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1.
Sci Transl Med ; 11(502)2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31341059

RESUMO

TYK2 is a nonreceptor tyrosine kinase involved in adaptive and innate immune responses. A deactivating coding variant has previously been shown to prevent receptor-stimulated activation of this kinase and provides high protection from several common autoimmune diseases but without immunodeficiency. An agent that recapitulates the phenotype of this deactivating coding variant may therefore represent an important advancement in the treatment of autoimmunity. BMS-986165 is a potent oral agent that similarly blocks receptor-stimulated activation of TYK2 allosterically and with high selectivity and potency afforded through optimized binding to a regulatory domain of the protein. Signaling and functional responses in human TH17, TH1, B cells, and myeloid cells integral to autoimmunity were blocked by BMS-986165, both in vitro and in vivo in a phase 1 clinical trial. BMS-986165 demonstrated robust efficacy, consistent with blockade of multiple autoimmune pathways, in murine models of lupus nephritis and inflammatory bowel disease, supporting its therapeutic potential for multiple immune-mediated diseases.

2.
J Immunol ; 202(5): 1540-1548, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30683702

RESUMO

IL-17A is a critical proinflammatory cytokine for the pathogenesis of asthma including neutrophilic pulmonary inflammation and airway hyperresponsiveness. In this study, by cell type-specific deletion of IL-17R and adaptor Act1, we demonstrated that IL-17R/Act1 exerts a direct impact on the contraction of airway smooth muscle cells (ASMCs). Mechanistically, IL-17A induced the recruitment of Rab35 (a small monomeric GTPase) and DennD1C (guanine nucleotide exchange factor [GEF]) to the IL-17R/Act1 complex in ASMCs, resulting in activation of Rab35. Rab35 knockdown showed that IL-17A-induced Rab35 activation was essential for protein kinase Cα (PKCα) activation and phosphorylation of fascin at Ser39 in ASMCs, allowing F-actin to interact with myosin to form stress fibers and enhance the contraction induced by methacholine. PKCα inhibitor or Rab35 knockdown indeed substantially reduced IL-17A-induced stress fiber formation in ASMCs and attenuated IL-17A-enhanced, methacholine-induced contraction of airway smooth muscle. Taken together, these data indicate that IL-17A promotes airway smooth muscle contraction via direct recruitment of Rab35 to IL-17R, followed by PKCα activation and stress fiber formation.


Assuntos
Interleucina-17/metabolismo , Músculo Liso/metabolismo , Proteína Quinase C-alfa/antagonistas & inibidores , Receptores de Interleucina-17/metabolismo , Fibras de Estresse/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Animais , Interleucina-17/antagonistas & inibidores , Interleucina-17/deficiência , Camundongos , Camundongos Knockout , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Proteína Quinase C-alfa/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Receptores de Interleucina-17/antagonistas & inibidores , Fibras de Estresse/efeitos dos fármacos , Proteínas rab de Ligação ao GTP/antagonistas & inibidores
3.
Artigo em Inglês | MEDLINE | ID: mdl-30553974

RESUMO

INTRODUCTION: Low intrinsic solubility leading to poor oral bioavailability is a common challenge in drug discovery that can often be overcome by formulation strategies, however, it remains a potential limitation that can pose challenges for early risk assessment and represent a significant obstacle to drug development. We identified a selective inhibitor (BMS-986126) of the IL-1 receptor-associated kinase 4 (IRAK4) with favorable properties as a lead candidate, but with unusually low intrinsic solubility of <1 µg/mL. METHODS: Conventional histopathology identified the issue of crystal formation in vivo. Subsequent investigative work included confocal Raman micro-spectroscopy, MALDI-MS, polarized light microscopy of fresh wet-mount tissue scrapings and transmission electron microscopy. RESULTS: BMS-986126 was advanced into a 2-week toxicology study in rats. The main finding in this study was minimal granulomatous inflammation in the duodenum, associated with the presence of birefringent crystals at the highest dosage of 100 mg/kg/day. Considering the safety margin, and the single location of the lesion, BMS-986126 was further progressed into IND-enabling toxicology studies where tolerability deteriorated with increasing dosing duration. Birefringent crystals and granulomatous inflammation were detected in multiple organs at dosages ≥20 mg/kg/day. Raman spectroscopy confirmed the identity of the crystals as BMS-986126. Therefore, follow up investigations were conducted to further characterize drug crystallization and to evaluate detection methods for their potential to reliably detect in vivo crystallization early. DISCUSSION: The purpose of our efforts was to identify critical factors influencing in vivo drug crystallization and to provide a preliminary assessment (based on one compound) which method would be best suited for identifying crystals. Results indicated a combination of methods was required to provide a complete assessment of drug crystallization and that a simple technique, scraping of freshly collected tissue followed by evaluation under polarizing light was suitable for detecting crystals. However, dosing for 2 weeks was required for crystals to grow to a clearly detectable size.


Assuntos
Cristalização , Quinases Associadas a Receptores de Interleucina-1/antagonistas & inibidores , Pirazóis/química , Piridinas/química , Animais , Disponibilidade Biológica , Descoberta de Drogas , Duodeno/patologia , Feminino , Quinases Associadas a Receptores de Interleucina-1/química , Macrófagos Alveolares/efeitos dos fármacos , Masculino , Cultura Primária de Células , Pirazóis/administração & dosagem , Pirazóis/farmacocinética , Piridinas/administração & dosagem , Piridinas/farmacocinética , Ratos , Ratos Sprague-Dawley , Risco , Solubilidade , Análise Espectral Raman
4.
J Clin Invest ; 128(12): 5399-5412, 2018 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-30372424

RESUMO

NLRP3 inflammasome plays a critical spatiotemporal role in the pathogenesis of experimental autoimmune encephalomyelitis (EAE). This study reports a mechanistic insight into noncanonical NLRP3 inflammasome activation in microglia for the effector stage of EAE. Microglia-specific deficiency of ASC (apoptosis-associated speck-like protein containing a C-terminal caspase-activation and recruitment [CARD] domain) attenuated T cell expansion and neutrophil recruitment during EAE pathogenesis. Mechanistically, TLR stimulation led to IRAKM-caspase-8-ASC complex formation, resulting in the activation of caspase-8 and IL-1ß release in microglia. Noncanonical inflammasome-derived IL-1ß produced by microglia in the CNS helped to expand the microglia population in an autocrine manner and amplified the production of inflammatory cytokines/chemokines. Furthermore, active caspase-8 was markedly increased in the microglia in the brain tissue from patients with multiple sclerosis. Taken together, our study suggests that microglia-derived IL-1ß via noncanonical caspase-8-dependent inflammasome is necessary for microglia to exert their pathogenic role during CNS inflammation.


Assuntos
Caspase 8/metabolismo , Encefalomielite Autoimune Experimental/enzimologia , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Microglia/metabolismo , Esclerose Múltipla/enzimologia , Transdução de Sinais , Animais , Caspase 8/genética , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/patologia , Feminino , Humanos , Inflamação/genética , Inflamação/metabolismo , Inflamação/patologia , Quinases Associadas a Receptores de Interleucina-1/genética , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Masculino , Camundongos , Camundongos Knockout , Microglia/patologia , Esclerose Múltipla/genética , Esclerose Múltipla/patologia , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo
5.
Front Immunol ; 9: 1613, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30123214

RESUMO

Agents targeting the PD1-PDL1 axis have transformed cancer therapy. Factors that influence clinical response to PD1-PDL1 inhibitors include tumor mutational burden, immune infiltration of the tumor, and local PDL1 expression. To identify peripheral correlates of the anti-tumor immune response in the absence of checkpoint blockade, we performed a retrospective study of circulating T cell subpopulations and matched tumor gene expression in melanoma and non-small cell lung cancer (NSCLC) patients. Notably, both melanoma and NSCLC patients whose tumors exhibited increased inflammatory gene transcripts presented high CD4+ and CD8+ central memory T cell (CM) to effector T cell (Eff) ratios in blood. Consequently, we evaluated CM/Eff T cell ratios in a second cohort of NSCLC. The data showed that high CM/Eff T cell ratios correlated with increased tumor PDL1 expression. Furthermore, of the 22 patients within this NSCLC cohort who received nivolumab, those with high CM/Eff T cell ratios, had longer progression-free survival (PFS) (median survival: 91 vs. 215 days). These findings show that by providing a window into the state of the immune system, peripheral T cell subpopulations inform about the state of the anti-tumor immune response and identify potential blood biomarkers of clinical response to checkpoint inhibitors in melanoma and NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/imunologia , Neoplasias Pulmonares/imunologia , Melanoma/imunologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Subpopulações de Linfócitos T/imunologia , Idoso , Antineoplásicos Imunológicos/uso terapêutico , Antígeno B7-H1/imunologia , Antígeno B7-H1/metabolismo , Biomarcadores Tumorais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Nivolumabe/uso terapêutico , Receptor de Morte Celular Programada 1/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Intervalo Livre de Progressão , Subpopulações de Linfócitos T/metabolismo
6.
Nat Commun ; 9(1): 2745, 2018 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-30013031

RESUMO

Although Act1 (adaptor for IL-17 receptors) is necessary for IL-17-mediated inflammatory responses, Act1- (but not Il17ra-, Il17rc-, or Il17rb-) deficient mice develop spontaneous SLE- and Sjögren's-like diseases. Here, we show that Act1 functions as a negative regulator in T and B cells via direct inhibition of STAT3. Mass spectrometry analysis detected an Act1-STAT3 complex, deficiency of Act1 (but not Il17ra-, Il17rc-, or Il17rb) results in hyper IL-23- and IL-21-induced STAT3 activation in T and B cells, respectively. IL-23R deletion or blockade of IL-21 ameliorates SLE- and Sjögren's-like diseases in Act1-/- mice. Act1 deficiency results in hyperactivated follicular Th17 cells with elevated IL-21 expression, which promotes T-B cell interaction for B cell expansion and antibody production. Moreover, anti-IL-21 ameliorates the SLE- and Sjögren's-like diseases in Act1-deficient mice. Thus, IL-21 blocking antibody might be an effective therapy for treating SLE- and Sjögren's-like syndrome in patients containing Act1 mutation.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Linfócitos B/imunologia , Interleucinas/genética , Lúpus Eritematoso Sistêmico/genética , Fator de Transcrição STAT3/genética , Síndrome de Sjogren/genética , Linfócitos T/imunologia , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Animais , Anticorpos Monoclonais/farmacologia , Linfócitos B/efeitos dos fármacos , Linfócitos B/patologia , Diferenciação Celular , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica , Interleucina-17/genética , Interleucina-17/imunologia , Interleucinas/antagonistas & inibidores , Interleucinas/imunologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/patologia , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Cultura Primária de Células , Receptores de Interleucina/deficiência , Receptores de Interleucina/genética , Receptores de Interleucina/imunologia , Receptores de Interleucina-17/deficiência , Receptores de Interleucina-17/genética , Receptores de Interleucina-17/imunologia , Fator de Transcrição STAT3/imunologia , Transdução de Sinais , Síndrome de Sjogren/tratamento farmacológico , Síndrome de Sjogren/imunologia , Síndrome de Sjogren/patologia , Baço , Linfócitos T/efeitos dos fármacos , Linfócitos T/patologia
7.
Arthritis Rheumatol ; 70(8): 1331-1342, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29534336

RESUMO

OBJECTIVE: To develop an objective, readily measurable pharmacodynamic biomarker of glucocorticoid (GC) activity. METHODS: Genes modulated by prednisolone were identified from in vitro studies using peripheral blood mononuclear cells from normal healthy volunteers. Using the criteria of a >2-fold change relative to vehicle controls and an adjusted P value cutoff of less than 0.05, 64 up-regulated and 18 down-regulated genes were identified. A composite score of the up-regulated genes was generated using a single-sample gene set enrichment analysis algorithm. RESULTS: GC gene signature expression was significantly elevated in peripheral blood leukocytes from normal healthy volunteers following oral administration of prednisolone. Expression of the signature increased in a dose-dependent manner, peaked at 4 hours postadministration, and returned to baseline levels by 48 hours after dosing. Lower expression was detected in normal healthy volunteers who received a partial GC receptor agonist, which is consistent with the reduced transactivation potential of this compound. In cohorts of patients with systemic lupus erythematosus and patients with rheumatoid arthritis, expression of the GC signature was negatively correlated with the percentages of peripheral blood lymphocytes and positively correlated with peripheral blood neutrophil counts, which is consistent with the known biology of the GC receptor. Expression of the signature largely agreed with reported GC use in these populations, although there was significant interpatient variability within the dose cohorts. CONCLUSION: The GC gene signature identified in this study represents a pharmacodynamic marker of GC exposure.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Glucocorticoides/administração & dosagem , Leucócitos Mononucleares/efeitos dos fármacos , Prednisolona/administração & dosagem , Administração Oral , Artrite Reumatoide/sangue , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/genética , Biomarcadores/sangue , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Feminino , Voluntários Saudáveis , Humanos , Contagem de Leucócitos , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Lúpus Eritematoso Sistêmico/genética , Masculino , Testes Farmacogenômicos , Regulação para Cima/efeitos dos fármacos
8.
J Immunol ; 199(11): 3849-3857, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-29070673

RESUMO

This study identifies a novel mechanism linking IL-17A with colon tissue repair and tumor development. Abrogation of IL-17A signaling in mice attenuated tissue repair of dextran sulfate sodium (DSS)-induced damage in colon epithelium and markedly reduced tumor development in an azoxymethane/DSS model of colitis-associated cancer. A novel IL-17A target gene, PLET1 (a progenitor cell marker involved in wound healing), was highly induced in DSS-treated colon tissues and tumors in an IL-17RC-dependent manner. PLET1 expression was induced in LGR5+ colon epithelial cells after DSS treatment. LGR5+PLET1+ marks a highly proliferative cell population with enhanced expression of IL-17A target genes. PLET1 deficiency impaired tissue repair of DSS-induced damage in colon epithelium and reduced tumor formation in an azoxymethane/DSS model of colitis-associated cancer. Our results suggest that IL-17A-induced PLET1 expression contributes to tissue repair and colon tumorigenesis.


Assuntos
Colite/imunologia , Colo/metabolismo , Neoplasias do Colo/imunologia , Células Epiteliais/imunologia , Interleucina-17/metabolismo , Proteínas da Gravidez/metabolismo , Animais , Azoximetano , Carcinogênese , Células Cultivadas , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Colite/induzido quimicamente , Colo/patologia , Neoplasias do Colo/induzido quimicamente , Sulfato de Dextrana , Regulação Neoplásica da Expressão Gênica , Interleucina-17/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas da Gravidez/genética , Receptores de Interleucina/genética , Cicatrização
9.
Elife ; 62017 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-28990926

RESUMO

Expression of inflammatory genes is determined in part by post-transcriptional regulation of mRNA metabolism but how stimulus- and transcript-dependent nuclear export influence is poorly understood. Here, we report a novel pathway in which LPS/TLR4 engagement promotes nuclear localization of IRAK2 to facilitate nuclear export of a specific subset of inflammation-related mRNAs for translation in murine macrophages. IRAK2 kinase activity is required for LPS-induced RanBP2-mediated IRAK2 sumoylation and subsequent nuclear translocation. Array analysis showed that an SRSF1-binding motif is enriched in mRNAs dependent on IRAK2 for nuclear export. Nuclear IRAK2 phosphorylates SRSF1 to reduce its binding to target mRNAs, which promotes the RNA binding of the nuclear export adaptor ALYREF and nuclear export receptor Nxf1 loading for the export of the mRNAs. In summary, LPS activates a nuclear function of IRAK2 that facilitates the assembly of nuclear export machinery to export selected inflammatory mRNAs to the cytoplasm for translation.


Assuntos
Transporte Ativo do Núcleo Celular , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Macrófagos/imunologia , RNA Mensageiro/metabolismo , Animais , Lipopolissacarídeos/metabolismo , Macrófagos/efeitos dos fármacos , Camundongos , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Fosforilação , Proteínas de Ligação a RNA/metabolismo , Fatores de Processamento de Serina-Arginina/metabolismo , Sumoilação
10.
Bioorg Med Chem Lett ; 27(21): 4908-4913, 2017 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-28947151

RESUMO

The identification of small molecule inhibitors of IRAK4 for the treatment of autoimmune diseases has been an area of intense research. We discovered novel 4,6-diaminonicotinamides which potently inhibit IRAK4. Optimization efforts were aided by X-ray crystal structures of inhibitors bound to IRAK4. Structure activity relationship (SAR) studies led to the identification of compound 29 which exhibited sub-micromolar potency in a LTA stimulated cellular assay.


Assuntos
Desenho de Drogas , Quinases Associadas a Receptores de Interleucina-1/antagonistas & inibidores , Niacinamida/química , Inibidores de Proteínas Quinases/química , Sítios de Ligação , Cristalografia por Raios X , Avaliação Pré-Clínica de Medicamentos , Humanos , Concentração Inibidora 50 , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Janus Quinase 3/química , Janus Quinase 3/metabolismo , Conformação Molecular , Simulação de Dinâmica Molecular , Niacinamida/metabolismo , Inibidores de Proteínas Quinases/metabolismo , Estrutura Terciária de Proteína , Relação Estrutura-Atividade
11.
Nat Commun ; 8: 15508, 2017 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-28561022

RESUMO

NOTCH1 signalling contributes to defective remyelination by impairing differentiation of oligodendrocyte progenitor cells (OPCs). Here we report that IL-17 stimulation induces NOTCH1 activation in OPCs, contributing to Th17-mediated demyelinating disease. Mechanistically, IL-17R interacts with NOTCH1 via the extracellular domain, which facilitates the cleavage of NOTHC1 intracellular domain (NICD1). IL-17-induced NOTCH1 activation results in the interaction of IL-17R adaptor Act1 with NICD1, followed by the translocation of the Act1-NICD1 complex into the nucleus. Act1-NICD1 are recruited to the promoters of several NOTCH1 target genes (including STEAP4, a metalloreductase important for inflammation and cell proliferation) that are specifically induced in the spinal cord by Th17 cells. A decoy peptide disrupting the IL-17RA-NOTCH1 interaction inhibits IL-17-induced NOTCH1 activation and attenuates Th17-mediated experimental autoimmune encephalitis (EAE). Taken together, these findings demonstrate critical crosstalk between the IL-17 and NOTCH1 pathway, regulating Th17-induced inflammatory and proliferative genes to promote demyelinating disease.


Assuntos
Encefalomielite Autoimune Experimental/imunologia , Interleucina-17/metabolismo , Esclerose Múltipla/imunologia , Células Precursoras de Oligodendrócitos/fisiologia , Receptor Notch1/imunologia , Células Th17/imunologia , Animais , Astrócitos , Diferenciação Celular/imunologia , Proliferação de Células/fisiologia , Técnicas de Cocultura , Feminino , Células HEK293 , Células HeLa , Humanos , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/genética , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/imunologia , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Interleucina-17/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Cultura Primária de Células , Ligação Proteica/imunologia , Domínios Proteicos/fisiologia , Receptor Notch1/genética , Receptor Notch1/metabolismo , Receptores de Interleucina-17/metabolismo , Remielinização/fisiologia , Transdução de Sinais/imunologia , Células Th1/imunologia , Células Th17/metabolismo , Peptídeos e Proteínas Associados a Receptores de Fatores de Necrose Tumoral/metabolismo
12.
J Immunol ; 198(3): 1308-1319, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28003376

RESUMO

The serine/threonine kinase IL-1R-associated kinase (IRAK)4 is a critical regulator of innate immunity. We have identified BMS-986126, a potent, highly selective inhibitor of IRAK4 kinase activity that demonstrates equipotent activity against multiple MyD88-dependent responses both in vitro and in vivo. BMS-986126 failed to inhibit assays downstream of MyD88-independent receptors, including the TNF receptor and TLR3. Very little activity was seen downstream of TLR4, which can also activate an MyD88-independent pathway. In mice, the compound inhibited cytokine production induced by injection of several different TLR agonists, including those for TLR2, TLR7, and TLR9. The compound also significantly suppressed skin inflammation induced by topical administration of the TLR7 agonist imiquimod. BMS-986126 demonstrated robust activity in the MRL/lpr and NZB/NZW models of lupus, inhibiting multiple pathogenic responses. In the MRL/lpr model, robust activity was observed with the combination of suboptimal doses of BMS-986126 and prednisolone, suggesting the potential for steroid sparing activity. BMS-986126 also demonstrated synergy with prednisolone in assays of TLR7- and TLR9-induced IFN target gene expression using human PBMCs. Lastly, BMS-986126 inhibited TLR7- and TLR9-dependent responses using cells derived from lupus patients, suggesting that inhibition of IRAK4 has the potential for therapeutic benefit in treating lupus.


Assuntos
Quinases Associadas a Receptores de Interleucina-1/antagonistas & inibidores , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Prednisolona/uso terapêutico , Animais , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fator 88 de Diferenciação Mieloide/fisiologia , Receptor 7 Toll-Like/fisiologia , Receptor Toll-Like 9/fisiologia
13.
JCI Insight ; 1(9): e87310, 2016 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-27699274

RESUMO

Systemic lupus erythematosus (SLE) is a complex systemic autoimmune disease driven by both innate and adaptive immune cells. African Americans tend to present with more severe disease at an earlier age compared with patients of European ancestry. In order to better understand the immunological differences between African American and European American patients, we analyzed the frequencies of B cell subsets and the expression of B cell activation markers from a total of 68 SLE patients and 69 normal healthy volunteers. We found that B cells expressing the activation markers CD86, CD80, PD1, and CD40L, as well as CD19+CD27-IgD- double-negative B cells, were enriched in African American patients vs. patients of European ancestry. In addition to increased expression of CD40L, surface levels of CD40 on B cells were lower, suggesting the engagement of the CD40 pathway. In vitro experiments confirmed that CD40L expressed by B cells could lead to CD40 activation and internalization on adjacent B cells. To conclude, these results indicate that, compared with European American patients, African American SLE patients present with a particularly active B cell component, possibly via the activation of the CD40/CD40L pathway. These data may help guide the development of novel therapies.


Assuntos
Linfócitos B/citologia , Lúpus Eritematoso Sistêmico/etnologia , Afro-Americanos , Antígenos de Superfície/análise , Antígeno B7-2/análise , Antígenos CD40/análise , Ligante de CD40/análise , Humanos , Fenótipo
14.
J Med Chem ; 58(10): 4278-90, 2015 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-25905990

RESUMO

An empirical approach to improve the microsomal stability and CYP inhibition profile of lead compounds 1a and 1b led to the identification of 5 (BMS-341) as a dissociated glucocorticoid receptor modulator. Compound 5 showed significant improvements in pharmacokinetic properties and, unlike compounds 1a-b, displayed a linear, dose-dependent pharmacokinetic profile in rats. When tested in a chronic model of adjuvant-induced arthritis in rat, the ED50 of 5 (0.9 mg/kg) was superior to that of both 1a and 1b (8 and 17 mg/kg, respectively).


Assuntos
Artrite Experimental/tratamento farmacológico , Inibidores das Enzimas do Citocromo P-450/farmacologia , Compostos Heterocíclicos com 3 Anéis/farmacologia , Receptores de Glucocorticoides/metabolismo , Tiadiazóis/farmacologia , Animais , Sangue/efeitos dos fármacos , Sangue/metabolismo , Técnicas de Química Sintética , Citocromo P-450 CYP3A , Inibidores do Citocromo P-450 CYP3A/química , Inibidores do Citocromo P-450 CYP3A/farmacologia , Inibidores das Enzimas do Citocromo P-450/química , Inibidores das Enzimas do Citocromo P-450/farmacocinética , Modelos Animais de Doenças , Cães , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/métodos , Estabilidade de Medicamentos , Compostos Heterocíclicos com 3 Anéis/química , Compostos Heterocíclicos com 3 Anéis/farmacocinética , Humanos , Masculino , Ratos Endogâmicos Lew , Receptores de Glucocorticoides/agonistas , Relação Estrutura-Atividade , Tiadiazóis/química , Tiadiazóis/farmacocinética , Fator de Transcrição AP-1/metabolismo
15.
Bioorg Med Chem Lett ; 24(15): 3268-73, 2014 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-24980053

RESUMO

Acylureas and acyclic imides are found to be excellent isosteres for 2-acylamino-1,3,4-thiadiazole in the azaxanthene-based series of glucocorticoid receptor (GR) agonists. The results reported herein show that primary acylureas maintain high affinity and selectivity for GR while providing improved CYP450 inhibition and pharmacokinetic profile over 2-acylamino-1,3,4-thiadiazoles. General methods for synthesis of a variety of acylureas and acyclic imides from a carboxylic acid were utilized and are described.


Assuntos
Descoberta de Drogas , Compostos Heterocíclicos com 3 Anéis/farmacologia , Receptores de Glucocorticoides/agonistas , Tiadiazóis/farmacologia , Ureia/farmacologia , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Compostos Heterocíclicos com 3 Anéis/síntese química , Compostos Heterocíclicos com 3 Anéis/química , Humanos , Modelos Moleculares , Estrutura Molecular , Estereoisomerismo , Relação Estrutura-Atividade , Tiadiazóis/química , Ureia/análogos & derivados , Ureia/química
16.
J Immunol ; 191(2): 640-9, 2013 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-23772036

RESUMO

IL-17, a major inflammatory cytokine plays a critical role in the pathogenesis of many autoimmune inflammatory diseases. In this study, we report a new function of RNA-binding protein HuR in IL-17-induced Act1-mediated chemokine mRNA stabilization. HuR deficiency markedly reduced IL-17-induced chemokine expression due to increased mRNA decay. Act1-mediated HuR polyubiquitination was required for the binding of HuR to CXCL1 mRNA, leading to mRNA stabilization. Although IL-17 induced the coshift of Act1 and HuR to the polysomal fractions in a sucrose gradient, HuR deficiency reduced the ratio of translation-active/translation-inactive IL-17-induced chemokine mRNAs. Furthermore, HuR deletion in distal lung epithelium attenuated IL-17-induced neutrophilia. In summary, HuR functions to couple receptor-proximal signaling to posttranscriptional machinery, contributing to IL-17-induced inflammation.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Quimiocina CXCL1/genética , Quimiocina CXCL5/genética , Proteínas ELAV/metabolismo , Interleucina-17/metabolismo , Estabilidade de RNA , Animais , Linhagem Celular , Proteínas ELAV/genética , Células HeLa , Humanos , Inflamação/imunologia , Pulmão/metabolismo , Camundongos , Camundongos Knockout , Ligação Proteica , RNA Mensageiro/metabolismo , Mucosa Respiratória/metabolismo , Transdução de Sinais , Ubiquitinação
17.
Nat Immunol ; 14(1): 72-81, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23202271

RESUMO

Act1 is an essential adaptor in interleukin 17 (IL-17)-mediated signaling and is recruited to the receptor for IL-17 after stimulation with IL-17. Here we found that Act1 was a 'client' protein of the molecular chaperone hsp90. The D10N variant of Act1 (Act1(D10N)) that is linked to susceptibility to psoriasis was defective in its interaction with hsp90, which resulted in a global loss of Act1 function. Act1-deficient mice modeled the mechanistic link between loss of Act1 function and susceptibility to psoriasis. Although Act1 was necessary for IL-17-mediated inflammation, Act1-deficient mice had a hyperactive response of the T(H)17 subset of helper T cells and developed spontaneous IL-22-dependent skin inflammation. In the absence of IL-17 signaling, IL-22 was the main contributor to skin inflammation, which provides a molecular mechanism for the association of Act1(D10N) with psoriasis susceptibility.


Assuntos
Conexina 43/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Chaperonas Moleculares/metabolismo , Fragmentos de Peptídeos/metabolismo , Psoríase/imunologia , Células Th17/imunologia , Animais , Linhagem Celular , Conexina 43/genética , Conexina 43/imunologia , Modelos Animais de Doenças , Predisposição Genética para Doença , Humanos , Interleucina-17/metabolismo , Camundongos , Camundongos Knockout , Chaperonas Moleculares/genética , Mutação/genética , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Polimorfismo Genético , Ligação Proteica/genética , Ligação Proteica/imunologia , Psoríase/genética , Transdução de Sinais
18.
Immunity ; 37(5): 800-12, 2012 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-23142783

RESUMO

Interleukin-1 (IL-1)-induced activation of the mTOR kinase pathway has major influences on Th17 cell survival, proliferation, and effector function. Via biochemical and genetic approaches, the kinases IKKi and GSK3α were identified as the critical intermediate signaling components for IL-1-induced AKT activation, which in turn activated mTOR. Although insulin-induced AKT activation is known to phosphorylate and inactivate GSK3α and GSK3ß, we found that GSK3α but not GSK3ß formed a constitutive complex to phosphorylate and suppress AKT activation, showing that a reverse action from GSK to AKT can take place. Upon IL-1 stimulation, IKKi was activated to mediate GSK3α phosphorylation at S21, thereby inactivating GSK3α to promote IL-1-induced AKT-mTOR activation. Thus, IKKi has a critical role in Th17 cell maintenance and/or proliferation through the GSK-AKT-mTOR pathway, implicating the potential of IKKi as a therapeutic target.


Assuntos
Quinase 3 da Glicogênio Sintase/metabolismo , Quinase I-kappa B/metabolismo , Interleucina-1/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Células Th17/metabolismo , Animais , Processos de Crescimento Celular/fisiologia , Ativação Enzimática , Quinase 3 da Glicogênio Sintase/imunologia , Glicogênio Sintase Quinase 3 beta , Insulina/imunologia , Insulina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Proto-Oncogênicas c-akt/imunologia , Transdução de Sinais , Serina-Treonina Quinases TOR/imunologia , Células Th17/citologia , Células Th17/enzimologia , Células Th17/imunologia
19.
J Med Chem ; 54(20): 7318-33, 2011 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-21899328

RESUMO

Structurally novel 5H-chromeno[2,3-b]pyridine (azaxanthene) selective glucocorticoid receptor (GR) modulators have been identified. A screening paradigm utilizing cellular assays of GR-mediated transrepression of proinflammatory transcription factors and transactivation of GR-dependent genes combined with three physiologically relevant assays of cytokine induction in human whole blood has allowed for the identification of high affinity, selective GR ligands that display a broad range of pharmacological profiles. Agonist efficacy in reporter assays can be tuned by halogenation of a pendent phenyl ring and correlates well with efficacy for cytokine inhibition in human whole blood. A hypothetical binding mode is proposed, invoking an expanded ligand binding pocket resembling that of arylpyrazole-bound GR structures. Two compounds of close structural similarity (35 and 37; BMS-776532 and BMS-791826, respectively) have been found to maintain distinct and consistent levels of partial agonist efficacy across several assays, displaying anti-inflammatory activity comparable to that of prednisolone 2 in suppressing cytokine production in whole blood and in rodent models of acute and chronic inflammation.


Assuntos
Anti-Inflamatórios não Esteroides/síntese química , Compostos Heterocíclicos com 3 Anéis/síntese química , Receptores de Glucocorticoides/agonistas , Tiadiazóis/síntese química , Fosfatase Alcalina/biossíntese , Animais , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Artrite Experimental/tratamento farmacológico , Linhagem Celular Tumoral , Agonismo Parcial de Drogas , Edema/tratamento farmacológico , Glutamato-Amônia Ligase/biossíntese , Compostos Heterocíclicos com 3 Anéis/química , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Técnicas In Vitro , Interleucina-1beta/sangue , Masculino , Modelos Moleculares , Ratos , Ratos Endogâmicos Lew , Ratos Sprague-Dawley , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Elementos de Resposta , Estereoisomerismo , Relação Estrutura-Atividade , Tiadiazóis/química , Tiadiazóis/farmacologia , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/metabolismo , Ativação Transcricional , Fator de Necrose Tumoral alfa/sangue , Tirosina Transaminase/biossíntese
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