RESUMO
Enteropeptidase, localized into the duodenum brush border, is a key enzyme catalyzing the conversion of pancreatic trypsinogen proenzyme to active trypsin, thereby regulating protein digestion and energy homeostasis. We report the discovery and pharmacological profiles of SCO-792, a novel inhibitor of enteropeptidase. A screen employing fluorescence resonance energy transfer was performed to identify enteropeptidase inhibitors. Inhibitory profiles were determined by in vitro assays. To evaluate the in vivo inhibitory effect on protein digestion, an oral protein challenge test was performed in rats. Our screen identified a series of enteropeptidase inhibitors, and compound optimization resulted in identification of SCO-792, which inhibited enteropeptidase activity in vitro, with IC 50 values of 4.6 and 5.4 nmol/L in rats and humans, respectively. In vitro inhibition of enteropeptidase by SCO-792 was potentiated by increased incubation time, and the calculated Kinact/KI was 82 000/mol/L s. An in vitro dissociation assay showed that SCO-792 had a dissociation half-life of almost 14 hour, with a calculated koff rate of 0.047/hour, which suggested that SCO-792 is a reversible enteropeptidase inhibitor. In normal rats, a ≤4 hour prior oral dose of SCO-792 effectively inhibited plasma elevation of branched-chain amino acids in an oral protein challenge test, which indicated that SCO-792 effectively inhibited protein digestion in vivo. In conclusion, our new screen system identified SCO-792 as a potent and reversible inhibitor against enteropeptidase. SCO-792 slowly dissociated from enteropeptidase in vitro and inhibited protein digestion in vivo. Further study using SCO-792 could reveal the effects of inhibiting enteropeptidase on biological actions.
Assuntos
Enteropeptidase/antagonistas & inibidores , Inibidores Enzimáticos/administração & dosagem , Bibliotecas de Moléculas Pequenas/administração & dosagem , Administração Oral , Animais , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Transferência Ressonante de Energia de Fluorescência , Humanos , Concentração Inibidora 50 , Ratos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologiaRESUMO
A lead compound A was identified previously as an stearoyl coenzyme A desaturase (SCD) inhibitor during research on potential treatments for obesity. This compound showed high SCD1 binding affinity, but a poor pharmacokinetic (PK) profile and limited chemical accessibility, making it suboptimal for use in anticancer research. To identify potent SCD1 inhibitors with more promising PK profiles, we newly designed a series of 'non-spiro' 4, 4-disubstituted piperidine derivatives based on molecular modeling studies. As a result, we discovered compound 1a, which retained moderate SCD1 binding affinity. Optimization around 1a was accelerated by analyzing Hansch-Fujita and Hammett constants to obtain 4-phenyl-4-(trifluoromethyl)piperidine derivative 1n. Fine-tuning of the azole moiety of 1n led to compound 1o (T-3764518), which retained nanomolar affinity and exhibited an excellent PK profile. Reflecting the good potency and PK profile, orally administrated compound 1o showed significant pharmacodynamic (PD) marker reduction (at 0.3mg/kg, bid) in HCT116 mouse xenograft model and tumor growth suppression (at 1mg/kg, bid) in 786-O mouse xenograft model. In conclusion, we identified a new series of SCD1 inhibitors, represented by compound 1o, which represents a promising new chemical tool suitable for the study of SCD1 biology as well as the potential development of novel anticancer therapies.
Assuntos
Antineoplásicos/química , Inibidores Enzimáticos/síntese química , Oxidiazóis/síntese química , Piridazinas/síntese química , Estearoil-CoA Dessaturase/antagonistas & inibidores , Animais , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/farmacocinética , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Células HCT116 , Humanos , Concentração Inibidora 50 , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Microssomos Hepáticos/metabolismo , Oxidiazóis/farmacocinética , Oxidiazóis/uso terapêutico , Oxidiazóis/toxicidade , Piperidinas/química , Piperidinas/metabolismo , Piperidinas/farmacologia , Ligação Proteica , Piridazinas/farmacocinética , Piridazinas/uso terapêutico , Piridazinas/toxicidade , Compostos de Espiro/química , Estearoil-CoA Dessaturase/metabolismo , Relação Estrutura-Atividade , Transplante HeterólogoRESUMO
Matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) is a powerful technique for visualizing the distribution of a wide range of biomolecules within tissue sections. However, methodology for visualizing a bioactive ellagitannin has not yet been established. This paper presents a novel in situ label-free MALDI-MSI technique for visualizing the distribution of strictinin, a bioactive ellagitannin found in green tea, within mammalian kidney after oral dosing. Among nine representative matrix candidates, 1,5-diaminonaphthalene (1,5-DAN), harmane, and ferulic acid showed higher sensitivity to strictinin spotted onto a MALDI sample plate. Of these, 1,5-DAN enables visualization of a two-dimensional image of strictinin directly spotted on mouse kidney sections with the highest sensitivity. Furthermore, 1,5-DAN-based MALDI-MSI could detect the unique distribution of orally dosed strictinin within kidney sections. This in situ label-free imaging technique will contribute to the localization analysis of strictinin and its biological mechanisms.
Assuntos
Camellia sinensis/metabolismo , Rim/química , Fenóis/química , Fenóis/metabolismo , Extratos Vegetais/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Animais , Rim/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Fenóis/administração & dosagem , Extratos Vegetais/administração & dosagem , Extratos Vegetais/metabolismo , Espectrometria de Massas em Tandem/métodosRESUMO
We investigated the effects of extracts of Benifuuki (a tea cultivar that contains methylated catechins such as epigallocatechin-3-O-(3-O-methyl) gallate (EGCG3"Me)) in mice fed a high-fat/high-sucrose (HF/HS) diet. This tea cultivar was then compared with an extract of Yabukita (a popular tea cultivar that lacks methylated catechins). For 6 weeks, C57BL/6J mice were fed either HF/HS diet with or without tea extracts from tea cultivars, which contained almost identical ingredients except for methylated catechins (i.e., Yabukita (0.2% and 1%) or Benifuuki (0.2% and 1%) extract powders). Supplementation with Benifuuki 0.2% markedly lowered plasma levels of TG and NEFAs compared with mice supplemented with Yabukita 0.2%. The diet containing Benifuuki 1% decreased adipose tissue weights, liver TG, and expression of lipogenic genes in the liver. These results suggested that Benifuuki had much greater lipid-lowering effects than Yabukita. Taken together, these data suggest that methylated catechins direct the strong lipid-lowering activity of Benifuuki.