RESUMO
While grouping/read-across is widely used to fill data gaps, chemical registration dossiers are often rejected due to weak category justifications based on structural similarity only. Metabolomics provides a route to robust chemical categories via evidence of shared molecular effects across source and target substances. To gain international acceptance, this approach must demonstrate high reliability, and best-practice guidance is required. The MetAbolomics ring Trial for CHemical groupING (MATCHING), comprising six industrial, government and academic ring-trial partners, evaluated inter-laboratory reproducibility and worked towards best-practice. An independent team selected eight substances (WY-14643, 4-chloro-3-nitroaniline, 17α-methyl-testosterone, trenbolone, aniline, dichlorprop-p, 2-chloroaniline, fenofibrate); ring-trial partners were blinded to their identities and modes-of-action. Plasma samples were derived from 28-day rat tests (two doses per substance), aliquoted, and distributed to partners. Each partner applied their preferred liquid chromatography-mass spectrometry (LC-MS) metabolomics workflows to acquire, process, quality assess, statistically analyze and report their grouping results to the European Chemicals Agency, to ensure the blinding conditions of the ring trial. Five of six partners, whose metabolomics datasets passed quality control, correctly identified the grouping of eight test substances into three categories, for both male and female rats. Strikingly, this was achieved even though a range of metabolomics approaches were used. Through assessing intrastudy quality-control samples, the sixth partner observed high technical variation and was unable to group the substances. By comparing workflows, we conclude that some heterogeneity in metabolomics methods is not detrimental to consistent grouping, and that assessing data quality prior to grouping is essential. We recommend development of international guidance for quality-control acceptance criteria. This study demonstrates the reliability of metabolomics for chemical grouping and works towards best-practice.
Assuntos
Espectrometria de Massa com Cromatografia Líquida , Metabolômica , Ratos , Masculino , Feminino , Animais , Reprodutibilidade dos Testes , Metabolômica/métodos , Fluxo de TrabalhoRESUMO
PURPOSE: Breast cancer is associated with adverse outcomes in patients with the metabolic syndrome phenotype. To study this further, we examined the relationship between serum metabolite levels and the components of metabolic syndrome with treatment outcomes in breast cancer. METHODS: A total of 88 women with measurable breast cancer were studied; their serum metabolites as assessed by (1)H nuclear magnetic resonance spectroscopy, blood pressure, lipids, glucose, body mass index and waist circumference were recorded and correlated with treatment response. RESULTS: We identified metabolic syndrome in approximately half of our cohort (42 patients) and observed a significant trend (P = 0.03) of increased incidence of metabolic syndrome in partial response (33.3%), stable disease (42.9%) and progressive disease groups (66.1%). High blood sugar predicted a poor response (P < 0.001). Logistic regression of metabonomic data demonstrated that high lactate (P = 0.03) and low alanine (P = 0.01) combined with high glucose (P = 0.01) were associated with disease progression. CONCLUSIONS: Metabolic syndrome is commonly observed in metastatic breast cancer and these patients have poorer outcomes. These data, which support our previous findings, suggest that high blood glucose as part of metabolic syndrome is associated with a poor response in breast cancer. They also validate new therapeutic approaches that focus on metabolism.
Assuntos
Neoplasias da Mama/sangue , Carcinoma Ductal de Mama/sangue , Síndrome Metabólica/sangue , Fenótipo , Adulto , Idoso , Idoso de 80 Anos ou mais , Glicemia , Pressão Sanguínea , Neoplasias da Mama/complicações , Neoplasias da Mama/tratamento farmacológico , Carcinoma Ductal de Mama/complicações , Carcinoma Ductal de Mama/tratamento farmacológico , Feminino , Humanos , Modelos Logísticos , Síndrome Metabólica/complicações , Pessoa de Meia-Idade , Análise Multivariada , Pós-Menopausa , Resultado do Tratamento , Triglicerídeos/sangueRESUMO
1. A study of the rates and routes of excretion of 3-fluoro-[U-(14)C]aniline following intraperitoneal administration to male bile-cannulated rats by liquid scintillation counting (LSC) gave a total recovery of approximately 90% in the 48 h following dosing, with the majority of the dose being excreted in the urine during the first 24 h (approximately 49%). 2. The total recovery as determined by (19)F-nuclear magnetic resonance ((19)F-NMR) was approximately 49%, with the majority of the dose excreted in the first 24 h (approximately 41%). The comparatively low recovery in comparison to that obtained from LSC was due to matrix effects in bile and a contribution from metabolic defluorination. 3. High-performance liquid chromatography with radiometric profiling of urine and bile revealed a complex pattern of metabolites with the bulk of the dose excreted as a single peak. 4. Ultra-performance liquid chromatography-orthogonal acceleration time of flight mass spectrometry profiling also showed a complex pattern of metabolites, detecting approximately 21 metabolites of 3-fluoroaniline (3-FA) with six of these detected only in urine and four solely in bile. 5. (19)F-NMR revealed the presence of the parent compound and 15 metabolites in urine collected during the first 24 h after -dosing. The matrix effects of bile on (19)F-NMR spectroscopy made metabolite profiling impractical for this biofluid. The major metabolite of 3-FA was identified as 2-fluoro-4-acetamidophenol-sulfate.
Assuntos
Alanina/análogos & derivados , Ductos Biliares/metabolismo , Radioisótopos de Carbono/metabolismo , Radioisótopos de Carbono/farmacocinética , Alanina/metabolismo , Alanina/farmacocinética , Animais , Bile/química , Radioisótopos de Carbono/urina , Cateterismo , Cromatografia Líquida de Alta Pressão , Fezes/química , Espectroscopia de Ressonância Magnética , Masculino , Espectrometria de Massas , Radiometria , Ratos , Ratos Wistar , Contagem de CintilaçãoRESUMO
The non-selective beta-adrenergic receptor antagonist propranolol [1-(isopropylamino)-3-(1-naphthoxy)-2-propanol] is metabolised extensively in vivo. Enumerating and identifying the many metabolites that result from multiple biotransformations provides a considerable analytical challenge, greatly aided by efficient chromatography coupled to sensitive mass spectrometric detection. Here the use of the newly introduced high-resolution technique of "ultra performance liquid chromatography" (UPLC) linked to quadrupole time-of-flight mass spectrometry (TOFMS) with simultaneous [(14)C]-radioflow detection was applied to rapid metabolite profiling. [14C]-propranolol, dosed intraperitoneally to rat at 25 mg kg(-1) and 200 microCi kg(-1) was used as a model compound for this evaluation. Some 14 metabolites were detected in the urine by this technique including a number of conjugated metabolites such as sulphates, several isobaric glucuronides and two novel di-glucuronides.
Assuntos
Antagonistas Adrenérgicos beta/farmacocinética , Antagonistas Adrenérgicos beta/urina , Cromatografia Líquida/métodos , Espectrometria de Massas/métodos , Propranolol/farmacocinética , Propranolol/urina , Antagonistas Adrenérgicos beta/análise , Animais , Biotransformação , Radioisótopos de Carbono/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Masculino , Estrutura Molecular , Propranolol/análise , Propranolol/química , Ratos , Ratos Wistar , Urinálise/métodosRESUMO
As end products of xenobiotic metabolism via glutathione conjugation, mercapturic acids (MCAs) can be used as markers to indicate exposure to allylic compounds as well as the rate and efficiency of their excretion. In addition, the formation of certain MCAs indicates metabolism via the known toxin acrolein, a strong electrophile. High-resolution 1H NMR spectroscopy has been employed to quantitatively measure the presence of MCAs in the urine of Sprague-Dawley rats, collected in the 8 h following 25 and 50 mgkg(-1) i.p. doses of allyl formate (AF), a model toxin. 3-Hydroxypropylmercapturic acid (HPMA) was found to be the only 1H NMR-observable MCA excreted in the urine, exhibiting a percentage recovery of approximately 20% at the 25 mgkg(-1) dose level, and approximately 30% at the 50 mgkg(-1) dose level.
Assuntos
Acetilcisteína/análogos & derivados , Ésteres do Ácido Fórmico/toxicidade , Acetilcisteína/urina , Animais , Biomarcadores/urina , Relação Dose-Resposta a Droga , Ésteres do Ácido Fórmico/administração & dosagem , Glutationa/metabolismo , Injeções Intraperitoneais , Fígado/efeitos dos fármacos , Fígado/patologia , Espectroscopia de Ressonância Magnética , Masculino , Necrose , Ratos , Ratos Sprague-Dawley , Fatores de TempoRESUMO
[14C]-5-chloro-1,3-benzodioxol-4-amine was administered intraperitoneally (i.p.) to bile duct-cannulated rats (Alpk:ApfSD, Wistar derived) at 25 mg kg-1 to determine the rates and routes of excretion of the compound and to investigate its metabolic fate. A total of 89.1% of the dose was excreted in the 48 h following administration, the majority being recovered in the urine during the first 12 h. The main metabolite in both urine and bile, detected by high-performance liquid chromatography (HPLC) with radioprofiling and mass spectrometry, was identified as a demethylenated monosulfate conjugate. Unchanged parent compound formed a major component of the radiolabel excreted in urine and, in addition to unchanged parent and demethylenated sulphate conjugate, a large number of minor metabolites were detected in urine and bile. The overall metabolic fate of 5-chloro-1,3-benzodioxol-4-amine in the rat was complex, with some similarities to previously studied methylenedioxyphenyl compounds.