RESUMEN
Signal transducers and activators of transcription 6 (STAT6) is an important transcription factor in interleukin (IL)-4 signaling pathway and a key regulator of the type 2 helper T (Th2) cell immune response. Therefore, STAT6 may be an excellent therapeutic target for allergic conditions, including asthma and atopic diseases. Previously, we reported 4-aminopyrimidine-5-carboxamide derivatives as STAT6 inhibitors. To search for novel STAT6 inhibitors, we synthesized fused bicyclic pyrimidine derivatives and identified a 7H-pyrrolo[2,3-d]pyrimidine derivative as a STAT6 inhibitor. Optimization of the pyrrolopyrimidine derivatives led to identification of 2-[4-(4-{[7-(3,5-difluorobenzyl)-7H-pyrrolo[2,3-d]pyrimidin-2-yl]amino}phenyl)piperazin-1-yl]acetamide (24, AS1810722) which showed potent STAT6 inhibition and a good CYP3A4 inhibition profile. Compound 24 also inhibited in vitro Th2 differentiation without affecting type 1 helper T (Th1) cell differentiation and eosinophil infiltration in an antigen-induced mouse asthmatic model after oral administration.
Asunto(s)
Pirimidinas/síntesis química , Pirroles/síntesis química , Factor de Transcripción STAT6/antagonistas & inhibidores , Administración Oral , Animales , Asma/tratamiento farmacológico , Citocromo P-450 CYP3A , Inhibidores del Citocromo P-450 CYP3A , Eosinófilos/efectos de los fármacos , Humanos , Inmunidad , Ratones , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Pirroles/farmacología , Pirroles/uso terapéutico , Relación Estructura-Actividad , Células Th2/efectos de los fármacosRESUMEN
YM-58483/BTP2 is a blocker of store-operated Ca2+ entry (SOCE), which regulates the activation of non-excitable cells such as lymphocytes. YM-58483 has been reported to inhibit cytokine production and proliferation in T cells, and to be useful as a probable medicinal candidate for treatment of bronchial asthma. The present study investigated the pharmacological profile and therapeutic potential of YM-58483 in relation to cell-mediated immune responses. In the mouse graft-versus-host disease (GVHD) model, YM-58483 (1-30 mg/kg, p.o.) and cyclosporine A (1-30 mg/kg, p.o.) inhibited donor anti-host cytotoxic T lymphocyte (CTL) activity and IFN-gamma production, and also reduced the number of donor T cells, especially donor CD8+ T cells, in the spleen. YM-58483 and cyclosporine A inhibited T cell proliferation in a one-way mixed lymphocyte reaction (MLR) with IC50 values of 330 and 12.7 nM, respectively. Additionally, YM-58483 (1-10 mg/kg, p.o.) and cyclosporine A (2, 10 mg/kg, p.o.) inhibited the sheep red blood cell (SRBC)-induced delayed type hypersensitivity (DTH) response. These results suggest that the inhibition of SOCE leads to the prevention of antigen-induced T cell responses, which participate in autoimmune diseases such as autoimmune hepatitis and rheumatoid arthritis.
Asunto(s)
Anilidas/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Enfermedad Injerto contra Huésped/inmunología , Hipersensibilidad Tardía/inmunología , Linfocitos T Citotóxicos/efectos de los fármacos , Células TH1/efectos de los fármacos , Tiadiazoles/farmacología , Animales , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Ciclosporina/farmacología , Enfermedad Injerto contra Huésped/metabolismo , Interferón gamma/biosíntesis , Interferón gamma/efectos de los fármacos , Interferón gamma/inmunología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Linfocitos T Citotóxicos/inmunología , Células TH1/inmunologíaRESUMEN
T helper (Th) 2 cells play a central role in the pathogenesis of allergic diseases such as allergic asthma, atopic dermatitis, and allergic rhinitis. We have found that YM-341619 hydrochloride, which suppressed IL-4-induced STAT6-dependent reporter gene expression, inhibited the differentiation of mouse spleen T cells into Th2 cells in vitro. YM-341619 suppressed the production of IL-4 and the expression of GATA-3 mRNA, a Th2 transcription factor, in T cells cultured with anti-CD3 antibody and anti-CD28 antibody in the presence of IL-4. In contrast, the production of IFN-gamma and the expression of T-bet mRNA, a Th1 transcription factor, in T cells cultured with anti-CD3 antibody in the presence of IL-12, were not effected by YM-341619. Orally administered YM-341619 (0.003-0.03 mg/kg) reduced the plasma IgE level of DNP-Ascaris-sensitized rats, but not the IgG(2a) level. YM-341619 suppressed IL-4 and IL-13 production in the splenocytes of these DNP-Ascaris-sensitized rats without augmenting IFN-gamma production. YM-341619 also dose-dependently suppressed eosinophil accumulation in the lung (0.003-3 mg/kg, p.o.) and airway hyperresponsiveness (0.3-3 mg/kg, p.o.) induced by repeated exposure to ovalbumin in ovalbumin-sensitized rats. These results suggest that YM-341619 has the ability to suppress allergen-induced Th2 responses by selectively inhibiting the differentiation of CD4(+) T cells into the Th2 subset.
Asunto(s)
Asma/tratamiento farmacológico , Hiperreactividad Bronquial/tratamiento farmacológico , Eosinofilia/tratamiento farmacológico , Morfolinas/farmacología , Pirimidinas/farmacología , Bazo/citología , Linfocitos T/citología , Células Th2/citología , Animales , Asma/inmunología , Diferenciación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Femenino , Factor de Transcripción GATA3/genética , Interferón gamma/biosíntesis , Interleucina-4/antagonistas & inhibidores , Interleucina-4/biosíntesis , Masculino , Ratones , Ratones Endogámicos C57BL , Morfolinas/uso terapéutico , Regiones Promotoras Genéticas , Pirimidinas/uso terapéutico , Ratas , Ratas Endogámicas BN , Ratas Wistar , Factor de Transcripción STAT6/genéticaRESUMEN
Signal transducers and activators of transcription 6 (STAT6) is a key regulator of the type 2 helper T (Th2) cell immune response and a potential therapeutic target for allergic diseases such as asthma and atopic diseases. To search for potent and orally bioavailable STAT6 inhibitors, we synthesized a series of 4-benzylaminopyrimidine-5-carboxamide derivatives and evaluated their STAT6 inhibitory activities. Among these compounds, 2-[(4-morpholin-4-ylphenyl)amino]-4-[(2,3,6-trifluorobenzyl)amino]pyrimidine-5-carboxamide (25y, YM-341619, AS1617612) showed potent STAT6 inhibition with an IC(50) of 0.70nM, and also inhibited Th2 differentiation in mouse spleen T cells induced by interleukin (IL)-4 with an IC(50) of 0.28 nM without affecting type 1 helper T (Th1) cell differentiation induced by IL-12. In addition, compound 25y showed an oral bioavailability of 25% in mouse.
Asunto(s)
Morfolinas/administración & dosificación , Morfolinas/farmacología , Pirimidinas/administración & dosificación , Pirimidinas/farmacología , Factor de Transcripción STAT6/antagonistas & inhibidores , Administración Oral , Animales , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Fenómenos Químicos , Química Física , Ratones , Ratones Endogámicos C57BL , Estructura Molecular , Morfolinas/química , Morfolinas/farmacocinética , Pirimidinas/química , Pirimidinas/farmacocinética , Factor de Transcripción STAT6/metabolismo , Relación Estructura-Actividad , Linfocitos T/citología , Linfocitos T/efectos de los fármacos , Linfocitos T/metabolismoRESUMEN
YM-58483/BTP-2, 4-methyl-4'-[3,5-bis(trifluoromethyl)-1H-pyrazol-1-yl]-1,2,3-thiadiazole-5-carboxanilide, blocks the store-operated Ca2+ entry (SOCE) that mediates the activation of non-excitable cells. This study investigated the pharmacological profile and therapeutic potential of YM-58483 as anti-asthma drug. YM-58483 inhibited DNP antigen-induced histamine release from and leukotrienes (LTs) production in IgE-primed RBL-2H3 cells, a rat basophilic leukemia cell line, with IC50 values of 460 and 310 nM, respectively. Prednisolone did not inhibit either of these responses. YM-58483 also inhibited phytohemagglutinin-P (PHA)-stimulated IL-5 and IL-13 production in human peripheral blood cells with IC50 values of 125 and 148 nM, respectively, which is approximately 5 times less potent than prednisolone. YM-58483 (30 mg/kg, p.o.) significantly suppressed ovalbumin (OVA)-induced bronchoconstriction in OVA-sensitized guinea pigs, whereas prednisolone did not. YM-58483 (3-30 mg/kg, p.o.) and prednisolone (100mg/kg, p.o.) both significantly and completely suppressed airway hyperresponsiveness (AHR) caused by OVA exposure. Since YM-58483 inhibits two major characteristic symptoms of bronchial asthma, namely bronchoconstriction and AHR via the suppression of inflammatory mediator and cytokine production, SOCE inhibition is a potential approach for treatment.
Asunto(s)
Anilidas/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Tiadiazoles/farmacología , Anilidas/uso terapéutico , Animales , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Células Sanguíneas/efectos de los fármacos , Células Sanguíneas/metabolismo , Hiperreactividad Bronquial/inmunología , Hiperreactividad Bronquial/prevención & control , Broncoconstricción/efectos de los fármacos , Calcio/metabolismo , Bloqueadores de los Canales de Calcio/uso terapéutico , Línea Celular , Dinitrofenoles/inmunología , Relación Dosis-Respuesta a Droga , Cobayas , Liberación de Histamina/efectos de los fármacos , Humanos , Técnicas In Vitro , Leucotrienos/biosíntesis , Masculino , Contracción Muscular/efectos de los fármacos , Músculo Liso/efectos de los fármacos , Músculo Liso/inmunología , Músculo Liso/fisiopatología , Ovalbúmina/inmunología , Fitohemaglutininas/inmunología , Prednisolona/farmacología , Prednisolona/uso terapéutico , Ratas , Albúmina Sérica Bovina/inmunología , Tiadiazoles/uso terapéutico , Tráquea/efectos de los fármacos , Tráquea/inmunología , Tráquea/fisiopatologíaRESUMEN
T cells play a regulatory role in the pathogenesis of various immune and allergic diseases, including human asthma. Recently, it was reported that a pyrazole derivative, YM-58483 (BTP2), potently inhibits Ca(2+) release-activated Ca(2+) (CRAC) channels and interleukin (IL)-2 production in T cells. We investigated the effects of YM-58483 on T helper type 2 (Th2) cytokine production in vitro and antigen-induced airway asthmatic responses in vivo. YM-58483 inhibited IL-4 and IL-5 production in a conalbumine-stimulated murine Th2 T cell clone (D10.G4.1), and IL-5 production in phytohemagglutinin-stimulated human whole blood cells with IC(50) values comparable to those reported for its CRAC channel inhibition (around 100 nM). YM-58483 inhibited antigen-induced eosinophil infiltration into airways, and decreased IL-4 and cysteinyl-leukotrienes content in inflammatory airways induced in actively sensitized Brown Norway rats. Furthermore, orally administered YM-58483 prevented antigen-induced late phase asthmatic bronchoconstriction and eosinophil infiltration in actively sensitized guinea pigs. These data suggest that the inhibition of Ca(2+) influx through CRAC channel leads to the prevention of antigen-induced airway inflammation, probably via the inhibition of Th2 cytokine production and inflammatory mediators release. YM-58483 may therefore be useful for treating airway inflammation in bronchial asthma.
Asunto(s)
Anilidas/farmacología , Asma/tratamiento farmacológico , Canales de Calcio/efectos de los fármacos , Calcio/metabolismo , Eosinofilia/prevención & control , Interleucina-4/antagonistas & inhibidores , Interleucina-5/antagonistas & inhibidores , Tiadiazoles/farmacología , Animales , Antígenos/inmunología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Cobayas , Humanos , Interleucina-4/biosíntesis , Interleucina-5/biosíntesis , Masculino , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratas , Ratas Endogámicas BNRESUMEN
The STAT6 (signal transducers and activators of transcription 6) protein is activated by interleukin (IL)-4 and IL-13, and plays an important role in T-helper cell 2 (Th2) differentiation. STAT6 might therefore be an excellent therapeutic target for various allergic conditions, including asthma and atopic diseases. We synthesized a series of 2-{[2-(4-hydroxyphenyl)ethyl]amino}pyrimidine-5-carboxamide derivatives and evaluated their STAT6 inhibitory activities. Among these compounds, 4-(benzylamino)-2-{[2-(3-chloro-4-hydroxyphenyl)ethyl]amino}pyrimidine-5-carboxamide (2t, AS1517499) showed potent STAT6 inhibition with an IC(50) value of 21 nM, and also inhibited IL-4-induced Th2 differentiation of mouse spleen T cells with an IC(50) value of 2.3 nM and without influencing T-helper cell 1 (Th1) differentiation induced by IL-12.
Asunto(s)
Etilaminas/síntesis química , Etilaminas/farmacología , Pirimidinas/síntesis química , Pirimidinas/farmacología , Factor de Transcripción STAT6/antagonistas & inhibidores , Animales , Diferenciación Celular/efectos de los fármacos , Línea Celular , Genes Reporteros , Humanos , Indicadores y Reactivos , Interferón gamma/biosíntesis , Interleucina-12/biosíntesis , Interleucina-4/biosíntesis , Isomerismo , Luciferasas/genética , Espectroscopía de Resonancia Magnética , Ratones , Ratones Endogámicos C57BL , Plásmidos , Relación Estructura-Actividad , Linfocitos T/efectos de los fármacos , Células Th2RESUMEN
In nonexcitable cells, Ca(2+) entry is mediated predominantly through the store depletion-dependent Ca(2+) channels called store-operated Ca(2+) (SOC) or Ca(2+) release-activated Ca(2+) channels. YM-58483, a pyrazole derivative, inhibited an anti-CD3 mAb-induced sustained Ca(2+) influx in acute T cell leukemia, Jurkat cells. But it did not affect an anti-CD3 mAb-induced transient intracellular Ca(2+) increase in Ca(2+)-free medium, nor anti-CD3 mAb-induced phosphorylation of phospholipase Cgamma1. It was suggested that YM-58483 inhibited Ca(2+) influx through SOC channels without affecting the TCR signal transduction cascade. Furthermore, YM-58483 inhibited thapsigargin-induced sustained Ca(2+) influx with an IC(50) value of 100 nM without affecting membrane potential. YM-58483 inhibited by 30-fold the Ca(2+) influx through SOC channels compared with voltage-operated Ca(2+) channels, while econazole inhibited both SOC channels and voltage-operated Ca(2+) channels with an equivalent range of IC(50) values. YM-58483 potently inhibited IL-2 production and NF-AT-driven promoter activity, but not AP-1-driven promoter activity in Jurkat cells. Moreover, this compound inhibited delayed-type hypersensitivity in mice with an ED(50) of 1.1 mg/kg. Therefore, we concluded that YM-58483 was a novel store-operated Ca(2+) entry blocker and a potent immunomodulator, and could be useful for the treatment of autoimmune diseases and chronic inflammation. Furthermore, YM-58483 would be a candidate for the study of capacitative Ca(2+) entry mechanisms through SOC/CRAC channels and for identification of putative Ca(2+) channel genes.