RESUMO
Immune clearance of Hepatitis B virus (HBV) is characterized by broad and robust antiviral T cell responses, while virus-specific T cells in chronic hepatitis B (CHB) are rare and exhibit immune exhaustion that includes programmed-death-1 (PD-1) expression on virus-specific T cells. Thus, an immunotherapy able to expand and activate virus-specific T cells may have therapeutic benefit for CHB patients. Like HBV-infected patients, woodchucks infected with woodchuck hepatitis virus (WHV) can have increased hepatic expression of PD-1-ligand-1 (PD-L1), increased PD-1 on CD8+ T cells, and a limited number of virus-specific T cells with substantial individual variation in these parameters. We used woodchucks infected with WHV to assess the safety and efficacy of anti-PD-L1 monoclonal antibody therapy (αPD-L1) in a variety of WHV infection states. Experimentally-infected animals lacked PD-1 or PD-L1 upregulation compared to uninfected controls, and accordingly, αPD-L1 treatment in lab-infected animals had limited antiviral effects. In contrast, animals with naturally acquired WHV infections displayed elevated PD-1 and PD-L1. In these same animals, combination therapy with αPD-L1 and entecavir (ETV) improved control of viremia and antigenemia compared to ETV treatment alone, but with efficacy restricted to a minority of animals. Pre-treatment WHV surface antigen (sAg) level was identified as a statistically significant predictor of treatment response, while PD-1 expression on peripheral CD8+ T cells, T cell production of interferon gamma (IFN-γ) upon in vitro antigen stimulation (WHV ELISPOT), and circulating levels of liver enzymes were not. To further assess the safety of this strategy, αPD-L1 was tested in acute WHV infection to model the risk of liver damage when the extent of hepatic infection and antiviral immune responses were expected to be the greatest. No significant increase in serum markers of hepatic injury was observed over those in infected, untreated control animals. These data support a positive benefit/risk assessment for blockade of the PD-1:PD-L1 pathway in CHB patients and may help to identify patient groups most likely to benefit from treatment. Furthermore, the efficacy of αPD-L1 in only a minority of animals, as observed here, suggests that additional agents may be needed to achieve a more robust and consistent response leading to full sAg loss and durable responses through anti-sAg antibody seroconversion.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Antígeno B7-H1/imunologia , Modelos Animais de Doenças , Hepatite B/terapia , Animais , Anticorpos Monoclonais/efeitos adversos , MarmotaRESUMO
Synthesis and biological activities of some quinolinone and dihydroquinolinone p38 MAP kinase inhibitors are reported. Modifications to the dihydroquinolinone pharmacophore revealed that dihydroquinolinone may be replaced with a quinolinone pharmacophore and lead to enhanced p38 inhibitory activity. From a study of C-7 substitutions by amino acid side chains, a very potent series of compounds in the p38 enzyme assays was identified. Translation of the in vitro activity into reasonable whole blood activity can be improved in this series of compounds by judicious modification of the physical properties at appropriate regions of the lead.
Assuntos
Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacologia , Piridazinas/química , Pirimidinas/química , Quinolonas/síntese química , Quinolonas/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Cristalografia por Raios X , Ciclização , Humanos , Estrutura Molecular , Quinolonas/química , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Identification and validation of novel drug targets continues to be a major bottleneck in drug development, particularly for polygenic complex diseases such as type 2 diabetes. Here, the authors describe an approach that allows researchers to rapidly identify and validate potential drug targets by combining chemical tools and RNA interference technology. As a proof-of-concept study, the known mechanism Sigma LOPAC library was used to screen for glucose-dependent insulin secretion (GDIS) in INS-1 832/13 cells. In addition to several mechanisms that are known to regulate GDIS (such as cyclic adenosine monophosphate-specific phosphodiesterases, adrenoceptors, and Ca(2+) channels), the authors find that several of the dopamine receptor (DRD) antagonists significantly enhance GDIS, whereas DRD agonists profoundly inhibit GDIS. Subsequent siRNA studies in the same cell line indicate that knockdown of DRD2 enhanced GDIS. Furthermore, selective DRD2 antagonists and agonists also enhance or suppress, respectively, GDIS in isolated rat islets. The data support that the approach described here offers a rapid and effective way for target identification and validation.
Assuntos
Biblioteca Gênica , Inativação Gênica , Glucose/farmacologia , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Redes e Vias Metabólicas/genética , RNA Interferente Pequeno/farmacologia , Animais , Células Cultivadas , Antagonistas de Dopamina/farmacologia , Inativação Gênica/fisiologia , Secreção de Insulina , Redes e Vias Metabólicas/efeitos dos fármacos , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores Dopaminérgicos/genética , Receptores Dopaminérgicos/metabolismoRESUMO
Chemistry was developed to synthesize the title series of compounds. The ability of these novel ligands to bind to the glucocorticoid receptor was investigated. These compounds were also tested in a series of functional assays and some were found to display the profile of a dissociated glucocorticoid. The SAR of the 6,5-bicyclic series differed markedly from the previously reported 6,6-series. Molecular modeling studies were employed to understand the conformational differences between the two series of compounds, which may explain their divergent activity. Two compounds were profiled in vivo and shown to reduce inflammation in a mouse model. An active metabolite is suspected in one case.
Assuntos
Antineoplásicos/farmacologia , Compostos Bicíclicos com Pontes/química , Glucocorticoides/química , Pirazóis/química , Receptores de Glucocorticoides/efeitos dos fármacos , Animais , Antineoplásicos/síntese química , Linhagem Celular Tumoral , Humanos , Ligantes , Camundongos , Modelos Químicos , Modelos Moleculares , Receptores de Glucocorticoides/metabolismo , Relação Estrutura-AtividadeRESUMO
Quinolinones and naphthyridinones with C7 N-t-butyl piperidine substituents were found to be potent p38 MAP kinase inhibitors. These compounds significantly suppress TNF-alpha release in both cellular and LPS-stimulated whole blood assays. They also displayed excellent PK profiles across three animal species. Quinolinone at 10 mpk showed comparable oral efficacy to that of dexamethasone at 1 mpk in a murine collagen-induced arthritis model.
Assuntos
Inibidores Enzimáticos/farmacologia , Naftiridinas/química , Piperidinas/química , Quinolonas/química , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Animais , Artrite Experimental , Colágeno/química , Dexametasona/química , Cães , Haplorrinos , Humanos , Concentração Inibidora 50 , Lipopolissacarídeos/metabolismo , Camundongos , Modelos Químicos , Ratos , Fatores de Tempo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/química , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
A novel series of selective ligands for the human glucocorticoid receptor is described. Structure-activity studies focused on variation of B-ring size, ketal ring size, and ketal substitution. These analogs were found to be potent and selective ligands for GR and have partial agonist profiles in functional assays for transactivation (TAT, GS) and transrepression (IL-6). Of these compounds, 27, 28, and 35 were evaluated further in a mouse LPS-induced TNF-alpha secretion model. Compound 28 had an ED(50) of 14.1 mg/kg compared with 0.5 mg/kg for prednisolone in the same assay.
Assuntos
Receptores de Glucocorticoides/metabolismo , Animais , Células Cultivadas , Humanos , Técnicas In Vitro , Ligantes , CamundongosRESUMO
A series of novel ligands for the glucocorticoid receptor containing two heterocycles were synthesized. These compounds were investigated for a dissociative profile using transrepression and transactivation assays. Several compounds were tested in vivo and showed the ability to reduce inflammation in a mouse.
Assuntos
Glucocorticoides/química , Compostos Heterocíclicos/química , Animais , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Linhagem Celular , Glucocorticoides/administração & dosagem , Glucocorticoides/farmacologia , Compostos Heterocíclicos/administração & dosagem , Compostos Heterocíclicos/farmacologia , Humanos , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/biossínteseRESUMO
A novel series of selective ligands for the human glucocorticoid receptor (hGR) are described. Preliminary structure-activity relationships were focused on substitution at C-1 and indicated a preference for 3-, 4-, and 5-substituted aromatic and benzylic groups. The resulting analogues, e.g., 18 and 34, exhibited excellent affinity for hGR (IC(50) 1.9 nM and 2.8 nM, respectively) and an interesting partial agonist profile in functional assays of transactivation (tyrosine aminotransferase, TAT, and glutamine synthetase, GS) and transrepression (IL-6). The most potent compounds described in this study were the tertiary alcohol derivatives 21 and 25. These candidates showed highly efficacious IL-6 inhibition versus dexamethasone. The thiophenyl analogue 25 was evaluated in vivo in the mouse LPS challenge model and showed an ED(50) = 4.0 mg/kg, compared to 0.5 mg/kg for prednisolone in the same assay.
Assuntos
Anti-Inflamatórios não Esteroides/síntese química , Indazóis/síntese química , Pirazóis/síntese química , Receptores de Glucocorticoides/metabolismo , Tiofenos/síntese química , Animais , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Linhagem Celular , Cristalografia por Raios X , Indução Enzimática , Feminino , Glutamato-Amônia Ligase/biossíntese , Glutamato-Amônia Ligase/genética , Humanos , Indazóis/química , Indazóis/farmacologia , Interleucina-6/antagonistas & inibidores , Ligantes , Camundongos , Camundongos Endogâmicos BALB C , Conformação Molecular , Isoformas de Proteínas/agonistas , Isoformas de Proteínas/metabolismo , Pirazóis/química , Pirazóis/farmacologia , Ensaio Radioligante , Receptores de Glucocorticoides/agonistas , Estereoisomerismo , Relação Estrutura-Atividade , Tiofenos/química , Tiofenos/farmacologia , Transcrição Gênica/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacos , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Tirosina Transaminase/biossíntese , Tirosina Transaminase/genéticaRESUMO
Imidazo[1,2-a]pyridyl N-arylpyridazinones were hybridized from the classic pyridinylimidazoles and the more recent dual hydrogen bond acceptors, resulting in a new structural class of selective p38 MAP kinase inhibitors.
Assuntos
Inibidores Enzimáticos/síntese química , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Piridazinas/síntese química , Animais , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Camundongos , Proteínas Quinases Ativadas por Mitógeno/química , Modelos Moleculares , Piridazinas/química , Piridazinas/farmacologia , Relação Estrutura-Atividade , Proteínas Quinases p38 Ativadas por MitógenoRESUMO
A new class of p38 antagonists based on 3,4-dihydropyrido[3,2,-d]pyrimidine scaffold has been developed. These inhibitors exhibit unprecedented selectivity towards p38 over other very closely related kinases. Compounds 25, 33, and 34 were identified as benchmark analogues for follow-up studies. They show good potency for enzyme inhibition and excellent functional activity.
Assuntos
Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Piridinas/síntese química , Piridinas/farmacologia , Pirimidinonas/síntese química , Pirimidinonas/farmacologia , Desenho de Fármacos , Indicadores e Reagentes , Relação Estrutura-Atividade , Proteínas Quinases p38 Ativadas por MitógenoRESUMO
The development of potent, orally bioavailable (in rat) and selective dihydroquinazolinone inhibitors of p38alpha MAP kinase is described. These analogues are hybrids of a pyridinylimidazole p38alpha inhibitor reported by Merck Research Laboratories and VX-745. Optimization of the C-5 phenyl and the C-7 piperidinyl substituents led to the identification of 15i which gave excellent suppression of TNF-alpha production in LPS-stimulated whole blood (IC(50)=10nM) and good oral exposure in rats (F=68%, AUCn PO=0.58 microM h).