RESUMEN
CUO246, a novel DNA gyrase/topoisomerase IV inhibitor, is active in vitro against a broad range of Gram-positive, fastidious Gram-negative, and atypical bacterial pathogens and retains activity against quinolone-resistant strains in circulation. The frequency of selection for single step mutants of wild-type S. aureus with reduced susceptibility to CUO246 was <4.64 × 10-9 at 4× and 8× MIC and remained low when using an isogenic QRDR mutant (<5.24 × 10-9 at 4× and 8× MIC). Biochemical assays indicated that CUO246 had potent inhibitory activity against both DNA gyrase (GyrAB) and topoisomerase IV (ParCE). Furthermore, CUO246 showed rapid bactericidal activity in time-kill assays and potent in vivo efficacy against S. aureus in a neutropenic murine thigh infection model. These results suggest that CUO246 may be useful in treating infections by various causative agents of acute skin and skin structure infections, respiratory tract infections, and sexually transmitted infections.
Asunto(s)
Girasa de ADN , Topoisomerasa de ADN IV , Animales , Ratones , Girasa de ADN/genética , Topoisomerasa de ADN IV/genética , Inhibidores de Topoisomerasa II/farmacología , ADN Bacteriano , Staphylococcus aureus , Pruebas de Sensibilidad Microbiana , Antibacterianos/farmacología , Antibacterianos/uso terapéuticoRESUMEN
Herein, we describe the discovery and optimization of a novel series that inhibits bacterial DNA gyrase and topoisomerase IV via binding to, and stabilization of, DNA cleavage complexes. Optimization of this series led to the identification of compound 25, which has potent activity against Gram-positive bacteria, a favorable in vitro safety profile, and excellent in vivo pharmacokinetic properties. Compound 25 was found to be efficacious against fluoroquinolone-sensitive Staphylococcus aureus infection in a mouse thigh model at lower doses than moxifloxacin. An X-ray crystal structure of the ternary complex formed by topoisomerase IV from Klebsiella pneumoniae, compound 25, and cleaved DNA indicates that this compound does not engage in a water-metal ion bridge interaction and forms no direct contacts with residues in the quinolone resistance determining region (QRDR). This suggests a structural basis for the reduced impact of QRDR mutations on antibacterial activity of 25 compared to fluoroquinolones.
Asunto(s)
Antibacterianos/farmacología , Girasa de ADN/metabolismo , Topoisomerasa de ADN IV/antagonistas & inhibidores , Diseño de Fármacos , Fluoroquinolonas/farmacología , Staphylococcus aureus/efectos de los fármacos , Inhibidores de Topoisomerasa II/farmacología , Animales , Antibacterianos/química , Farmacorresistencia Bacteriana/efectos de los fármacos , Ratones , Inhibidores de Topoisomerasa II/químicaRESUMEN
Since their discovery over 5 decades ago, quinolone antibiotics have found enormous success as broad spectrum agents that exert their activity through dual inhibition of bacterial DNA gyrase and topoisomerase IV. Increasing rates of resistance, driven largely by target-based mutations in the GyrA/ParC quinolone resistance determining region, have eroded the utility and threaten the future use of this vital class of antibiotics. Herein we describe the discovery and optimization of a series of 4-(aminomethyl)quinolin-2(1H)-ones, exemplified by 34, that inhibit bacterial DNA gyrase and topoisomerase IV and display potent activity against ciprofloxacin-resistant Gram-negative pathogens. X-ray crystallography reveals that 34 occupies the classical quinolone binding site in the topoisomerase IV-DNA cleavage complex but does not form significant contacts with residues in the quinolone resistance determining region.
Asunto(s)
Antibacterianos/farmacología , Farmacorresistencia Bacteriana/efectos de los fármacos , Fluoroquinolonas/farmacología , Bacterias Gramnegativas/efectos de los fármacos , Inhibidores de Topoisomerasa II/farmacología , Antibacterianos/síntesis química , Antibacterianos/metabolismo , Antibacterianos/toxicidad , Sitios de Unión , Línea Celular Tumoral , Girasa de ADN/metabolismo , Topoisomerasa de ADN IV/antagonistas & inhibidores , Topoisomerasa de ADN IV/química , Fluoroquinolonas/síntesis química , Fluoroquinolonas/metabolismo , Fluoroquinolonas/toxicidad , Bacterias Gramnegativas/enzimología , Humanos , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Relación Estructura-Actividad , Inhibidores de Topoisomerasa II/síntesis química , Inhibidores de Topoisomerasa II/metabolismo , Inhibidores de Topoisomerasa II/toxicidadRESUMEN
The lipopolysaccharide biosynthesis pathway is considered an attractive drug target against the rising threat of multi-drug-resistant Gram-negative bacteria. Here, we report two novel small-molecule inhibitors (compounds 1 and 2) of the acyltransferase LpxA, the first enzyme in the lipopolysaccharide biosynthesis pathway. We show genetically that the antibacterial activities of the compounds against efflux-deficient Escherichia coli are mediated by LpxA inhibition. Consistently, the compounds inhibited the LpxA enzymatic reaction in vitro. Intriguingly, using biochemical, biophysical, and structural characterization, we reveal two distinct mechanisms of LpxA inhibition; compound 1 is a substrate-competitive inhibitor targeting apo LpxA, and compound 2 is an uncompetitive inhibitor targeting the LpxA/product complex. Compound 2 exhibited more favorable biological and physicochemical properties than compound 1 and was optimized using structural information to achieve improved antibacterial activity against wild-type E. coli. These results show that LpxA is a promising antibacterial target and imply the advantages of targeting enzyme/product complexes in drug discovery.
Asunto(s)
Aciltransferasas/antagonistas & inhibidores , Antibacterianos/farmacología , Inhibidores Enzimáticos/farmacología , Imidazoles/farmacología , Pirazoles/farmacología , Aciltransferasas/metabolismo , Antibacterianos/metabolismo , Cristalografía por Rayos X , Inhibidores Enzimáticos/metabolismo , Escherichia coli/efectos de los fármacos , Escherichia coli/enzimología , Imidazoles/metabolismo , Pruebas de Sensibilidad Microbiana , Unión Proteica , Pirazoles/metabolismoRESUMEN
RAS oncogenes have been implicated in >30% of human cancers, all representing high unmet medical need. The exquisite dependency on CRAF kinase in KRAS mutant tumors has been established in genetically engineered mouse models and human tumor cells. To date, many small molecule approaches are under investigation to target CRAF, yet kinase-selective and cellular potent inhibitors remain challenging to identify. Herein, we describe 14 (RAF709) [ Aversa , Biaryl amide compounds as kinase inhibitors and their preparation . WO 2014151616, 2014 ], a selective B/C RAF inhibitor, which was developed through a hypothesis-driven approach focusing on drug-like properties. A key challenge encountered in the medicinal chemistry campaign was maintaining a balance between good solubility and potent cellular activity (suppression of pMEK and proliferation) in KRAS mutant tumor cell lines. We investigated the small molecule crystal structure of lead molecule 7 and hypothesized that disruption of the crystal packing would improve solubility, which led to a change from N-methylpyridone to a tetrahydropyranyl oxy-pyridine derivative. 14 proved to be soluble, kinase selective, and efficacious in a KRAS mutant xenograft model.
Asunto(s)
2,2'-Dipiridil/análogos & derivados , Antineoplásicos/farmacología , Benzamidas/farmacología , Quinasas raf/antagonistas & inhibidores , Proteínas ras/genética , 2,2'-Dipiridil/química , 2,2'-Dipiridil/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/farmacocinética , Benzamidas/química , Cristalografía por Rayos X , Perros , Diseño de Fármacos , Descubrimiento de Drogas , Estabilidad de Medicamentos , Humanos , Concentración 50 Inhibidora , Ratones , Terapia Molecular Dirigida , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Proteínas Proto-Oncogénicas B-raf/química , Proteínas Proto-Oncogénicas p21(ras)/genética , Ratas , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
ATR, a protein kinase in the PIKK family, plays a critical role in the cell DNA-damage response and is an attractive anticancer drug target. Several potent and selective inhibitors of ATR have been reported showing significant antitumor efficacy, with most advanced ones entering clinical trials. However, due to the absence of an experimental ATR structure, the determinants contributing to ATR inhibitors' potency and specificity are not well understood. Here we present the mutations in the ATP-binding site of PI3Kα to progressively transform the pocket to mimic that of ATR. The generated PI3Kα mutants exhibit significantly improved affinity for selective ATR inhibitors in multiple chemical classes. Furthermore, we obtained the X-ray structures of the PI3Kα mutants in complex with the ATR inhibitors. The crystal structures together with the analysis on the inhibitor affinity profile elucidate the roles of individual amino acid residues in the binding of ATR inhibitors, offering key insights for the binding mechanism and revealing the structure features important for the specificity of ATR inhibitors. The ability to obtain structural and binding data for these PI3Kα mutants, together with their ATR-like inhibitor binding profiles, makes these chimeric PI3Kα proteins valuable model systems for structure-based inhibitor design.
Asunto(s)
Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas de la Ataxia Telangiectasia Mutada/antagonistas & inhibidores , Sitios de Unión , Fosfatidilinositol 3-Quinasa Clase I , Cristalografía por Rayos X , Modelos Moleculares , Proteínas Mutantes/química , Fosfatidilinositol 3-Quinasas/química , Unión Proteica , Conformación ProteicaRESUMEN
RAS mutations lead to a constitutively active oncogenic protein that signals through multiple effector pathways. In this chemical biology study, we describe a novel coupled biochemical assay that measures activation of the effector BRAF by prenylated KRASG12V in a lipid-dependent manner. Using this assay, we discovered compounds that block biochemical and cellular functions of KRASG12V with low single-digit micromolar potency. We characterized the structural basis for inhibition using NMR methods and showed that the compounds stabilized the inactive conformation of KRASG12V. Determination of the biophysical affinity of binding using biolayer interferometry demonstrated that the potency of inhibition matches the affinity of binding only when KRAS is in its native state, namely post-translationally modified and in a lipid environment. The assays we describe here provide a first-time alignment across biochemical, biophysical, and cellular KRAS assays through incorporation of key physiological factors regulating RAS biology, namely a negatively charged lipid environment and prenylation, into the in vitro assays. These assays and the ligands we discovered are valuable tools for further study of KRAS inhibition and drug discovery.
Asunto(s)
Lípidos/química , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Animales , Línea Celular , Línea Celular Tumoral , Humanos , Espectroscopía de Resonancia Magnética , PrenilaciónRESUMEN
The discovery of inhibitors targeting novel allosteric kinase sites is very challenging. Such compounds, however, once identified could offer exquisite levels of selectivity across the kinome. Herein we report our structure-based optimization strategy of a dibenzodiazepine hit 1, discovered in a fragment-based screen, yielding highly potent and selective inhibitors of PAK1 such as 2 and 3. Compound 2 was cocrystallized with PAK1 to confirm binding to an allosteric site and to reveal novel key interactions. Compound 3 modulated PAK1 at the cellular level and due to its selectivity enabled valuable research to interrogate biological functions of the PAK1 kinase.
RESUMEN
Two scaffolds based on 5,6-fused heterocyclic backbones were designed and synthesized as Raf kinase inhibitors. The scaffolds were assessed for in vitro pan-Raf inhibition, activity in cell proliferation and target modulation assays, and pharmacokinetic parameters.
Asunto(s)
Amidas/síntesis química , Diseño de Fármacos , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Compuestos Heterocíclicos/síntesis química , Quinasas raf/antagonistas & inhibidores , Amidas/química , Amidas/farmacología , Sitios de Unión , Proliferación Celular , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/química , Compuestos Heterocíclicos/química , Compuestos Heterocíclicos/farmacología , Modelos Moleculares , Estructura Molecular , Relación Estructura-ActividadRESUMEN
A series of arylaminobenzimidazoles was designed and synthesized as Raf kinase inhibitors. Exploration of the structure-activity relationship resulted in compounds that are potent in vitro and show desirable in vivo properties.
Asunto(s)
Bencimidazoles/síntesis química , Bencimidazoles/farmacología , Diseño de Fármacos , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Quinasas raf/antagonistas & inhibidores , Administración Oral , Animales , Bencimidazoles/administración & dosificación , Bencimidazoles/química , Disponibilidad Biológica , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Ratones , Modelos Moleculares , Estructura Molecular , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/química , Estructura Terciaria de Proteína , Estereoisomerismo , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de Xenoinjerto , Quinasas raf/metabolismoRESUMEN
PURPOSE: Chk1 kinase is a critical regulator of both S and G(2)-M phase cell cycle checkpoints in response to DNA damage. This study aimed to evaluate the biochemical, cellular, and antitumor effects of a novel Chk1 inhibitor, CHIR124. EXPERIMENTAL DESIGN: CHIR-124 was evaluated for its ability to abrogate cell cycle checkpoints, to potentiate cytotoxicity, and to inhibit Chk1-mediated signaling induced by topoisomerase I poisons in human tumor cell line and xenograft models. RESULTS: CHIR-124 is a quinolone-based small molecule that is structurally unrelated to other known inhibitors of Chk1. It potently and selectively inhibits Chk1 in vitro (IC(50) = 0.0003 micromol/L). CHIR-124 interacts synergistically with topoisomerase poisons (e.g., camptothecin or SN-38) in causing growth inhibition in several p53-mutant solid tumor cell lines as determined by isobologram or response surface analysis. CHIR-124 abrogates the SN-38-induced S and G(2)-M checkpoints and potentiates apoptosis in MDA-MD-435 breast cancer cells. The abrogation of the G(2)-M checkpoint and induction of apoptosis by CHIR-124 are enhanced by the loss of p53. We have also shown that CHIR-124 treatment can restore the level of cdc25A protein, which is normally targeted by Chk1 for degradation following DNA damage, indicating that Chk1 signaling is suppressed in the presence of CHIR-124. Finally, in an orthotopic breast cancer xenograft model, CHIR-124 potentiates the growth inhibitory effects of irinotecan by abrogating the G(2)-M checkpoint and increasing tumor apoptosis. CONCLUSIONS: CHIR-124 is a novel and potent Chk1 inhibitor with promising antitumor activities when used in combination with topoisomerase I poisons.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Sinergismo Farmacológico , Proteínas Quinasas/metabolismo , Quinolinas/administración & dosificación , Quinuclidinas/administración & dosificación , Inhibidores de Topoisomerasa I , Animales , Antineoplásicos/farmacología , Apoptosis , Línea Celular Tumoral , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , Inhibidores Enzimáticos/farmacología , Humanos , Concentración 50 Inhibidora , Ratones , Ratones SCID , Modelos Químicos , Trasplante de Neoplasias , Distribución AleatoriaRESUMEN
A series of 2-pyrimidyl-5-amidothiophenes has been synthesized and evaluated for AKT inhibition. SAR studies resulted in potent inhibitors of AKT with IC(50) values as low as single digit nanomolar as represented by compound 2aa. Compound 2aa showed cellular activity including antiproliferation and downstream target modulation. Selectivity profile is described. A co-crystal of 2aa with PKA is determined and discussed.
Asunto(s)
Química Farmacéutica/métodos , Inhibidores Enzimáticos/farmacología , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Tiofenos/química , Tiofenos/síntesis química , Línea Celular Tumoral , Proliferación Celular , Cristalización , Cristalografía por Rayos X , Humanos , Concentración 50 Inhibidora , Modelos Químicos , Modelos Moleculares , Unión Proteica , Relación Estructura-ActividadRESUMEN
The 3-benzimidazol-2-yl-1H-indazole scaffold was developed as an alternate scaffold for our receptor tyrosine kinase (RTK) inhibitor program. In exploring the SAR of this series, it was discovered that a subset of these compounds potently inhibit the enzyme c-ABL. The SAR of these compounds is described.
Asunto(s)
Proliferación Celular/efectos de los fármacos , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Indazoles/síntesis química , Indazoles/farmacología , Proteínas Proto-Oncogénicas c-abl/antagonistas & inhibidores , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Bencimidazoles/síntesis química , Bencimidazoles/farmacología , Células Cultivadas , Humanos , Relación Estructura-ActividadRESUMEN
3-Benzimidazol-2-yl-1H-indazole analogs were developed as inhibitors of receptor tyrosine kinases (RTK). The synthesis and SAR of this series is reported.
Asunto(s)
Bencimidazoles/síntesis química , Bencimidazoles/farmacología , Indazoles/síntesis química , Indazoles/farmacología , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Animales , Bencimidazoles/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Relación Dosis-Respuesta a Droga , Diseño de Fármacos , Indazoles/química , Ratones , Ratones Desnudos , Modelos Moleculares , Estructura Molecular , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
CHK-1 is one of the key enzymes regulating checkpoints in cellular growth cycles. Novel 4-(amino-alkylamino)-3-benzimidazole-quinolinones were prepared and assayed for their ability to inhibit CHK-1. These compounds are potent cell permeable CHK-1 inhibitors and showed synergistic effect with a DNA-damaging agent, camptothecin.
Asunto(s)
Bencimidazoles/química , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas/metabolismo , Quinolonas/química , Quinolonas/farmacología , Alquilación , Aminación , Línea Celular Tumoral , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , Cristalografía por Rayos X , Humanos , Concentración 50 Inhibidora , Modelos Moleculares , Estructura Molecular , Inhibidores de Proteínas Quinasas/química , Proteínas Quinasas/química , Quinolonas/síntesis química , Relación Estructura-ActividadRESUMEN
Herein are described a series of novel heterocyclic analogs of the 4-amino-3-benzimidazol-2-ylhydroquinolin-2-one scaffold. These compounds are potent inhibitors of receptor tyrosine kinases and exhibit favorable pharmacokinetic profiles. The synthesis and SAR of these compounds are described.
Asunto(s)
Bencimidazoles/síntesis química , Inhibidores Enzimáticos/síntesis química , Compuestos Heterocíclicos/síntesis química , Hidroquinonas/síntesis química , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Bencimidazoles/farmacología , Diseño de Fármacos , Inhibidores Enzimáticos/farmacología , Compuestos Heterocíclicos/farmacología , Hidroquinonas/farmacología , Relación Estructura-ActividadRESUMEN
Insulin resistance plays a central role in the development of type 2 diabetes, but the precise defects in insulin action remain to be elucidated. Glycogen synthase kinase 3 (GSK-3) can negatively regulate several aspects of insulin signaling, and elevated levels of GSK-3 have been reported in skeletal muscle from diabetic rodents and humans. A limited amount of information is available regarding the utility of highly selective inhibitors of GSK-3 for the modification of insulin action under conditions of insulin resistance. In the present investigation, we describe novel substituted aminopyrimidine derivatives that inhibit human GSK-3 potently (K(i) < 10 nmol/l) with at least 500-fold selectivity against 20 other protein kinases. These low molecular weight compounds activated glycogen synthase at approximately 100 nmol/l in cultured CHO cells transfected with the insulin receptor and in primary hepatocytes isolated from Sprague-Dawley rats, and at 500 nmol/l in isolated type 1 skeletal muscle of both lean Zucker and ZDF rats. It is interesting that these GSK-3 inhibitors enhanced insulin-stimulated glucose transport in type 1 skeletal muscle from the insulin-resistant ZDF rats but not from insulin-sensitive lean Zucker rats. Single oral or subcutaneous doses of the inhibitors (30-48 mg/kg) rapidly lowered blood glucose levels and improved glucose disposal after oral or intravenous glucose challenges in ZDF rats and db/db mice, without causing hypoglycemia or markedly elevating insulin. Collectively, our results suggest that these selective GSK-3 inhibitors may be useful as acute-acting therapeutics for the treatment of the insulin resistance of type 2 diabetes.