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1.
Bioorg Med Chem ; 28(22): 115723, 2020 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-33007547

RESUMO

Myeloperoxidase (MPO) is a heme peroxidase found in neutrophils, monocytes and macrophages that efficiently catalyzes the oxidation of endogenous chloride into hypochlorous acid for antimicrobial activity. Chronic MPO activation can lead to indiscriminate protein modification causing tissue damage, and has been associated with chronic inflammatory diseases, atherosclerosis, and acute cardiovascular events. Triazolopyrimidine 5 is a reversible MPO inhibitor; however it suffers from poor stability in acid, and is an irreversible inhibitor of the DNA repair protein methyl guanine methyl transferase (MGMT). Structure-based drug design was employed to discover benzyl triazolopyridines with improved MPO potency, as well as acid stability, no reactivity with MGMT, and selectivity against thyroid peroxidase (TPO). Structure-activity relationships, a crystal structure of the MPO-inhibitor complex, and acute in vivo pharmacodynamic data are described herein.


Assuntos
Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Peroxidase/antagonistas & inibidores , Piridinas/farmacologia , Triazóis/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Peroxidase/metabolismo , Piridinas/síntese química , Piridinas/química , Relação Estrutura-Atividade , Triazóis/síntese química , Triazóis/química
2.
Vet Pathol ; 55(2): 331-340, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29338616

RESUMO

C-terminal Src kinase (Csk) is one of the critical negative regulators of the Src family of kinases. The Src family of kinases are nonreceptor tyrosine kinases that regulate inflammation, cell proliferation, motility, and adhesion. To investigate potential histologic lesions associated with systemic loss of Csk gene activity in adult mice, conditional Csk-knockout mice were examined. Cre-mediated systemic excision of Csk induced by tamoxifen treatment resulted in multiorgan inflammation. Specifically, induction of Csk gene excision with three days of tamoxifen treatment resulted in greater than 90% gene excision. Strikingly, these mice developed enteritis that ranged from minimal and suppurative to severe, fibrinonecrosuppurative and hemorrhagic. Other inflammatory lesions included suppurative pneumonia, gastritis, and myocarditis, and increased numbers of inflammatory cells within the hepatic parenchyma. When tamoxifen treatment was reduced from three days to one day in an effort to lower the level of Csk gene excision and limit lesion development, the mice developed severe suppurative to pyogranulomatous pneumonia and minimal to mild suppurative enteritis. Lesions observed secondary to Csk gene excision suggest important roles for Csk in downregulating the proinflammatory activity of the Src family of kinases and limiting neutrophil-mediated inflammation.


Assuntos
Inflamação/veterinária , Camundongos Knockout/metabolismo , Supuração/veterinária , Quinases da Família src/metabolismo , Animais , Southern Blotting , Proteína Tirosina Quinase CSK , Feminino , Expressão Gênica , Inflamação/metabolismo , Inflamação/patologia , Masculino , Supuração/metabolismo , Supuração/patologia
3.
ACS Med Chem Lett ; 7(12): 1207-1212, 2016 Dec 08.
Artigo em Inglês | MEDLINE | ID: mdl-27994765

RESUMO

Introducing a uniquely substituted phenyl sulfone into a series of biphenyl imidazole liver X receptor (LXR) agonists afforded a dramatic potency improvement for induction of ATP binding cassette transporters, ABCA1 and ABCG1, in human whole blood. The agonist series demonstrated robust LXRß activity (>70%) with low partial LXRα agonist activity (<25%) in cell assays, providing a window between desired blood cell ABCG1 gene induction in cynomolgus monkeys and modest elevation of plasma triglycerides for agonist 15. The addition of polarity to the phenyl sulfone also reduced binding to the plasma protein, human α-1-acid glycoprotein. Agonist 15 was selected for clinical development based on the favorable combination of in vitro properties, excellent pharmacokinetic parameters, and a favorable lipid profile.

4.
Cell Metab ; 24(2): 223-33, 2016 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-27508871

RESUMO

The development of LXR agonists for the treatment of coronary artery disease has been challenged by undesirable properties in animal models. Here we show the effects of an LXR agonist on lipid and lipoprotein metabolism and neutrophils in human subjects. BMS-852927, a novel LXRß-selective compound, had favorable profiles in animal models with a wide therapeutic index in cynomolgus monkeys and mice. In healthy subjects and hypercholesterolemic patients, reverse cholesterol transport pathways were induced similarly to that in animal models. However, increased plasma and hepatic TG, plasma LDL-C, apoB, apoE, and CETP and decreased circulating neutrophils were also evident. Furthermore, similar increases in LDL-C were observed in normocholesterolemic subjects and statin-treated patients. The primate model markedly underestimated human lipogenic responses and did not predict human neutrophil effects. These studies demonstrate both beneficial and adverse LXR agonist clinical responses and emphasize the importance of further translational research in this area.


Assuntos
Movimento Celular , Imidazóis/efeitos adversos , Imidazóis/farmacologia , Metabolismo dos Lipídeos , Lipoproteínas/metabolismo , Receptores X do Fígado/agonistas , Neutrófilos/metabolismo , Transportador 1 de Cassete de Ligação de ATP/genética , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Tecido Adiposo/metabolismo , Adolescente , Adulto , Animais , Movimento Celular/efeitos dos fármacos , Colesterol/sangue , Colesterol/metabolismo , Voluntários Saudáveis , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Hipercolesterolemia/sangue , Hipercolesterolemia/tratamento farmacológico , Imidazóis/uso terapêutico , Contagem de Leucócitos , Lipoproteínas/sangue , Macaca fascicularis , Macrófagos/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Sistema Fagocitário Mononuclear/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Triglicerídeos/sangue , Adulto Jovem
5.
J Pharmacol Exp Ther ; 352(2): 305-14, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25467132

RESUMO

Liver X Receptors (LXRs) α and ß are nuclear hormone receptors that regulate multiple genes involved in reverse cholesterol transport (RCT) and are potential drug targets for atherosclerosis. However, full pan agonists also activate lipogenic genes, resulting in elevated plasma and hepatic lipids. We report the pharmacology of BMS-779788 [2-(2-(1-(2-chlorophenyl)-1-methylethyl)-1-(3'-(methylsulfonyl)-4-biphenylyl)-1H-imidazol-4-yl)-2-propanol], a potent partial LXR agonist with LXRß selectivity, which has an improved therapeutic window in the cynomolgus monkey compared with a full pan agonist. BMS-779788 induced LXR target genes in blood in vivo with an EC50 = 610 nM, a value similar to its in vitro blood gene induction potency. BMS-779788 was 29- and 12-fold less potent than the full agonist T0901317 in elevating plasma triglyceride and LDL cholesterol, respectively, with similar results for plasma cholesteryl ester transfer protein and apolipoprotein B. However, ABCA1 and ABCG1 mRNA inductions in blood, which are critical for RCT, were comparable. Increased liver triglyceride was observed after 7-day treatment with BMS-779788 at the highest dose tested and was nearly identical to the dose response for plasma triglyceride, consistent with the central role of liver LXR in these lipogenic effects. Dose-dependent increases in biliary cholesterol and decreases in phospholipid and bile acid occurred in BMS-779788-treated animals, similar to LXR agonist effects reported in mouse. In summary, BMS-779788, a partial LXRß selective agonist, has decreased lipogenic potential compared with a full pan agonist in cynomolgus monkeys, with similar potency in the induction of genes known to stimulate RCT. This provides support in nonhuman primates for improving LXR agonist therapeutic windows by limiting LXRα activity.


Assuntos
Anticolesterolemiantes/farmacologia , Imidazóis/farmacologia , Fígado/efeitos dos fármacos , Receptores Nucleares Órfãos/agonistas , Sulfonas/farmacologia , Transportadores de Cassetes de Ligação de ATP/sangue , Transportadores de Cassetes de Ligação de ATP/genética , Animais , Anticolesterolemiantes/administração & dosagem , Anticolesterolemiantes/sangue , Relação Dose-Resposta a Droga , Agonismo Parcial de Drogas , Imidazóis/administração & dosagem , Imidazóis/sangue , Lipídeos/sangue , Lipogênese/efeitos dos fármacos , Fígado/metabolismo , Receptores X do Fígado , Macaca fascicularis , Masculino , Sulfonas/administração & dosagem , Sulfonas/sangue , Triglicerídeos/metabolismo
6.
J Med Chem ; 57(5): 2013-32, 2014 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-24521299

RESUMO

The biphenyl derivatives 2 and 3 are prototypes of a novel class of NS5A replication complex inhibitors that demonstrate high inhibitory potency toward a panel of clinically relevant HCV strains encompassing genotypes 1-6. However, these compounds exhibit poor systemic exposure in rat pharmacokinetic studies after oral dosing. The structure-activity relationship investigations that improved the exposure properties of the parent bis-phenylimidazole chemotype, culminating in the identification of the highly potent NS5A replication complex inhibitor daclatasvir (33) are described. An element critical to success was the realization that the arylglycine cap of 2 could be replaced with an alkylglycine derivative and still maintain the high inhibitory potency of the series if accompanied with a stereoinversion, a finding that enabled a rapid optimization of exposure properties. Compound 33 had EC50 values of 50 and 9 pM toward genotype-1a and -1b replicons, respectively, and oral bioavailabilities of 38-108% in preclinical species. Compound 33 provided clinical proof-of-concept for the NS5A replication complex inhibitor class, and regulatory approval to market it with the NS3/4A protease inhibitor asunaprevir for the treatment of HCV genotype-1b infection has recently been sought in Japan.


Assuntos
Antivirais/farmacologia , Inibidores Enzimáticos/farmacologia , Hepacivirus/efeitos dos fármacos , Imidazóis/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Animais , Antivirais/química , Antivirais/farmacocinética , Área Sob a Curva , Carbamatos , Cães , Descoberta de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Hepacivirus/enzimologia , Hepacivirus/fisiologia , Imidazóis/química , Imidazóis/farmacocinética , Espectroscopia de Ressonância Magnética , Pirrolidinas , Ratos , Espectrometria de Massas por Ionização por Electrospray , Relação Estrutura-Atividade , Valina/análogos & derivados
7.
Bioorg Med Chem Lett ; 23(11): 3157-61, 2013 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-23632269

RESUMO

Calcitonin gene-related peptide (CGRP) receptor antagonists have been shown to be efficacious as abortive migraine therapeutics with the absence of cardiovascular liabilities that are associated with triptans. Herein, we report the discovery of a highly potent CGRP receptor antagonist, BMS-742413, with the potential to provide rapid onset of action through intranasal delivery. The compound displays excellent aqueous solubility, oxidative stability, and toxicological profile. BMS-742413 has good intranasal bioavailability in the rabbit and shows a robust, dose-dependent inhibition of CGRP-induced increases in marmoset facial blood flow.


Assuntos
Amidas/química , Antagonistas do Receptor do Peptídeo Relacionado ao Gene de Calcitonina , Indazóis/química , Quinolonas/química , Administração Intranasal , Amidas/farmacologia , Amidas/uso terapêutico , Animais , Células CACO-2 , Callithrix , Vasos Coronários/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Face/irrigação sanguínea , Humanos , Indazóis/farmacologia , Indazóis/uso terapêutico , Transtornos de Enxaqueca/tratamento farmacológico , Quinolonas/farmacologia , Quinolonas/uso terapêutico , Coelhos , Ratos , Receptores de Peptídeo Relacionado com o Gene de Calcitonina/metabolismo , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/patologia
8.
Am J Pathol ; 178(3): 1117-25, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21356363

RESUMO

Chronic cholestatic liver injury induced by cholestasis in rodents is associated with hepatic fibrin deposition, and we found evidence of fibrin deposition in livers of patients with cholestasis. Key components of the fibrinolytic pathway modulate cholestatic liver injury by regulating activation of hepatocyte growth factor. However, the exact role of hepatic fibrin deposition in chronic cholestasis is not known. We tested the hypothesis that fibrinogen (Fbg) deficiency worsens liver injury induced by cholestasis. Fbg-deficient mice (Fbgα(-/-) mice) and heterozygous control mice (Fbgα(+/-) mice) were fed either the control diet or a diet containing 0.025% α-naphthylisothiocyanate (ANIT), which selectively injures bile duct epithelial cells in the liver, for 2 weeks. Hepatic fibrin and collagen deposits were evident in livers of heterozygous control mice fed the ANIT diet. Complete Fbg deficiency was associated with elevated serum bile acids, periportal necrosis, and increased serum alanine aminotransferase activity in mice fed the ANIT diet. Fbg deficiency was associated with enhanced hepatic expression of the transcription factor early growth response-1 (Egr-1) and enhanced induction of genes encoding the Egr-1-regulated proinflammatory chemokines monocyte chemotactic protein-1, KC growth-regulated protein, and macrophage inflammatory protein-2. Interestingly, peribiliary collagen deposition was not evident near necrotic areas in Fbg-deficient mice. The results suggest that in this model of chronic cholestasis, fibrin constrains the release of bile constituents from injured intrahepatic bile ducts, thereby limiting the progression of hepatic inflammation and hepatocellular injury.


Assuntos
Afibrinogenemia/complicações , Afibrinogenemia/metabolismo , Colestase/complicações , Colestase/metabolismo , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Fibrinogênio/metabolismo , Fígado/patologia , 1-Naftilisotiocianato/administração & dosagem , Afibrinogenemia/patologia , Idoso , Animais , Ductos Biliares/patologia , Colestase/induzido quimicamente , Colestase/patologia , Doença Crônica , Colágeno/metabolismo , Dieta , Modelos Animais de Doenças , Comportamento Alimentar , Feminino , Fibrina/deficiência , Fibrina/metabolismo , Regulação da Expressão Gênica , Humanos , Hiperplasia , Inflamação/complicações , Inflamação/genética , Inflamação/patologia , Fígado/metabolismo , Cirrose Hepática/complicações , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Neutrófilos/metabolismo , Xenobióticos
9.
Xenobiotica ; 41(2): 144-54, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21043805

RESUMO

2-Bromoethanamine (BEA) causes renal papillary necrosis (RPN) in rats after a single dose and has been widely used as a model compound for studying the lesion. Although the metabolism of BEA may be an important determinant of toxicity, the metabolic fate of the compound has not been fully elucidated. To date, the only identified BEA metabolites are aziridine, 2-oxazolidone and 5-hydroxy-2-oxazolidone. In this study, stable isotope labelling (SIL) of BEA analogs ((¹³C and ²H) were used to differentiate generated BEA metabolites from endogenous molecules which enabled the accurate liquid chromatography mass spectrometry detection of more than 180 novel metabolites. BEA metabolism was evaluated in rats after acute administration of a non-toxic dose (50 mg/kg) and a toxic dose (250 mg/kg) that caused frank RPN and polyuria. Newly identified metabolites include three carbamoylation products, two mercapturic acids and a group of amino acid conjugates. Overall, the results indicate that BEA metabolism is very complex, suggest the potential formation of reactive intermediates and establish that BEA is subject to conjugation with glutathione. The results also demonstrate the utility and sensitivity of the SIL approach for identification of metabolites from small, reactive compounds.


Assuntos
Carbamatos/metabolismo , Etilaminas/urina , Glutationa/metabolismo , Marcação por Isótopo/métodos , Aminoácidos/metabolismo , Animais , Etilaminas/química , Etilaminas/toxicidade , Rim/efeitos dos fármacos , Rim/patologia , Masculino , Ratos , Ratos Sprague-Dawley , Fatores de Tempo
10.
Toxicol Appl Pharmacol ; 250(3): 256-62, 2011 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-21070799

RESUMO

Cholestatic liver injury induced by alpha-naphthylisothiocyanate (ANIT) is provoked by injury to intrahepatic bile ducts and the progression of hepatic necrosis requires the procoagulant protein tissue factor (TF) and extrahepatic cells including neutrophils. Recent studies have shown that myeloid cell TF contributes to neutrophil activation. We tested the hypothesis that myeloid cell TF contributes to neutrophil activation in ANIT-treated mice. TF activity in liver homogenates increased significantly in TF(flox/flox) mice treated with ANIT, but not in TF(flox/flox)/LysMCre mice (TF(ΔMyeloid) mice), which have reduced TF expression in monocytes/macrophages and neutrophils. Myeloid cell-specific TF deficiency did not alter expression of the chemokines KC or MIP-2 but reduced hepatic neutrophil accumulation in ANIT-treated mice at 48 h as indicated by tissue myeloperoxidase (MPO) activity. Myeloid cell TF deficiency significantly reduced CD11b expression by blood neutrophils in ANIT-treated mice, and this was associated with reduced plasma MPO protein levels, an index of neutrophil degranulation. However, myeloid cell-specific TF deficiency had no effect on ANIT-induced coagulation cascade activation. The increase in serum ALT and ALP activities in ANIT-treated mice was reduced by myeloid cell TF deficiency (p<0.05), but the myeloid cell TF deficiency did not reduce hepatic necrosis at 48 h, as determined by histopathology and morphometry. The results suggest that myeloid cell TF contributes to neutrophil CD11b expression during cholestasis by a coagulation-independent pathway. However, the resultant reduction in neutrophil accumulation/activation is insufficient to substantially reduce ANIT hepatotoxicity, suggesting that myeloid cell TF is only one of many factors modulating hepatic necrosis during cholestasis.


Assuntos
1-Naftilisotiocianato/toxicidade , Antígeno CD11b/biossíntese , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Neutrófilos/efeitos dos fármacos , Tromboplastina/metabolismo , Xenobióticos/toxicidade , Animais , Coagulação Sanguínea/efeitos dos fármacos , Doença Hepática Induzida por Substâncias e Drogas/sangue , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Quimiocinas/biossíntese , Colestase/induzido quimicamente , Colestase/metabolismo , Molécula 1 de Adesão Intercelular/biossíntese , Fígado/efeitos dos fármacos , Fígado/enzimologia , Masculino , Camundongos , Células Mieloides/metabolismo , Ativação de Neutrófilo , Neutrófilos/metabolismo , Peroxidase/metabolismo
11.
Toxicol Sci ; 118(2): 510-20, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20823374

RESUMO

Chronic analgesic abuse has been shown to induce severe renal injury characterized by renal papillary necrosis (RPN), an injury detectable at late stage. While direct toxicity of the drug may exist, the molecular mechanisms underlying analgesics induction of RPN remain unknown. A major limitation to study the pathogenesis of RPN is the required chronic exposure before detection of injury. Here, we employed 2-bromoethanamine (BEA) to simulate rapid papillary toxicity using inner medullary collecting duct (IMCD3) cells. Although exposure to 10µM BEA had no effect on cellular viability under isotonic conditions, a 50% loss in cell viability was observed in the first 24 h when cells were subjected to sublethal hypertonic stress and nearly complete cell death after 48 h suggesting that BEA exerts cytotoxicity only under hypertonic conditions. Because TonEBP is a transcription factor critical for cell survival during hypertonic conditions, we undertook experiments to examine the effect of BEA on TonEBP expression and activity. Exposure of cells to 10µM BEA resulted in a substantial reduction in TonEBP protein expression after 24 h. In addition, TonEBP was not translocated to the nucleus in BEA-treated IMCD3 cells under acute hypertonic stress for transcription of target genes essential for osmolyte accumulation. Finally, we found a substantial decrease in TonEBP expression in medullary kidney tissues of mice injected with a single ip dose of BEA. Our data suggest that TonEBP is a potential target for BEA leading to the process of papillary necrosis in the settings of hypertonic stress.


Assuntos
Etilaminas/toxicidade , Medula Renal/efeitos dos fármacos , Necrose Papilar Renal/induzido quimicamente , Túbulos Renais Coletores/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Expressão Gênica/efeitos dos fármacos , Soluções Hipertônicas/farmacologia , Medula Renal/metabolismo , Medula Renal/patologia , Necrose Papilar Renal/metabolismo , Necrose Papilar Renal/patologia , Túbulos Renais Coletores/metabolismo , Túbulos Renais Coletores/patologia , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/metabolismo , Estresse Fisiológico/fisiologia , Fatores de Transcrição/genética
13.
Endocrinology ; 151(9): 4123-32, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20610571

RESUMO

The biology of IGF-IR/IR signaling was studied in normal mouse embryonic fibroblasts (MEFs) that were either wild type (wt), heterozygous (het), or null for the IGF-IR. The ability of IGF-I, IGF-II, or insulin to stimulate serum-starved MEFs was characterized by gene expression profiling and biochemical analyses for activation of downstream signals. Each genotypic group of MEFs exhibited distinct patterns of expression both while resting and in response to stimulation. The insulin receptor (IR) pathway in IGF-IR null MEFs was hypersensitive to insulin ligand stimulation resulting in greater AKT phosphorylation than in wt or het MEFs stimulated with the same ligand. Interestingly, the IR pathway hypersensitivity in IGF-IR null MEFs occurred with no observed changes in the levels of IR isoforms A or B. A new small molecule IGF-IR inhibitor (BMS-754807), having equipotent activity against both IGF-IR and IR, proved effective in suppressing both AKT and ERK phosphorylation from both the IGF-IR and IR pathways by all three ligands tested in wt, het, and null MEFs. The use of a dual IGF-IR/IR inhibitor addresses concerns about the use of growth inhibiting therapies directed against the IGF-IR receptor in certain cancers. Lastly, comparison of the antiproliferative effects (IC(50)s) of various compounds in wt vs. null MEFs demonstrates that genetically characterized MEFs provide a simple and inexpensive tool with which to define compounds as having mostly on-target or off-target IGF-IR activities because off-target compounds affect both wt and null MEFs equally.


Assuntos
Proliferação de Células/efeitos dos fármacos , Pirazóis/farmacologia , Receptor IGF Tipo 1/antagonistas & inibidores , Receptor de Insulina/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Triazinas/farmacologia , Animais , Western Blotting , Células Cultivadas , Análise por Conglomerados , Embrião de Mamíferos/citologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Perfilação da Expressão Gênica , Insulina/farmacologia , Fator de Crescimento Insulin-Like I/farmacologia , Fator de Crescimento Insulin-Like II/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Knockout , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor IGF Tipo 1/genética , Receptor IGF Tipo 1/metabolismo , Receptor de Insulina/genética , Receptor de Insulina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
14.
J Toxicol Environ Health A ; 73(1): 58-73, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-19953420

RESUMO

Acetaminophen (APAP)-induced hepatotoxicity accounts for nearly half of acute liver failure cases in the United States. The doses that produce hepatotoxicity vary considerably and many risk factors have been proposed, including liver inflammation from viral hepatitis. Interestingly, inflammatory stress from another stimulus, bacterial endotoxin (lipopolysaccharide, LPS), renders the liver more sensitive to hepatotoxicity from numerous xenobiotic agents. The purpose of these studies was to test the hypothesis that inflammation induced by LPS or infection with reovirus increases sensitivity to APAP-induced liver injury. For LPS-induced inflammation, C57BL/6J mice were treated with either saline or LPS (44 x 10(6) EU/kg, ip) 2 h before treatment with APAP (100-400 mg/kg, ip) or saline. No elevation in serum alanine aminotransferase (ALT) activity was observed in mice that received vehicle or LPS alone. LPS co-treatment produced a leftward shift of the dose-response curve for APAP-induced hepatotoxicity and led to significantly greater tumor necrosis factor-alpha (TNF) production than APAP alone. Reovirus serotype 1 (10(8) PFU, iv) induced inflammation in Balb/c mice as evidenced by increases in hepatic mRNAs for macrophage inhibitory protein-2, interleukin-6, and TNF. Co-administration of reovirus and APAP at doses of 450 and 700 mg/kg (2 h after reovirus) led to increases in serum ALT activity, whereas neither reovirus nor APAP alone produced liver injury. Consistent with the increases in serum ALT activity, histopathologic examination revealed centrilobular necrosis with marked neutrophilic accumulation only in livers of mice treated with LPS/APAP or with reovirus/APAP. The results suggest that normally noninjurious doses of APAP are rendered hepatotoxic by modest inflammation, whether bacterial or viral in origin.


Assuntos
Acetaminofen/efeitos adversos , Fígado/efeitos dos fármacos , Infecções por Reoviridae/imunologia , Animais , Citocinas/biossíntese , Relação Dose-Resposta a Droga , Feminino , Glutationa/metabolismo , Inflamação/induzido quimicamente , Inflamação/microbiologia , Inflamação/virologia , Lipopolissacarídeos/farmacologia , Fígado/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Neutrófilos/efeitos dos fármacos , Fator de Necrose Tumoral alfa/biossíntese
15.
J Proteome Res ; 9(1): 59-69, 2010 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19445528

RESUMO

Tissue injury and repair are often overlapping consequences of disease or toxic exposure, but are not often considered as distinct processes in molecular studies. To establish the systemic metabolic response to liver regeneration, the partial hepatectomy (PH) model has been studied in the rat by an integrated metabonomics strategy, utilizing (1)H NMR spectroscopy of urine, liver and serum. Male Sprague-Dawley rats were subjected to either surgical removal of approximately two-thirds of the liver, sham operated (SO) surgery, or no treatment (n = 10/group) and samples collected over a 7 day period. A number of urinary metabolic perturbations were observed in PH rats compared with SO and control animals, including elevated levels of taurine, hypotaurine, creatine, guanidinoacetic acid, betaine, dimethylglycine and bile acids. Serum betaine and creatine were also elevated after PH, while levels of triglyceride were reduced. In the liver, triglycerides, cholesterol, alanine and betaine were elevated after PH, while choline and its derivatives were reduced. Upon examining the dynamic pattern of urinary response (the 'metabolic trajectory'), several metabolites could be categorized into groups likely to reflect perturbations to different processes such as dietary intake or hepatic 1-carbon metabolism. Several of the urinary perturbations observed during the regenerative phase of the PH model have also been observed after exposure to liver toxins, indicating that hepatic regeneration may make a contribution to the systemic alterations in metabolism associated with hepatotoxicity. The observed changes in 1-carbon and lipid metabolism are consistent with the proposed role of these pathways in the activation of a regenerative response and provide further evidence regarding the utility of urinary NMR profiles in the detection of liver-specific pathology. Biofluid (1)H NMR-based metabolic profiling provides new insight into the role of metabolism of liver regeneration, and suggests putative biomarkers for the noninvasive monitoring of the regeneration process.


Assuntos
Regeneração Hepática/fisiologia , Fígado/fisiologia , Metabolômica/métodos , Ressonância Magnética Nuclear Biomolecular/métodos , Animais , Biomarcadores/análise , Análise Química do Sangue , Peso Corporal , Hepatectomia , Histocitoquímica , Fígado/química , Fígado/metabolismo , Fígado/cirurgia , Masculino , Tamanho do Órgão , Ratos , Ratos Sprague-Dawley , Urina/química
16.
Mol Cancer Ther ; 8(12): 3341-9, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19996272

RESUMO

BMS-754807 is a potent and reversible inhibitor of the insulin-like growth factor 1 receptor/insulin receptor family kinases (Ki, <2 nmol/L). It is currently in phase I development for the treatment of a variety of human cancers. BMS-754807 effectively inhibits the growth of a broad range of human tumor types in vitro, including mesenchymal (Ewing's, rhabdomyosarcoma, neuroblastoma, and liposarcoma), epithelial (breast, lung, pancreatic, colon, gastric), and hematopoietic (multiple myeloma and leukemia) tumor cell lines (IC50, 5-365 nmol/L); the compound caused apoptosis in a human rhabdomyosarcoma cell line, Rh41, as shown by an accumulation of the sub-G1 fraction, as well as by an increase in poly ADP ribose polymerase and Caspase 3 cleavage. BMS-754807 is active in vivo in multiple (epithelial, mesenchymal, and hematopoietic) xenograft tumor models with tumor growth inhibition ranging from 53% to 115% and at a minimum effective dose of as low as 6.25 mg/kg dosed orally daily. Combination studies with BMS-754807 have been done on multiple human tumor cell types and showed in vitro synergies (combination index, <1.0) when combined with cytotoxic, hormonal, and targeted agents. The combination of cetuximab and BMS-754807 in vivo, at multiple dose levels, resulted in improved clinical outcome over single agent treatment. These data show that BMS-754807 is an efficacious, orally active growth factor 1 receptor/insulin receptor family-targeted kinase inhibitor that may act in combination with a wide array of established anticancer agents.


Assuntos
Pirazóis/farmacologia , Receptor IGF Tipo 1/antagonistas & inibidores , Receptor de Insulina/antagonistas & inibidores , Triazinas/farmacologia , Animais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Humanizados , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Western Blotting , Caspase 3/metabolismo , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cetuximab , Relação Dose-Resposta a Droga , Teste de Tolerância a Glucose , Humanos , Concentração Inibidora 50 , Camundongos , Camundongos Nus , Estrutura Molecular , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologia , Poli(ADP-Ribose) Polimerases/metabolismo , Pirazóis/administração & dosagem , Pirazóis/farmacocinética , Receptor IGF Tipo 1/metabolismo , Receptor de Insulina/metabolismo , Triazinas/administração & dosagem , Triazinas/farmacocinética , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
17.
J Proteome Res ; 8(11): 5175-87, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19821561

RESUMO

The amino sugar galactosamine (galN) induces alterations in the hepatic uridine nucleotide pool and has been widely used as a model of human viral hepatitis. Histopathological and clinical chemistry analyses of a cohort of rats following administration of galN revealed extreme interindividual variability in the extent of the toxic response which enabled classification of 'responder' and 'non-responder' phenotypes. An integrative metabolic profiling approach was applied to characterize biomarkers of exposure to galN in urine, serum, feces and liver from responders and non-responders. The presence of N-acetylglucosamine and galN in the urine correlated with the occurrence and extent of toxic response. Conversely, the novel identification of galN-pyrazines in the feces of non-responders and their virtual absence in the feces of responders suggests an alternative means of distribution and metabolism of galN in non-responders. The absence of the UDP-hexosamines in the liver of non-responders further supports differential metabolism of galN and suggests an ability of non-responders to avoid UDP-glucose depletion. An observed disturbance of gut microbial derived metabolites in the urine and feces of non-responders may suggest a role of the microflora in reducing the effective dose of galN. This systems level metabonomic approach has provided new mechanistic insights into differential response to galN and is widely applicable to the study of interindividual variation in metabolism for any xenobiotic intervention.


Assuntos
Biomarcadores/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Galactosamina , Animais , Biomarcadores/química , Doença Hepática Induzida por Substâncias e Drogas/patologia , Fezes/química , Galactosamina/metabolismo , Galactosamina/toxicidade , Humanos , Fígado/química , Fígado/metabolismo , Fígado/patologia , Masculino , Estrutura Molecular , Ressonância Magnética Nuclear Biomolecular , Fenótipo , Ratos , Ratos Sprague-Dawley , Urina/química
18.
Magn Reson Chem ; 47 Suppl 1: S26-35, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19639609

RESUMO

The time-related metabolic effects of 1-cyano-2-hydroxy-3-butene (CHB, crambene), a naturally occurring nitrile and experimental model toxin causing exocrine pancreatitis, have been investigated in rats using high-resolution NMR spectroscopy of urine and serum in combination with pattern recognition analysis. Rats were administered CHB subcutaneously in two doses, 15 mg/kg dose (n = 10) and 150 mg/kg (n = 10), and conventional histopathology and clinical chemistry assessments were performed. Urine samples were collected at - 16 and 0, 8, 24, 48, 72, 96, 120, 144 and 168 h postdosing and serum samples were collected at 48 and 168 h postdosing; these were analyzed using a range of 1D and 2D NMR spectroscopic methods. The metabolic profile perturbations seen throughout the time-course of the study are described, and the application of the spectral correlation technique Statistical TOtal Correlation SpectroscopY (STOCSY) to detect both structural and novel toxicological connectivities between xenobiotic and endogenous metabolite signals is illustrated for the first time. As a result, it is suggested that the STOCSY approach may be of wider application in the identification of toxic versus nontoxic metabolites in drug metabolism studies.


Assuntos
Alcenos/intoxicação , Metabolômica , Nitrilas/intoxicação , Pâncreas Exócrino , Pancreatite/sangue , Pancreatite/urina , Animais , Peso Corporal , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Espectroscopia de Ressonância Magnética , Masculino , Estrutura Molecular , Tamanho do Órgão , Pâncreas Exócrino/patologia , Ratos , Ratos Sprague-Dawley , Padrões de Referência
19.
Am J Physiol Gastrointest Liver Physiol ; 296(4): G840-9, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19179621

RESUMO

Separation of concentrated bile acids from hepatic parenchymal cells is a key function of the bile duct epithelial cells (BDECs) that form intrahepatic bile ducts. Using coimmunostaining, we found that tissue factor (TF), the principal activator of coagulation, colocalized with cytokeratin 19, a marker of BDECs in the adult mouse liver. BDEC injury induced by xenobiotics such as alpha-naphthylisothiocyanate (ANIT) causes cholestasis, inflammation, and hepatocellular injury. We tested the hypothesis that acute ANIT-induced cholestatic hepatitis is associated with TF-dependent activation of coagulation and determined the role of TF in ANIT hepatotoxicity. Treatment of mice with ANIT (60 mg/kg) caused multifocal hepatic necrosis and significantly increased serum biomarkers of cholestasis and hepatic parenchymal cell injury. ANIT treatment also significantly increased liver TF expression and activity. ANIT-induced activation of the coagulation cascade was shown by increased plasma thrombin-antithrombin levels and significant deposition of fibrin within the necrotic foci. ANIT-induced coagulation and liver injury were reduced in low-TF mice, which express 1% of normal TF levels. The results indicate that ANIT-induced liver injury is accompanied by TF-dependent activation of the coagulation cascade and that TF contributes to the progression of injury during acute cholestatic hepatitis.


Assuntos
1-Naftilisotiocianato/toxicidade , Coagulação Sanguínea/fisiologia , Doença Hepática Induzida por Substâncias e Drogas/patologia , Colestase/induzido quimicamente , Tromboplastina/metabolismo , Animais , Regulação da Expressão Gênica/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tromboplastina/genética
20.
Artigo em Inglês | MEDLINE | ID: mdl-18472313

RESUMO

Urinary metabolic perturbations associated with acute and chronic acetaminophen-induced hepatotoxicity were investigated using nuclear magnetic resonance (NMR) spectroscopy and ultra performance liquid chromatography/mass spectrometry (UPLC/MS) metabonomics approaches to determine biomarkers of hepatotoxicity. Acute and chronic doses of acetaminophen (APAP) were administered to male Sprague-Dawley rats. NMR and UPLC/MS were able to detect both drug metabolites and endogenous metabolites simultaneously. The principal component analysis (PCA) of NMR or UPLC/MS spectra showed that metabolic changes observed in both acute and chronic dosing of acetaminophen were similar. Histopathology and clinical chemistry studies were performed and correlated well with the PCA analysis and magnitude of metabolite changes. Depletion of antioxidants (e.g. ferulic acid), trigonelline, S-adenosyl-L-methionine, and energy-related metabolites indicated that oxidative stress was caused by acute and chronic acetaminophen administration. Similar patterns of metabolic changes in response to acute or chronic dosing suggest similar detoxification and recovery mechanisms following APAP administration.


Assuntos
Acetaminofen/intoxicação , Doença Hepática Induzida por Substâncias e Drogas , Cromatografia Líquida de Alta Pressão/métodos , Biologia Computacional/métodos , Hepatopatias/urina , Espectrometria de Massas/métodos , Metabolismo , Ressonância Magnética Nuclear Biomolecular/métodos , Ratos Sprague-Dawley/urina , Animais , Fígado/patologia , Hepatopatias/patologia , Masculino , Necrose/patologia , Necrose/urina , Ratos
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