203368 were identified in rat liver microsomes and the metabolic pathways include hydroxylation (M1-3), N-oxidation (M4), N-deisopropylation (M5), N,N-dealkylation (M6), and oxidation and dehydrogenation (M7). Only five metabolites (M2, M3, and M5-M7) were detected in human liver microsomes. In the conventional approach using extracted ion monitoring for values of mass increase or decrease by known metabolic reactions, only five metabolites (M1-M5) were found in rat liver microsomes, whereas three metabolites (M2, M3, and M5) were found in human liver microsomes. This study revealed that nontargeted metabolomics combined with high-resolution mass spectrometry and multivariate analysis could be a more efficient tool for drug metabolite identification than the conventional approach. These results might also be useful for understanding the pharmacokinetics and metabolism of DN203368 in animals and humans. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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10 pages, 1116 KiB

Open AccessArticle

Quantification of Teicoplanin Using the HPLC-UV Method for Clinical Applications in Critically Ill Patients in Korea

by Jaeok Lee, Eun-Kyoung Chung, Sung-Wook Kang, Hwa-Jeong Lee and Sandy-Jeong Rhie

Pharmaceutics 2021, 13(4), 572; https://doi.org/10.3390/pharmaceutics13040572 - 17 Apr 2021

Cited by 3 | Viewed by 2084

Abstract

A high-performance liquid chromatography-ultraviolet detector (HPLC-UV) method has been used to quantify teicoplanin concentrations in human plasma. However, the limited analytical accuracy of previously bioanalytical methods for teicoplanin has given rise to uncertainty due to the use of an external standard. In this [...] Read more.

A high-performance liquid chromatography-ultraviolet detector (HPLC-UV) method has been used to quantify teicoplanin concentrations in human plasma. However, the limited analytical accuracy of previously bioanalytical methods for teicoplanin has given rise to uncertainty due to the use of an external standard. In this study, an internal standard (IS), polymyxin B, was applied to devise a precise, accurate, and feasible HPLC-UV method. The deproteinized plasma sample containing teicoplanin and an IS of acetonitrile was chromatographed on a C18 column with an acidic mobile phase consisting of NaH₂PO₄ buffer and acetonitrile (78:22, v/v) by isocratic elution and detection at 220 nm. The linearity was in the range 7.8–500 mg/L calculated by the ratio of the teicoplanin signal to the IS signal. This analytical method, validated by FDA guidelines with ICH Q2 (R1), was successfully applied to analyze the plasma samples of patients in the intensive care unit for treating serious resistant bacterial infectious diseases, such as those by methicillin-resistant Staphylococcus aureus and Enterococcus faecalis. The methods suggested the potential for use in routine clinical practice for therapeutic drug monitoring of teicoplanin, providing both improved accuracy and a wide range of linearity from lower than steady-state trough concentrations (10 mg/L) to much higher concentrations. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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12 pages, 2161 KiB

Open AccessArticle

Urinary Metabolomic Profiling after Administration of Corydalis Tuber and Pharbitis Seed Extract in Healthy Korean Volunteers

by Hyeon-Cheol Jeong, Jung Eun Park, Yohan Seo, Min-Gul Kim and Kwang-Hee Shin

Pharmaceutics 2021, 13(4), 522; https://doi.org/10.3390/pharmaceutics13040522 - 09 Apr 2021

Cited by 2 | Viewed by 1655

Abstract

Pharmacometabolomics is a useful tool to identify biomarkers that can assess and predict response after drug administration. The primary purpose of pharmacometabolomics is to better understand the mechanisms and pathways of a drug by searching endogenous metabolites that have significantly changed after drug [...] Read more.

Pharmacometabolomics is a useful tool to identify biomarkers that can assess and predict response after drug administration. The primary purpose of pharmacometabolomics is to better understand the mechanisms and pathways of a drug by searching endogenous metabolites that have significantly changed after drug administration. DA-9701, a prokinetic agent, consists of Pharbitis seed and Corydalis tube extract and it is known to improve the gastrointestinal motility. Although the overall mechanism of action of DA-9701 remains unclear, its active ingredients, corydaline and chlorogenic acid, act as a 5-HT3 and D2 receptor antagonist and 5-HT4 receptor agonist. To determine the significant metabolites after the administration of DA-9701, a qualitative analysis was carried out using ultra-high performance liquid chromatography coupled with orbitrap mass spectrometer followed by a multivariate analysis. Seven candidates were selected and a statistical analysis of fold change was performed over time. Our study concluded that all the seven selected metabolites were commonly involved in lipid metabolism and purine metabolism. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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20 pages, 2613 KiB

Open AccessArticle

Transcriptome Analysis Identifies Novel Mechanisms Associated with the Antitumor Effect of Chitosan-Stabilized Selenium Nanoparticles

by Hector Estevez, Estefania Garcia-Calvo, Jose Rivera-Torres, María Vallet-Regí, Blanca González and Jose L. Luque-Garcia

Pharmaceutics 2021, 13(3), 356; https://doi.org/10.3390/pharmaceutics13030356 - 08 Mar 2021

Cited by 8 | Viewed by 2018

Abstract

Selenium nanoparticles (SeNPs) have been receiving special attention in recent years due to their antioxidant capacity and antitumor properties. However, the mechanisms associated with these properties remain to be elucidated. For this reason, a global transcriptome analysis has been designed in this work [...] Read more.

Selenium nanoparticles (SeNPs) have been receiving special attention in recent years due to their antioxidant capacity and antitumor properties. However, the mechanisms associated with these properties remain to be elucidated. For this reason, a global transcriptome analysis has been designed in this work and it was carried out using human hepatocarcinoma cells and chitosan-stabilized SeNPs (Ch-SeNPs) to identify new targets and pathways related to the antitumor mechanisms associated with Ch-SeNPs. The results obtained confirm the alteration of the cell cycle and the effect of Ch-SeNPs on different tumor suppressors and other molecules involved in key mechanisms related to cancer progression. Furthermore, we demonstrated the antioxidant properties of these nanoparticles and their capacity to induce senescence, which was further confirmed through the measurement of β-galactosidase activity. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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17 pages, 4036 KiB

Open AccessFeature PaperArticle

In Vitro Metabolism of DWP16001, a Novel Sodium-Glucose Cotransporter 2 Inhibitor, in Human and Animal Hepatocytes

by Ju-Hyun Kim, Dong Kyun Kim, Won-Gu Choi, Hye-Young Ji, Ji-Soo Choi, Im-Sook Song, Sangkyu Lee and Hye Suk Lee

Pharmaceutics 2020, 12(9), 865; https://doi.org/10.3390/pharmaceutics12090865 - 11 Sep 2020

Cited by 16 | Viewed by 2995

Abstract

DWP16001 is currently in a phase 2 clinical trial as a novel anti-diabetes drug for the treatment of type 2 diabetes by selective inhibition of sodium-glucose cotransporter 2. This in vitro study was performed to compare the metabolism of DWP16001 in human, dog, [...] Read more.

DWP16001 is currently in a phase 2 clinical trial as a novel anti-diabetes drug for the treatment of type 2 diabetes by selective inhibition of sodium-glucose cotransporter 2. This in vitro study was performed to compare the metabolism of DWP16001 in human, dog, monkey, mouse, and rat hepatocytes, and the drug-metabolizing enzymes responsible for the metabolism of DWP16001 were characterized using recombinant human cytochrome 450 (CYP) and UDP-glucuronosyltransferase (UGT) enzymes expressed from cDNAs. The hepatic extraction ratio of DWP16001 in five species ranged from 0.15 to 0.56, suggesting that DWP16001 may be subject to species-dependent and weak-to-moderate hepatic metabolism. Five phase I metabolites (M1–M5) produced by oxidation as well as three DWP16001 glucuronides (U1–U3) and two hydroxy-DWP16001 (M1) glucuronides (U4, U5), were identified from hepatocytes incubated with DWP16001 by liquid chromatography-high resolution mass spectrometry. In human hepatocytes, M1, M2, M3, U1, and U2 were identified. Formation of M1 and M2 from DWP16001 was catalyzed by CYP3A4 and CYP2C19. M3 was produced by hydroxylation of M1, while M4 was produced by hydroxylation of M2; both hydroxylation reactions were catalyzed by CYP3A4. The formation of U1 was catalyzed by UGT2B7, but UGT1A4, UGT1A9, and UGT2B7 contributed to the formation of U2. In conclusion, DWP16001 is a substrate for CYP3A4, CYP2C19, UGT1A4, UGT1A9, and UGT2B7 enzymes. Overall, DWP16001 is weakly metabolized in human hepatocytes, but there is a potential for the pharmacokinetic modulation and drug–drug interactions, involved in the responsible metabolizing enzymes of DWP16001 in humans. Full article

(This article belongs to the Special Issue Bioanalysis and Metabolomics)

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