RESUMO
OBJECTIVES: We have previously reported using gene-deficient mice that the interleukin (IL)-23p19 subunit is required for the development of innate immune-driven arthritic pain and disease. We aimed to explore here, using a number of in vivo approaches, how the IL-23p19 subunit can mechanistically control arthritic pain and disease in a T- and B- lymphocyte-independent manner. METHODS: We used the zymosan-induced arthritis (ZIA) model in wild-type and Il23p19-/- mice, by a radiation chimera approach, and by single cell RNAseq and qPCR analyses, to identify the IL23p19-expressing and IL-23-responding cell type(s) in the inflamed joints. This model was also utilized to investigate the efficacy of IL-23p19 subunit blockade with a neutralizing monoclonal antibody (mAb). A novel IL-23-driven arthritis model was established, allowing the identification of putative downstream mediators of IL-23 in the control of pain and disease. Pain and arthritis were assessed by relative static weight distribution and histology, respectively. RESULTS: We present evidence that (i) IL-23p19+ non-bone marrow-derived macrophages are required for the development of ZIA pain and disease, (ii) prophylactic and therapeutic blockade of the IL-23p19 subunit ameliorate ZIA pain and disease and (iii) systemically administered IL-23 can induce arthritic pain and disease in a manner dependent on TNF, GM-CSF, CCL17 and cyclooxygenase activity, but independently of lymphocytes, CGRP, NGF and substance P. CONCLUSIONS: The data presented should aid IL-23 targeting both in the choice of inflammatory disease to be treated and the design of clinical trials.
Assuntos
Artrite Experimental , Camundongos Endogâmicos C57BL , Camundongos Knockout , Animais , Artrite Experimental/imunologia , Artrite Experimental/patologia , Camundongos , Interleucina-23/metabolismo , Interleucina-23/imunologia , Subunidade p19 da Interleucina-23/imunologia , Subunidade p19 da Interleucina-23/antagonistas & inibidores , Subunidade p19 da Interleucina-23/genética , Dor/etiologia , Zimosan , Masculino , Inflamação/imunologia , Inflamação/metabolismoRESUMO
The interleukin (IL)-23 pathway is a pathogenic driver in psoriasis, psoriatic arthritis, and inflammatory bowel disease. Currently, no oral therapeutics selectively target this pathway. JNJ-77242113 is a peptide targeting the IL-23 receptor with high affinity (KD: 7.1 pM). In human cells, JNJ-77242113 potently and selectively inhibited proximal IL-23 signaling (IC50: 5.6 pM) without impacting IL-12 signaling. JNJ-77242113 inhibited IL-23-induced interferon (IFN)γ production in NK cells, and in blood from healthy donors and psoriasis patients (IC50: 18.4, 11 and 9 pM, respectively). In a rat trinitrobenzene sulfonic acid-induced colitis model, oral JNJ-77242113 attenuated disease parameters at doses ≥ 0.3 mg/kg/day. Pharmacologic activity beyond the gastrointestinal tract was also demonstrated. In blood from rats receiving oral JNJ-77242113, dose-dependent inhibition of ex vivo IL-23-stimulated IL-17A production was observed. In an IL-23-induced rat skin inflammation model, JNJ-77242113 inhibited IL-23-induced skin thickening and IL-17A, -17F and -22 gene induction. Oral dosing of JNJ-77242113 in healthy human volunteers inhibited ex vivo IL-23-stimulated IFNγ production in whole blood. Thus, JNJ-77242113 provided selective, systemic IL-23 pathway inhibition in preclinical models which translated to pharmacodynamic activity in healthy human volunteers, supporting the potential for JNJ-77242113 as a selective oral therapy for IL-23-driven immune-mediated diseases.
Assuntos
Interleucina-23 , Receptores de Interleucina , Animais , Humanos , Ratos , Interleucina-23/metabolismo , Administração Oral , Receptores de Interleucina/metabolismo , Masculino , Transdução de Sinais/efeitos dos fármacos , Psoríase/tratamento farmacológico , Psoríase/induzido quimicamente , Colite/tratamento farmacológico , Colite/induzido quimicamente , Modelos Animais de Doenças , Peptídeos/farmacologia , Peptídeos/administração & dosagem , Feminino , Ratos Sprague-DawleyRESUMO
Interleukin (IL)-23 is implicated in the pathogenesis of several inflammatory diseases and is usually linked with helper T cell (Th17) biology. However, there is some data linking IL-23 with innate immune biology in such diseases. We therefore examined the effects of IL-23p19 genetic deletion and/or neutralization on in vitro macrophage activation and in an innate immune-driven peritonitis model. We report that endogenous IL-23 was required for maximal macrophage activation by zymosan as determined by pro-inflammatory cytokine production, including a dramatic upregulation of granulocyte-colony stimulating factor (G-CSF). Furthermore, both IL-23p19 genetic deletion and neutralization in zymosan-induced peritonitis (ZIP) led to a specific reduction in the neutrophil numbers, as well as a reduction in the G-CSF levels in exudate fluids. We conclude that endogenous IL-23 can contribute significantly to macrophage activation during an inflammatory response, mostly likely via an autocrine/paracrine mechanism; of note, endogenous IL-23 can directly up-regulate macrophage G-CSF expression, which in turn is likely to contribute to the regulation of IL-23-dependent neutrophil number and function during an inflammatory response, with potential significance for IL-23 targeting particularly in neutrophil-associated inflammatory diseases.
Assuntos
Inflamação , Interleucina-23 , Células Mieloides , Neutrófilos , Zimosan , Animais , Inflamação/metabolismo , Inflamação/imunologia , Interleucina-23/metabolismo , Camundongos , Neutrófilos/metabolismo , Neutrófilos/imunologia , Células Mieloides/metabolismo , Peritonite/metabolismo , Peritonite/imunologia , Camundongos Endogâmicos C57BL , Fator Estimulador de Colônias de Granulócitos/metabolismo , Ativação de Macrófagos , Macrófagos/metabolismo , Macrófagos/imunologia , Subunidade p19 da Interleucina-23/metabolismo , Subunidade p19 da Interleucina-23/genética , Camundongos KnockoutRESUMO
The nuclear receptor retinoid-related orphan receptor gamma t (RORγt) plays a critical role in driving Th17 cell differentiation and expansion, as well as IL-17 production in innate and adaptive immune cells. The IL-23/IL-17 axis is implicated in several autoimmune and inflammatory diseases, and biologics targeting IL-23 and IL-17 have shown significant clinical efficacy in treating psoriasis and psoriatic arthritis. JNJ-61803534 is a potent RORγt inverse agonist, selectively inhibiting RORγt-driven transcription versus closely-related family members, RORα and RORß. JNJ-61803534 inhibited IL-17A production in human CD4+ T cells under Th17 differentiation conditions, but did not inhibit IFNγ production under Th1 differentiation conditions, and had no impact on in vitro differentiation of regulatory T cells (Treg), nor on the suppressive activity of natural Tregs. In the mouse collagen-induced arthritis model, JNJ-61803534 dose-dependently attenuated inflammation, achieving ~ 90% maximum inhibition of clinical score. JNJ-61803534 significantly inhibited disease score in the imiquimod-induced mouse skin inflammation model, and dose-dependently inhibited the expression of RORγt-regulated genes, including IL-17A, IL-17F, IL-22 and IL-23R. Preclinical 1-month toxicity studies in rats and dogs identified doses that were well tolerated supporting progression into first-in-human studies. An oral formulation of JNJ-61803534 was studied in a phase 1 randomized double-blind study in healthy human volunteers to assess safety, pharmacokinetics, and pharmacodynamics. The compound was well tolerated in single ascending doses (SAD) up to 200 mg, and exhibited dose-dependent increases in exposure upon oral dosing, with a plasma half-life of 164 to 170 h. In addition, dose-dependent inhibition of ex vivo stimulated IL-17A production in whole blood was observed, demonstrating in vivo target engagement. In conclusion, JNJ-61803534 is a potent and selective RORγt inhibitor that exhibited acceptable preclinical safety and efficacy, as well as an acceptable safety profile in a healthy volunteer SAD study, with clear evidence of a pharmacodynamic effect in humans.
Assuntos
Anti-Inflamatórios/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/antagonistas & inibidores , Linfócitos T Reguladores/efeitos dos fármacos , Células Th17/efeitos dos fármacos , Animais , Anti-Inflamatórios/uso terapêutico , Linfócitos T CD4-Positivos/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Método Duplo-Cego , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Interleucinas/genética , Interleucinas/metabolismo , Camundongos , Dermatopatias/tratamento farmacológico , Dermatopatias/metabolismo , Linfócitos T Reguladores/metabolismo , Células Th17/metabolismoRESUMO
BACKGROUND: Psoriasis is an inflammatory, IL-17-driven skin disease in which autoantigen-induced CD8+ T cells have been identified as pathogenic drivers. OBJECTIVE: Our study focused on comprehensively characterizing the phenotypic variation of CD8+ T cells in psoriatic lesions. METHODS: We used single-cell RNA sequencing to compare CD8+ T-cell transcriptomic heterogeneity between psoriatic and healthy skin. RESULTS: We identified 11 transcriptionally diverse CD8+ T-cell subsets in psoriatic and healthy skin. Among several inflammatory subsets enriched in psoriatic skin, we observed 2 Tc17 cell subsets that were metabolically divergent, were developmentally related, and expressed CXCL13, which we found to be a biomarker of psoriasis severity and which achieved comparable or greater accuracy than IL17A in a support vector machine classifier of psoriasis and healthy transcriptomes. Despite high coinhibitory receptor expression in the Tc17 cell clusters, a comparison of these cells with melanoma-infiltrating CD8+ T cells revealed upregulated cytokine, cytolytic, and metabolic transcriptional activity in the psoriatic cells that differed from an exhaustion program. CONCLUSION: Using high-resolution single-cell profiling in tissue, we have uncovered the diverse landscape of CD8+ T cells in psoriatic and healthy skin, including 2 nonexhausted Tc17 cell subsets associated with disease severity.
Assuntos
Autoimunidade , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Psoríase/etiologia , Psoríase/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Estudos de Casos e Controles , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Memória Imunológica , Imunofenotipagem , Interleucina-17/biossíntese , Neoplasias/genética , Neoplasias/imunologia , Análise de Célula ÚnicaRESUMO
The nuclear receptor retinoic acid receptor-related orphan receptor gamma t (RORγt) is a transcription factor that drives Th17 cell differentiation and IL-17 production in both innate and adaptive immune cells. The IL-23/IL-17 pathway is implicated in major autoimmune and inflammatory diseases. RORγt lies at the core of this pathway and represents an attractive opportunity for intervention with small molecule therapeutics. Despite diverse chemical series having been reported, combining high potency and nuclear receptor selectivity with good physicochemical properties remains a challenging endeavor in the field of RORγt drug discovery. We recently described the discovery and evaluation of a new class of potent and selective RORγt inverse agonists based on a thiazole scaffold. Herein we describe the successful optimization of this class by incorporation of an additional amide moiety at the 4-position of the thiazole core. In several optimization cycles, we have reduced human PXR activation, improved solubility, and increased potency while maintaining nuclear receptor selectivity. X-ray crystallographic analysis of compound 1g bound in the sterol binding site of the ligand binding domain of RORγt was largely consistent with an earlier structure, guiding further insight into the molecular mechanism for RORγt inhibition with this series. Compound 1g is orally bioavailable, potent in a human whole blood assay and proved to be efficacious in an ex-vivo IL-17A assay, and was selected for preclinical evaluation.
Assuntos
Amidas/química , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/agonistas , Bibliotecas de Moléculas Pequenas/química , Tiazóis/química , Doenças Autoimunes/tratamento farmacológico , Sítios de Ligação , Cristalografia por Raios X , Avaliação Pré-Clínica de Medicamentos , Humanos , Inflamação/tratamento farmacológico , Interleucina-17/química , Modelos Moleculares , Estrutura Molecular , Ligação Proteica , Bibliotecas de Moléculas Pequenas/farmacologia , Relação Estrutura-Atividade , Tiazóis/farmacologiaRESUMO
Starting from previously identified thiazole-2-carboxamides exemplified by compound 1/6, two new series of RORγt inverse agonists with significantly improved aqueous solubility, ADME parameters and oral PK properties were discovered. These scaffolds were identified from a bioisosteric amide replacement approach. Amongst the variety of heterocycles explored, a 1,3,4-oxadiazole led to compounds with the best overall profile for SAR development and in vivo exploration. In an ex vivo mouse PD model, concentration dependent efficacy was demonstrated and compounds 3/5 and 6/3 were profiled in a 5-day rat tolerability study.
Assuntos
Amidas/farmacologia , Descoberta de Drogas , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/agonistas , Oxidiazóis/farmacologia , Tiazóis/farmacologia , Administração Oral , Amidas/administração & dosagem , Amidas/química , Animais , Relação Dose-Resposta a Droga , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Oxidiazóis/administração & dosagem , Oxidiazóis/química , Ratos , Relação Estrutura-Atividade , Tiazóis/administração & dosagem , Tiazóis/químicaRESUMO
We have previously reported the syntheses of a series of 3,6-disubstituted quinolines as modulators of the retinoic acid receptor-related orphan receptor gamma t (RORγt). These molecules are potent binders but are high molecular weight and they exhibited poor solubility at pH 2 and pH 7. This manuscript details our efforts at improving physical chemical properties for this series of compounds by increasing the diversity at the 3-position (i.e. introducing heteroatoms and lowering the molecular weight). These efforts have led to molecules which are potent binders with improved solubility.
Assuntos
Agonismo Inverso de Drogas , Quinolinas/agonistas , Animais , Humanos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
We identified 6-substituted quinolines as modulators of the retinoic acid receptor-related orphan receptor gamma t (RORγt). The synthesis of this class of RORγt modulators is reported, and optimization of the substituents at the quinoline 6-position that produced compounds with high affinity for the receptor is detailed. This effort identified molecules that act as potent, full inverse agonists in a RORγt-driven cell-based reporter assay. The X-ray crystal structures of two full inverse agonists from this chemical series bound to the RORγt ligand binding domain are disclosed, and we highlight the interaction of a hydrogen-bond acceptor on the 6-position substituent of the inverse agonist with Glu379:NH as a conserved binding contact.
Assuntos
Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/agonistas , Quinolinas/farmacologia , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Humanos , Ligantes , Modelos Moleculares , Estrutura Molecular , Quinolinas/síntese química , Quinolinas/química , Relação Estrutura-AtividadeRESUMO
A high-throughput screen of the ligand binding domain of the nuclear receptor retinoic acid-related orphan receptor gamma t (RORγt) employing a thermal shift assay yielded a quinoline tertiary alcohol hit. Optimization of the 2-, 3- and 4-positions of the quinoline core using structure-activity relationships and structure-based drug design methods led to the discovery of a series of modulators with improved RORγt inhibitory potency and inverse agonism properties.
Assuntos
Desenho de Fármacos , Agonismo Inverso de Drogas , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/antagonistas & inibidores , Quinolinas/química , Quinolinas/farmacologia , Humanos , Simulação de Acoplamento Molecular , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Relação Estrutura-Atividade , Células Th17/efeitos dos fármacosRESUMO
The IL-23/IL-17 pathway is implicated in autoimmune diseases, particularly psoriasis, where biologics targeting IL-23 and IL-17 have shown significant clinical efficacy. Retinoid-related orphan nuclear receptor gamma t (RORγt) is required for Th17 differentiation and IL-17 production in adaptive and innate immune cells. We identified JNJ-54271074, a potent and highly-selective RORγt inverse agonist, which dose-dependently inhibited RORγt-driven transcription, decreased co-activator binding and promoted interaction with co-repressor protein. This compound selectively blocked Th17 differentiation, significantly reduced IL-17A production from memory T cells, and decreased IL-17A- and IL-22-producing human and murine γδ and NKT cells. In a murine collagen-induced arthritis model, JNJ-54271074 dose-dependently suppressed joint inflammation. Furthermore, JNJ-54271074 suppressed IL-17A production in human PBMC from rheumatoid arthritis patients. RORγt-deficient mice showed decreased IL-23-induced psoriasis-like skin inflammation and cytokine gene expression, consistent with dose-dependent inhibition in wild-type mice through oral dosing of JNJ-54271074. In a translational model of human psoriatic epidermal cells and skin-homing T cells, JNJ-54271074 selectively inhibited streptococcus extract-induced IL-17A and IL-17F. JNJ-54271074 is thus a potent, selective RORγt modulator with therapeutic potential in IL-23/IL-17 mediated autoimmune diseases.
Assuntos
Artrite Reumatoide/tratamento farmacológico , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Peptídeos Cíclicos/administração & dosagem , Psoríase/tratamento farmacológico , Células Th17/efeitos dos fármacos , Administração Oral , Animais , Artrite Reumatoide/genética , Artrite Reumatoide/metabolismo , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Interleucina-17/metabolismo , Interleucinas/metabolismo , Camundongos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Peptídeos Cíclicos/farmacologia , Psoríase/genética , Psoríase/metabolismo , Células Th17/citologia , Células Th17/metabolismo , Transcrição Gênica , Interleucina 22RESUMO
The RAR-related orphan receptor gamma t (RORγt) is a nuclear receptor required for generating IL-17-producing CD4(+) Th17 T cells, which are essential in host defense and may play key pathogenic roles in autoimmune diseases. Oxysterols elicit profound effects on immune and inflammatory responses as well as on cholesterol and lipid metabolism. Here, we describe the identification of several naturally occurring oxysterols as RORγt agonists. The most potent and selective activator for RORγt is 7ß, 27-dihydroxycholesterol (7ß, 27-OHC). We show that these oxysterols reverse the inhibitory effect of an RORγt antagonist, ursolic acid, in RORγ- or RORγt-dependent cell-based reporter assays. These ligands bind directly to recombinant RORγ ligand binding domain (LBD), promote recruitment of a coactivator peptide, and reduce binding of a corepressor peptide to RORγ LBD. In primary cells, 7ß, 27-OHC and 7α, 27-OHC enhance the differentiation of murine and human IL-17-producing Th17 cells in an RORγt-dependent manner. Importantly, we showed that Th17, but not Th1 cells, preferentially produce these two oxysterols. In vivo, administration of 7ß, 27-OHC in mice enhanced IL-17 production. Mice deficient in CYP27A1, a key enzyme in generating these oxysterols, showed significant reduction of IL-17-producing cells, including CD4(+) and γδ(+) T cells, similar to the deficiency observed in RORγt knockout mice. Our results reveal a previously unknown mechanism for selected oxysterols as immune modulators and a direct role for CYP27A1 in generating these RORγt agonist ligands, which we propose as RORγt endogenous ligands, driving both innate and adaptive IL-17-dependent immune responses.
Assuntos
Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/agonistas , Esteróis/farmacologia , Células Th17/citologia , Animais , Diferenciação Celular , Colestanotriol 26-Mono-Oxigenase/metabolismo , Interleucina-17/biossíntese , Ligantes , Camundongos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Esteróis/metabolismoRESUMO
Leukotrienes (LT's) are known to play a physiological role in inflammatory immune response. Leukotriene A(4) hydrolase (LTA(4)H) is a cystolic enzyme that stereospecifically catalyzes the transformation of LTA(4) to LTB(4). LTB(4) is a known pro-inflammatory mediator. This paper describes the identification and synthesis of substituted benzofurans as LTH(4)H inhibitors. The benzofuran series demonstrated reduced mouse and human whole blood LTB(4) levels in vitro and led to the identification one analog for advanced profiling. Benzofuran 28 showed dose responsive target engagement and provides a useful tool to explore a LTA(4)H inhibitor for the treatment of inflammatory diseases, such as asthma and inflammatory bowel disease (IBD).
Assuntos
Benzofuranos/química , Inibidores Enzimáticos/química , Epóxido Hidrolases/antagonistas & inibidores , Animais , Benzofuranos/farmacologia , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Humanos , Concentração Inibidora 50 , Camundongos , Estrutura Molecular , Ratos , Ratos Sprague-DawleyRESUMO
Previously, benzthiazole containing LTA(4)H inhibitors were discovered that were potent (1-3), but were associated with the potential for a hERG liability. Utilizing medicinal chemistry first principles (e.g., introducing rigidity, lowering cLogD) a new benzthiazole series was designed, congeners of 1-3, which led to compounds 7a, 7c, 12a-d which exhibited LTA(4)H IC(50)=3-6 nM and hERG Dofetilide Binding IC(50)=8.9-> >10 µM.
Assuntos
Compostos Aza/farmacologia , Benzotiazóis/farmacologia , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Animais , Compostos Aza/síntese química , Compostos Aza/química , Benzotiazóis/síntese química , Benzotiazóis/química , Relação Dose-Resposta a Droga , Desenho de Fármacos , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Epóxido Hidrolases/metabolismo , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Humanos , Camundongos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
In recent years, the classic paradigm of Th1/Th2 CD4(+) T cell-mediated immunity has evolved to include the IL-17A-producing Th17 subset, a distinct proinflammatory CD4(+) T cell lineage. Accumulating evidence suggests that IL-17A and the Th17 pathway may play an important role in the pathology of psoriasis and in other immune-mediated inflammatory diseases. This review summarizes the preclinical and clinical evidence implicating Th17 cells in psoriasis and the therapeutic approaches, approved or under investigation, to target this pathway in psoriasis.
Assuntos
Psoríase/imunologia , Psoríase/metabolismo , Transdução de Sinais , Células Th17/imunologia , Células Th17/metabolismo , Animais , Humanos , Fatores Imunológicos/farmacologia , Fatores Imunológicos/uso terapêutico , Terapia de Alvo Molecular , Psoríase/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Células Th17/efeitos dos fármacosRESUMO
RATIONALE: Allergic asthma is characterized by reversible airway obstruction, lung inflammation, and airway hyperresponsiveness (AHR). Previous studies using leukotriene B(4) (LTB(4)) receptor 1-deficient mice and adoptive transfer experiments have suggested that LTB(4) plays a role in lung inflammation and AHR. OBJECTIVES: In this study, we used a leukotriene A(4) hydrolase (LTA(4)H) inhibitor as a pharmacological tool to directly examine the role of LTB(4) in a mast cell-dependent murine model of allergic airway inflammation. METHODS: We used the forced oscillation technique to test the effects of an LTA(4)H inhibitor dosed during the challenge phase on AHR. Lung tissue and lavage were collected for analysis. MEASUREMENTS AND MAIN RESULTS: Treatment with an LTA(4)H inhibitor improved multiple parameters encompassing AHR and lung function. Significant decreases in inflammatory leukocytes, cytokines, and mucin were observed in the lung lumen. Serum levels of antigen-specific IgE and IgG1 were also decreased. Labeled antigen uptake by lung dendritic cells and subsequent trafficking to draining lymph nodes and the lung were decreased on LTA(4)H inhibitor treatment. Provocatively, inhibition of LTA(4)H increased lipoxin A(4) levels in lung lavage fluid. CONCLUSIONS: These data suggest that LTB(4) plays a key role in driving lung inflammation and AHR. Mechanistically, we provide evidence that inhibition of LTA(4)H, affects recruitment of both CD4(+) and CD8(+) T cells, as well as trafficking of dendritic cells to draining lymph nodes, and may beneficially modulate other pro- and antiinflammatory eicosanoids in the lung. Inhibition of LTA(4)H is thus a potential therapeutic strategy that could modulate key aspects of asthma.
Assuntos
Asma/fisiopatologia , Hiper-Reatividade Brônquica/fisiopatologia , Epóxido Hidrolases/antagonistas & inibidores , Leucotrieno B4/fisiologia , Animais , Líquido da Lavagem Broncoalveolar/química , Células Cultivadas , Citocinas/análise , Imunoglobulina E/sangue , Leucotrieno B4/análise , Mastócitos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Mucinas/análise , Ovalbumina/imunologiaRESUMO
Leukotriene A4 hydrolase (LTA4H) is a ubiquitously expressed enzyme that catalyzes the final step in the synthesis of leukotriene B4 (LTB4), a potent proinflammatory lipid mediator derived from arachidonic acid. Although LTB4 was identified 30 years ago, several recent findings have refocused attention on this mediator as a target for inflammatory and autoimmune diseases. While LTB4 was once thought to be a chemoattractant and activator only of leukocytes mediating acute, innate inflammatory responses, LTB4 receptors have since been discovered on multiple cell types, including T-lymphocytes and antigen-presenting dendritic cells. Thus, the inhibition of LTB4 synthesis demonstrates potential for targeting chronic, autoimmune-driven inflammation. In addition to genetic data in animals and humans linking the LTB4 pathway to cardiovascular disease, variants in the LTA4H gene have been linked with susceptibility to asthma. Several companies have initiated drug discovery efforts to identify potent, selective LTA4H inhibitors. Selected molecules have demonstrated oral efficacy in preclinical models of asthma, inflammatory bowel disease and arthritis, suggesting therapeutic potential for multiple indications. This review focuses on developments with therapeutic relevance for inhibitors of LTA4H as anti-inflammatory drugs, and particularly in the treatment of respiratory disease.
Assuntos
Anti-Inflamatórios/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Inflamação/tratamento farmacológico , Animais , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/imunologia , Células Dendríticas/imunologia , Desenho de Fármacos , Inibidores Enzimáticos/farmacologia , Humanos , Inflamação/imunologia , Leucotrieno B4/antagonistas & inibidores , Leucotrieno B4/biossíntese , Doenças Respiratórias/tratamento farmacológico , Doenças Respiratórias/imunologia , Linfócitos T/imunologiaRESUMO
LTA 4H is a ubiquitously distributed 69 kDa zinc-containing cytosolic enzyme with both hydrolase and aminopeptidase activity. As a hydrolase, LTA 4H stereospecifically catalyzes the transformation of the unstable epoxide LTA 4 to the diol LTB 4, a potent chemoattractant and activator of neutrophils and a chemoattractant of eosinophils, macrophages, mast cells, and T cells. Inhibiting the formation of LTB 4 is expected to be beneficial in the treatment of inflammatory diseases such as inflammatory bowel disease (IBD), asthma, and atherosclerosis. We developed a pharmacophore model using a known inhibitor manually docked into the active site of LTA 4H to identify a subset of compounds for screening. From this work we identified a series of benzoxazole, benzthiazole, and benzimidazole inhibitors. SAR studies resulted in the identification of several potent inhibitors with an appropriate cross-reactivity profile and excellent PK/PD properties. Our efforts focused on further profiling JNJ 27265732, which showed encouraging efficacy in a disease model relevant to IBD.
Assuntos
Anti-Inflamatórios/farmacologia , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Administração Oral , Animais , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/química , Catálise , Cães , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/química , Humanos , Espectroscopia de Ressonância Magnética , Camundongos , Relação Estrutura-AtividadeRESUMO
Leukotriene A(4) hydrolase (LTA(4)H) catalyzes production of the proinflammatory lipid mediator, leukotriene (LT) B(4), which is implicated in a number of inflammatory diseases. We have identified a potent and selective inhibitor of both the epoxide hydrolase and aminopeptidase activities of recombinant human LTA(4)H (IC(50), approximately 10 nM). In a murine model of arachidonic acid-induced ear inflammation, the LTA(4)H inhibitor, JNJ-26993135 (1-[4-(benzothiazol-2-yloxy)-benzyl]-piperidine-4-carboxylic acid), dose-dependently inhibited ex vivo LTB(4) production in blood, in parallel with dose-dependent inhibition of neutrophil influx (ED(50), 1-3 mg/kg) and ear edema. In murine whole blood and in zymosan-induced peritonitis, JNJ-26993135 selectively inhibited LTB(4) production, without affecting cysteinyl leukotriene production, while maintaining or increasing production of the anti-inflammatory mediator, lipoxin (LX) A(4). The 5-lipoxygenase (5-LO) inhibitor zileuton showed inhibition of LTB(4), LTC(4), and LXA(4) production. Although zileuton inhibited LTB(4) production in the peritonitis model more effectively than the LTA(4)H inhibitor, the influx of neutrophils into the peritoneum after 1 and 2 h was significantly higher in zileuton- versus JNJ-26993135-treated animals. This difference may have been mediated by the increased LXA(4) levels in the presence of the LTA(4)H inhibitor. The selective inhibition of LTB(4) production by JNJ-26993135, while increasing levels of the anti-inflammatory mediator, LXA(4), may translate to superior therapeutic efficacy versus 5-LO or 5-LO-activating protein inhibitors in LTB(4)-mediated inflammatory diseases.