RESUMEN
OBJECTIVE: Bruton's tyrosine kinase (BTK) plays a critical role in B cell development and function. We recently described a selective BTK inhibitor, RN486, that blocks B cell receptor (BCR) and Fcγ receptor signaling and is efficacious in animal models of arthritis. The aim of this study was to examine the potential efficacy of BTK in systemic lupus erythematosus (SLE), using an NZB × NZW mouse model of spontaneous SLE. METHODS: Mice received RN486 or its vehicle (administered in chow) at a final concentration of 30 mg/kg for 8 weeks, starting at 32 weeks of age. RESULTS: The administration of RN486 completely stopped disease progression, as determined by histologic and functional analyses of glomerular nephritis. The efficacy was associated with striking inhibition of B cell activation, as demonstrated by a significant reduction in CD69 expression in response to BCR crosslinking. RN486 markedly reduced the secretion of IgG anti-double-stranded DNA (anti-dsDNA) secretion, as determined by enzyme-linked immunosorbent and enzyme-linked immunospot assays. Flow cytometric analysis demonstrated depletion of CD138(high) B220(low) plasma cells in the spleen. RN486 inhibited secretion of IgG anti-dsDNA but not IgM anti-dsDNA, suggesting that pharmacologic blockade of BTK resembles the reported transgenic expression of low levels of endogenous BTK in B cells. In addition, RN486 may also impact the effector function of autoantibodies, as evidenced by a significant reduction in immune complex-mediated activation of human monocytes in vitro and down-regulation of the expression of macrophage-related and interferon-inducible genes in both the kidneys and spleens of treated mice. CONCLUSION: Collectively, our data suggest that BTK inhibitors may simultaneously target autoantibody-producing and effector cells in SLE, thus constituting a promising therapeutic alternative for this disease.
Asunto(s)
Linfocitos B/patología , Glomerulonefritis/tratamiento farmacológico , Glomérulos Renales/patología , Lupus Eritematoso Sistémico/tratamiento farmacológico , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Agammaglobulinemia Tirosina Quinasa , Animales , Complejo Antígeno-Anticuerpo/metabolismo , Antígenos CD/metabolismo , Antígenos de Diferenciación de Linfocitos T/metabolismo , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Regulación hacia Abajo , Glomerulonefritis/metabolismo , Glomerulonefritis/patología , Glomérulos Renales/efectos de los fármacos , Glomérulos Renales/metabolismo , Lectinas Tipo C/metabolismo , Lupus Eritematoso Sistémico/metabolismo , Lupus Eritematoso Sistémico/patología , Activación de Linfocitos/efectos de los fármacos , Ratones , Ratones Endogámicos NZB , Receptores de IgG/metabolismoRESUMEN
In a discovery effort to find safe and effective DGAT-1 inhibitors, we have identified 2-phenyloxazole 4-carboxamide 1 as a conformationally constrained analog of a hydrazide hit, which was previously identified from high-throughput screening. Further optimization of this series has led to chemically more stable 2-phenyloxazole-based DGAT-1 inhibitor 25 with improved solubility, cell-based activity, and pharmacokinetic properties. Compound 25 also demonstrated in vivo efficacy in a diet-induced obesity (DIO) rat model.
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Diacilglicerol O-Acetiltransferasa/antagonistas & inhibidores , Descubrimiento de Drogas , Inhibidores Enzimáticos , Oxazoles/química , Oxazoles/farmacología , Administración Oral , Animales , Peso Corporal , Diacilglicerol O-Acetiltransferasa/química , Modelos Animales de Enfermedad , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Humanos , Concentración 50 Inhibidora , Ratones , Estructura Molecular , Obesidad/tratamiento farmacológico , Oxazoles/uso terapéutico , Ratas , Solubilidad , Relación Estructura-ActividadRESUMEN
INTRODUCTION: Spleen tyrosine kinase (SYK) is a key integrator of intracellular signals triggered by activated immunoreceptors, including Bcell receptors (BCR) and Fc receptors, which are important for the development and function of lymphoid cells. Given the clinical efficacy of Bcell depletion in the treatment of rheumatoid arthritis and multiple sclerosis, pharmacological modulation of Bcells using orally active small molecules that selectively target SYK presents an attractive alternative therapeutic strategy. METHODS: A SYK inhibitor was developed and assayed in various in vitro systems and in the mouse model of collagen-induced arthritis (mCIA). RESULTS: A novel ATP-competitive inhibitor of SYK, 6-[(1R,2S)-2-Amino-cyclohexylamino]-4-(5,6-dimethyl-pyridin-2-ylamino)-pyridazine-3-carboxylic acid amide, designated RO9021, with an adequate kinase selectivity profile and oral bioavailability, was developed. In addition to suppression of BCR signaling in human peripheral blood mononuclear cells (PBMC) and whole blood, FcγR signaling in human monocytes, and FcϵR signaling in human mast cells, RO9021 blocked osteoclastogenesis from mouse bone marrow macrophages in vitro. Interestingly, Toll-like Receptor (TLR) 9 signaling in human Bcells was inhibited by RO9021, resulting in decreased levels of plasmablasts, immunoglobulin (Ig) M and IgG upon B-cell differentiation. RO9021 also potently inhibited type I interferon production by human plasmacytoid dendritic cells (pDC) upon TLR9 activation. This effect is specific to TLR9 as RO9021 did not inhibit TLR4- or JAK-STAT-mediated signaling. Finally, oral administration of RO9021 inhibited arthritis progression in the mCIA model, with observable pharmacokinetics (PK)-pharmacodynamic (PD) correlation. CONCLUSIONS: Inhibition of SYK kinase activity impinges on various innate and adaptive immune responses. RO9021 could serve as a starting point for the development of selective SYK inhibitors for the treatment of inflammation-related and autoimmune-related disorders.
Asunto(s)
Inmunidad Adaptativa/efectos de los fármacos , Inmunidad Innata/efectos de los fármacos , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/farmacología , Administración Oral , Aminopiridinas/química , Aminopiridinas/farmacología , Animales , Artritis Experimental/metabolismo , Artritis Experimental/prevención & control , Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/metabolismo , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Linfocitos B/metabolismo , Línea Celular Tumoral , Células Cultivadas , Citometría de Flujo , Humanos , Péptidos y Proteínas de Señalización Intracelular/química , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Células Jurkat , Macrófagos/citología , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Modelos Moleculares , Estructura Molecular , Osteoclastos/citología , Osteoclastos/efectos de los fármacos , Osteoclastos/metabolismo , Unión Proteica , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/química , Estructura Terciaria de Proteína , Proteínas Tirosina Quinasas/química , Proteínas Tirosina Quinasas/metabolismo , Piridazinas/química , Piridazinas/farmacología , Transducción de Señal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/administración & dosificación , Bibliotecas de Moléculas Pequeñas/química , Quinasa SykRESUMEN
3-[4-((1S,2S,3R,5S,7S)-5-Hydroxyadamantan-2-ylcarbamoyl)benzyl]-4-oxo-1-phenyl-1,4-dihydro-[1,8]naphthyridine-2-carboxylic acid methyl ester (4) was identified as a novel, druglike and selective quinolone pan JNK inhibitor. In this communication, some of the structure-activity relationship of the azaquinolone analogues leading to 4 is discussed. The focus is on how changes at the amide functionality affected the biochemical potency, cellular potency, metabolic properties, and solubility of this class of JNK inhibitors. Optimization of these properties led to the identification of the adamantyl analogue, 4. 4 achieved proof of mechanism in both rat and mouse TNF-α challenge models.
RESUMEN
Diacylglycerol acyltransferase-1 (DGAT-1) is the enzyme that catalyzes the final and committed step of triglyceride formation, namely, the acylation of diacylglycerol with acyl coenzyme A. DGAT-1 deficient mice demonstrate resistance to weight gain on high fat diet, improved insulin sensitivity, and reduced liver triglyceride content. Inhibition of DGAT-1 thus represents a potential novel approach for the treatment of obesity, dyslipidemia, and metabolic syndrome. In this communication, we report the identification of the lead structure 6 and our lead optimization efforts culminating in the discovery of potent, selective, and orally efficacious carboxylic acid derivatives of 2-phenyl-5-trifluoromethyloxazole-4-carboxamides. In particular, compound 29 (DGAT-1 enzyme assay, IC(50) = 57 nM; CHO-K1 cell triglyceride formation assay, EC(50) = 0.5 µM) demonstrated dose dependent inhibition of weight gain in diet induced obese (DIO) rats (0.3, 1, and 3 mg/kg, p.o., qd) during a 21-day efficacy study. Furthermore, compound 29 demonstrated improved glucose tolerance determined by an oral glucose tolerance test (OGTT).