RESUMEN
Novel conformationally constrained BET bromodomain inhibitors have been developed. These inhibitors were optimized in two similar, yet distinct chemical series, the 6-methyl-1H-pyrrolo[2,3-c]pyridin-7(6H)-ones (A) and the 1-methyl-1H-pyrrolo[2,3-c]pyridin-7(6H)-ones (B). Each series demonstrated excellent activity in binding and cellular assays, and lead compounds from each series demonstrated significant efficacy in in vivo tumor xenograft models.
Asunto(s)
Proteínas Nucleares/antagonistas & inhibidores , Piridonas/química , Factores de Transcripción/antagonistas & inhibidores , Animales , Sitios de Unión , Proteínas de Ciclo Celular , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Evaluación Preclínica de Medicamentos , Semivida , Humanos , Ratones , Microsomas/metabolismo , Simulación de Dinámica Molecular , Mieloma Múltiple/tratamiento farmacológico , Proteínas Nucleares/metabolismo , Estructura Terciaria de Proteína , Piridonas/farmacocinética , Piridonas/farmacología , Piridonas/uso terapéutico , Relación Estructura-Actividad , Factores de Transcripción/metabolismo , Trasplante HeterólogoRESUMEN
Members of the BET family of bromodomain containing proteins have been identified as potential targets for blocking proliferation in a variety of cancer cell lines. A two-dimensional NMR fragment screen for binders to the bromodomains of BRD4 identified a phenylpyridazinone fragment with a weak binding affinity (1, Ki = 160 µM). SAR investigation of fragment 1, aided by X-ray structure-based design, enabled the synthesis of potent pyridone and macrocyclic pyridone inhibitors exhibiting single digit nanomolar potency in both biochemical and cell based assays. Advanced analogs in these series exhibited high oral exposures in rodent PK studies and demonstrated significant tumor growth inhibition efficacy in mouse flank xenograft models.
Asunto(s)
Compuestos Macrocíclicos/química , Compuestos Macrocíclicos/farmacología , Piridonas/química , Piridonas/farmacología , Animales , Cristalografía por Rayos X , Descubrimiento de Drogas , Compuestos Macrocíclicos/farmacocinética , Estructura Molecular , Piridonas/farmacocinética , Ratas , Relación Estructura-ActividadRESUMEN
Histone deacetylases (HDACs) are a family of enzymes involved in transcription regulation. HDACs are known to play key roles in the regulation of cell proliferation; consequently, inhibition of HDACs has become an interesting approach for anti-cancer therapy. However, expression of mammalian HDACs has proven to be difficult. All attempts to express these HDACs in E.coli, Pichia and baculovirus systems were unsuccessful. Here we present the stable expression of human recombinant His-tagged HDAC1 and HDAC3 in mammalian cells. Full-length human genes for HDAC1 and HDAC3 were cloned into the pcDNA 3.1 vector containing a N-terminal His-tag with an enterokinase cleavage site. Recombinant HDAC enzyme activity was only detected after nickel affinity purification due to high activity of endogenous HDACs; and removal of the His-tag increased activity 2-4 fold. Western blots demonstrated the nickel affinity purified rhHDAC1 preparation also contained endogenous HDAC2 and HDAC3; likewise, rhHDAC3 preparation contained endogenous HDAC1 and HDAC2. Therefore, the active HDAC preparation is actually a multi-protein and a multi-HDAC containing complex. This provides one explanation for the similar IC50 values exhibited by SAHA and MS-275 against nuclear HDACs and rhHDAC1 and 3 preparations. These results demonstrate that recombinant forms of the HDACs can be over-expressed in mammalian cells, isolated as active multi-protein complexes that contain multiple HDAC enzymes, and caution must be used when determining HDAC inhibitor in vitro selectivity.
Asunto(s)
Regulación Enzimológica de la Expresión Génica , Histona Desacetilasas/biosíntesis , Complejos Multienzimáticos/biosíntesis , Proteínas Recombinantes de Fusión/biosíntesis , Western Blotting , División Celular , Clonación Molecular , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Células HeLa , Histona Desacetilasa 1 , Histona Desacetilasas/genética , Histona Desacetilasas/farmacología , Humanos , Complejos Multienzimáticos/genética , Complejos Multienzimáticos/farmacología , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/farmacología , Especificidad por Sustrato , TransfecciónRESUMEN
Alpha-keto ester and amides were found to be potent inhibitors of histone deacetylase. Nanomolar inhibitors against the isolated enzyme and sub-micromolar inhibitors of cellular proliferation were obtained. The alpha-keto amide 30 also exhibited significant anti-tumor effects in an in vivo tumor model.
Asunto(s)
Amidas/química , Amidas/farmacología , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Inhibidores de Histona Desacetilasas , Animales , Línea Celular Tumoral , Histona Desacetilasas/metabolismo , Humanos , Ratones , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
A novel series of sulfone N-formylhydroxylamines (retrohydroxamates) have been investigated as matrix metalloproteinases (MMP) inhibitors. The substitution of the ether linkage of ABT-770 (5) with a sulfone group 13a led to a substantial increase in activity against MMP-9 but was accompanied by a loss of selectivity for inhibition of MMP-2 and -9 over MMP-1 and diminished oral exposure. Replacement of the biphenyl P1' substituent with a phenoxyphenyl group provided compounds that are highly selective for inhibition of MMP-2 and -9 over MMP-1. Optimization of the substituent adjacent to the retrohydroxamate center in this series led to the clinical candidate ABT-518 (6), a highly potent, selective, orally bioavailable MMP inhibitor that has been shown to significantly inhibit tumor growth in animal cancer models.
