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1.
Proc Natl Acad Sci U S A ; 117(46): 28992-29000, 2020 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-33139537

RESUMO

Fibroblast growth factor 21 (FGF21) controls metabolic organ homeostasis and eating/drinking behavior via FGF receptor 1/Klothoß (FGFR1/KLB) complexes expressed in adipocytes, pancreatic acinar cells, and the nervous system in mice. Chronic administration of recombinant FGF21 or engineered variants improves metabolic health in rodents, nonhuman primates, and humans; however, the rapid turnover of these molecules limits therapeutic utility. Here we show that the bispecific anti-FGFR1/KLB agonist antibody BFKB8488A induced marked weight loss in obese cynomolgus monkeys while elevating serum adiponectin and the adipose expression of FGFR1 target genes, demonstrating its action as an FGF21 mimetic. In a randomized, placebo-controlled, single ascending-dose study in overweight/obese human participants, subcutaneous BFKB8488A injection caused transient body weight reduction, sustained improvement in cardiometabolic parameters, and a trend toward reduction in preference for sweet taste and carbohydrate intake. These data suggest that specific activation of the FGFR1/KLB complex in humans can be used as therapy for obesity-related metabolic defects.


Assuntos
Preferências Alimentares , Obesidade/tratamento farmacológico , Obesidade/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Adiponectina/sangue , Tecido Adiposo/metabolismo , Adolescente , Adulto , Idoso , Animais , Anticorpos/uso terapêutico , Biomarcadores/sangue , Peso Corporal , Feminino , Fatores de Crescimento de Fibroblastos , Homeostase , Humanos , Macaca fascicularis , Masculino , Camundongos , Pessoa de Meia-Idade , Redução de Peso , Adulto Jovem
2.
Int J Clin Pharmacol Ther ; 59(3): 198-201, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33210993

RESUMO

OBJECTIVE: To assess the effect of green tea intake on the pharmacokinetics of the ß-blocker celiprolol. MATERIALS AND METHOD: In an open-label crossover study, 3 healthy subjects were given water or a green tea beverage daily for 3 days. On day 4, each subject received a single oral dose of 200 mg celiprolol with water or green tea. Serum and urinary concentrations of celiprolol were measured for up to 24 hours. RESULTS: Green tea intake decreased the area under the serum concentration-time curve and urinary excretion of celiprolol by 98.6 and 98.0%, respectively. CONCLUSION: Green tea intake might have a negative impact on the clinical effectiveness of celiprolol.


Assuntos
Celiprolol , Chá , Antagonistas Adrenérgicos beta , Estudos Cross-Over , Voluntários Saudáveis , Humanos
3.
J Clin Pharm Ther ; 44(1): 119-121, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30311244

RESUMO

WHAT IS KNOWN AND OBJECTIVE: Multi-drug combinations often make chemotherapy difficult owing to drug-drug interactions (DDIs). We report a rare and difficult-to-treat case due to DDIs between drugs for Mycobacterium avium complex (MAC) infection and antiepileptic drugs. CASE DESCRIPTION: A 70-year-old Japanese woman was diagnosed as having pulmonary MAC disease. She had a history of symptomatic epilepsy, which was successfully treated with phenytoin and phenobarbital. Serum phenytoin concentrations increased upon the initiation of MAC infection treatment. WHAT IS NEW AND CONCLUSION: We evaluated DDIs and adjusted the dosage of drugs by monitoring the serum drug level.


Assuntos
Antibacterianos/farmacologia , Anticonvulsivantes/farmacocinética , Fenitoína/farmacocinética , Idoso , Antibacterianos/administração & dosagem , Anticonvulsivantes/administração & dosagem , Relação Dose-Resposta a Droga , Interações Medicamentosas , Epilepsia/tratamento farmacológico , Feminino , Humanos , Complexo Mycobacterium avium/isolamento & purificação , Infecção por Mycobacterium avium-intracellulare/tratamento farmacológico , Fenobarbital/administração & dosagem , Fenitoína/administração & dosagem
4.
J Am Soc Nephrol ; 29(1): 69-80, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-28993502

RESUMO

The bone-derived hormone fibroblast growth factor-23 (FGF-23) activates complexes composed of FGF receptors (FGFRs), including FGFR1, and α-Klotho in the kidney distal tubule (DT), leading to increased sodium retention and hypertension. However, the role of FGFR1 in regulating renal processes linked to hypertension is unclear. Here, we investigated the effects of selective FGFR1 loss in the DT. Conditional knockout (cKO) of FGFR1 in the DT (FGFR1DT-cKO mice) resulted in left ventricular hypertrophy (LVH) and decreased kidney expression of α-Klotho in association with enhanced BP, decreased expression of angiotensin converting enzyme 2, and increased expression of the Na+-K+-2Cl- cotransporter. Notably, recombinant FGF-23 administration similarly decreased the kidney expression of α-Klotho and induced LVH in mice. Pharmacologic activation of FGFR1 with a monoclonal anti-FGFR1 antibody (R1MAb1) normalized BP and significantly attenuated LVH in the Hyp mouse model of excess FGF-23, but did not induce a response in FGFR1DT-cKO mice. The hearts of FGFR1DT-cKO mice showed increased expression of the transient receptor potential cation channel, subfamily C, member 6 (TRPC6), consistent with cardiac effects of soluble Klotho deficiency. Moreover, administration of recombinant soluble Klotho lowered BP in the Hyp mice. Thus, FGFR1 in the DT regulates systemic hemodynamic responses opposite to those predicted by the actions of FGF-23. These cardiovascular effects appear to be mediated by paracrine FGF control of kidney FGFR1 and subsequent regulation of soluble Klotho and TRPC6. FGFR1 in the kidney may provide a new molecular target for treating hypertension.


Assuntos
Pressão Sanguínea/efeitos dos fármacos , Pressão Sanguínea/genética , Hipertensão/genética , Hipertrofia Ventricular Esquerda/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Enzima de Conversão de Angiotensina 2 , Animais , Anticorpos Monoclonais/farmacologia , Feminino , Fator de Crescimento de Fibroblastos 23 , Fatores de Crescimento de Fibroblastos/farmacologia , Glucuronidase/genética , Glucuronidase/metabolismo , Fatores Imunológicos/farmacologia , Túbulos Renais Distais , Proteínas Klotho , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miocárdio/metabolismo , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , RNA Mensageiro/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia , Proteínas Recombinantes/farmacologia , Simportadores de Cloreto de Sódio-Potássio/genética , Simportadores de Cloreto de Sódio-Potássio/metabolismo , Canais de Cátion TRPC/genética , Canais de Cátion TRPC/metabolismo , Canal de Cátion TRPC6
5.
J Biol Chem ; 291(11): 5986-5996, 2016 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-26797127

RESUMO

FGF21 is a stress-induced hormone with potent anti-obesity, insulin-sensitizing, and hepatoprotective properties. Although proteolytic cleavage of recombinant human FGF21 in preclinical species has been observed previously, the regulation of endogenously produced FGF21 is not well understood. Here we identify fibroblast activation protein (FAP) as the enzyme that cleaves and inactivates human FGF21. A selective chemical inhibitor, immunodepletion, or genetic deletion of Fap stabilized recombinant human FGF21 in serum. In addition, administration of a selective FAP inhibitor acutely increased circulating intact FGF21 levels in cynomolgus monkeys. On the basis of our findings, we propose selective FAP inhibition as a potential therapeutic approach to increase endogenous FGF21 activity for the treatment of obesity, type 2 diabetes, non-alcoholic steatohepatitis, and related metabolic disorders.


Assuntos
Fatores de Crescimento de Fibroblastos/metabolismo , Gelatinases/metabolismo , Proteínas de Membrana/metabolismo , Serina Endopeptidases/metabolismo , Sequência de Aminoácidos , Animais , Endopeptidases , Fatores de Crescimento de Fibroblastos/química , Gelatinases/genética , Deleção de Genes , Células HEK293 , Humanos , Macaca fascicularis , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Dados de Sequência Molecular , Proteólise , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Serina Endopeptidases/genética
6.
J Biol Chem ; 290(40): 24166-77, 2015 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-26272615

RESUMO

Fibroblast growth factors receptors (FGFRs) are thought to initiate intracellular signaling cascades upon ligand-induced dimerization of the extracellular domain. Although the existence of unliganded FGFR1 dimers on the surface of living cells has been proposed, this notion remains rather controversial. Here, we employed time-resolved Förster resonance energy transfer combined with SNAP- and ACP-tag labeling in COS7 cells to monitor dimerization of full-length FGFR1 at the cell-surface with or without the coreceptor ßKlotho. Using this approach we observed homodimerization of unliganded FGFR1 that is independent of its surface density. The homo-interaction signal observed for FGFR1 was indeed as robust as that obtained for epidermal growth factor receptor (EGFR) and was further increased by the addition of activating ligands or pathogenic mutations. Mutational analysis indicated that the kinase and the transmembrane domains, rather than the extracellular domain, mediate the ligand-independent FGFR1 dimerization. In addition, we observed a formation of a higher order ligand-independent complex by the c-spliced isoform of FGFR1 and ßKlotho. Collectively, our approach provides novel insights into the assembly and dynamics of the full-length FGFRs on the cell surface.


Assuntos
Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/química , Animais , Células COS , Membrana Celular/metabolismo , Chlorocebus aethiops , Análise Mutacional de DNA , Ensaio de Imunoadsorção Enzimática , Receptores ErbB/metabolismo , Fatores de Crescimento de Fibroblastos/química , Citometria de Fluxo , Transferência Ressonante de Energia de Fluorescência , Humanos , Proteínas Klotho , Ligantes , Proteínas de Membrana/metabolismo , Mutação , Fosforilação , Ligação Proteica , Multimerização Proteica , Transdução de Sinais , Relação Estrutura-Atividade
7.
Sci Rep ; 9(1): 8833, 2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31222070

RESUMO

In response to environmental and nutrient stress, adipose tissues must establish a new homeostatic state. Here we show that cold exposure of obese mice triggers an adaptive tissue remodeling in visceral adipose tissue (VAT) that involves extracellular matrix deposition, angiogenesis, sympathetic innervation, and adipose tissue browning. Obese VAT is predominated by pro-inflammatory M1 macrophages; cold exposure induces an M1-to-M2 shift in macrophage composition and dramatic changes in macrophage gene expression in both M1 and M2 macrophages. Antibody-mediated CSF1R blocking prevented the cold-induced recruitment of adipose tissue M2 macrophages, suggesting the role of CSF1R signaling in the process. These cold-induced effects in obese VAT are phenocopied by an administration of the FGF21-mimetic antibody, consistent with its action to stimulate sympathetic nerves. Collectively, these studies illuminate adaptive visceral adipose tissue plasticity in obese mice in response to cold stress and antibody-based metabolic therapy.


Assuntos
Adaptação Fisiológica , Anticorpos/farmacologia , Resposta ao Choque Frio , Gordura Intra-Abdominal/fisiologia , Animais , Movimento Celular , Fatores de Crescimento de Fibroblastos/imunologia , Macrófagos/citologia , Macrófagos/imunologia , Camundongos , Camundongos Obesos , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/imunologia , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Células Estromais/fisiologia
8.
FEBS Lett ; 582(1): 2-9, 2008 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-18023286

RESUMO

Nuclear receptors (NR) are a superfamily of ligand-activated transcription factors that regulate development, reproduction, and metabolism of lipids, drugs and energy. The importance of this family of proteins in metabolic disease is exemplified by NR ligands used in the clinic or under exploratory development for the treatment of diabetes mellitus, dyslipidemia, hypercholesterolemia, or other metabolic abnormalities. Genetic studies in humans and rodents support the notion that NRs control a wide variety of metabolic processes by regulating the expression of genes encoding key enzymes, transporters and other proteins involved in metabolic homeostasis. Current knowledge of complex NR metabolic networks is summarized here.


Assuntos
Doenças Metabólicas/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Humanos , Conformação Proteica , Receptores Citoplasmáticos e Nucleares/química , Triglicerídeos/metabolismo
9.
Toxicol Appl Pharmacol ; 232(2): 337-50, 2008 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-18692084

RESUMO

Cytochromes P450 (P450s) and glutathione S-transferases (GSTs) constitute two important enzyme families involved in carcinogen metabolism. Generally, P450s play activation or detoxifying roles while GSTs act primarily as detoxifying enzymes. We previously demonstrated that oral administration of the linear furanocoumarins, isopimpinellin and imperatorin, modulated P450 and GST activities in various tissues of mice. The purpose of the present study was to compare a broader range of naturally occurring coumarins (simple coumarins, and furanocoumarins of the linear and angular type) for their abilities to modulate hepatic drug-metabolizing enzymes when administered orally to mice. We now report that all of the different coumarins tested (coumarin, limettin, auraptene, angelicin, bergamottin, imperatorin and isopimpinellin) induced hepatic GST activities, whereas the linear furanocoumarins possessed the greatest abilities to induce hepatic P450 activities, in particular P450 2B and 3A. In both cases, this corresponded to an increase in protein expression of the enzymes. Induction of P4502B10, 3A11, and 2C9 by xenobiotics often is a result of activation of the pregnane X receptor (PXR) and/or constitutive androstane receptor (CAR). Using a pregnane X receptor reporter system, our results demonstrated that isopimpinellin activated both PXR and its human ortholog SXR by recruiting coactivator SRC-1 in transfected cells. In CAR transfection assays, isopimpinellin counteracted the inhibitory effect of androstanol on full-length mCAR, a Gal4-mCAR ligand-binding domain fusion, and restored coactivator binding. Orally administered isopimpinellin induced hepatic mRNA expression of Cyp2b10, Cyp3a11, and GSTain CAR(+/+) wild-type mice. In contrast, the induction of Cyp2b10 mRNA by isopimpinellin was attenuated in the CAR(-/-) mice, suggesting that isopimpinellin induces Cyp2b10 via the CAR receptor. Overall, the current data indicate that naturally occurring coumarins have diverse activities in terms of inducing various xenobiotic metabolizing enzymes based on their chemical structure.


Assuntos
Cumarínicos/administração & dosagem , Fígado/efeitos dos fármacos , Fígado/enzimologia , Animais , Cumarínicos/química , Cumarínicos/toxicidade , Sistema Enzimático do Citocromo P-450/metabolismo , Relação Dose-Resposta a Droga , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos SENCAR , Camundongos Knockout
10.
J Pharmacol Toxicol Methods ; 94(Pt 1): 19-25, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29660397

RESUMO

INTRODUCTION: bFKB1 is a humanized bispecific IgG1 antibody, created by conjoining an anti-Fibroblast Growth Factor Receptor 1 (FGFR1) half-antibody to an anti-Klothoß (KLB) half-antibody, using the knobs-into-holes strategy. bFKB1 acts as a highly selective agonist for the FGFR1/KLB receptor complex and is intended to ameliorate obesity-associated metabolic defects by mimicking the activity of the hormone FGF21. An important aspect of the biologics product manufacturing process is to establish meaningful product specifications regarding the tolerable levels of impurities that copurify with the drug product. The aim of the current study was to determine acceptable levels of product-related impurities for bFKB1. METHODS: To determine the tolerable levels of these impurities, we dosed obese mice with bFKB1 enriched with various levels of either HMW impurities or anti-FGFR1-related impurities, and measured biomarkers for KLB-independent FGFR1 signaling. RESULTS: Here, we show that product-related impurities of bFKB1, in particular, high molecular weight (HMW) impurities and anti-FGFR1-related impurities, when purposefully enriched, stimulate FGFR1 in a KLB-independent manner. By taking this approach, the tolerable levels of product-related impurities were successfully determined. DISCUSSION: Our study demonstrates a general pharmacology-guided approach to setting a product specification for a bispecific antibody whose homomultimer-related impurities could lead to undesired biological effects.


Assuntos
Anticorpos/química , Anticorpos/farmacologia , Animais , Células CHO , Linhagem Celular , Cricetulus , Modelos Animais de Doenças , Combinação de Medicamentos , Células HEK293 , Humanos , Imunoglobulina G/química , Masculino , Proteínas de Membrana/química , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Peso Molecular , Obesidade/tratamento farmacológico , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/química , Transdução de Sinais/efeitos dos fármacos
11.
Horm Mol Biol Clin Investig ; 30(2)2017 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-28525362

RESUMO

Fibroblast growth factor 21 (FGF21) analogs and FGF21 receptor agonists (FGF21RAs) that mimic FGF21 ligand activity constitute the new "FGF21-class" of anti-obesity and anti-diabetic molecules that improve insulin sensitivity, ameliorate hepatosteatosis and promote weight loss. The metabolic actions of FGF21-class proteins in obese mice are attributed to stimulation of brown fat thermogenesis and increased secretion of adiponectin. The therapeutic utility of this class of molecules is being actively investigated in clinical trials for the treatment of type 2 diabetes and non-alcoholic steatohepatitis (NASH). This review is focused on various FGF21-class molecules, their molecular designs and the preclinical and clinical activities. These molecules include modified FGF21 as well as agonistic antibodies against the receptor for FGF21, namely the complex of FGF receptor 1 (FGFR1) and the obligatory coreceptor ßKlotho (KLB). In addition, a novel approach to increase endogenous FGF21 activity by inhibiting the FGF21-degrading protease fibroblast activation protein (FAP) is discussed.


Assuntos
Fatores de Crescimento de Fibroblastos/metabolismo , Obesidade/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/agonistas , Animais , Metabolismo Energético/efeitos dos fármacos , Fatores de Crescimento de Fibroblastos/farmacologia , Fatores de Crescimento de Fibroblastos/uso terapêutico , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Obesidade/tratamento farmacológico , Obesidade/genética , Estabilidade Proteica , Transdução de Sinais/efeitos dos fármacos , Transcrição Gênica
12.
Sci Rep ; 7(1): 12524, 2017 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-28970566

RESUMO

Fibroblast Activation Protein (FAP) is a membrane-bound serine protease whose expression is often elevated in activated fibroblasts associated with tissue remodeling in various common diseases such as cancer, arthritis and fibrosis. Like the closely related dipeptidyl peptidase DPPIV, the extracellular domain of FAP can be released into circulation as a functional enzyme, and limited studies suggest that the circulating level of FAP correlates with the degree of tissue fibrosis. Here we describe a novel homogeneous fluorescence intensity assay for circulating FAP activity based on a recently identified natural substrate, FGF21. This assay is unique in that it can effectively distinguish endopeptidase activity of FAP from that of other related enzymes such as prolyl endopeptidase (PREP) and was validated using Fap-deficient mice. Structural modeling was used to elucidate the mechanistic basis for the observed specificity in substrate recognition by FAP, but not by DPPIV or PREP. Finally, the assay was used to detect elevated FAP activity in human patients diagnosed with liver cirrhosis and to determine the effectiveness of a chemical inhibitor for FAP in mice. We propose that the assay presented here could thus be utilized for diagnosis of FAP-related pathologies and for the therapeutic development of FAP inhibitors.


Assuntos
Fatores de Crescimento de Fibroblastos/genética , Fibrose/genética , Gelatinases/genética , Cirrose Hepática/genética , Proteínas de Membrana/genética , Serina Endopeptidases/genética , Animais , Endopeptidases/genética , Fibroblastos/enzimologia , Fibroblastos/metabolismo , Fibrose/enzimologia , Fibrose/patologia , Regulação Enzimológica da Expressão Gênica , Humanos , Cirrose Hepática/enzimologia , Cirrose Hepática/patologia , Camundongos , Prolil Oligopeptidases , Especificidade por Substrato
13.
Protein Eng Des Sel ; 30(9): 627-637, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28985411

RESUMO

Bispecific antibodies offer a clinically validated platform for drug discovery. In generating functionally active bispecific antibodies, it is necessary to identify a unique parental antibody pair to merge into a single molecule. However, technologies that allow high-throughput production of bispecific immunoglobulin Gs (BsIgGs) for screening purposes are limited. Here, we describe a novel bispecific antibody format termed tethered-variable CLBsIgG (tcBsIgG) that allows robust production of intact BsIgG in a single cell line, concurrently ensuring cognate light chain pairing and preserving key antibody structural and functional properties. This technology is broadly applicable in the generation of BsIgG from a variety of antibody isotypes, including human BsIgG1, BsIgG2 and BsIgG4. The practicality of the tcBsIgG platform is demonstrated by screening BsIgGs generated from FGF21-mimetic anti-Klotho-ß agonistic antibodies in a combinatorial manner. This screen identified multiple biepitopic combinations with enhanced agonistic activity relative to the parental monoclonal antibodies, thereby demonstrating that biepitopic antibodies can acquire enhanced functionality compared to monospecific parental antibodies. By design, the tcBsIgG format is amenable to high-throughput production of large panels of bispecific antibodies and thus can facilitate the identification of rare BsIgG combinations to enable the discovery of molecules with improved biological function.


Assuntos
Anticorpos Biespecíficos/biossíntese , Anticorpos Monoclonais/biossíntese , Ensaios de Triagem em Larga Escala , Imunoglobulina G/biossíntese , Engenharia de Proteínas/métodos , Animais , Anticorpos Biespecíficos/química , Anticorpos Biespecíficos/genética , Anticorpos Monoclonais/química , Anticorpos Monoclonais/genética , Células CHO , Clonagem Molecular , Cricetulus , Fatores de Crescimento de Fibroblastos/genética , Fatores de Crescimento de Fibroblastos/imunologia , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Células HEK293 , Humanos , Imunoglobulina G/química , Imunoglobulina G/genética , Proteínas Klotho , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
14.
Cell Metab ; 26(5): 709-718.e3, 2017 Nov 07.
Artigo em Inglês | MEDLINE | ID: mdl-28988823

RESUMO

Despite the different physiologic functions of FGF19 and FGF21 as hormonal regulators of fed and fasted metabolism, their pharmacologic administration causes similar increases in energy expenditure, weight loss, and enhanced insulin sensitivity in obese animals. Here, in genetic loss-of-function studies of the shared co-receptor ß-Klotho, we show that these pharmacologic effects are mediated through a common, tissue-specific pathway. Surprisingly, FGF19 and FGF21 actions in liver and adipose tissue are not required for their longer-term weight loss and glycemic effects. In contrast, ß-Klotho in neurons is essential for both FGF19 and FGF21 to cause weight loss and lower glucose and insulin levels. We further show an FGF21 mimetic antibody that activates the FGF receptor 1/ß-Klotho complex also requires neuronal ß-Klotho for its metabolic effects. These studies highlight the importance of the nervous system in mediating the beneficial weight loss and glycemic effects of endocrine FGF drugs.


Assuntos
Anticorpos Monoclonais/farmacologia , Glicemia/efeitos dos fármacos , Fatores de Crescimento de Fibroblastos/farmacologia , Proteínas de Membrana/agonistas , Sistema Nervoso/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/agonistas , Redução de Peso , Tecido Adiposo/metabolismo , Animais , Resistência à Insulina , Proteínas Klotho , Fígado/metabolismo , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Obesos , Neurônios/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia
15.
MAbs ; 9(8): 1379-1388, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28895785

RESUMO

Target receptor levels can influence pharmacokinetics (PK) or pharmacodynamics (PD) of monoclonal antibodies (mAbs), and can affect drug development of this class of molecules. We generated an effector-less humanized bispecific antibody that selectively activates fibroblast growth factor receptor (FGFR)1 and ßKlotho receptor, a FGF21 receptor complex highly expressed in both white and brown adipocytes. The molecule shows cross-species binding with comparable equilibrium binding affinity (Kd) for human, cynomolgus monkey, and mouse FGFR1/ßKlotho. To understand the PK/PD relationship in non-obese and obese animals, we evaluated the adipose tissue distribution of the antibody, serum exposures, and an associated PD marker (high-molecular-weight adiponectin), in both non-obese and obese mice and monkeys. Antibody uptake into fat tissue was found to be higher on a per gram basis in non-obese animals compared to obese animals. Since obesity has been reported to be associated with reduced expression of FGFR1 and ßKlotho receptor in white adipose tissues in mice, our results suggest that the distribution in adipose tissues was influenced by target expression levels. Even so, the overall dose-normalized serum exposures were comparable between non-obese and obese mice and monkeys, suggesting that adipose tissue uptake plays a limited role in overall systemic PK determination. It remains to be determined if and how obesity and receptor expression in humans influence the PK and PD profile of this novel therapeutic candidate.


Assuntos
Tecido Adiposo/metabolismo , Anticorpos Monoclonais/farmacocinética , Avaliação Pré-Clínica de Medicamentos/métodos , Obesidade/metabolismo , Adiponectina/sangue , Adiponectina/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Células CHO , Cricetinae , Cricetulus , Dieta Hiperlipídica/efeitos adversos , Feminino , Fatores de Crescimento de Fibroblastos/antagonistas & inibidores , Fatores de Crescimento de Fibroblastos/imunologia , Fatores de Crescimento de Fibroblastos/metabolismo , Macaca fascicularis , Masculino , Camundongos Endogâmicos C57BL , Obesidade/etiologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Distribuição Tecidual
16.
Artigo em Inglês | MEDLINE | ID: mdl-28808580

RESUMO

BACKGROUND: As of 2014, community pharmacies in Japan are approved by the Ministry of Health, Labour and Welfare to measure lipid panel, HbA1c, glucose, ALT, AST and γ-GTP, but not to screen for influenza virus. We provided influenza virus screening tests at a community pharmacy to triage people with symptoms suggestive of influenza. Participants were given appropriate advice on how to prevent the spread of and safeguard against influenza. We subsequently evaluated the effects of community pharmacy-based influenza virus screening and prevention measures. METHODS: Local residents with symptoms suggestive of influenza participated in this study. Influenza virus screening tests using nasal samples were provided to the pharmacy, and we assessed samples for the presence of influenza virus. The study consisted of a preliminary interview, informed consent, and screening test on Day 1, and mail-in survey on Day 14. RESULTS: A total 52 local residents participated in the study. The number of participants and influenza virus positive results followed the same trend as the influenza epidemic in the study area. Influenza virus was found in 28.8% of samples. There was no significant difference between the appearance ratios of subjective symptoms among influenza-positive and influenza-negative groups. The percentages of participants who were first screened at the pharmacy, and those who were first screened at a clinic and then tested again at the pharmacy, were 71.2% (37/52) and 28.8% (15/52), respectively. In the latter group, 14 of 15 were negative by screening at the clinic, and one was diagnosed with influenza without testing. Subsequently, 46.8% (7/15) of participants tested positive for influenza by pharmacy-based screening. According to the mail-in survey, all influenza-positive (100%, 7/7) and 35.3% (6/17) of influenza-negative participants visited the clinic after being tested at the community pharmacy; test results between the community pharmacy and clinic were consistent. A total 64.7% (11/17) of symptomatic participants who tested negative recovered spontaneously at home. CONCLUSIONS: Implementation of influenza virus screening followed by provision of appropriate advice for both influenza-positive and influenza-negative participants at the community pharmacy showed a significant effect on improving the health of the local community.

17.
Mol Metab ; 6(11): 1454-1467, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-29107292

RESUMO

OBJECTIVE: Fibroblast Growth Factor 21 (FGF21) is a potent stimulator of brown fat thermogenesis that improves insulin sensitivity, ameliorates hepatosteatosis, and induces weight loss by engaging the receptor complex comprised of Fibroblast Growth Factor Receptor 1 (FGFR1) and the requisite coreceptor ßKlotho. Previously, recombinant antibody proteins that activate the FGFR1/ßKlotho complex were proposed to act as an FGF21-mimetic; however, in vivo action of these engineered proteins has not been well studied. METHODS: We investigated the mechanism by which anti-FGFR1/ßKlotho bispecific antibody (bFKB1) stimulates thermogenesis in UCP1-expressing brown adipocytes using genetically engineered mice. Anti-FGFR1 agonist antibody was also used to achieve brown adipose tissue restricted activation in transgenic mice. RESULTS: Studies with global Ucp1-deficient mice and adipose-specific Fgfr1 deficient mice demonstrated that bFKB1 acts on targets distal to adipocytes and indirectly stimulates brown adipose thermogenesis in a UCP1-independent manner. Using a newly developed transgenic system, we also show that brown adipose tissue restricted activation of a transgenic FGFR1 expressed under the control of Ucp1 promoter does not stimulate energy expenditure. Finally, consistent with its action as a FGF21 mimetic, bFBK1 suppresses intake of saccharin-containing food and alcohol containing water in mice. CONCLUSIONS: Collectively, we propose that FGFR1/ßKlotho targeted therapy indeed mimics the action of FGF21 in vivo and stimulates UCP1-independent brown fat thermogenesis through receptors outside of adipocytes and likely in the nervous system.


Assuntos
Proteínas de Membrana/imunologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia , Termogênese/fisiologia , Adipócitos Marrons/metabolismo , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Animais , Anticorpos/metabolismo , Metabolismo Energético/fisiologia , Fatores de Crescimento de Fibroblastos/metabolismo , Proteínas Klotho , Proteínas de Membrana/agonistas , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Proteínas Mitocondriais/metabolismo , Obesidade/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/agonistas , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Termogênese/genética , Proteína Desacopladora 1/genética , Proteína Desacopladora 1/metabolismo , Redução de Peso
18.
MAbs ; 8(8): 1487-1497, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27680183

RESUMO

Bispecific antibodies have shown promise in the clinic as medicines with novel mechanisms of action. Lack of efficient production of bispecific IgGs, however, has limited their rapid advancement. Here, we describe a single-reactor process using mammalian cell co-culture production to efficiently produce a bispecific IgG with 4 distinct polypeptide chains without the need for parallel processing of each half-antibody or additional framework mutations. This method resembles a conventional process, and the quality and yield of the monoclonal antibodies are equal to those produced using parallel processing methods. We demonstrate the application of the approach to diverse bispecific antibodies, and its suitability for production of a tissue specific molecule targeting fibroblast growth factor receptor 1 and klotho ß that is being developed for type 2 diabetes and other obesity-linked disorders.


Assuntos
Anticorpos Biespecíficos/biossíntese , Reatores Biológicos , Técnicas de Cocultura/métodos , Imunoglobulina G/biossíntese , Animais , Anticorpos Biespecíficos/imunologia , Células CHO , Cricetinae , Cricetulus , Humanos , Imunoglobulina G/imunologia , Proteínas Klotho , Mamíferos , Proteínas de Membrana/imunologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/imunologia
19.
Artigo em Inglês | MEDLINE | ID: mdl-27144011

RESUMO

BACKGROUND: Osteoporosis is estimated to afflict over 200 million people worldwide and healthcare professionals are needed to successfully intervene. The aim of this study was to assess cognitive changes in students pertaining to the primary prevention of osteoporosis after measuring their bone density and having them participate in a simulated health class during pharmacy school. METHODS: Third year pharmacy students participated in the training program, which consisted of measuring their bone density using quantitative ultrasound and preparing educational materials and conducting a simulated health class. The students' knowledge concerning the prevention and education on osteoporosis was surveyed using questionnaires before and after the training. RESULTS: The bone area ratio (BAR) in 24 % of the students was evaluated as category 4 (slightly low) or 5 (low or caution). Regression analysis indicated a significant relationship between the BAR and amount of exercise reported in both males (p = 0.005) and females (p = 0.004). The student-made educational materials were prepared in line with the requirements of the Japanese 2011 guidelines. The student response rates for the importance of food, exercise, and the bone density measurement in youth were significantly increased after the training (p < 0.001 in all). More than 95 % of students reported that the program was useful, improved their understanding, and important, with 94 % satisfied with the experience. CONCLUSIONS: This experience-based educational program combining measuring the bone density and the preparation and presentation of a simulated health class appeared to improve the awareness and understanding of osteoporosis prevention in pharmacy students.

20.
EBioMedicine ; 2(7): 730-43, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26288846

RESUMO

Dissipating excess calories as heat through therapeutic stimulation of brown adipose tissues (BAT) has been proposed as a potential treatment for obesity-linked disorders. Here, we describe the generation of a humanized effector-less bispecific antibody that activates fibroblast growth factor receptor (FGFR) 1/ßKlotho complex, a common receptor for FGF21 and FGF19. Using this molecule, we show that antibody-mediated activation of FGFR1/ßKlotho complex in mice induces sustained energy expenditure in BAT, browning of white adipose tissue, weight loss, and improvements in obesity-associated metabolic derangements including insulin resistance, hyperglycemia, dyslipidemia and hepatosteatosis. In mice and cynomolgus monkeys, FGFR1/ßKlotho activation increased serum high-molecular-weight adiponectin, which appears to contribute over time by enhancing the amplitude of the metabolic benefits. At the same time, insulin sensitization by FGFR1/ßKlotho activation occurs even before the onset of weight loss in a manner that is independent of adiponectin. Together, selective activation of FGFR1/ßKlotho complex with a long acting therapeutic antibody represents an attractive approach for the treatment of type 2 diabetes and other obesity-linked disorders through enhanced energy expenditure, insulin sensitization and induction of high-molecular-weight adiponectin.


Assuntos
Tecido Adiposo Marrom/metabolismo , Anticorpos Biespecíficos/farmacologia , Insulina/farmacologia , Proteínas de Membrana/agonistas , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/agonistas , Adiponectina/metabolismo , Tecido Adiposo Marrom/efeitos dos fármacos , Animais , Linhagem Celular , Metabolismo Energético/efeitos dos fármacos , Fatores de Crescimento de Fibroblastos/farmacologia , Células HEK293 , Humanos , Proteínas Klotho , Macaca fascicularis , Masculino , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Obesos , Ligação Proteica/efeitos dos fármacos , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Termogênese/efeitos dos fármacos , Redução de Peso/efeitos dos fármacos
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