RESUMO
Structure-activity relationship (SAR) studies on a highly potent series of arylamide FMS inhibitors were carried out with the aim of improving FMS kinase selectivity, particularly over KIT. Potent compound 17r (FMS IC50 0.7 nM, FMS cell IC50 6.1 nM) was discovered that had good PK properties and a greater than fivefold improvement in selectivity for FMS over KIT kinase in a cellular assay relative to the previously reported clinical candidate 4. This improved selectivity was manifested in vivo by no observed decrease in circulating reticulocytes, a measure of bone safety, at the highest studied dose. Compound 17r was highly active in a mouse pharmacodynamic model and demonstrated disease-modifying effects in a dose-dependent manner in a strep cell wall-induced arthritis model of rheumatoid arthritis in rats.
Assuntos
Amidas/farmacologia , Compostos Heterocíclicos/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Amidas/síntese química , Amidas/química , Animais , Compostos Heterocíclicos/síntese química , Compostos Heterocíclicos/química , Masculino , Camundongos , Modelos Moleculares , Estrutura Molecular , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Ratos , Ratos Sprague-Dawley , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismo , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
The inflammatory response associated with the activation of C-C chemokine receptor CCR2 via it's interaction with the monocyte chemoattractant protein-1 (MCP-1, CCL2) has been implicated in many disease states, including rheumatoid arthritis, multiple sclerosis, atherosclerosis, asthma and neuropathic pain. Small molecule antagonists of CCR2 have been efficacious in animal models of inflammatory disease, and have been advanced into clinical development. The necessity to attenuate hERG binding appears to be a common theme for many of the CCR2 antagonist scaffolds appearing in the literature, presumably due the basic hydrophobic motif present in all of these molecules. Following the discovery of a novel cyclohexyl azetidinylamide CCR2 antagonist scaffold, replacement of the amide bond with heterocyclic rings was explored as a strategy for reducing hERG binding and improving pharmacokinetic properties.
Assuntos
Acetamidas/química , Acetamidas/farmacologia , Azetidinas/química , Azetidinas/farmacologia , Receptores CCR2/antagonistas & inibidores , Animais , Humanos , CamundongosRESUMO
Complement C1s protease inhibitors have potential utility in the treatment of diseases associated with activation of the classical complement pathway such as humorally mediated graft rejection, ischemia-reperfusion injury (IRI), vascular leak syndrome, and acute respiratory distress syndrome (ARDS). The utility of biphenylsulfonyl-thiophene-carboxamidine small-molecule C1s inhibitors are limited by their poor in vivo pharmacokinetic properties. Pegylation of a potent analog has provided compounds with good potency and good in vivo pharmacokinetic properties.
Assuntos
Amidas/química , Complemento C1s/antagonistas & inibidores , Desenho de Fármacos , Polietilenoglicóis/química , Inibidores de Proteases/síntese química , Tiofenos/química , Animais , Complemento C1s/metabolismo , Meia-Vida , Inibidores de Proteases/química , Inibidores de Proteases/farmacocinética , RatosRESUMO
As a result of further SAR studies on a piperidinyl piperidine scaffold, we report the discovery of compound 44, a potent, orally bioavailable CCR2 antagonist. While having some in vitro hERG activity, this molecule was clean in an in vivo model of QT prolongation. In addition, it showed excellent efficacy when dosed orally in a transgenic murine model of acute inflammation.
Assuntos
Amidas/química , Anti-Inflamatórios/química , Receptores CCR2/antagonistas & inibidores , Doença Aguda , Administração Oral , Amidas/farmacologia , Amidas/uso terapêutico , Animais , Anti-Inflamatórios/farmacocinética , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Avaliação Pré-Clínica de Medicamentos , Humanos , Inflamação/tratamento farmacológico , Camundongos , Camundongos Transgênicos , Ratos , Receptores CCR2/metabolismo , Relação Estrutura-AtividadeRESUMO
A class of potent inhibitors of colony-stimulating factor-1 receptor (CSF-1R or FMS), as exemplified by 8 and 21, was optimized to improve pharmacokinetic and pharmacodynamic properties and potential toxicological liabilities. Early stage absorption, distribution, metabolism, and excretion assays were employed to ensure the incorporation of druglike properties resulting in the selection of several compounds with good activity in a pharmacodynamic screening assay in mice. Further investigation, utilizing the type II collagen-induced arthritis model in mice, culminated in the selection of anti-inflammatory development candidate JNJ-28312141 (23, FMS IC(50) = 0.69 nM, cell assay IC(50) = 2.6 nM). Compound 23 also demonstrated efficacy in rat adjuvant and streptococcal cell wall-induced models of arthritis and has entered phase I clinical trials.
Assuntos
Anti-Inflamatórios não Esteroides/síntese química , Imidazóis/síntese química , Piperidinas/síntese química , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Animais , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Artrite Experimental/tratamento farmacológico , Artrite Experimental/etiologia , Artrite Experimental/patologia , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular , Proliferação de Células/efeitos dos fármacos , Cristalografia por Raios X , Feminino , Humanos , Imidazóis/farmacocinética , Imidazóis/farmacologia , Técnicas In Vitro , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Microssomos Hepáticos/metabolismo , Modelos Moleculares , Piperidinas/farmacocinética , Piperidinas/farmacologia , Conformação Proteica , Ratos , Ratos Endogâmicos Lew , Receptor de Fator Estimulador de Colônias de Macrófagos/química , Solubilidade , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
A novel series of 4-azetidinyl-1-aryl-cyclohexanes containing indazole or benzoisoxazole moiety have been identified as potent CCR2 antagonists with high selectivity versus hERG.
Assuntos
Indazóis/química , Indazóis/farmacologia , Oxazóis/química , Oxazóis/farmacologia , Receptores CCR2/antagonistas & inibidores , Animais , Linhagem Celular , Cicloexanos/síntese química , Cicloexanos/química , Cicloexanos/farmacocinética , Cicloexanos/farmacologia , Cães , Desenho de Fármacos , Humanos , Indazóis/síntese química , Indazóis/farmacocinética , Oxazóis/síntese química , Oxazóis/farmacocinética , Ratos , Receptores CCR2/metabolismo , Relação Estrutura-Atividade , Transativadores/metabolismo , Regulador Transcricional ERGRESUMO
A series of 4-azetidinyl-1-aryl-cyclohexanes as potent CCR2 antagonists with high selectivity over activity for the hERG potassium channel is discovered through divergent SARs of CCR2 and hERG.
Assuntos
Azetidinas/farmacologia , Descoberta de Drogas , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Hipuratos/farmacologia , Receptores CCR2/antagonistas & inibidores , Azetidinas/síntese química , Azetidinas/química , Relação Dose-Resposta a Droga , Canais de Potássio Éter-A-Go-Go/metabolismo , Hipuratos/síntese química , Hipuratos/química , Humanos , Estrutura Molecular , Receptores CCR2/metabolismo , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Chymases (EC 3.4.21.39) are mast cell serine proteinases that are variably expressed in different species and, in most cases, display either chymotryptic or elastolytic substrate specificity. Given that chymase inhibitors have emerged as potential therapeutic agents for treating various inflammatory, allergic, and cardiovascular disorders, it is important to understand interspecies differences of the enzymes as well as the behavior of inhibitors with them. We have expressed chymases from humans, macaques, dogs, sheep (MCP2 and MCP3), guinea pigs, and hamsters (HAM1 and HAM2) in baculovirus-infected insect cells. The enzymes were purified and characterized with kinetic constants by using chromogenic substrates. We evaluated in vitro the potency of five nonpeptide inhibitors, originally targeted against human chymase. The inhibitors exhibited remarkable cross-species variation of sensitivity, with the greatest potency observed against human and macaque chymases, with K(i) values ranging from approximately 0.4 to 72nM. Compounds were 10-300-fold less potent, and in some instances ineffective, against chymases from the other species. The X-ray structure of one of the potent phosphinate inhibitors, JNJ-18054478, complexed with human chymase was solved at 1.8A resolution to further understand the binding mode. Subtle variations in the residues in the active site that are already known to influence chymase substrate specificity can also strongly affect the compound potency. The results are discussed in the context of selecting a suitable animal model to study compounds ultimately targeted for human chymase.
Assuntos
Quimases/metabolismo , Animais , Baculoviridae/metabolismo , Cricetinae , Cães , Cobaias , Humanos , Macaca , Mastócitos/enzimologia , Mastócitos/metabolismo , Serina Proteases , Ovinos , Especificidade por Substrato , Raios XRESUMO
During efforts to improve the bioavailability of FMS kinase inhibitors 1 and 2, a series of saturated and aromatic 4-heterocycles of reduced basicity were prepared and evaluated in an attempt to also improve the cardiovascular safety profile over lead arylamide 1, which possessed ion channel activity. The resultant compounds retained excellent potency and exhibited diminished ion channel activity.
Assuntos
Amidas/farmacologia , Compostos Heterocíclicos/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Amidas/síntese química , Amidas/química , Relação Dose-Resposta a Droga , Compostos Heterocíclicos/síntese química , Compostos Heterocíclicos/química , Modelos Moleculares , Estrutura Molecular , Oxirredução , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
We have identified RWJ-671818 (8) as a novel, low molecular weight, orally active inhibitor of human alpha-thrombin (K(i) = 1.3 nM) that is potentially useful for the acute and chronic treatment of venous and arterial thrombosis. In a rat deep venous thrombosis model used to assess antithrombotic efficacy, oral administration of 8 at 30 and 50 mg/kg reduced thrombus weight by 87 and 94%, respectively. In an anesthetized rat antithrombotic model, where electrical stimulation of the carotid artery created a thrombus, 8 prolonged occlusion time 2- and 3-fold at 0.1 and 1.0 mg/kg, i.v., respectively, and more than doubled activated clotting time and activated partial thromboplastin time at the higher dose. This compound had excellent oral bioavailability of 100% in dogs with an estimated half-life of approximately 3 h. On the basis of its noteworthy preclinical data, 8 was advanced into human clinical trials and successfully progressed through phase 1 studies.
Assuntos
Anticoagulantes/síntese química , Fibrinolíticos/síntese química , Guanidinas/síntese química , Pirazinas/síntese química , Trombina/antagonistas & inibidores , Motivos de Aminoácidos , Animais , Anticoagulantes/farmacocinética , Anticoagulantes/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Células CACO-2 , Cristalografia por Raios X , Citocromo P-450 CYP3A , Inibidores do Citocromo P-450 CYP3A , Cães , Método Duplo-Cego , Eletrocardiografia , Feminino , Fibrinolíticos/farmacocinética , Fibrinolíticos/farmacologia , Guanidinas/farmacocinética , Guanidinas/farmacologia , Cobaias , Frequência Cardíaca/efeitos dos fármacos , Hemodinâmica/efeitos dos fármacos , Humanos , Técnicas In Vitro , Masculino , Microssomos Hepáticos/metabolismo , Modelos Moleculares , Pirazinas/farmacocinética , Pirazinas/farmacologia , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/antagonistas & inibidores , Relação Estrutura-Atividade , Trombina/química , Trombose Venosa/sangue , Trombose Venosa/tratamento farmacológicoRESUMO
There is increasing evidence that tumor-associated macrophages promote the malignancy of some cancers. Colony-stimulating factor-1 (CSF-1) is expressed by many tumors and is a growth factor for macrophages and mediates osteoclast differentiation. Herein, we report the efficacy of a novel orally active CSF-1 receptor (CSF-1R) kinase inhibitor, JNJ-28312141, in proof of concept studies of solid tumor growth and tumor-induced bone erosion. H460 lung adenocarcinoma cells did not express CSF-1R and were not growth inhibited by JNJ-28312141 in vitro. Nevertheless, daily p.o. administration of JNJ-28312141 caused dose-dependent suppression of H460 tumor growth in nude mice that correlated with marked reductions in F4/80(+) tumor-associated macrophages and with increased plasma CSF-1, a possible biomarker of CSF-1R inhibition. Furthermore, the tumor microvasculature was reduced in JNJ-28312141-treated mice, consistent with a role for macrophages in tumor angiogenesis. In separate studies, JNJ-28312141 was compared with zoledronate in a model in which MRMT-1 mammary carcinoma cells inoculated into the tibias of rats led to severe cortical and trabecular bone lesions. Both agents reduced tumor growth and preserved bone. However, JNJ-28312141 reduced the number of tumor-associated osteoclasts superior to zoledronate. JNJ-28312141 exhibited additional activity against FMS-related receptor tyrosine kinase-3 (FLT3). To more fully define the therapeutic potential of this new agent, JNJ-28312141 was evaluated in a FLT3-dependent acute myeloid leukemia tumor xenograft model and caused tumor regression. In summary, this novel CSF-1R/FLT3 inhibitor represents a new agent with potential therapeutic activity in acute myeloid leukemia and in settings where CSF-1-dependent macrophages and osteoclasts contribute to tumor growth and skeletal events.
Assuntos
Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/secundário , Imidazóis/farmacologia , Leucemia Mieloide Aguda/tratamento farmacológico , Piperidinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Tirosina Quinase 3 Semelhante a fms/antagonistas & inibidores , Animais , Neoplasias Ósseas/enzimologia , Neoplasias Ósseas/patologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Feminino , Humanos , Imuno-Histoquímica , Leucemia Mieloide Aguda/enzimologia , Leucemia Mieloide Aguda/patologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/patologia , Neoplasias Mamárias Experimentais/tratamento farmacológico , Neoplasias Mamárias Experimentais/enzimologia , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Nus , Osteoclastos/efeitos dos fármacos , Osteoclastos/patologia , Ratos , Ratos Sprague-Dawley , Receptor de Fator Estimulador de Colônias de Macrófagos/sangue , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismo , Especificidade por Substrato , Ensaios Antitumorais Modelo de Xenoenxerto , Tirosina Quinase 3 Semelhante a fms/metabolismoRESUMO
A series of pyrido[2,3-d]pyrimidin-5-ones has been synthesized and evaluated as inhibitors of the kinase domain of macrophage colony-stimulating factor-1 receptor (FMS). FMS inhibitors may be useful in treating rheumatoid arthritis and other chronic inflammatory diseases. Structure-based optimization of the lead amide analogue 10 led to hydroxamate analogue 37, which possessed excellent potency and an improved pharmacokinetic profile. During the chronic phase of streptococcal cell wall-induced arthritis in rats, compound 37 (10, 3, and 1 mg/kg) was highly effective at reversing established joint swelling. In an adjuvant-induced arthritis model in rats, 37 prevented joint swelling partially at 10 mg/kg. In this model, osteoclastogenesis and bone erosion were prevented by low doses (1 or 0.33 mg/kg) that had minimal impact on inflammation. These data underscore the potential of FMS inhibitors to prevent erosions and reduce symptoms in rheumatoid arthritis.
Assuntos
Anti-Inflamatórios/química , Artrite Reumatoide/tratamento farmacológico , Pirimidinonas/química , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Animais , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Reabsorção Óssea/prevenção & controle , Modelos Animais de Doenças , Inflamação/tratamento farmacológico , Osteoclastos/efeitos dos fármacos , Farmacocinética , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Estrutura Terciária de Proteína , Pirimidinonas/farmacologia , Pirimidinonas/uso terapêutico , Ratos , Relação Estrutura-AtividadeRESUMO
An anti-inflammatory 1,2,4-phenylenetriamine-containing series of FMS inhibitors with a potential to form reactive metabolites was transformed into a series with equivalent potency by incorporation of carbon-based replacement groups. Structure-based modeling provided the framework to efficiently effect this transformation and restore potencies to previous levels. This optimization removed a risk factor for potential idiosyncratic drug reactions.
Assuntos
Anti-Inflamatórios/farmacologia , Desenho de Fármacos , Inibidores Enzimáticos/farmacologia , Fenilenodiaminas/farmacologia , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Anti-Inflamatórios/síntese química , Anti-Inflamatórios/química , Simulação por Computador , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Ligação de Hidrogênio , Concentração Inibidora 50 , Modelos Moleculares , Estrutura Molecular , Fenilenodiaminas/síntese química , Fenilenodiaminas/química , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
2-Cyano-6-fluorophenylacetamide was explored as a novel P2 scaffold in the design of thrombin inhibitors. Optimization around this structural motif culminated in 14, which is a potent thrombin inhibitor (K(i)=1.2nM) that exhibits robust efficacy in canine anticoagulation and thrombosis models upon oral administration.
Assuntos
Acetamidas , Motivos de Aminoácidos , Anticoagulantes/administração & dosagem , Desenho de Fármacos , Nitrilas , Trombina/antagonistas & inibidores , Trombose/tratamento farmacológico , Acetamidas/síntese química , Acetamidas/farmacocinética , Acetamidas/uso terapêutico , Administração Oral , Animais , Anticoagulantes/síntese química , Anticoagulantes/farmacocinética , Sítios de Ligação , Modelos Animais de Doenças , Cães , Relação Dose-Resposta a Droga , Haplorrinos , Humanos , Ligação de Hidrogênio , Modelos Químicos , Nitrilas/síntese química , Nitrilas/farmacocinética , Nitrilas/uso terapêutico , Ratos , Relação Estrutura-AtividadeRESUMO
A series of novel acylsulfonamide, acylsulfamide, and sulfonylurea bioisosteres of carboxylic acids were prepared as CXCR2 antagonists. Structure-activity relationships are reported for these series. One potent orally bioavailable inhibitor had excellent PK properties and was active in a lung injury model in hyperoxia-exposed newborn rats.
Assuntos
Ácidos Carboxílicos/química , Ácidos Carboxílicos/farmacologia , Pulmão/efeitos dos fármacos , Receptores de Interleucina-8B/antagonistas & inibidores , Sulfonamidas/química , Ácidos Sulfônicos/química , Compostos de Sulfonilureia/química , Administração Oral , Animais , Animais Recém-Nascidos , Disponibilidade Biológica , Lavagem Broncoalveolar , Ácidos Carboxílicos/farmacocinética , Quimiotaxia/efeitos dos fármacos , Sistema Enzimático do Citocromo P-450/metabolismo , Humanos , Hiperóxia , Pulmão/metabolismo , Lesão Pulmonar , Estrutura Molecular , Neutrófilos/metabolismo , Coelhos , Ratos , Relação Estrutura-AtividadeRESUMO
A series of 3,4,6-substituted 2-quinolones has been synthesized and evaluated as inhibitors of the kinase domain of macrophage colony-stimulating factor-1 receptor (FMS). The fully optimized compound, 4-(4-ethyl-phenyl)-3-(2-methyl-3H-imidazol-4-yl)-2-quinolone-6-carbonitrile 21b, has an IC(50) of 2.5 nM in an in vitro assay and 5.0 nM in a bone marrow-derived macrophage cellular assay. Inhibition of FMS signaling in vivo was also demonstrated in a mouse pharmacodynamic model.
Assuntos
Macrófagos/efeitos dos fármacos , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacologia , Quinolonas/síntese química , Quinolonas/farmacologia , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Administração Oral , Animais , Disponibilidade Biológica , Proliferação de Células/efeitos dos fármacos , Polarização de Fluorescência , Genes fos/genética , Fator Estimulador de Colônias de Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Quinolonas/farmacocinética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Baço/metabolismo , Relação Estrutura-AtividadeRESUMO
Complement activation has been implicated in disease states such as hereditary angioedema, ischemia-reperfusion injury, acute respiratory distress syndrome, and acute transplant rejection. Even though the complement cascade provides several protein targets for potential therapeutic intervention only two complement inhibitors have been approved so far for clinical use including anti-C5 antibodies for the treatment of paroxysmal nocturnal hemoglobinuria and purified C1-esterase inhibitor replacement therapy for the control of hereditary angioedema flares. In the present study, optimization of potency and physicochemical properties of a series of thiophene amidine-based C1s inhibitors with potential utility as intravenous agents for the inhibition of the classical pathway of complement is described.
Assuntos
Complemento C1s/antagonistas & inibidores , Compostos Heterocíclicos/química , Compostos Heterocíclicos/farmacologia , Animais , Sítios de Ligação , Meia-Vida , Modelos Moleculares , Estrutura Molecular , Ratos , Relação Estrutura-AtividadeRESUMO
The optimization of the arylamide lead 2 resulted in identification of a highly potent series of 2,4-disubstituted arylamides. Compound 8 (FMS kinase IC(50)=0.0008 microM) served as a proof-of-concept candidate in a collagen-induced model of arthritis in mice.
Assuntos
Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Receptor de Fator Estimulador de Colônias de Macrófagos/antagonistas & inibidores , Animais , Artrite/induzido quimicamente , Artrite/tratamento farmacológico , Colágeno , Relação Dose-Resposta a Droga , Camundongos , Modelos Moleculares , Estrutura Molecular , Relação Estrutura-Atividade , Fatores de TempoRESUMO
Divergence of substrate specificity within the context of a common structural framework represents an important mechanism by which new enzyme activity naturally evolves. We present enzymological and x-ray structural data for hamster chymase-2 (HAM2) that provides a detailed explanation for the unusual hydrolytic specificity of this rodent alpha-chymase. In enzymatic characterization, hamster chymase-1 (HAM1) showed typical chymase proteolytic activity. In contrast, HAM2 exhibited atypical substrate specificity, cleaving on the carboxyl side of the P1 substrate residues Ala and Val, characteristic of elastolytic rather than chymotryptic specificity. The 2.5-A resolution crystal structure of HAM2 complexed to the peptidyl inhibitor MeOSuc-Ala-Ala-Pro-Ala-chloromethylketone revealed a narrow and shallow S1 substrate binding pocket that accommodated only a small hydrophobic residue (e.g. Ala or Val). The different substrate specificities of HAM2 and HAM1 are explained by changes in four S1 substrate site residues (positions 189, 190, 216, and 226). Of these, Asn(189), Val(190), and Val(216) form an easily identifiable triplet in all known rodent alpha-chymases that can be used to predict elastolytic specificity for novel chymase-like sequences. Phylogenetic comparison defines guinea pig and rabbit chymases as the closest orthologs to rodent alpha-chymases.
Assuntos
Quimases/química , Quimases/metabolismo , Sequência de Aminoácidos , Animais , Baculoviridae/genética , Sítios de Ligação/genética , Linhagem Celular , Quimases/genética , Cricetinae , Cristalografia por Raios X , Eletroforese em Gel de Poliacrilamida , Vetores Genéticos/genética , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Filogenia , Ligação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Homologia de Sequência de Aminoácidos , Especificidade por SubstratoRESUMO
We have developed a novel series of potent and selective factor Xa inhibitors that employ a key 7-fluoroindazolyl moiety. The 7-fluoro group on the indazole scaffold replaces the carbonyl group of an amide that is found in previously reported factor Xa inhibitors. The structure of a factor Xa cocrystal containing 7-fluoroindazole 51a showed the 7-fluoro atom hydrogen-bonding with the N-H of Gly216 (2.9 A) in the peptide backbone. Thus, the 7-fluoroindazolyl moiety not only occupied the same space as the carbonyl group of an amide found in prior factor Xa inhibitors but also maintained a hydrogen bond interaction with the protein's beta-sheet domain. The structure-activity relationship for this series was consistent with this finding, as the factor Xa inhibitory potencies were about 60-fold greater (DeltaDelta G approximately 2.4 kcal/mol) for the 7-fluoroindazoles 25a and 25c versus the corresponding indazoles 25b and 25d. Highly convergent synthesis of these factor Xa inhibitors is also described.