RESUMO
Glycogen synthase kinase-3 plays an essential role in multiple biochemical pathways in the cell, particularly in regards to energy regulation. As such, Glycogen synthase kinase-3 is an attractive target for pharmacological intervention in a variety of disease states, particularly non-insulin dependent diabetes mellitus. However, due to homology with other crucial kinases, such as the cyclin-dependent protein kinase CDC2, developing compounds that are both potent and selective is challenging. A novel series of derivatives of 5-nitro-N2-(2-(pyridine-2ylamino)ethyl)pyridine-2,6-diamine were synthesized and have been shown to potently inhibit glycogen synthase kinase-3 (GSK3). Potency in the low nanomolar range was obtained along with remarkable selectivity. The compounds activate glycogen synthase in insulin receptor-expressing CHO-IR cells and in primary rat hepatocytes, and have acceptable pharmacokinetics and pharmacodynamics to allow for oral dosing. The X-ray co-crystal structure of human GSK3-ß in complex with compound 2 is reported and provides insights into the structural determinants of the series responsible for its potency and selectivity.
Assuntos
Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Inibidores de Proteínas Quinases/química , Piridinas/química , Animais , Sítios de Ligação , Cristalografia por Raios X , Avaliação Pré-Clínica de Medicamentos , Quinase 3 da Glicogênio Sintase/metabolismo , Meia-Vida , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Concentração Inibidora 50 , Simulação de Dinâmica Molecular , Inibidores de Proteínas Quinases/metabolismo , Inibidores de Proteínas Quinases/farmacocinética , Estrutura Terciária de Proteína , Piridinas/metabolismo , Piridinas/farmacocinética , Ratos , Relação Estrutura-AtividadeRESUMO
The success of hit-finding campaigns relies on many factors, including the quality and diversity of the set of compounds that is selected for screening. This paper presents a generalized workflow that guides compound selections from large compound archives with opportunities to bias the selections with available knowledge in order to improve hit quality while still effectively sampling the accessible chemical space. An optional flag in the workflow supports an explicit complement design function where diversity selections complement a given core set of compounds. Results from three project applications as well as a literature case study exemplify the effectiveness of the approach, which is available as a KNIME workflow named Biased Complement Diversity (BCD).
Assuntos
Descoberta de Drogas/métodos , Animais , Antibacterianos/farmacologia , Antimaláricos/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Bactérias Gram-Negativas/efeitos dos fármacos , Infecções por Bactérias Gram-Negativas/tratamento farmacológico , Ensaios de Triagem em Larga Escala/métodos , Humanos , Malária Falciparum/tratamento farmacológico , Plasmodium falciparum/efeitos dos fármacos , Mapas de Interação de Proteínas/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Fluxo de TrabalhoRESUMO
In an effort to identify new antidiabetic agents, we have discovered a novel family of (5-imidazol-2-yl-4-phenylpyrimidin-2-yl)[2-(2-pyridylamino)ethyl]amine analogues which are inhibitors of human glycogen synthase kinase 3 (GSK3). We developed efficient synthetic routes to explore a wide variety of substitution patterns and convergently access a diverse array of analogues. Compound 1 (CHIR-911, CT-99021, or CHIR-73911) emerged from an exploration of heterocycles at the C-5 position, phenyl groups at C-4, and a variety of differently substituted linker and aminopyridine moieties attached at the C-2 position. These compounds exhibited GSK3 IC50s in the low nanomolar range and excellent selectivity. They activate glycogen synthase in insulin receptor-expressing CHO-IR cells and primary rat hepatocytes. Evaluation of lead compounds 1 and 2 (CHIR-611 or CT-98014) in rodent models of type 2 diabetes revealed that single oral doses lowered hyperglycemia within 60 min, enhanced insulin-stimulated glucose transport, and improved glucose disposal without increasing insulin levels.
Assuntos
Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Quinases da Glicogênio Sintase/antagonistas & inibidores , Hipoglicemiantes/síntese química , Hipoglicemiantes/farmacologia , Pirimidinas/farmacologia , Animais , Células CHO , Cromatografia Líquida de Alta Pressão , Cricetulus , Cristalografia por Raios X , Inibidores Enzimáticos/metabolismo , Humanos , Hipoglicemiantes/metabolismo , Espectrometria de Massas , Espectroscopia de Prótons por Ressonância Magnética , Pirimidinas/química , Pirimidinas/metabolismo , Ratos , Relação Estrutura-AtividadeRESUMO
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.
Assuntos
Lipídeos/química , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Animais , Linhagem Celular , Linhagem Celular Tumoral , Humanos , Espectroscopia de Ressonância Magnética , PrenilaçãoRESUMO
CSN5 is the zinc metalloprotease subunit of the COP9 signalosome (CSN), which is an important regulator of cullin-RING E3 ubiquitin ligases (CRLs). CSN5 is responsible for the cleavage of NEDD8 from CRLs, and blocking deconjugation of NEDD8 traps the CRLs in a hyperactive state, thereby leading to auto-ubiquitination and ultimately degradation of the substrate recognition subunits. Herein, we describe the discovery of azaindoles as a new class of CSN5 inhibitors, which interact with the active-site zinc ion of CSN5 through an unprecedented binding mode. The best compounds inhibited CSN5 with nanomolar potency, led to degradation of the substrate recognition subunit Skp2 in cells, and reduced the viability of HCT116 cells.
Assuntos
Complexo do Signalossomo COP9/antagonistas & inibidores , Indóis/metabolismo , Zinco/metabolismo , Sítios de Ligação , Complexo do Signalossomo COP9/genética , Complexo do Signalossomo COP9/metabolismo , Domínio Catalítico , Proliferação de Células/efeitos dos fármacos , Cristalografia por Raios X , Transferência Ressonante de Energia de Fluorescência , Células HCT116 , Humanos , Indóis/química , Indóis/farmacologia , Simulação de Acoplamento Molecular , Proteína NEDD8/química , Proteína NEDD8/metabolismo , Subunidades Proteicas/antagonistas & inibidores , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Proteínas Quinases Associadas a Fase S/química , Proteínas Quinases Associadas a Fase S/metabolismo , Zinco/químicaRESUMO
The discovery of a highly potent and selective small molecule inhibitor 9 for in vitro target validation of MNK1/2 kinases is described. The aminopyrazine benzimidazole series was derived from an HTS hit and optimized by utilization of a docking model, conformation analysis, and binding pocket comparison against antitargets.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Sítios de Ligação , Células CACO-2/efeitos dos fármacos , Desenho de Fármacos , Ensaios de Triagem em Larga Escala/métodos , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/química , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Simulação de Acoplamento Molecular , Permeabilidade , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/metabolismo , Solubilidade , Relação Estrutura-AtividadeRESUMO
Phospoinositide-3-kinases (PI3K) are important oncology targets due to the deregulation of this signaling pathway in a wide variety of human cancers. A series of 2-morpholino, 4-substituted, 6-(3-hydroxyphenyl) pyrimidines have been reported as potent inhibitors of PI3Ks. Herein, we describe the structure-guided optimization of these pyrimidines with a focus on replacing the phenol moiety, while maintaining potent target inhibition and improving in vivo properties. A series of 2-morpholino, 4-substituted, 6-heterocyclic pyrimidines, which potently inhibit PI3K, were discovered. Within this series a compound, 17, was identified with suitable pharmacokinetic (PK) properties, which allowed for the establishment of a PI3K PK/pharmacodynamic-efficacy relationship as determined by in vivo inhibition of AKT(Ser473) phosphorylation and tumor growth inhibition in a mouse A2780 tumor xenograft model.
RESUMO
Phosphoinositide-3-kinases (PI3Ks) are important oncology targets due to the deregulation of this signaling pathway in a wide variety of human cancers. Herein we describe the structure guided optimization of a series of 2-morpholino, 4-substituted, 6-heterocyclic pyrimidines where the pharmacokinetic properties were improved by modulating the electronics of the 6-position heterocycle, and the overall druglike properties were fine-tuned further by modification of the 4-position substituent. The resulting 2,4-bismorpholino 6-heterocyclic pyrimidines are potent class I PI3K inhibitors showing mechanism modulation in PI3K dependent cell lines and in vivo efficacy in tumor xenograft models with PI3K pathway deregulation (A2780 ovarian and U87MG glioma). These efforts culminated in the discovery of 15 (NVP-BKM120), currently in Phase II clinical trials for the treatment of cancer.
RESUMO
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.