RESUMO
A series of novel, potent 4-aminothienopyridine B-Raf kinase inhibitors was designed and synthesized using knowledge-based design. Compounds 5f, and 6k exhibited not only excellent potency in both enzyme assay (IC(50)=5.1, 16.6 nM) and cellular assay (IC(50)=0.2, 0.2 µM), but also had an outstanding selectivity profile against other kinases.
Assuntos
Aminopiridinas/química , Compostos de Fenilureia/química , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Tienopiridinas/química , Aminopiridinas/síntese química , Aminopiridinas/metabolismo , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , Compostos de Fenilureia/síntese química , Compostos de Fenilureia/metabolismo , Ligação Proteica , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/metabolismo , Proteínas Proto-Oncogênicas B-raf/metabolismo , Relação Estrutura-Atividade , Tienopiridinas/síntese química , Tienopiridinas/metabolismoRESUMO
A potent series of inhibitors against the B-Raf(V600E) kinase have been developed that show excellent activity in cellular assays and good oral bioavailability in rats. The key structural features of the series are an arylsulfonamide headgroup, a thiazole core, and a fluorine ortho to the sulfonamide nitrogen.
Assuntos
Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Sulfonamidas/química , Administração Oral , Substituição de Aminoácidos , Animais , Sítios de Ligação , Domínio Catalítico , Flúor/química , Microssomos Hepáticos/metabolismo , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacocinética , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Ratos , Relação Estrutura-AtividadeRESUMO
Analysis of the x-ray crystal structure of mono-substituted acetylenic thienopyrimidine 6 complexed with the ErbB family enzyme ErbB-4 revealed a covalent bond between the terminal carbon of the acetylene moiety and the sulfhydryl group of Cys-803 at the solvent interface. The identification of this covalent adduct suggested that acetylenic thienopyrimidine 6 and related analogs might also be capable of forming an analogous covalent adduct with EGFR, which has a conserved cysteine (797) near the ATP binding pocket. To test this hypothesis, we treated a truncated, catalytically competent form of EGFR (678-1020) with a structurally related propargylic amine (8). An investigation of the resulting complex by mass spectrometry revealed the formation of a covalent complex of thienopyrimidine 8 with Cys-797 of EGFR. This finding enabled us to readily assess the irreversibility of various inhibitors and also facilitated a structure-activity relationship understanding of the covalent modifying potential and biological activity of a series of acetylenic thienopyrimidine compounds with potent antitumor activity. Several ErbB family enzyme and cell potent 6-ethynyl thienopyrimidine kinase inhibitors were found to form covalent adducts with EGFR.
Assuntos
Alcinos/metabolismo , Compostos de Anilina/metabolismo , Receptores ErbB/metabolismo , Modelos Moleculares , Pirimidinas/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Feminino , Isatina/análogos & derivados , Isatina/metabolismo , Espectrometria de Massas , Camundongos , Camundongos SCID , Estrutura Molecular , Pirimidinas/toxicidade , Receptores Proteína Tirosina Quinases/metabolismo , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Optimization of the amino acid residue of a series of anthranilimide-based glycogen phosphorylase inhibitors is described leading to the identification of serine and threonine ether analogs. t-Butylthreonine analog 20 displayed potent in vitro inhibition of GPa, low potential for P450 inhibition, and excellent pharmacokinetic properties.
Assuntos
Hidrocarboneto de Aril Hidroxilases/antagonistas & inibidores , Química Farmacêutica/métodos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Glicogênio Fosforilase/antagonistas & inibidores , Imidas/química , Serina/química , Treonina/química , ortoaminobenzoatos/química , Animais , Hidrocarboneto de Aril Hidroxilases/química , Cristalografia por Raios X , Citocromo P-450 CYP2C9 , Desenho de Fármacos , Glicina/química , Glicogênio Fosforilase/metabolismo , Humanos , Concentração Inibidora 50 , Fígado/enzimologia , Modelos Químicos , RatosRESUMO
Aniline 'headgroups' were synthesized and incorporated into an alkynyl thienopyrimidine series of EGFR and ErbB-2 inhibitors. Potent inhibition of enzyme activity and cellular proliferation was observed. In certain instances, protein binding was reduced and oral exposure was found to be somewhat improved relative to compounds containing the reference aniline.
Assuntos
Compostos de Anilina/síntese química , Receptores ErbB/antagonistas & inibidores , Inibidores de Proteínas Quinases/síntese química , Pirimidinas/síntese química , Receptor ErbB-2/antagonistas & inibidores , Administração Oral , Compostos de Anilina/administração & dosagem , Compostos de Anilina/farmacologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Receptores ErbB/metabolismo , Inibidores do Crescimento/administração & dosagem , Inibidores do Crescimento/síntese química , Inibidores do Crescimento/farmacologia , Humanos , Camundongos , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/administração & dosagem , Pirimidinas/farmacologia , Receptor ErbB-2/metabolismoRESUMO
Key binding interactions of the anthranilimide based glycogen phosphorylase a (GPa) inhibitor 2 from X-ray crystallography studies are described. This series of compounds bind to the AMP site of GP. Using the binding information the core and the phenyl urea moieties were optimized. This work culminated in the identification of compounds with single nanomolar potency as well as in vivo efficacy in a diabetic model.
Assuntos
Diabetes Mellitus Tipo 2/tratamento farmacológico , Glicogênio Fosforilase/antagonistas & inibidores , Hipoglicemiantes/síntese química , ortoaminobenzoatos/síntese química , ortoaminobenzoatos/farmacologia , Animais , Glicemia/análise , Técnicas de Química Combinatória , Cristalografia por Raios X , Modelos Animais de Doenças , Hipoglicemiantes/sangue , Hipoglicemiantes/química , Hipoglicemiantes/farmacologia , Camundongos , Conformação Molecular , Estrutura Molecular , Relação Estrutura-Atividade , Ureia/farmacologia , ortoaminobenzoatos/sangue , ortoaminobenzoatos/químicaRESUMO
Two new series of potent and selective dual EGFR/ErbB-2 kinase inhibitors derived from novel thienopyrimidine cores have been identified. Isomeric thienopyrimidine cores were evaluated as isosteres for a 4-anilinoquinazoline core and several analogs containing the thieno[3,2-d]pyrimidine core showed anti-proliferative activity with IC(50) values less than 1 microM against human tumor cells in vitro.
Assuntos
Antineoplásicos/farmacologia , Química Farmacêutica/métodos , Receptores ErbB/química , Pirimidinas/química , Receptor ErbB-2/antagonistas & inibidores , Antineoplásicos/síntese química , Linhagem Celular Tumoral , Proliferação de Células , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/farmacologia , Humanos , Concentração Inibidora 50 , Lapatinib , Modelos Químicos , Conformação Molecular , Quinazolinas/farmacologiaRESUMO
Optimization of the amino acid residue within a series of anthranilimide-based glycogen phosphorylase inhibitors is described. These studies culminated in the identification of anthranilimides 16 and 22 which displayed potent in vitro inhibition of GPa in addition to reduced inhibition of CYP2C9 and excellent pharmacokinetic properties.
Assuntos
Hidrocarboneto de Aril Hidroxilases/antagonistas & inibidores , Ácidos Carboxílicos/química , Química Farmacêutica/métodos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Glicogênio Fosforilase/antagonistas & inibidores , Imidas/farmacologia , ortoaminobenzoatos/farmacologia , Animais , Hidrocarboneto de Aril Hidroxilases/química , Cristalografia por Raios X , Citocromo P-450 CYP2C9 , Cães , Desenho de Fármacos , Glicina/química , Glicogênio Fosforilase/metabolismo , Humanos , Imidas/química , Concentração Inibidora 50 , Fígado/enzimologia , Conformação Molecular , Ratos , ortoaminobenzoatos/químicaRESUMO
A novel class of pyrrolidinyl-acetyleneic thieno[3,2-d]pyrimidines has been identified which potently inhibit the EGFR and ErbB-2 receptor tyrosine kinases. Synthetic modifications of the pyrrolidine carbamate moiety result in a range of effects on enzyme and cellular potency. In addition, the impact of the absolute stereochemical configuration on cellular potency and oral mouse pharmacokinetics is described.
Assuntos
Antineoplásicos/química , Receptores ErbB/antagonistas & inibidores , Pirimidinas/farmacologia , Pirrolidinas/farmacologia , Receptor ErbB-2/antagonistas & inibidores , Administração Oral , Animais , Camundongos , Farmacocinética , Pirimidinas/síntese química , Pirrolidinas/síntese química , Relação Estrutura-AtividadeRESUMO
The optimization of imidazo[1,2-a]pyridine inhibitors as potent and selective inhibitors of IGF-1R is presented. Further optimization of oral exposure in mice is also discussed. Detailed selectivity, in vitro activity, and in vivo PK profiles of an optimized compound is also highlighted.
Assuntos
Química Farmacêutica/métodos , Piridinas/química , Receptor IGF Tipo 1/antagonistas & inibidores , Receptor IGF Tipo 1/química , Administração Oral , Compostos de Anilina/química , Animais , Linhagem Celular Tumoral , Cristalografia por Raios X , Desenho de Fármacos , Humanos , Concentração Inibidora 50 , Camundongos , Modelos Químicos , Conformação Molecular , Piridinas/síntese química , Piridinas/farmacologia , Receptor de Insulina/metabolismoRESUMO
The synthesis of a 7-azaindole series of novel, potent B-Raf kinase inhibitors using knowledge-based design was carried out. Compound 6h exhibits not only excellent potency in both the enzyme assay (IC(50)=2.5 nM) and the cellular assay (IC(50)=63 nM), but also has an outstanding selectivity profile against other kinases.
Assuntos
Química Farmacêutica/métodos , Indóis/farmacologia , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/metabolismo , Desenho de Fármacos , Humanos , Concentração Inibidora 50 , Modelos Biológicos , Modelos Químicos , Estrutura Molecular , Fosfotransferases/metabolismo , Inibidores de Proteínas Quinases/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
A novel class of substituted pyrrolidinyl-acetylenic thieno[3,2-d]pyrimidines has been identified that are potent and selective inhibitors of both EGFR/ErbB-2 receptor tyrosine kinases. The inhibitors are found to display a range of enzyme and cellular potency and also to display a varying level of covalent modification of the kinase targets. Selected molecules, including compound 15h, were found to be potent in enzymatic and cellular assays while also demonstrating exposure in the mouse from an oral dose.
Assuntos
Receptores ErbB/antagonistas & inibidores , Pirimidinas/farmacologia , Receptor ErbB-2/antagonistas & inibidores , Animais , Linhagem Celular , Camundongos , Ligação Proteica , Pirimidinas/química , Relação Estrutura-AtividadeRESUMO
GW572016 (Lapatinib) is a tyrosine kinase inhibitor in clinical development for cancer that is a potent dual inhibitor of epidermal growth factor receptor (EGFR, ErbB-1) and ErbB-2. We determined the crystal structure of EGFR bound to GW572016. The compound is bound to an inactive-like conformation of EGFR that is very different from the active-like structure bound by the selective EGFR inhibitor OSI-774 (Tarceva) described previously. Surprisingly, we found that GW572016 has a very slow off-rate from the purified intracellular domains of EGFR and ErbB-2 compared with OSI-774 and another EGFR selective inhibitor, ZD-1839 (Iressa). Treatment of tumor cells with these inhibitors results in down-regulation of receptor tyrosine phosphorylation. We evaluated the duration of the drug effect after washing away free compound and found that the rate of recovery of receptor phosphorylation in the tumor cells reflected the inhibitor off-rate from the purified intracellular domain. The slow off-rate of GW572016 correlates with a prolonged down-regulation of receptor tyrosine phosphorylation in tumor cells. The differences in the off-rates of these drugs and the ability of GW572016 to inhibit ErbB-2 can be explained by the enzyme-inhibitor structures.
Assuntos
Inibidores Enzimáticos/química , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/química , Quinazolinas/química , Sequência de Aminoácidos , Sítios de Ligação , Linhagem Celular Tumoral , Cristalografia por Raios X , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Receptores ErbB/metabolismo , Cloridrato de Erlotinib , Humanos , Cinética , Lapatinib , Modelos Moleculares , Dados de Sequência Molecular , Proteínas Oncogênicas v-erbB/antagonistas & inibidores , Conformação Proteica , Estrutura Secundária de Proteína , Quinazolinas/metabolismo , Quinazolinas/farmacologia , Especificidade por SubstratoRESUMO
Glycogen synthase kinase 3 regulates glycogen synthase, the rate-determining enzyme for glycogen synthesis. Liver and muscle glycogen synthesis is defective in type 2 diabetics, resulting in elevated plasma glucose levels. Inhibition of GSK-3 could potentially be an effective method to control plasma glucose levels in type 2 diabetics. Structure-activity studies on a N-phenyl-4-pyrazolo[1,5-b]pyridazin-3-ylpyrimidin-2-amine series have led to the identification of potent and selective compounds with good cellular efficacy. Molecular modeling studies have given insights into the mode of binding of these inhibitors. Since the initial leads were also potent inhibitors of CDK-2/CDK-4, an extensive SAR was performed at various positions of the pyrazolo[1,5-b]pyridazin core to afford potent GSK-3 inhibitors that were highly selective over CDK-2. In addition, these inhibitors also exhibited very good cell efficacy and functional response. A representative example was shown to have good oral exposure levels, extending their utility in an in vivo setting. These inhibitors provide a viable lead series in the discovery of new therapies for the treatment of type 2 diabetes.
Assuntos
Aminas/química , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Piridazinas/química , Piridazinas/farmacologia , Animais , Sítios de Ligação , Linhagem Celular , Inibidores Enzimáticos/síntese química , Quinase 3 da Glicogênio Sintase/metabolismo , Humanos , Concentração Inibidora 50 , Modelos Moleculares , Estrutura Molecular , Estrutura Terciária de Proteína , Piridazinas/síntese química , Ratos , Relação Estrutura-AtividadeRESUMO
Hyperactive signaling of the MAP kinase pathway resulting from the constitutively active B-Raf(V600E) mutated enzyme has been observed in a number of human tumors, including melanomas. Herein we report the discovery and biological evaluation of GSK2118436, a selective inhibitor of Raf kinases with potent in vitro activity in oncogenic B-Raf-driven melanoma and colorectal carcinoma cells and robust in vivo antitumor and pharmacodynamic activity in mouse models of B-Raf(V600E) human melanoma. GSK2118436 was identified as a development candidate, and early clinical results have shown significant activity in patients with B-Raf mutant melanoma.
RESUMO
Recent results from clinical trials with the BRAF inhibitors GSK2118436 (dabrafenib) and PLX4032 (vemurafenib) have shown encouraging response rates; however, the duration of response has been limited. To identify determinants of acquired resistance to GSK2118436 and strategies to overcome the resistance, we isolated GSK2118436 drug-resistant clones from the A375 BRAF(V600E) and the YUSIT1 BRAF(V600K) melanoma cell lines. These clones also showed reduced sensitivity to the allosteric mitogen-activated protein/extracellular signal-regulated kinase (MEK) inhibitor GSK1120212 (trametinib). Genetic characterization of these clones identified an in-frame deletion in MEK1 (MEK1(K59del)) or NRAS mutation (NRAS(Q61K) and/or NRAS(A146T)) with and without MEK1(P387S) in the BRAF(V600E) background and NRAS(Q61K) in the BRAF(V600K) background. Stable knockdown of NRAS with short hairpin RNA partially restored GSK2118436 sensitivity in mutant NRAS clones, whereas expression of NRAS(Q61K) or NRAS(A146T) in the A375 parental cells decreased sensitivity to GSK2118436. Similarly, expression of MEK1(K59del), but not MEK1(P387S), decreased sensitivity of A375 cells to GSK2118436. The combination of GSK2118436 and GSK1120212 effectively inhibited cell growth, decreased ERK phosphorylation, decreased cyclin D1 protein, and increased p27(kip1) protein in the resistant clones. Moreover, the combination of GSK2118436 or GSK1120212 with the phosphoinositide 3-kinase/mTOR inhibitor GSK2126458 enhanced cell growth inhibition and decreased S6 ribosomal protein phosphorylation in these clones. Our results show that NRAS and/or MEK mutations contribute to BRAF inhibitor resistance in vitro, and the combination of GSK2118436 and GSK1120212 overcomes this resistance. In addition, these resistant clones respond to the combination of GSK2126458 with GSK2118436 or GSK1120212. Clinical trials are ongoing or planned to test these combinations.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Melanoma/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Apoptose/efeitos dos fármacos , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Humanos , Imidazóis/administração & dosagem , Imidazóis/farmacologia , Indóis/administração & dosagem , Indóis/farmacologia , MAP Quinase Quinase 1/antagonistas & inibidores , Melanoma/enzimologia , Oximas/administração & dosagem , Oximas/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Fosforilação , Inibidores de Proteínas Quinases/administração & dosagem , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Sulfonamidas/administração & dosagem , Sulfonamidas/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , VemurafenibRESUMO
Anilinoalkynylpyrimidines were prepared and evaluated as dual EGFR/ErbB2 kinase inhibitors. A preference was found for substituted phenyl and heteroaromatic rings attached to the alkyne. In addition, the presence of a potential hydrogen bond donor appended to this ring was favored. Selected molecules in the series demonstrated some activity against human tumor cell lines.
Assuntos
Alcinos/química , Receptores ErbB/antagonistas & inibidores , Pirimidinas/farmacologia , Receptor ErbB-2/antagonistas & inibidores , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cristalografia por Raios X , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Modelos Moleculares , Estrutura Molecular , Pirimidinas/síntese química , Pirimidinas/química , Relação Estrutura-AtividadeRESUMO
The identification of the kinase or kinases targeted by protein kinase inhibitors is a critical challenge in validating their use as therapeutic agents or molecular probes. Here, to address this problem, we describe a chemical genomics strategy that uses a direct comparison between microarray transcriptional signatures elicited by an inhibitor of unknown specificity and those elicited by highly specific pharmacological inhibition of engineered candidate kinase targets. By using this approach, we have identified two cyclin-dependent kinases, Cdk1 and Pho85, as the targets of the inhibitor GW400426 in Saccharomyces cerevisiae. We demonstrate that simultaneous inhibition of Cdk1 and Pho85, and not inhibition of either kinase alone, by GW400426 controls the expression of specific transcripts involved in polarized cell growth, thus revealing a cellular process that is uniquely sensitive to the multiplex inhibition of these two kinases. Our results suggest that the cellular responses induced by multiplex protein kinase inhibitors may be an emergent property that cannot be understood fully by considering only the sum of individual inhibitor-kinase interactions.
Assuntos
Proteína Quinase CDC2/antagonistas & inibidores , Quinases Ciclina-Dependentes/antagonistas & inibidores , Perfilação da Expressão Gênica , Inibidores de Proteínas Quinases/farmacologia , Proteínas de Saccharomyces cerevisiae/antagonistas & inibidores , Genômica , Análise de Sequência com Séries de OligonucleotídeosRESUMO
A novel series of [1-(1H-benzimidazol-7-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl] arylhydrazones was synthesized and shown to potently inhibit glycogen synthase kinase-3 (GSK-3). In light of detailed structure-activity relationships and structural knowledge of the GSK-3 binding pocket, a benzimidazole substituent was incorporated onto the pyrazolopyrimidine core resulting in improved potency over previous analogs. More importantly, these derivatives show low nanomolar efficacy for stimulating glycogen synthesis in vitro and therefore may be useful in the treatment of type 2 diabetes mellitus.
Assuntos
Inibidores Enzimáticos/farmacologia , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Hidrazonas/farmacologia , Animais , Linhagem Celular , Cães , Inibidores Enzimáticos/química , Hidrazonas/química , Modelos MolecularesRESUMO
A series of [1-aryl-1H-pyrazolo[3,4-d]pyrimidin-4-yl]arylhydrazones were discovered as novel inhibitors glycogen synthase kinase-3 (GSK-3). Based on initial modeling a detailed SAR was constructed. Modification of the interior binding aryl ring (Ar(1)) determined this to be a tight binding region with little room for modification. As predicted from the model, a large variety of modifications could be incorporated into the hydrazone aryl ring. This work led to GSK-3 inhibitors in the low nano-molar range.