Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 6.855
Filtrar
1.
Commun Biol ; 4(1): 193, 2021 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-33564093

RESUMO

SARS-CoV-2 Nsp15 is a uridine-specific endoribonuclease with C-terminal catalytic domain belonging to the EndoU family that is highly conserved in coronaviruses. As endoribonuclease activity seems to be responsible for the interference with the innate immune response, Nsp15 emerges as an attractive target for therapeutic intervention. Here we report the first structures with bound nucleotides and show how the enzyme specifically recognizes uridine moiety. In addition to a uridine site we present evidence for a second base binding site that can accommodate any base. The structure with a transition state analog, uridine vanadate, confirms interactions key to catalytic mechanisms. In the presence of manganese ions, the enzyme cleaves unpaired RNAs. This acquired knowledge was instrumental in identifying Tipiracil, an FDA approved drug that is used in the treatment of colorectal cancer, as a potential anti-COVID-19 drug. Using crystallography, biochemical, and whole-cell assays, we demonstrate that Tipiracil inhibits SARS-CoV-2 Nsp15 by interacting with the uridine binding pocket in the enzyme's active site. Our findings provide new insights for the development of uracil scaffold-based drugs.


Assuntos
Antivirais/farmacologia , /virologia , Endorribonucleases/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Pirrolidinas/farmacologia , /enzimologia , Timina/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Células A549 , Antivirais/química , Antivirais/farmacocinética , Domínio Catalítico , Cristalografia por Raios X , Endorribonucleases/química , Endorribonucleases/metabolismo , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Humanos , Ligantes , Modelos Moleculares , Conformação Proteica , Pirrolidinas/química , Pirrolidinas/farmacocinética , Timina/química , Timina/farmacocinética , Uridina/metabolismo , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/metabolismo
2.
Antiviral Res ; 187: 105020, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33515606

RESUMO

The 3-chymotrypsin-like cysteine protease (3CLpro) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is considered a major target for the discovery of direct antiviral agents. We previously reported the evaluation of SARS-CoV-2 3CLpro inhibitors in a novel self-assembled monolayer desorption ionization mass spectrometry (SAMDI-MS) enzymatic assay (Gurard-Levin et al., 2020). The assay was further improved by adding the rhinovirus HRV3C protease to the same well as the SARS-CoV-2 3CLpro enzyme. High substrate specificity for each enzyme allowed the proteases to be combined in a single assay reaction without interfering with their individual activities. This novel duplex assay was used to profile a diverse set of reference protease inhibitors. The protease inhibitors were grouped into three categories based on their relative potency against 3CLpro and HRV3C including those that are: equipotent against 3CLpro and HRV3C (GC376 and calpain inhibitor II), selective for 3CLpro (PF-00835231, calpain inhibitor XII, boceprevir), and selective for HRV3C (rupintrivir). Structural analysis showed that the combination of minimal interactions, conformational flexibility, and limited bulk allows GC376 and calpain inhibitor II to potently inhibit both enzymes. In contrast, bulkier compounds interacting more tightly with pockets P2, P3, and P4 due to optimization for a specific target display a more selective inhibition profile. Consistently, the most selective viral protease inhibitors were relatively weak inhibitors of human cathepsin L. Taken together, these results can guide the design of cysteine protease inhibitors that are either virus-specific or retain a broad antiviral spectrum against coronaviruses and rhinoviruses.


Assuntos
Antivirais/farmacologia , Inibidores de Proteases/farmacologia , Rhinovirus/efeitos dos fármacos , /efeitos dos fármacos , Antivirais/química , Sítios de Ligação , Catepsina L/metabolismo , Descoberta de Drogas , Glicoproteínas/farmacologia , Humanos , Cinética , Modelos Moleculares , Inibidores de Proteases/química , Pirrolidinas/farmacologia
3.
Medicine (Baltimore) ; 100(2): e24110, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-33466182

RESUMO

BACKGROUND: Gastric cancer is a global health problem with high incidence rate and mortality rate. Due to the limitations of traditional chemotherapy drugs, such as patient intolerance, low efficacies and serious adverse effects, trifluridine/tipiracil has been considered to be a promising treatment for patients with heavily pretreated metastatic gastric cancer. However, the relevant systematic review has not been occurring. The presentation of this protocol is to scientifically evaluate the efficacy and safety of trifluridine/tipiracil in patients with highly pretreated metastatic gastric cancer. METHODS: The protocol followed Preferred Reporting Items for Systematic Reviews and Meta-Analyses Protocols. We will systematically search MEDLINE, PubMed, Embase, Cochrane Library, Web of Science, China National Knowledge Infrastructure Database (CNKI), VIP Chinese Science and Technology Periodical Database (VIP), Wan Fang Database up to November 1, 2020 to identify published articles. Using the Cochrane risk assessment tool to assess the methodological quality of the RCTs, and all included studies will be analyzed according to the criteria in the Cochrane Handbook. Review Manager 5.3 software will be used for literature quality evaluation and data analysis. RESULTS: Objective to evaluate the efficacy and safety of trifluridine/tipiracil in patients with heavily pretreated metastatic gastric cancer by analyzing the eligible data extracted under limited conditions. CONCLUSION: This study provides clear evidence to evaluate the effectiveness and safety of trifluridine/tipiracil for patients with highly pretreated metastatic gastric cancer, and the findings will also be published in a peer-reviewed journal. ETHICS AND DISSEMINATION: No ethical statement will be required for this study because there is no direct involvement of human. This review will be published in a peer-reviewed journal as a conference report or an article. REGISTRATION: OSF registration number: DOI 10.17605/OSF.IO/6MF5U.


Assuntos
Protocolos Clínicos , Pirrolidinas/uso terapêutico , Neoplasias Gástricas/tratamento farmacológico , Timina/uso terapêutico , Trifluridina/uso terapêutico , Adolescente , Adulto , Idoso , Combinação de Medicamentos , Humanos , Metanálise como Assunto , Pessoa de Meia-Idade , Pirrolidinas/farmacologia , Neoplasias Gástricas/fisiopatologia , Revisões Sistemáticas como Assunto , Timina/farmacologia , Trifluridina/farmacologia
4.
Metabolism ; 114: 154409, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33096076

RESUMO

BACKGROUND AND OBJECTIVES: The gut-liver axis plays an important role in the pathogenesis of nonalcoholic steatohepatitis (NASH), and increased intestinal permeability causes transfer of endotoxin to the liver, which activates the immune response, ultimately leading to hepatic inflammation. Nuclear receptor Rev-erbα is a critical regulator of circadian rhythm, cellular metabolism, and inflammatory responses. However, the role and mechanism of Rev-erbα in gut barrier function and NASH remain unclear. In the present study, we investigated the involvement of Rev-erbα in the regulation of intestinal permeability and the treatment of NASH. METHODS AND RESULTS: The expression of tight junction-related genes and Rev-erbs decreased in the jejunum, ileum and colon of mice with high cholesterol, high fat diet (CL)-induced NASH. Chromatin immunoprecipitation analysis indicated that REV-ERBα directly bound to the promoters of tight junction genes to regulate intestinal permeability. Pharmacological activation of REV-ERBα by SR9009 protected against lipopolysaccharide-induced increased intestinal permeability both in vitro and in vivo, and these effects were associated with the activation of autophagy and decreased apoptotic signaling of epithelial cells. In addition, the chronopharmacological effects of SR9009 were more potent at Zeitgeber time 0 (ZT0) than at ZT12, which was contrary to the rhythm of Rev-erbs in the gastrointestinal tract. The administration of SR9009 attenuated hepatic lipid accumulation, insulin resistance, inflammation, and fibrosis in mice with CL diet-induced NASH, which might be partly attributed to the enhancement of intestinal barrier function. CONCLUSION: Chronopharmacological activation of REV-ERBα might be a potential strategy to treat intestinal barrier dysfunction-related disorders and NASH.


Assuntos
Mucosa Intestinal/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Pirrolidinas/uso terapêutico , Receptores Citoplasmáticos e Nucleares/agonistas , Proteínas Repressoras/agonistas , Tiofenos/uso terapêutico , Junções Íntimas/efeitos dos fármacos , Animais , Autofagia/efeitos dos fármacos , Glicemia , Células CACO-2 , Colesterol/sangue , Humanos , Insulina/sangue , Mucosa Intestinal/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Hepatopatia Gordurosa não Alcoólica/metabolismo , Permeabilidade/efeitos dos fármacos , Pirrolidinas/farmacologia , Receptores Citoplasmáticos e Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tiofenos/farmacologia , Junções Íntimas/metabolismo , Triglicerídeos/sangue
5.
PLoS One ; 15(12): e0232864, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33373369

RESUMO

Activation of the kappa opioid receptor (KOR) contributes to the aversive properties of stress, and modulates key neuronal circuits underlying many neurobehavioral disorders. KOR agonists directly inhibit ventral tegmental area (VTA) dopaminergic neurons, contributing to aversive responses (Margolis et al. 2003, 2006); therefore, selective KOR antagonists represent a novel therapeutic approach to restore circuit function. We used whole cell electrophysiology in acute rat midbrain slices to evaluate pharmacological properties of four novel KOR antagonists: BTRX-335140, BTRX-395750, PF-04455242, and JNJ-67953964. Each compound concentration-dependently reduced the outward current induced by the KOR selective agonist U-69,593. BTRX-335140 and BTRX-395750 fully blocked U-69,593 currents (IC50 = 1.2 ± 0.9 and 1.2 ± 1.3 nM, respectively). JNJ-67953964 showed an IC50 of 3.0 ± 4.6 nM. PF-04455242 exhibited partial antagonist activity asymptoting at 55% blockade (IC50 = 6.7 ± 15.1 nM). In 3/8 of neurons, 1 µM PF-04455242 generated an outward current independent of KOR activation. BTRX-335140 (10 nM) did not affect responses to saturating concentrations of the mu opioid receptor (MOR) agonist DAMGO or the delta opioid receptor (DOR) agonist DPDPE, while JNJ-67953964 (10 nM) partially blocked DAMGO and DPDPE responses. Importantly, BTRX-335140 (10 nM) rapidly washed out with complete recovery of U-69,593 responses within 10 min. Collectively, we show electrophysiological evidence of key differences amongst KOR antagonists that could impact their therapeutic potential and have not been observed using recombinant systems. The results of this study demonstrate the value of characterizing compounds in native neuronal tissue and within circuits implicated in the neurobehavioral disorders of interest.


Assuntos
Neurônios Dopaminérgicos/efeitos dos fármacos , Potenciais da Membrana/efeitos dos fármacos , Receptores Opioides kappa/antagonistas & inibidores , Analgésicos Opioides/farmacologia , Animais , Benzamidas/farmacologia , Compostos de Bifenilo/farmacologia , Neurônios Dopaminérgicos/metabolismo , Eletrofisiologia , D-Penicilina (2,5)-Encefalina/farmacologia , Masculino , Mesencéfalo/metabolismo , Antagonistas de Entorpecentes/farmacologia , Oxidiazóis/farmacologia , Técnicas de Patch-Clamp/métodos , Piperidinas/farmacologia , Pirrolidinas/farmacologia , Quinolinas/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores Opioides kappa/metabolismo , Receptores Opioides mu/metabolismo , Sulfonamidas/farmacologia , Área Tegmentar Ventral/efeitos dos fármacos
6.
PLoS One ; 15(10): e0236000, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33002003

RESUMO

Non-alcoholic fatty liver disease (NAFLD) affects a significant number of people worldwide and currently there are no pharmacological treatments. NAFLD often presents with obesity, insulin resistance, and in some cases cardiovascular diseases. There is a clear need for treatment options to alleviate this disease since it often progresses to much more the much more severe non-alcoholic steatohepatitis (NASH). The REV-ERB nuclear receptor is a transcriptional repressor that regulates physiological processes involved in the development of NAFLD including lipogenesis and inflammation. We hypothesized that pharmacologically activating REV-ERB would suppress the progression of fatty liver in a mouse model of NASH. Using REV-ERB agonist SR9009 in a mouse NASH model, we demonstrate the beneficial effects of REV-ERB activation that led to an overall improvement of hepatic health by suppressing hepatic fibrosis and inflammatory response.


Assuntos
Hepatopatia Gordurosa não Alcoólica , Membro 1 do Grupo D da Subfamília 1 de Receptores Nucleares/efeitos dos fármacos , Pirrolidinas/farmacologia , Tiofenos/farmacologia , Animais , Modelos Animais de Doenças , Humanos , Inflamação/metabolismo , Lipogênese/fisiologia , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/patologia , Camundongos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/patologia , Obesidade/metabolismo
7.
Nat Commun ; 11(1): 4417, 2020 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-32887884

RESUMO

COVID-19 was declared a pandemic on March 11 by WHO, due to its great threat to global public health. The coronavirus main protease (Mpro, also called 3CLpro) is essential for processing and maturation of the viral polyprotein, therefore recognized as an attractive drug target. Here we show that a clinically approved anti-HCV drug, Boceprevir, and a pre-clinical inhibitor against feline infectious peritonitis (corona) virus (FIPV), GC376, both efficaciously inhibit SARS-CoV-2 in Vero cells by targeting Mpro. Moreover, combined application of GC376 with Remdesivir, a nucleotide analogue that inhibits viral RNA dependent RNA polymerase (RdRp), results in sterilizing additive effect. Further structural analysis reveals binding of both inhibitors to the catalytically active side of SARS-CoV-2 protease Mpro as main mechanism of inhibition. Our findings may provide critical information for the optimization and design of more potent inhibitors against the emerging SARS-CoV-2 virus.


Assuntos
Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Prolina/análogos & derivados , Inibidores de Proteases/farmacologia , Pirrolidinas/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Animais , Antivirais/farmacologia , Betacoronavirus/enzimologia , Sítios de Ligação/efeitos dos fármacos , Domínio Catalítico , Chlorocebus aethiops , Cristalografia por Raios X , Cisteína Endopeptidases/química , Cisteína Endopeptidases/metabolismo , Modelos Animais de Doenças , Ensaios de Triagem em Larga Escala , Modelos Moleculares , Pandemias , Prolina/farmacologia , /química , Células Vero , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/efeitos dos fármacos
8.
Nat Commun ; 11(1): 4659, 2020 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-32938936

RESUMO

The αvß6 integrin plays a key role in the activation of transforming growth factor-ß (TGFß), a pro-fibrotic mediator that is pivotal to the development of idiopathic pulmonary fibrosis (IPF). We identified a selective small molecule αvß6 RGD-mimetic, GSK3008348, and profiled it in a range of disease relevant pre-clinical systems. To understand the relationship between target engagement and inhibition of fibrosis, we measured pharmacodynamic and disease-related end points. Here, we report, GSK3008348 binds to αvß6 with high affinity in human IPF lung and reduces downstream pro-fibrotic TGFß signaling to normal levels. In human lung epithelial cells, GSK3008348 induces rapid internalization and lysosomal degradation of the αvß6 integrin. In the murine bleomycin-induced lung fibrosis model, GSK3008348 engages αvß6, induces prolonged inhibition of TGFß signaling and reduces lung collagen deposition and serum C3M, a marker of IPF disease progression. These studies highlight the potential of inhaled GSK3008348 as an anti-fibrotic therapy.


Assuntos
Butiratos/farmacologia , Fibrose Pulmonar Idiopática/tratamento farmacológico , Integrinas/antagonistas & inibidores , Naftiridinas/farmacologia , Pirazóis/farmacologia , Pirrolidinas/farmacologia , Administração por Inalação , Animais , Antígenos de Neoplasias/metabolismo , Bleomicina/toxicidade , Butiratos/administração & dosagem , Butiratos/metabolismo , Butiratos/farmacocinética , Colágeno/metabolismo , Modelos Animais de Doenças , Células Epiteliais/efeitos dos fármacos , Humanos , Fibrose Pulmonar Idiopática/induzido quimicamente , Fibrose Pulmonar Idiopática/patologia , Integrinas/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Simulação de Acoplamento Molecular , Naftiridinas/administração & dosagem , Naftiridinas/metabolismo , Naftiridinas/farmacocinética , Pirazóis/administração & dosagem , Pirazóis/metabolismo , Pirazóis/farmacocinética , Pirrolidinas/administração & dosagem , Pirrolidinas/metabolismo , Pirrolidinas/farmacocinética , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Tomografia Computadorizada de Emissão de Fóton Único , Fator de Crescimento Transformador beta/metabolismo , Pesquisa Médica Translacional
9.
Nat Commun ; 11(1): 4279, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855410

RESUMO

Plasma and tumor caveolin-1 (Cav-1) are linked with disease progression in prostate cancer. Here we report that metabolomic profiling of longitudinal plasmas from a prospective cohort of 491 active surveillance (AS) participants indicates prominent elevations in plasma sphingolipids in AS progressors that, together with plasma Cav-1, yield a prognostic signature for disease progression. Mechanistic studies of the underlying tumor supportive onco-metabolism reveal coordinated activities through which Cav-1 enables rewiring of cancer cell lipid metabolism towards a program of 1) exogenous sphingolipid scavenging independent of cholesterol, 2) increased cancer cell catabolism of sphingomyelins to ceramide derivatives and 3) altered ceramide metabolism that results in increased glycosphingolipid synthesis and efflux of Cav-1-sphingolipid particles containing mitochondrial proteins and lipids. We also demonstrate, using a prostate cancer syngeneic RM-9 mouse model and established cell lines, that this Cav-1-sphingolipid program evidences a metabolic vulnerability that is targetable to induce lethal mitophagy as an anti-tumor therapy.


Assuntos
Caveolina 1/metabolismo , Neoplasias da Próstata/metabolismo , Esfingolipídeos/metabolismo , Idoso , Animais , Caveolina 1/sangue , Caveolina 1/genética , Linhagem Celular Tumoral , Ceramidas/metabolismo , Intervalo Livre de Doença , Regulação Neoplásica da Expressão Gênica , Glicoesfingolipídeos/biossíntese , Humanos , Lipídeos/sangue , Masculino , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Estudos Prospectivos , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/mortalidade , Neoplasias da Próstata/patologia , Pirrolidinas/farmacologia , Esfingomielinas/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Nat Commun ; 11(1): 4282, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855413

RESUMO

The main protease, Mpro (or 3CLpro) in SARS-CoV-2 is a viable drug target because of its essential role in the cleavage of the virus polypeptide. Feline infectious peritonitis, a fatal coronavirus infection in cats, was successfully treated previously with a prodrug GC376, a dipeptide-based protease inhibitor. Here, we show the prodrug and its parent GC373, are effective inhibitors of the Mpro from both SARS-CoV and SARS-CoV-2 with IC50 values in the nanomolar range. Crystal structures of SARS-CoV-2 Mpro with these inhibitors have a covalent modification of the nucleophilic Cys145. NMR analysis reveals that inhibition proceeds via reversible formation of a hemithioacetal. GC373 and GC376 are potent inhibitors of SARS-CoV-2 replication in cell culture. They are strong drug candidates for the treatment of human coronavirus infections because they have already been successful in animals. The work here lays the framework for their use in human trials for the treatment of COVID-19.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Coronavirus Felino/efeitos dos fármacos , Inibidores de Proteases/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Células A549 , Animais , Antivirais/química , Betacoronavirus/enzimologia , Sítios de Ligação , Chlorocebus aethiops , Coronavirus Felino/enzimologia , Cristalografia por Raios X , Cisteína Endopeptidases/química , Efeito Citopatogênico Viral/efeitos dos fármacos , Reposicionamento de Medicamentos , Humanos , Concentração Inibidora 50 , Estrutura Molecular , Pró-Fármacos , Inibidores de Proteases/química , Pirrolidinas/química , Pirrolidinas/farmacologia , Vírus da SARS/efeitos dos fármacos , Vírus da SARS/enzimologia , Células Vero , Proteínas não Estruturais Virais/química , Replicação Viral/efeitos dos fármacos
11.
Artigo em Inglês | MEDLINE | ID: mdl-32669265

RESUMO

The coronavirus (CoV) disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome CoV-2 (SARS-CoV-2) is a health threat worldwide. Viral main protease (Mpro, also called 3C-like protease [3CLpro]) is a therapeutic target for drug discovery. Herein, we report that GC376, a broad-spectrum inhibitor targeting Mpro in the picornavirus-like supercluster, is a potent inhibitor for the Mpro encoded by SARS-CoV-2, with a half-maximum inhibitory concentration (IC50) of 26.4 ± 1.1 nM. In this study, we also show that GC376 inhibits SARS-CoV-2 replication with a half-maximum effective concentration (EC50) of 0.91 ± 0.03 µM. Only a small portion of SARS-CoV-2 Mpro was covalently modified in the excess of GC376 as evaluated by mass spectrometry analysis, indicating that improved inhibitors are needed. Subsequently, molecular docking analysis revealed that the recognition and binding groups of GC376 within the active site of SARS-CoV-2 Mpro provide important new information for the optimization of GC376. Given that sufficient safety and efficacy data are available for GC376 as an investigational veterinary drug, expedited development of GC376, or its optimized analogues, for treatment of SARS-CoV-2 infection in human is recommended.


Assuntos
Antivirais/química , Betacoronavirus/efeitos dos fármacos , Cisteína Endopeptidases/química , Inibidores de Proteases/química , Pirrolidinas/química , Proteínas não Estruturais Virais/química , Motivos de Aminoácidos , Animais , Antivirais/farmacologia , Betacoronavirus/patogenicidade , Domínio Catalítico , Chlorocebus aethiops , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Expressão Gênica , Simulação de Acoplamento Molecular , Inibidores de Proteases/farmacologia , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Pirrolidinas/farmacologia , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Termodinâmica , Células Vero , Proteínas não Estruturais Virais/antagonistas & inibidores , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/efeitos dos fármacos
12.
Am J Physiol Lung Cell Mol Physiol ; 319(3): L497-L512, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32697651

RESUMO

Hyperoxia (HO)-induced lung injury contributes to bronchopulmonary dysplasia (BPD) in preterm newborns. Intractable wheezing seen in BPD survivors is associated with airway remodeling (AWRM). Sphingosine kinase 1 (SPHK1)/sphingosine-1-phosphate (S1P) signaling promotes HO-mediated neonatal BPD; however, its role in the sequela of AWRM is not known. We noted an increased concentration of S1P in tracheal aspirates of neonatal infants with severe BPD, and earlier, demonstrated that Sphk1-/- mice showed protection against HO-induced BPD. The role of SPHK1/S1P in promoting AWRM following exposure of neonates to HO was investigated in a murine model. Therapy using PF543, the specific SPHK1 inhibitor, during neonatal HO reduced alveolar simplification followed by reduced AWRM in adult mice. This was associated with reduced airway hyperreactivity to intravenous methacholine. Neonatal HO exposure was associated with increased expression of SPHK1 in lung tissue of adult mice, which was reduced with PF543 therapy in the neonatal stage. This was accompanied by amelioration of HO-induced reduction of E-cadherin in airway epithelium. This may be suggestive of arrested partial epithelial mesenchymal transition (EMT) induced by HO. In vitro studies using human primary airway epithelial cells (HAEpCs) showed that SPHK1 inhibition or deletion restored HO-induced reduction in E-cadherin and reduced formation of mitochondrial reactive oxygen species (mtROS). Blocking mtROS with MitoTempo attenuated HO-induced partial EMT of HAEpCs. These results collectively support a therapeutic role for PF543 in preventing HO-induced BPD in neonates and the long-term sequela of AWRM, thus conferring a long-term protection resulting in improved lung development and function.


Assuntos
Remodelação das Vias Aéreas/efeitos dos fármacos , Displasia Broncopulmonar/tratamento farmacológico , Hiperóxia/tratamento farmacológico , Metanol/análogos & derivados , Pirrolidinas/farmacologia , Animais , Animais Recém-Nascidos , Displasia Broncopulmonar/induzido quimicamente , Modelos Animais de Doenças , Hiperóxia/induzido quimicamente , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Metanol/farmacologia , Camundongos Knockout , Fosfotransferases (Aceptor do Grupo Álcool)/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos
13.
J Nanosci Nanotechnol ; 20(12): 7311-7323, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-32711596

RESUMO

We started a study on the molecular docking of six potential pharmacologically active inhibitors compounds that can be used clinically against the COVID-19 virus, in this case, remdesivir, ribavirin, favipiravir, galidesivir, hydroxychloroquine and chloroquine interacting with the main COVID-19 protease in complex with a COVID-19 N3 protease inhibitor. The highest values of affinity energy found in order from highest to lowest were chloroquine (CHL), hydroxychloroquine (HYC), favipiravir (FAV), galidesivir (GAL), remdesivir (REM) and ribavirin (RIB). The possible formation of hydrogen bonds, associations through London forces and permanent electric dipole were analyzed. The values of affinity energy obtained for the hydroxychloroquine ligands was -9.9 kcal/mol and for the chloroquine of -10.8 kcal/mol which indicate that the coupling contributes to an effective improvement of the affinity energies with the protease. Indicating that, the position chosen to make the substitutions may be a pharmacophoric group, and cause changes in the protease.


Assuntos
Antivirais/química , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/enzimologia , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Cisteína Endopeptidases/química , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Proteínas não Estruturais Virais/química , Adenina/administração & dosagem , Adenina/análogos & derivados , Adenina/química , Adenina/farmacologia , Adenosina/análogos & derivados , Monofosfato de Adenosina/administração & dosagem , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/química , Monofosfato de Adenosina/farmacologia , Alanina/administração & dosagem , Alanina/análogos & derivados , Alanina/química , Alanina/farmacologia , Amidas/administração & dosagem , Amidas/química , Amidas/farmacologia , Antivirais/administração & dosagem , Sítios de Ligação , Cloroquina/administração & dosagem , Cloroquina/química , Cloroquina/farmacologia , Interações Medicamentosas , Humanos , Ligação de Hidrogênio , Hidroxicloroquina/administração & dosagem , Hidroxicloroquina/química , Hidroxicloroquina/farmacologia , Ligantes , Simulação de Acoplamento Molecular , Nanotecnologia , Pandemias , Inibidores de Proteases/administração & dosagem , Pirazinas/administração & dosagem , Pirazinas/química , Pirazinas/farmacologia , Pirrolidinas/administração & dosagem , Pirrolidinas/química , Pirrolidinas/farmacologia , Ribavirina/administração & dosagem , Ribavirina/química , Ribavirina/farmacologia , Eletricidade Estática
14.
Cell Res ; 30(8): 678-692, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32541865

RESUMO

A new coronavirus SARS-CoV-2, also called novel coronavirus 2019 (2019-nCoV), started to circulate among humans around December 2019, and it is now widespread as a global pandemic. The disease caused by SARS-CoV-2 virus is called COVID-19, which is highly contagious and has an overall mortality rate of 6.35% as of May 26, 2020. There is no vaccine or antiviral available for SARS-CoV-2. In this study, we report our discovery of inhibitors targeting the SARS-CoV-2 main protease (Mpro). Using the FRET-based enzymatic assay, several inhibitors including boceprevir, GC-376, and calpain inhibitors II, and XII were identified to have potent activity with single-digit to submicromolar IC50 values in the enzymatic assay. The mechanism of action of the hits was further characterized using enzyme kinetic studies, thermal shift binding assays, and native mass spectrometry. Significantly, four compounds (boceprevir, GC-376, calpain inhibitors II and XII) inhibit SARS-CoV-2 viral replication in cell culture with EC50 values ranging from 0.49 to 3.37 µM. Notably, boceprevir, calpain inhibitors II and XII represent novel chemotypes that are distinct from known substrate-based peptidomimetic Mpro inhibitors. A complex crystal structure of SARS-CoV-2 Mpro with GC-376, determined at 2.15 Å resolution with three protomers per asymmetric unit, revealed two unique binding configurations, shedding light on the molecular interactions and protein conformational flexibility underlying substrate and inhibitor binding by Mpro. Overall, the compounds identified herein provide promising starting points for the further development of SARS-CoV-2 therapeutics.


Assuntos
Antivirais/farmacologia , Betacoronavirus/enzimologia , Infecções por Coronavirus/metabolismo , Glicoproteínas/farmacologia , Pneumonia Viral/metabolismo , Prolina/análogos & derivados , Pirrolidinas/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Células A549 , Animais , Antivirais/química , Células CACO-2 , Chlorocebus aethiops , Infecções por Coronavirus/virologia , Cisteína Endopeptidases/química , Cisteína Endopeptidases/metabolismo , Descoberta de Drogas/métodos , Humanos , Concentração Inibidora 50 , Cinética , Pandemias , Pneumonia Viral/virologia , Prolina/farmacologia , Conformação Proteica , Pirrolidinas/química , Células Vero , Proteínas não Estruturais Virais/química , Proteínas não Estruturais Virais/metabolismo
15.
Psychopharmacology (Berl) ; 237(8): 2317-2326, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32382782

RESUMO

RATIONALE: Prolinol aryl ethers and their rigidified analogues pyrrolidinyl benzodioxanes have a high affinity for mammalian α4ß2 nicotinic acetylcholine receptors (nAChRs). Electrophysiological studies have shown that the former are full agonists and the latter partial agonists or antagonists of human α4ß2 receptors, but their in vivo effects are unknown. OBJECTIVES AND METHODS: As α4ß2 nAChRs play an important role in the cognition and the rewarding effects of nicotine, we tested the effects of two full agonists and one antagonist on spatial learning, memory and attention in zebrafish using a T-maze task and virtual object recognition test (VORT). The effect of a partial agonist in reducing nicotine-induced conditioned place preference (CPP) was also investigated. RESULTS: In comparison with the vehicle alone, the full agonists MCL-11 and MCL-28 induced a significant cognitive enhancement as measured by the reduced running time in the T-maze and increased attention as measured by the increased discrimination index in the VORT. MCL-11 was 882 times more potent than nicotine. The two compounds were characterised by an inverted U-shaped dose-response curve, and their effects were blocked by the co-administration of the antagonist MCL-117, which alone had no effect. The partial agonist MCL-54 induced CPP and had an inverted U-shaped dose-response curve similar to that of nicotine but blocked the reinforcing effect of co-administered nicotine. Binding studies showed that all of the compounds have a higher affinity for heteromeric [3H]-epibatidine receptors than [125I]-αBungarotoxin receptors. MCL-11 was the most selective of heteromeric receptors. CONCLUSIONS: These behavioural studies indicate that full agonist prolinol aryl ethers are very active in increasing spatial learning, memory and attention in zebrafish. The benzodioxane partial agonist MCL-54 reduced nicotine-induced CPP, and the benzodioxane antagonist MCL-117 blocked all agonist-induced activities.


Assuntos
Aprendizagem em Labirinto/efeitos dos fármacos , Agonistas Nicotínicos/metabolismo , Antagonistas Nicotínicos/metabolismo , Pirrolidinas/metabolismo , Receptores Nicotínicos/metabolismo , Animais , Relação Dose-Resposta a Droga , Éteres/metabolismo , Éteres/farmacologia , Feminino , Masculino , Aprendizagem em Labirinto/fisiologia , Morfinanos/metabolismo , Morfinanos/farmacologia , Nicotina/metabolismo , Nicotina/farmacologia , Agonistas Nicotínicos/farmacologia , Antagonistas Nicotínicos/farmacologia , Pirrolidinas/farmacologia , Pirrolidinonas/metabolismo , Pirrolidinonas/farmacologia , Peixe-Zebra
16.
Am J Physiol Heart Circ Physiol ; 318(6): H1487-H1508, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32357113

RESUMO

Cell-autonomous circadian clocks have emerged as temporal orchestrators of numerous biological processes. For example, the cardiomyocyte circadian clock modulates transcription, translation, posttranslational modifications, ion homeostasis, signaling cascades, metabolism, and contractility of the heart over the course of the day. Circadian clocks are composed of more than 10 interconnected transcriptional modulators, all of which have the potential to influence the cardiac transcriptome (and ultimately cardiac processes). These transcriptional modulators include BMAL1 and REV-ERBα/ß; BMAL1 induces REV-ERBα/ß, which in turn feeds back to inhibit BMAL1. Previous studies indicate that cardiomyocyte-specific BMAL1-knockout (CBK) mice exhibit a dysfunctional circadian clock (including decreased REV-ERBα/ß expression) in the heart associated with abnormalities in cardiac mitochondrial function, metabolism, signaling, and contractile function. Here, we hypothesized that decreased REV-ERBα/ß activity is responsible for distinct phenotypical alterations observed in CBK hearts. To test this hypothesis, CBK (and littermate control) mice were administered with the selective REV-ERBα/ß agonist SR-9009 (100 mg·kg-1·day-1 for 8 days). SR-9009 administration was sufficient to normalize cardiac glycogen synthesis rates, cardiomyocyte size, interstitial fibrosis, and contractility in CBK hearts (without influencing mitochondrial complex activities, nor normalizing substrate oxidation and Akt/mTOR/GSK3ß signaling). Collectively, these observations highlight a role for REV-ERBα/ß as a mediator of a subset of circadian clock-controlled processes in the heart.


Assuntos
Relógios Circadianos/fisiologia , Ritmo Circadiano/fisiologia , Miocárdio/metabolismo , Membro 1 do Grupo D da Subfamília 1 de Receptores Nucleares/agonistas , Fatores de Transcrição ARNTL/metabolismo , Animais , Ritmo Circadiano/efeitos dos fármacos , Expressão Gênica , Regulação da Expressão Gênica , Coração/efeitos dos fármacos , Camundongos , Camundongos Knockout , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Pirrolidinas/farmacologia , Tiofenos/farmacologia
17.
J Pharmacol Exp Ther ; 374(2): 273-282, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385092

RESUMO

Methylenedioxypyrovalerone (MDPV) is an abused synthetic cathinone, commonly referred to as a "bath salt." Because the dopamine (DA) transporter (DAT) and vesicular monoamine transporter-2 (VMAT-2) are key regulators of both the abuse and neurotoxic potential of structurally and behaviorally related agents, the impact of MDPV on these transporters was investigated. Results revealed that a single in vivo MDPV administration rapidly (within 1 hour) and reversibly increased both rat striatal DAT and VMAT-2 activity, as assessed via [3H]DA uptake in synaptosomes and synaptic vesicles, respectively, prepared from treated rats. There was no evidence of an MDPV-induced increase in plasmalemmal membrane DAT surface expression. Plasma concentrations of MDPV increased dose-dependently as assessed 1 hour after 2.5 and 5.0 mg/kg (s.c.) administration and returned to levels less than 10 ng/ml by 18 hours after 2.5 mg/kg (s.c.). Neither pretreatment with a D1 receptor (SCH23390), a D2 receptor (eticlopride), nor a nicotinic receptor (mecamylamine) antagonist attenuated the MDPV-induced increase in DAT activity. In contrast, eticlopride pretreatment attenuated both the MDPV-induced increase in VMAT-2-mediated DA uptake and an associated increase in cytoplasmic-associated vesicle VMAT-2 immunoreactivity. SCH23390 did not attenuate the MDPV-induced increase in VMAT-2 activity. Repeated MDPV injections did not cause persistent DAergic deficits, as assessed 7 to 8 days later. The impact of MDPV on striatal and hippocampal serotonergic assessments was minimal. Taken together, these data contribute to a growing pharmacological rubric for evaluating the ever-growing list of designer cathinone-related stimulants. The profile of MDPV compared with related psychostimulants is discussed. SIGNIFICANCE STATEMENT: Pharmacological characterization of the synthetic cathinone, 3,4-methylenedioxypyrovalerone (MDPV; commonly referred to as a "bath salt"), is critical for understanding the abuse liability and neurotoxic potential of this and related agents. Accordingly, the impact of MDPV on monoaminergic neurons is described and compared with that of related psychostimulants.


Assuntos
Benzodioxóis/farmacologia , Estimulantes do Sistema Nervoso Central/farmacologia , Drogas Desenhadas/farmacologia , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Pirrolidinas/farmacologia , Transtornos Relacionados ao Uso de Substâncias/metabolismo , Proteínas Vesiculares de Transporte de Monoamina/metabolismo , Animais , Benzodioxóis/farmacocinética , Temperatura Corporal/efeitos dos fármacos , Estimulantes do Sistema Nervoso Central/farmacocinética , Drogas Desenhadas/farmacocinética , Dopamina/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Feminino , Masculino , Neostriado/efeitos dos fármacos , Neostriado/metabolismo , Pirrolidinas/farmacocinética , Ratos , Ratos Sprague-Dawley
18.
J Pharmacol Exp Ther ; 374(1): 211-222, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32345620

RESUMO

The clinical use of first-generation phosphoinositide 3-kinase (PI3K)δ inhibitors in B-cell malignancies is hampered by hepatotoxicity, requiring dose reduction, treatment interruption, and/or discontinuation of therapy. In addition, potential molecular mechanisms by which resistance to this class of drugs occurs have not been investigated. Parsaclisib (INCB050465) is a potent and selective next-generation PI3Kδ inhibitor that differs in structure from first-generation PI3Kδ inhibitors and has shown encouraging anti-B-cell tumor activity and reduced hepatotoxicity in phase 1/2 clinical studies. Here, we present preclinical data demonstrating parsaclisib as a potent inhibitor of PI3Kδ with over 1000-fold selectivity against other class 1 PI3K isozymes. Parsaclisib directly blocks PI3K signaling-mediated cell proliferation in B-cell lines in vitro and in vivo and indirectly controls tumor growth by lessening immunosuppression through regulatory T-cell inhibition in a syngeneic lymphoma model. Diffuse large B-cell lymphoma cell lines overexpressing MYC were insensitive to proliferation blockade via PI3Kδ signaling inhibition by parsaclisib, but their proliferative activities were reduced by suppression of MYC gene transcription. Molecular structure analysis of the first- and next-generation PI3Kδ inhibitors combined with clinical observation suggests that hepatotoxicity seen with the first-generation inhibitors could result from a structure-related off-target effect. Parsaclisib is currently being evaluated in multiple phase 2 clinical trials as a therapy against various hematologic malignancies of B-cell origin (NCT03126019, NCT02998476, NCT03235544, NCT03144674, and NCT02018861). SIGNIFICANCE STATEMENT: The preclinical properties described here provide the mechanism of action and support clinical investigations of parsaclisib as a therapy for B-cell malignancies. MYC overexpression was identified as a resistance mechanism to parsaclisib in DLBCL cells, which may be useful in guiding further translational studies for the selection of patients with DLBCL who might benefit from PI3Kδ inhibitor treatment in future trials. Hepatotoxicity associated with first-generation PI3Kδ inhibitors may be an off-target effect of that class of compounds.


Assuntos
Fígado/efeitos dos fármacos , Linfoma/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase/efeitos adversos , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Pirazóis/efeitos adversos , Pirazóis/farmacologia , Pirimidinas/efeitos adversos , Pirimidinas/farmacologia , Pirrolidinas/efeitos adversos , Pirrolidinas/farmacologia , Animais , Antineoplásicos/efeitos adversos , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Humanos , Fatores Imunológicos/efeitos adversos , Fatores Imunológicos/farmacologia , Camundongos , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Asia Pac J Clin Oncol ; 16 Suppl 1: 3-12, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32348018

RESUMO

Trifluridine/tipiracil is available on the Australian Pharmaceutical Benefits Scheme for the treatment of patients with metastatic colorectal cancer (mCRC) previously treated with, or not considered candidates for, fluoropyrimidine-, oxaliplatin- and irinotecan-based chemotherapies, anti-vascular endothelial growth factor agents and anti-epidermal growth factor receptor agents. This article reviews trifluridine/tipiracil clinical data and presents practical information on its use in the management of refractory mCRC in Australia. Whereas the primary mechanism of action of fluoropyrimidines such as fluorouracil (5-FU) and capecitabine is enzyme inhibition of nucleotide synthesis, trifluridine/tipiracil primarily acts by incorporation into DNA, resulting in DNA dysfunction. Trifluridine/tipiracil has activity in patients with 5-FU-resistant tumors and can be considered in patients with prior intolerance or toxicity to 5-FU. In the pivotal phase III RECOURSE trial evaluating trifluridine/tipiracil in chemotherapy-refractory mCRC, efficacy benefits were observed across all a priori prognostic subgroups including those defined by age (≥65 and ≥75 years), geographical origin, primary tumor site or KRAS status. Trifluridine/tipiracil therapy benefits appropriately selected patients who have an ECOG performance status of 0 or 1, with no more than mild hepatic impairment or mild-to-moderate renal impairment, and who are capable of adhering to oral therapy safely. Appropriate dosing, monitoring for adverse events and effective management of side effects are essential.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Pirrolidinas/uso terapêutico , Qualidade de Vida/psicologia , Trifluridina/uso terapêutico , Uracila/análogos & derivados , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Austrália , Neoplasias Colorretais/patologia , Combinação de Medicamentos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Pirrolidinas/farmacologia , Timina , Trifluridina/farmacologia , Uracila/farmacologia , Uracila/uso terapêutico
20.
Gene ; 748: 144668, 2020 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-32334025

RESUMO

KMN-159 is the lead compound from a series of novel difluorolactam prostanoid EP4 receptor agonists aimed at inducing local bone formation while avoiding the inherent side effects of systemic EP4 activation. KMN-159 is a potent, selective small molecule possessing pharmacokinetic properties amenable to local administration. Unfractionated rat bone marrow cells (BMCs) were treated once at plating with escalating doses of KMN-159 (1 pM to 10 µM). The resulting elevated alkaline phosphatase (ALP) levels measured 9 days post-dose are consistent with increased osteoblastic differentiation and exposure to KMN-159 at low nanomolar concentrations for as little as 30 min was sufficient to induce complete osteoblast differentiation of the BMCs from both sexes and regardless of age. ALP induction was blocked by an EP4 receptor antagonist but not by EP1 or EP2 receptor antagonists and was not induced by EP2 or EP3 receptor agonists. Addition of BMCs to plates coated with KMN-159 24 days earlier resulted in ALP activation, highlighting the chemical stability of the compound. The expression of phenotype markers such as ALP, type I collagen, and osteocalcin was significantly elevated throughout the osteoblastic differentiation timecourse initiated by KMN-159 stimulation. An increased number of tartrate-resistant acid phosphatase-positive cells was observed KMN-159 or PGE2 treated BMCs but only in the presence of exogenous receptor activator of nuclear factor kappa-Β ligand (RANKL). No change in the number of adipocytes was observed. KMN-159 also increased bone healing in a rat calvarial defect model with a healing rate equivalent to recombinant human bone morphogenetic protein-2. Our studies show that KMN-159 is able to stimulate osteoblastic differentiation with a very short time of exposure, supporting its potential as a therapeutic candidate for augmenting bone mass.


Assuntos
Células da Medula Óssea/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Ácidos Heptanoicos/farmacologia , Osteoblastos/efeitos dos fármacos , Pirrolidinas/farmacologia , Receptores de Prostaglandina E Subtipo EP4/agonistas , Fosfatase Alcalina/metabolismo , Animais , Ativação Enzimática , Feminino , Células HEK293 , Humanos , Osteoblastos/citologia , Osteoblastos/enzimologia , Ratos , Ratos Sprague-Dawley
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...