RESUMEN
SLE is a complex autoimmune inflammatory disease characterized by pathogenic autoantibody production as a consequence of uncontrolled T-B cell activity and immune-complex deposition in various organs, including kidney, leading to tissue damage and function loss. There is a high unmet need for better treatment options other than corticosteroids and immunosuppressants. Phosphoinositol-3 kinase δ (PI3Kδ) is a promising target in this respect as it is essential in mediating B- and T-cell function in mouse and human. We report the identification of selective PI3Kδ inhibitors that blocked B-, T-, and plasmacytoid dendritic cell activities in human peripheral blood and in primary cell co-cultures (BioMAP(®)) without detecting signs of undesired toxicity. In an IFNα-accelerated mouse SLE model, our PI3Kδ inhibitors blocked nephritis development, whether administered at the onset of autoantibody appearance or the onset of proteinuria. Disease amelioration correlated with normalized immune cell numbers in the spleen, reduced immune-complex deposition as well as reduced inflammation, fibrosis, and tissue damage in the kidney. Improvements were similar to those achieved with a frequently prescribed drug for lupus nephritis, the potent immunosuppressant mycophenolate mofetil. Finally, we established a pharmacodynamics/pharmacokinetic/efficacy model that revealed that a sustained PI3Kδ inhibition of 50% is sufficient to achieve full efficacy in our disease model. These data demonstrate the therapeutic potential of PI3Kδ inhibitors in SLE and lupus nephritis.
RESUMEN
This chapter describes assays that focus on the characterization of compounds identified in high--throughput screening campaigns, and the subsequent medicinal chemistry programs. They cover methods to determine potency in buffer, the effect of whole blood on the compounds' activity and finally the pharmacokinetic (PK)/pharmacodynamic (PD) -relationship of the compounds in a rodent species.
Asunto(s)
Descubrimiento de Drogas/métodos , Ensayos Analíticos de Alto Rendimiento , Receptores de Quimiocina/antagonistas & inhibidores , Animales , Automatización de Laboratorios , Técnicas de Cultivo de Célula , Ensayos de Migración Celular , Células Cultivadas , Quimiocinas/metabolismo , Quimiotaxis/efectos de los fármacos , Espectroscopía Dieléctrica , Relación Dosis-Respuesta a Droga , Ensayo de Inmunoadsorción Enzimática , Humanos , Ligandos , Farmacocinética , Unión Proteica , Receptores de Quimiocina/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
New phenoxyacetic acid antagonists of CRTH2 are described. Following the discovery of a hit compound by a focused screening, high protein binding was identified as its main weakness. Optimization aimed at reducing serum protein binding led to the identification of several compounds that showed not only excellent affinities for the receptor (41 compounds with K(i) < 10 nM) but also excellent potencies in a human whole blood assay (IC(50) < 100 nM; PGD2-induced eosinophil shape change). Additional optimization of the PK characteristics led to the identification of several compounds suitable for in vivo testing. Of these, 19k and 19s were tested in two different pharmacological models (acute FITC-mediated contact hypersensitivity and ovalbumin-induced eosinophilia models) and found to be active after oral dosing (10 and 30 mg/kg).
Asunto(s)
Acetatos/síntesis química , Alquinos/síntesis química , Antialérgicos/síntesis química , Antiinflamatorios/síntesis química , Receptores Inmunológicos/antagonistas & inhibidores , Receptores de Prostaglandina/antagonistas & inhibidores , Sulfonas/síntesis química , Acetatos/farmacocinética , Acetatos/farmacología , Administración Oral , Alquinos/farmacocinética , Alquinos/farmacología , Animales , Antialérgicos/farmacocinética , Antialérgicos/farmacología , Antiinflamatorios/farmacocinética , Antiinflamatorios/farmacología , Unión Competitiva , Proteínas Sanguíneas/metabolismo , Células CACO-2 , Permeabilidad de la Membrana Celular , Forma de la Célula , Quimiotaxis de Leucocito , Dermatitis por Contacto/tratamiento farmacológico , Eosinófilos/efectos de los fármacos , Eosinófilos/patología , Eosinófilos/fisiología , Femenino , Células HEK293 , Humanos , Ratones , Ratones Endogámicos BALB C , Microsomas Hepáticos/metabolismo , Ovalbúmina/inmunología , Fenoxiacetatos , Unión Proteica , Eosinofilia Pulmonar/tratamiento farmacológico , Eosinofilia Pulmonar/inmunología , Ensayo de Unión Radioligante , Ratas , Relación Estructura-Actividad , Sulfonas/farmacocinética , Sulfonas/farmacologíaRESUMEN
Antagonism of the CRTH2 receptor represents a very attractive target for a variety of allergic diseases. Most CRTH2 antagonists known to date possess a carboxylic acid moiety, which is essential for binding. However, potential acid metabolites O-acyl glucuronides might be linked to idiosynchratic toxicity in humans. In this communication, we describe a new series of compounds that lack the carboxylic acid moiety. Compounds with high affinity (K i < 10 nM) for the receptor have been identified. Subsequent optimization succeeded in reducing the high metabolic clearance of the first compounds in human and rat liver microsomes. At the same time, inhibition of the CYP isoforms was optimized, giving rise to stable compounds with an acceptable CYP inhibition profile (IC50 CYP2C9 and 2C19 > 1 µM). Taken together, these data show that compounds devoid of carboxylic acid groups could represent an interesting alternative to current CRTH2 antagonists in development.
RESUMEN
New spiroindolinone antagonists of CRTH2 are described. Following identification of insufficient stability in human plasma as an important liability of the lead compounds, replacement of the spirosuccinimide core with a spirohydantoin or spiropyrrolidinone structure has yielded a compound that is fully stable in human plasma and with good potency in a human whole blood assay (IC50 = 69 nM) but shows a much lower oral bioavailability (6-9% in rodents) than the earlier compounds. Successive optimization aimed at restoring an acceptable oral bioavailability has yielded compound (S)-17a, which exhibits both stability in human plasma and a good oral bioavailability in rat (37%) and mouse (39%). This compound is also active in a mouse model of ovalbumin-induced lung inflammation following oral dosing at 30 mg/kg.
RESUMEN
A novel chemical class of potent chemoattractant receptor-homologous expressed on Th2 lymphocytes (CRTH2 or DP2) antagonists is reported. An initial and moderately potent spiro-indolinone compound ( 5) was found during a high-throughput screening campaign. Structure-activity relationship (SAR) investigation around the carboxylic acid group revealed that changes in this part of the molecule could lead to a reversal of functional activity, yielding weakly potent agonists. SAR investigation of the succinimide functional group led to the discovery of several single-digit nanomolar antagonists. The potency of these compounds was confirmed in a human eosinophil chemotaxis assay. Moreover, compounds ( R)- 58 and ( R)- 71 were shown to possess pharmacokinetic properties suitable for development as an orally bioavailable drug.