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1.
Int J Mol Sci ; 22(13)2021 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-34206780

RESUMO

Vascular calcification (VC) is a risk factor for cardiovascular events and mortality in chronic kidney disease (CKD). Several components influence the occurrence of VC, among which inflammation. A novel uremic toxin, lanthionine, was shown to increase intracellular calcium in endothelial cells and may have a role in VC. A group of CKD patients was selected and divided into patients with a glomerular filtration rate (GFR) of <45 mL/min/1.73 m2 and ≥45 mL/min/1.73 m2. Total Calcium Score (TCS), based on the Agatston score, was assessed as circulating lanthionine and a panel of different cytokines. A hemodialysis patient group was also considered. Lanthionine was elevated in CKD patients, and levels increased significantly in hemodialysis patients with respect to the two CKD groups; in addition, lanthionine increased along with the increase in TCS, starting from one up to three. Interleukin IL-6, IL-8, and Eotaxin were significantly increased in patients with GFR < 45 mL/min/1.73 m2 with respect to those with GFR ≥ 45 mL/min/1.73 m2. IL-1b, IL-7, IL-8, IL-12, Eotaxin, and VEGF increased in calcified patients with respect to the non-calcified. IL-8 and Eotaxin were elevated both in the low GFR group and in the calcified group. We propose that lanthionine, but also IL-8 and Eotaxin, in particular, are a key feature of VC of CKD, with possible marker significance.


Assuntos
Alanina/análogos & derivados , Citocinas/sangue , Insuficiência Renal Crônica/metabolismo , Sulfetos/sangue , Calcificação Vascular/metabolismo , Adulto , Alanina/sangue , Biomarcadores/sangue , Feminino , Taxa de Filtração Glomerular , Humanos , Masculino , Insuficiência Renal Crônica/sangue , Insuficiência Renal Crônica/complicações , Calcificação Vascular/sangue , Calcificação Vascular/etiologia
2.
Nat Metab ; 3(3): 394-409, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33758419

RESUMO

Both obesity and sarcopenia are frequently associated in ageing, and together may promote the progression of related conditions such as diabetes and frailty. However, little is known about the pathophysiological mechanisms underpinning this association. Here we show that systemic alanine metabolism is linked to glycaemic control. We find that expression of alanine aminotransferases is increased in the liver in mice with obesity and diabetes, as well as in humans with type 2 diabetes. Hepatocyte-selective silencing of both alanine aminotransferase enzymes in mice with obesity and diabetes retards hyperglycaemia and reverses skeletal muscle atrophy through restoration of skeletal muscle protein synthesis. Mechanistically, liver alanine catabolism driven by chronic glucocorticoid and glucagon signalling promotes hyperglycaemia and skeletal muscle wasting. We further provide evidence for amino acid-induced metabolic cross-talk between the liver and skeletal muscle in ex vivo experiments. Taken together, we reveal a metabolic inter-tissue cross-talk that links skeletal muscle atrophy and hyperglycaemia in type 2 diabetes.


Assuntos
Alanina/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Hiperglicemia/metabolismo , Fígado/metabolismo , Músculo Esquelético/patologia , Atrofia Muscular/metabolismo , Alanina/sangue , Alanina Transaminase/sangue , Animais , Glicemia/metabolismo , Diabetes Mellitus Tipo 2/patologia , Modelos Animais de Doenças , Homeostase , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/metabolismo
3.
Artigo em Inglês | MEDLINE | ID: mdl-33756448

RESUMO

Remdesivir, formerly GS-5734, has recently become the first antiviral drug approved by the U.S. Food and Drug Administration (FDA) to treat COVID-19, the disease caused by SARS-CoV-2. Therapeutic dosing and pharmacokinetic studies require a simple, sensitive, and selective validated assay to quantify drug concentrations in clinical samples. Therefore, we developed a rapid and sensitive LC-MS/MS assay for the quantification of remdesivir in human plasma with its deuterium-labeled analog, remdesivir-2H5, as the internal standard. Chromatographic separation was achieved on a Phenomenex® Synergi™ HPLC Fusion-RP (100 × 2 mm, 4 µm) column by gradient elution. Excellent accuracy and precision (<5.2% within-run variations and. <9.8% between-run variations) were obtained over the range of 0.5-5000 ng/mL. The assay met the FDA Bioanalytical Guidelines for selectivity and specificity, and low inter-matrix lot variability (<2.7%) was observed for extraction efficiency (77%) and matrix effect (123%) studies. Further, stability tests showed that the analyte does not degrade under working conditions, nor during freezing and thawing processes.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Antivirais/sangue , COVID-19/tratamento farmacológico , Monitoramento de Medicamentos/métodos , Espectrometria de Massas em Tandem/métodos , Monofosfato de Adenosina/sangue , Alanina/sangue , Cromatografia Líquida de Alta Pressão/economia , Cromatografia Líquida de Alta Pressão/métodos , Monitoramento de Medicamentos/economia , Feminino , Humanos , Limite de Detecção , Masculino , Espectrometria de Massas em Tandem/economia
4.
Pharmacol Res Perspect ; 9(2): e00743, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33710753

RESUMO

Both antiviral treatment with remdesivir and hemoadsorption using a CytoSorb® adsorption device are applied in the treatment of severe COVID-19. The CytoSorb® adsorber consists of porous polymer beads that adsorb a broad range of molecules, including cytokines but also several therapeutic drugs. In this study, we evaluated whether remdesivir and its main active metabolite GS-441524 would be adsorbed by CytoSorb® . Serum containing remdesivir or GS-441524 was circulated in a custom-made system containing a CytoSorb® device. Concentrations of remdesivir and GS-441524 before and after the adsorber were analyzed by liquid chromatography-tandem mass spectrometry. Measurements of remdesivir in the outgoing tube after the adsorber indicated almost complete removal of remdesivir by the device. In the reservoir, concentration of remdesivir showed an exponential decay and was not longer detectable after 60 mins. GS-441524 showed a similar exponential decay but, unlike remdesivir, it reached an adsorption-desorption equilibrium at ~48 µg/L. Remdesivir and its main active metabolite GS-441524 are rapidly eliminated from the perfusate by the CytoSorb® adsorber device in vitro. This should be considered in patients for whom both therapies are indicated, and simultaneous application should be avoided. In general, plasma levels of therapeutic drugs should be closely monitored under concurrent CytoSorb® therapy.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , COVID-19/terapia , Hemoperfusão/instrumentação , Monofosfato de Adenosina/sangue , Monofosfato de Adenosina/farmacocinética , Alanina/sangue , Alanina/farmacocinética , Análise Química do Sangue , COVID-19/sangue , Cromatografia Líquida , Terapia Combinada , Furanos/sangue , Furanos/farmacocinética , Hemoperfusão/efeitos adversos , Humanos , Pirróis/sangue , Pirróis/farmacocinética , Espectrometria de Massas em Tandem , Triazinas/sangue , Triazinas/farmacocinética
5.
J Pharm Biomed Anal ; 196: 113935, 2021 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-33548872

RESUMO

BACKGROUND: The present COVID-19 pandemic has prompted worldwide repurposing of drugs. The aim of the present work was to develop and validate a two-dimensional isotope-dilution liquid chromatrography tandem mass spectrometry (ID-LC-MS/MS) method for accurate quantification of remdesivir and its active metabolite GS-441524, chloroquine, hydroxychloroquine, lopinavir, ritonavir, favipiravir and azithromycin in serum; drugs that have gained attention for repurposing in the treatment of COVID-19. METHODS: Following protein precipitation, samples were separated with a two-dimensional ultra-high performance liquid chromatography (2D-UHPLC) setup, consisting of an online solid phase extraction (SPE) coupled to an analytical column. For quantification, stable isotope-labelled analogues were used as internal standards for all analytes. The method was validated on the basis of the European Medicines Agency bioanalytical method validation protocol. RESULTS: Detuning of lopinavir and ritonavir allowed simultaneous quantification of all analytes with different concentration ranges and sensitivity with a uniform injection volume of 5 µL. The method provided robust validation results with inaccuracy and imprecision values of ≤ 9.59 % and ≤ 11.1 % for all quality controls. CONCLUSION: The presented method is suitable for accurate and simultaneous quantification of remdesivir, its metabolite GS-441525, chloroquine, hydroxychloroquine, lopinavir, ritonavir, favipiravir and azithromycin in human serum. The quantitative assay may be an efficient tool for the therapeutic drug monitoring of these potential drug candidates in COVID-19 patients in order to increase treatment efficacy and safety.


Assuntos
Antivirais/sangue , Antivirais/uso terapêutico , COVID-19/sangue , COVID-19/tratamento farmacológico , Isótopos/química , SARS-CoV-2/efeitos dos fármacos , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/sangue , Alanina/análogos & derivados , Alanina/sangue , Amidas/sangue , Azitromicina/sangue , Cloroquina/sangue , Cromatografia Líquida/métodos , Furanos/sangue , Humanos , Hidroxicloroquina/sangue , Lopinavir/sangue , Pandemias/prevenção & controle , Pirazinas/sangue , Pirróis/sangue , Ritonavir/sangue , Espectrometria de Massas em Tandem/métodos , Triazinas/sangue
6.
J Clin Invest ; 131(2)2021 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-33463549

RESUMO

Mitochondrial disorders represent a large collection of rare syndromes that are difficult to manage both because we do not fully understand biochemical pathogenesis and because we currently lack facile markers of severity. The m.3243A>G variant is the most common heteroplasmic mitochondrial DNA mutation and underlies a spectrum of diseases, notably mitochondrial encephalomyopathy lactic acidosis and stroke-like episodes (MELAS). To identify robust circulating markers of m.3243A>G disease, we first performed discovery proteomics, targeted metabolomics, and untargeted metabolomics on plasma from a deeply phenotyped cohort (102 patients, 32 controls). In a validation phase, we measured concentrations of prioritized metabolites in an independent cohort using distinct methods. We validated 20 analytes (1 protein, 19 metabolites) that distinguish patients with MELAS from controls. The collection includes classic (lactate, alanine) and more recently identified (GDF-15, α-hydroxybutyrate) mitochondrial markers. By mining untargeted mass-spectra we uncovered 3 less well-studied metabolite families: N-lactoyl-amino acids, ß-hydroxy acylcarnitines, and ß-hydroxy fatty acids. Many of these 20 analytes correlate strongly with established measures of severity, including Karnofsky status, and mechanistically, nearly all markers are attributable to an elevated NADH/NAD+ ratio, or NADH-reductive stress. Our work defines a panel of organelle function tests related to NADH-reductive stress that should enable classification and monitoring of mitochondrial disease.


Assuntos
Síndrome MELAS/sangue , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alanina/sangue , Biomarcadores/sangue , Criança , Pré-Escolar , Feminino , Fator 15 de Diferenciação de Crescimento/sangue , Humanos , Hidroxibutiratos/sangue , Ácido Láctico/sangue , Síndrome MELAS/genética , Masculino , Pessoa de Meia-Idade , Mutação , Índice de Gravidade de Doença
7.
Anal Biochem ; 617: 114118, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33508271

RESUMO

Remdesivir (RDV) is a phosphoramidate prodrug designed to have activity against a broad spectrum of viruses. Following IV administration, RDV is rapidly distributed into cells and tissues and simultaneously metabolized into GS-441524 and GS-704277 in plasma. LC-MS/MS methods were validated for determination of the 3 analytes in human plasma that involved two key aspects to guarantee their precision, accuracy and robustness. First, instability issues of the analytes were overcome by diluted formic acid (FA) treatment of the plasma samples. Secondly, a separate injection for each analyte was performed with different ESI modes and organic gradients to achieve sensitivity and minimize carryover. Chromatographic separation was achieved on an Acquity UPLC HSS T3 column (2.1 × 50 mm, 1.8 µm) with a run time of 3.4 min. The calibration ranges were 4-4000, 2-2000, and 2-2000 ng/mL, respectively for RDV, GS-441524 and GS-704277. The intraday and interday precision (%CV) across validation runs at 3 QC levels for all 3 analytes was less than 6.6%, and the accuracy was within ±11.5%. The long-term storage stability in FA-treated plasma was established to be 392, 392 and 257 days at -70 °C, respectively for RDV, GS-441524 and GS-704277. The validated method was successfully applied in COVID-19 related clinical studies.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Antivirais/sangue , Monitoramento de Medicamentos/métodos , Furanos/sangue , Pirróis/sangue , Espectrometria de Massas em Tandem/métodos , Triazinas/sangue , Monofosfato de Adenosina/sangue , Alanina/sangue , COVID-19/tratamento farmacológico , Cromatografia Líquida de Alta Pressão/métodos , Humanos , Limite de Detecção
8.
J Pediatr ; 229: 175-181.e1, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33039387

RESUMO

OBJECTIVE: To validate our previously identified candidate metabolites, and to assess the ability of these metabolites to predict hypoxic-ischemic encephalopathy (HIE) both individually and combined with clinical data. STUDY DESIGN: Term neonates with signs of perinatal asphyxia, with and without HIE, and matched controls were recruited prospectively at birth from 2 large maternity units. Umbilical cord blood was collected for later batch metabolomic analysis by mass spectroscopy along with clinical details. The optimum selection of clinical and metabolites features with the ability to predict the development of HIE was determined using logistic regression modelling and machine learning techniques. Outcome of HIE was determined by clinical Sarnat grading and confirmed by electroencephalogram grade at 24 hours. RESULTS: Fifteen of 27 candidate metabolites showed significant alteration in infants with perinatal asphyxia or HIE when compared with matched controls. Metabolomic data predicted the development of HIE with an area under the curve of 0.67 (95% CI, 0.62-0.71). Lactic acid and alanine were the primary metabolite predictors for the development of HIE, and when combined with clinical data, gave an area under the curve of 0.96 (95% CI, 0.92-0.95). CONCLUSIONS: By combining clinical and metabolic data, accurate identification of infants who will develop HIE is possible shortly after birth, allowing early initiation of therapeutic hypothermia.


Assuntos
Sangue Fetal/metabolismo , Hipóxia-Isquemia Encefálica/diagnóstico , Alanina/sangue , Índice de Apgar , Asfixia Neonatal/complicações , Biomarcadores/sangue , Estudos de Casos e Controles , Eletroencefalografia , Humanos , Recém-Nascido , Ácido Láctico/sangue , Modelos Logísticos , Aprendizado de Máquina , Metabolômica , Valor Preditivo dos Testes , Estudos Prospectivos , Ressuscitação , Sensibilidade e Especificidade
10.
Sci Rep ; 10(1): 17616, 2020 10 19.
Artigo em Inglês | MEDLINE | ID: mdl-33077739

RESUMO

Currently, type 2 diabetes mellitus (T2DM) and obesity are major global public health issues, and their prevalence in the United Arab Emirates (UAE) are among the highest in the world. In 2019, The UAE diabetes national prevalence was 15.4%. In recent years there has been a considerable investigation of predictive biomarkers associated with these conditions. This study analysed fasting (8 h) blood samples from an obese, normoglycemic cohort and an obese, T2DM cohort of UAE nationals, employing clinical chemistry analysis, 1D 1H NMR and mass spectroscopy (FIA-MS/MS and LC-MS/MS) techniques. The novel findings reported for the first time in a UAE population revealed significant differences in a number of metabolites in the T2DM cohort. Metabolic fingerprints identified by NMR included BCAAs, trimethylamine N-oxide, ß-hydroxybutyrate, trimethyl uric acid, and alanine. A targeted MS approach showed significant differences in lysophosphatidylcholines, phosphatidylcholines, acylcarnitine, amino acids and sphingomyelins; Lyso.PC.a.C18.0, PC.ae.C34.2, C3.DC..C4.OH, glutamine and SM.C16.1, being the most significant metabolites. Pearson's correlation studies showed associations between these metabolites and the clinical chemistry parameters across both cohorts. This report identified differences in metabolites in response to T2DM in agreement with many published population studies. This contributes to the global search for a bank of metabolite biomarkers that can predict the advent of T2DM and give insight to its pathogenic mechanisms.


Assuntos
Biomarcadores/sangue , Diabetes Mellitus Tipo 2/diagnóstico , Obesidade/complicações , Ácido 3-Hidroxibutírico/sangue , Adulto , Alanina/sangue , Aminoácidos de Cadeia Ramificada/sangue , Cromatografia Líquida , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/etiologia , Feminino , Humanos , Masculino , Metabolômica , Metilaminas/sangue , Pessoa de Meia-Idade , Obesidade/sangue , Espectrometria de Massas em Tandem , Emirados Árabes Unidos , Ácido Úrico/sangue , Adulto Jovem
11.
Sci Rep ; 10(1): 17223, 2020 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-33057167

RESUMO

Systemic metabolic changes after renal transplantation reflect the key processes that are related to graft accommodation. In order to describe and better understand these changes, the 1HNMR based metabolomics approach was used. The changes of 47 metabolites in the serum samples of 19 individuals were interpreted over time with respect to their levels prior to transplantation. Considering the specific repeated measures design of the experiments, data analysis was mainly focused on the multiple analyses of variance (ANOVA) methods such as ANOVA simultaneous component analysis and ANOVA-target projection. We also propose here the combined use of ANOVA and classification and regression trees (ANOVA-CART) under the assumption that a small set of metabolites the binary splits on which may better describe the graft accommodation processes over time. This assumption is very important for developing a medical protocol for evaluating a patient's health state. The results showed that besides creatinine, which is routinely used to monitor renal activity, the changes in levels of hippurate, mannitol and alanine may be associated with the changes in renal function during the post-transplantation recovery period. Specifically, the level of hippurate (or histidine) is more sensitive to any short-term changes in renal activity than creatinine.


Assuntos
Nível de Saúde , Hipuratos/sangue , Falência Renal Crônica/diagnóstico , Transplante de Rim , Metabolômica/métodos , Monitorização Fisiológica/métodos , Alanina/sangue , Biomarcadores/sangue , Humanos , Falência Renal Crônica/sangue , Falência Renal Crônica/cirurgia , Espectroscopia de Ressonância Magnética , Manitol/sangue , Sensibilidade e Especificidade
12.
Virology ; 550: 61-69, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32882638

RESUMO

The world is in the midst of a pandemic caused by a novel coronavirus and is desperately searching for possible treatments. The antiviral remdesivir has shown some effectiveness against SARS-CoV-2 in vitro and in a recent animal study. We use data from a study of remdesivir in rhesus macaques to fit a viral kinetics model in an effort to determine the most appropriate mathematical descripton of the effect of remdesivir. We find statistically significant differences in the viral decay rate and use this to inform a possible mathematical formulation of the effect of remdesivir. Unfortunately, this model formulation suggests that the application of remdesivir will lengthen SARS-CoV-2 infections, putting into question its potential clinical benefit.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Antivirais/farmacocinética , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Modelos Estatísticos , Pneumonia Viral/tratamento farmacológico , Monofosfato de Adenosina/sangue , Monofosfato de Adenosina/farmacocinética , Alanina/sangue , Alanina/farmacocinética , Animais , Antivirais/sangue , Betacoronavirus/crescimento & desenvolvimento , Betacoronavirus/patogenicidade , COVID-19 , Infecções por Coronavirus/sangue , Infecções por Coronavirus/virologia , Modelos Animais de Doenças , Esquema de Medicação , Humanos , Inflamação , Macaca mulatta , Pandemias , Pneumonia Viral/sangue , Pneumonia Viral/virologia , SARS-CoV-2 , Carga Viral , Replicação Viral
13.
J Chromatogr Sci ; 58(9): 789-795, 2020 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-32776090

RESUMO

A simple, precise, rapid and accurate reversed-phase high-performance liquid chromatography (RP-HPLC) method was developed and validated for analysis of safinamide mesylate (SAF) in presence of its basic degradate, and co-administered drugs levodopa and ondansetron. The mobile phase consisted of acetonitrile and 20 mM potassium dihyrogen orthophosphate buffer having pH = 5 (40: 60 v/v). Quantification was achieved with ultraviolet detector at 226 nm. The linear range was 0.5-10 µg/mL with mean recovery ± SD of 99.72 ± 1.59. The peak purity of SAF in pharmaceutical preparation spiked with its degradate and co-administered drugs revealed symmetry factor (999.8) within the calculated threshold (>998.1). The suggested method was validated in compliance with the International Conference on Harmonization (ICH) guidelines and statistically compared with the manufacturer HPLC method with no significant difference in terms of accuracy and precision. The assay method was successfully used to estimate SAF in tablets with good percentage recoveries. The high sensitivity (lower than Cmax of the drug 0.65 µg/mL) of the proposed HPLC method enabled the determination of SAF in presence of its basic degradate and co-administered drug, ondansetron in human plasma with acceptable accuracy. The suggested HPLC method could be used in Quality Control (QC) lab for analysis of the studied drug in pharmaceutical preparation.


Assuntos
Alanina/análogos & derivados , Benzilaminas/sangue , Cromatografia Líquida de Alta Pressão/métodos , Cromatografia de Fase Reversa/métodos , Levodopa/sangue , Ondansetron/sangue , Alanina/sangue , Alanina/química , Benzilaminas/química , Humanos , Levodopa/química , Limite de Detecção , Modelos Lineares , Ondansetron/química , Reprodutibilidade dos Testes , Comprimidos
14.
Biopharm Drug Dispos ; 41(6): 268-272, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32557753

RESUMO

Rebamipide is used widely in East Asia for the treatment of gastric ulcers, acute gastritis, and exacerbated chronic gastritis. The objective of this study was to investigate the pharmacokinetic (PK) properties of rebamipide following single oral administration in rats and dogs. Eleven rats and dogs received single oral administrations of rebamipide (35 mg/kg and 100 mg, respectively). Blood samples were collected according to the assigned schedule, and the plasma concentration of rebamipide was determined using liquid chromatography-tandem mass spectrometry. A double-peak phenomenon was observed in the PK profile of rebamipide in rats. In contrast, rebamipide showed a conventional PK profile without double peaks in dogs. The half-life of rebamipide in rats (12.85 ± 7.86 h) was longer than that in dogs (5.62 ± 2.24 h), and the apparent total clearance (Clt /F) of rebamipide in rats (3.32 ± 1.18 L/h) was lower than that in dogs (105.01 ± 42.37 L/h). Simple allometric approaches showed that the correlation between body weight and Clt /F (R2 = 0.9287) among rats, dogs, and humans appeared satisfactory. This finding will help not only in understanding the pharmacology of rebamipide but also in establishing a strategy for in vivo evaluation of novel rebamipide formulations.


Assuntos
Alanina/análogos & derivados , Fármacos Gastrointestinais/farmacocinética , Quinolonas/farmacocinética , Administração Oral , Alanina/sangue , Alanina/farmacocinética , Animais , Cães , Fármacos Gastrointestinais/sangue , Meia-Vida , Masculino , Quinolonas/sangue , Ratos , Ratos Sprague-Dawley , Especificidade da Espécie
15.
Clin Chem Lab Med ; 58(9): 1461-1468, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32573468

RESUMO

Objectives: A method based on liquid chromatography coupled to triple quadrupole mass spectrometry detection using 50 µL of plasma was developed and fully validated for quantification of remdesivir and its active metabolites GS-441524. Methods: A simple protein precipitation was carried out using 75 µL of methanol containing the internal standard (IS) remdesivir-13C6 and 5 µL ZnSO4 1 M. After separation on Kinetex® 2.6 µm Polar C18 100A LC column (100 × 2.1 mm i.d.), both compounds were detected by a mass spectrometer with electrospray ionization in positive mode. The ion transitions used were m/z 603.3 â†’ m/z 200.0 and m/z 229.0 for remdesivir, m/z 292.2 â†’ m/z 173.1 and m/z 147.1 for GS-441524 and m/z 609.3 â†’ m/z 206.0 for remdesivir-13C6. Results: Calibration curves were linear in the 1-5000 µg/L range for remdesivir and 5-2500 for GS-441524, with limit of detection set at 0.5 and 2 µg/L and limit of quantification at 1 and 5 µg/L, respectively. Precisions evaluated at 2.5, 400 and 4000 µg/L for remdesivir and 12.5, 125, 2000 µg/L for GS-441524 were lower than 14.7% and accuracy was in the [89.6-110.2%] range. A slight matrix effect was observed, compensated by IS. Higher stability of remdesivir and metabolite was observed on NaF-plasma. After 200 mg IV single administration, remdesivir concentration decrease rapidly with a half-life less than 1 h while GS-441524 appeared rapidly and decreased slowly until H24 with a half-life around 12 h. Conclusions: This method would be useful for therapeutic drug monitoring of these compounds in Covid-19 pandemic.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Antivirais/sangue , Betacoronavirus , Cromatografia Líquida/métodos , Infecções por Coronavirus/sangue , Monitoramento de Medicamentos/métodos , Furanos/sangue , Pneumonia Viral/sangue , Pirróis/sangue , Espectrometria de Massas em Tandem/métodos , Triazinas/sangue , Monofosfato de Adenosina/sangue , Monofosfato de Adenosina/farmacocinética , Alanina/sangue , Alanina/farmacocinética , Antivirais/farmacocinética , COVID-19 , Estabilidade de Medicamentos , Feminino , Furanos/farmacocinética , Humanos , Limite de Detecção , Masculino , Pessoa de Meia-Idade , Pandemias , Pirróis/farmacocinética , Reprodutibilidade dos Testes , SARS-CoV-2 , Triazinas/farmacocinética
16.
AAPS J ; 22(4): 77, 2020 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-32458279

RESUMO

Remdesivir is one of the most promising drugs to treat COVID-19 based on the following facts: remdesivir has a broad-spectrum antiviral mechanism of action; it demonstrated in vitro activity against SARS-CoV-2 and in vivo efficacy in animal models against the similar coronavirus MERS-CoV; its safety profile has been tested in Ebola patients and in compassionate use in COVID-19 patients. Currently, remdesivir is being investigated in ten randomized controlled trials against COVID-19. The dose regimen of remdesivir is an IV loading dose of 200 mg on day 1 followed by daily IV maintenance doses of 100 mg for 5-9 days. Based on our data analysis, however, remdesivir with IV administration alone is unlikely to achieve excellent clinical efficacy. This analysis is based on the following observations: plasma exposures of remdesivir and its active metabolite are unlikely to be correlated with its clinical efficacy; remdesivir and its active metabolites are unlikely to be adequate in the lung to kill the SARS-CoV-2 virus. Even if remdesivir demonstrates benefits in the current randomized controlled trials, its efficacy may be limited. We suggest that a combination of an IV and pulmonary delivery dose regimen should be studied immediately to realize a potentially more effective antiviral therapy against COVID-19. Graphical abstract.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Monofosfato de Adenosina/administração & dosagem , Monofosfato de Adenosina/sangue , Monofosfato de Adenosina/química , Monofosfato de Adenosina/uso terapêutico , Administração por Inalação , Administração Intravenosa , Alanina/administração & dosagem , Alanina/sangue , Alanina/química , Alanina/uso terapêutico , Animais , COVID-19 , Humanos , Pandemias , SARS-CoV-2
17.
J Antimicrob Chemother ; 75(7): 1772-1777, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32361744

RESUMO

BACKGROUND: Remdesivir has received significant attention for its potential application in the treatment of COVID-19, caused by SARS-CoV-2. Remdesivir has already been tested for Ebola virus disease treatment and found to have activity against SARS and MERS coronaviruses. The remdesivir core contains GS-441524, which interferes with RNA-dependent RNA polymerases alone. In non-human primates, following IV administration, remdesivir is rapidly distributed into PBMCs and converted within 2 h to the active nucleoside triphosphate form, while GS-441524 is detectable in plasma for up to 24 h. Nevertheless, remdesivir pharmacokinetics and pharmacodynamics in humans are still unexplored, highlighting the need for a precise analytical method for remdesivir and GS-441524 quantification. OBJECTIVES: The validation of a reliable UHPLC-MS/MS method for remdesivir and GS-441524 quantification in human plasma. METHODS: Remdesivir and GS-441524 standards and quality controls were prepared in plasma from healthy donors. Sample preparation consisted of protein precipitation, followed by dilution and injection into the QSight 220 UHPLC-MS/MS system. Chromatographic separation was obtained through an Acquity HSS T3 1.8 µm, 2.1 × 50 mm column, with a gradient of water and acetonitrile with 0.05% formic acid. The method was validated using EMA and FDA guidelines. RESULTS: Analyte stability has been evaluated and described in detail. The method successfully fulfilled the validation process and it was demonstrated that, when possible, sample thermal inactivation could be a good choice in order to improve biosafety. CONCLUSIONS: This method represents a useful tool for studying remdesivir and GS-441524 clinical pharmacokinetics, particularly during the current COVID-19 outbreak.


Assuntos
Monofosfato de Adenosina/análogos & derivados , Trifosfato de Adenosina/análogos & derivados , Alanina/análogos & derivados , Cromatografia Líquida de Alta Pressão/métodos , Doença pelo Vírus Ebola/tratamento farmacológico , Espectrometria de Massas em Tandem/métodos , Monofosfato de Adenosina/análise , Monofosfato de Adenosina/sangue , Monofosfato de Adenosina/farmacocinética , Trifosfato de Adenosina/análise , Trifosfato de Adenosina/sangue , Trifosfato de Adenosina/farmacocinética , Alanina/análise , Alanina/sangue , Alanina/farmacocinética , Betacoronavirus , COVID-19 , Infecções por Coronavirus/tratamento farmacológico , Humanos , Pandemias , Pneumonia Viral/tratamento farmacológico , SARS-CoV-2 , Sensibilidade e Especificidade
18.
Ann Rheum Dis ; 79(4): 499-506, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32079570

RESUMO

OBJECTIVES: The differential diagnosis of seronegative rheumatoid arthritis (negRA) and psoriasis arthritis (PsA) is often difficult due to the similarity of symptoms and the unavailability of reliable clinical markers. Since chronic inflammation induces major changes in the serum metabolome and lipidome, we tested whether differences in serum metabolites and lipids could aid in improving the differential diagnosis of these diseases. METHODS: Sera from negRA and PsA patients with established diagnosis were collected to build a biomarker-discovery cohort and a blinded validation cohort. Samples were analysed by proton nuclear magnetic resonance. Metabolite concentrations were calculated from the spectra and used to select the variables to build a multivariate diagnostic model. RESULTS: Univariate analysis demonstrated differences in serological concentrations of amino acids: alanine, threonine, leucine, phenylalanine and valine; organic compounds: acetate, creatine, lactate and choline; and lipid ratios L3/L1, L5/L1 and L6/L1, but yielded area under the curve (AUC) values lower than 70%, indicating poor specificity and sensitivity. A multivariate diagnostic model that included age, gender, the concentrations of alanine, succinate and creatine phosphate and the lipid ratios L2/L1, L5/L1 and L6/L1 improved the sensitivity and specificity of the diagnosis with an AUC of 84.5%. Using this biomarker model, 71% of patients from a blinded validation cohort were correctly classified. CONCLUSIONS: PsA and negRA have distinct serum metabolomic and lipidomic signatures that can be used as biomarkers to discriminate between them. After validation in larger multiethnic cohorts this diagnostic model may become a valuable tool for a definite diagnosis of negRA or PsA patients.


Assuntos
Artrite Psoriásica/sangue , Artrite Reumatoide/sangue , Acetatos/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Alanina/sangue , Aminoácidos/sangue , Artrite Psoriásica/diagnóstico , Artrite Reumatoide/diagnóstico , Artrite Reumatoide/imunologia , Colina/sangue , Creatina/sangue , Diagnóstico Diferencial , Feminino , Humanos , Ácido Láctico/sangue , Lipidômica , Lipídeos/sangue , Masculino , Metaboloma , Metabolômica , Pessoa de Meia-Idade , Fosfocreatina/sangue , Espectroscopia de Prótons por Ressonância Magnética , Ácido Succínico/sangue
19.
J Pharm Biomed Anal ; 180: 113033, 2020 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-31841796

RESUMO

Mild cognitive impairment (MCI) is a transition phase between healthy individuals and Alzheimer's disease (AD). Therefore, diagnosis of MCI at early stage will help to delay or prevent its progression to disease. In the present study, we aim to identify the metabolic biomarkers, which can help in the diagnosis of MCI. We have screened 2000 elderly individuals from north India, out of which 200 were identified as MCI. We continued our study on 10 MCI individuals who regularly participated in the follow-up. The age and gender matched 10 healthy individuals were taken as control. These control and MCI individuals were subjected to neuropsychological examination such as Hindi mental state examination (HMSE) and Montreal cognitive assessment (MOCA) followed by 1H Nuclear Magnetic Resonance (NMR) analysis. Remarkable changes were noted between control and MCI individuals at metabolic level. In silico study showed the involvement of eight metabolites in MCI. We found higher level of lactate, N-acetyl aspartate, histidine and lower level of formate, choline, alanine, creatinine and glucose in blood plasma of MCI individuals compared to control. Further, In silico study showed that choline might be directly associated with MCI or AD. Such In silico study with quantitative metabolite analysis of plasma could be used as diagnostic biomarkers for the identification of MCI.


Assuntos
Biomarcadores/sangue , Biomarcadores/metabolismo , Disfunção Cognitiva/diagnóstico , Idoso , Alanina/sangue , Alanina/metabolismo , Glicemia/análise , Glicemia/metabolismo , Coleta de Amostras Sanguíneas , Colina/sangue , Colina/metabolismo , Simulação por Computador , Creatinina/sangue , Creatinina/metabolismo , Progressão da Doença , Feminino , Formiatos/sangue , Formiatos/metabolismo , Histidina/sangue , Histidina/metabolismo , Humanos , Índia , Ácido Láctico/sangue , Ácido Láctico/metabolismo , Espectroscopia de Ressonância Magnética , Metaboloma , Pessoa de Meia-Idade , Testes Neuropsicológicos
20.
Drug Metab Pharmacokinet ; 35(1): 131-138, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31727576

RESUMO

The present study aimed to characterize the relationships between plasma lactate, plasma alanine, monocarboxylate transporter (MCT) polymorphisms, and indices of diabetes in patients with type 2 diabetes (T2D) in Japan. Eighty-three patients with T2D were prospectively enrolled. The gluconeogenesis and glycogenolysis are enhanced and uptake of glucose is decreased in the T2D liver. Since the liver plays an important role in maintaining glucose metabolism, we examined the relationships between liver enzymes and indices of diabetes. Some studies have reported that MCT1 (SLC16A1) polymorphism causes metabolic diseases. In addition, a high frequency of MCT1 polymorphism was reported in a healthy Japanese population. However, little is known about the relationships between T2D and MCT polymorphisms. Plasma l-lactate concentration positively correlated with indices of diabetes (fasting plasma glucose [FPG] and hemoglobin A1c [HbA1c]) and with the liver enzymes alanine aminotransferase (ALT) and gamma-glutamyl transpeptidase (γ-GTP). MCT1 polymorphisms were associated with all of these markers. We identified no significant correlations between d-lactate or alanine concentrations and any of these markers, but a significant association was observed between l-lactate, a marker of oxidative capacity, and indices of diabetes. We conclude that plasma l-lactate concentration may represent a predictor of the progression or severity of T2D.


Assuntos
Alanina/sangue , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/genética , Ácido Láctico/sangue , Transportadores de Ácidos Monocarboxílicos/genética , Idoso , DNA/genética , Diabetes Mellitus Tipo 2/diagnóstico , Feminino , Variação Genética/genética , Humanos , Japão , Masculino , Pessoa de Meia-Idade , Transportadores de Ácidos Monocarboxílicos/metabolismo , Estudos Prospectivos
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