RESUMO
The design and optimization of a novel isoxazole S(1) linker for renin inhibitor is described herein. This effort culminated in the identification of compound 18, an orally bioavailable, sub-nanomolar renin inhibitor even in the presence of human plasma. When compound 18 was found to inhibit CYP3A4 in a time dependent manner, two strategies were pursued that successfully delivered equipotent compounds with minimal TDI potential.
Assuntos
Anti-Hipertensivos/química , Desenho de Fármacos , Isoxazóis/química , Isoxazóis/síntese química , Renina/antagonistas & inibidores , Administração Oral , Animais , Anti-Hipertensivos/síntese química , Anti-Hipertensivos/farmacologia , Domínio Catalítico , Ativação Enzimática/efeitos dos fármacos , Humanos , Isoxazóis/farmacologia , Estrutura Molecular , Ratos , Relação Estrutura-AtividadeRESUMO
The discovery and SAR of a series of potent renin inhibitors possessing a novel 3,4-diarylpiperidine scaffold are described herein. The resulting compound 38 exhibit low nanomolar plasma renin IC(50), had a clean CYP 3A4 profile and displayed micromolar affinity for the hERG channel. Furthermore, it was found to be efficacious in the double transgenic rat hypertension model and show good to moderate oral bioavailability in two animal species.
Assuntos
Anti-Hipertensivos/química , Anti-Hipertensivos/uso terapêutico , Hipertensão/tratamento farmacológico , Piperidinas/química , Piperidinas/uso terapêutico , Renina/antagonistas & inibidores , Animais , Anti-Hipertensivos/farmacocinética , Anti-Hipertensivos/farmacologia , Disponibilidade Biológica , Citocromo P-450 CYP3A/metabolismo , Inibidores do Citocromo P-450 CYP3A , Cães , Canais de Potássio Éter-A-Go-Go/metabolismo , Humanos , Piperidinas/farmacocinética , Piperidinas/farmacologia , Ratos , Ratos Sprague-Dawley , Ratos Transgênicos , Renina/metabolismo , Relação Estrutura-AtividadeRESUMO
An oral bioavailability issue encountered during the course of lead optimization in the renin program is described herein. The low F(po) of pyridone analogs was shown to be caused by a combination of poor passive permeability and gut efflux transport. Substitution of pyridone ring for a more lipophilic moiety (logD>1.7) had minimal effect on rMdr1a transport but led to increased passive permeability (P(app)>10 × 10(-6) cm/s), which contributed to overwhelm gut transporters and increase rat F(po). LogD and in vitro passive permeability determination were found to be key in guiding SAR and improve oral exposure of renin inhibitors.
Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Permeabilidade da Membrana Celular/efeitos dos fármacos , Piperidinas/farmacologia , Renina/antagonistas & inibidores , Subfamília B de Transportador de Cassetes de Ligação de ATP/deficiência , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Administração Oral , Animais , Disponibilidade Biológica , Transporte Biológico/efeitos dos fármacos , Relação Dose-Resposta a Droga , Camundongos , Camundongos Knockout , Estrutura Molecular , Piperidinas/administração & dosagem , Piperidinas/química , Ratos , Renina/metabolismo , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
The design and optimization of a novel series of renin inhibitor is described herein. Strategically, by committing the necessary resources to the development of synthetic sequences and scaffolds that were most amenable for late stage structural diversification, even as the focus of the SAR campaign moved from one end of the molecule to another, highly potent renin inhibitors could be rapidly identified and profiled.
Assuntos
Álcoois/síntese química , Anti-Hipertensivos/síntese química , Anti-Hipertensivos/uso terapêutico , Desenho de Fármacos , Hipertensão/tratamento farmacológico , Piperidinas/síntese química , Renina/antagonistas & inibidores , Álcoois/química , Álcoois/uso terapêutico , Animais , Anti-Hipertensivos/química , Estrutura Molecular , Piperidinas/química , Piperidinas/uso terapêutico , Ratos , Renina/química , Relação Estrutura-AtividadeRESUMO
An SAR campaign aimed at decreasing the overall lipophilicity of renin inhibitors such as 1 is described herein. It was found that replacement of the northern appendage in 1 with an N-methyl pyridone and subsequent re-optimization of the benzyl amide handle afforded compounds with in vitro and in vivo profiles suitable for further profiling. An unexpected CV toxicity in dogs observed with compound 20 led to the employment of a time and resource sparing rodent model for in vivo screening of key compounds. This culminated in the identification of compound 31 as an optimized renin inhibitor.
Assuntos
Desenho de Fármacos , Hipertensão/tratamento farmacológico , Piperidinas/síntese química , Piridonas/síntese química , Renina/antagonistas & inibidores , Animais , Cães , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Concentração Inibidora 50 , Modelos Moleculares , Estrutura Molecular , Piperidinas/química , Piperidinas/uso terapêutico , Piridonas/química , Piridonas/uso terapêutico , Ratos , Relação Estrutura-AtividadeRESUMO
The incorporation of a carboxylic acid within in a series of 3-amido-4-aryl substituted piperidines (represented by general structure 32) led to the discovery of potent, zwitterionic, renin inhibitors with improved off-target profiles (CYP3A4 time-dependent inhibition and hERG affinity) relative to analogous non-zwitterionic inhibitors of the past (i.e., 3). Strategies to address the oral absorption of these zwitterions are also discussed within.
Assuntos
Inibidores de Proteases/síntese química , Renina/antagonistas & inibidores , Administração Oral , Animais , Domínio Catalítico , Simulação por Computador , Cães , Avaliação Pré-Clínica de Medicamentos , Humanos , Piperidinas/síntese química , Piperidinas/química , Piperidinas/farmacocinética , Inibidores de Proteases/química , Inibidores de Proteases/farmacocinética , Ratos , Ratos Sprague-Dawley , Renina/metabolismo , Relação Estrutura-AtividadeRESUMO
Time-dependent inhibitors of CYPs have the potential to perpetrate drug-drug interactions in the clinical setting. After finding that several leading compounds in a novel series of substituted amino propanamide renin inhibitors inactivated CYP3A4 in an NADPH-dependent and time-dependent manner, a search to identify the cause of this liability was initiated. Extensive SAR revealed that the amide bridge present in compound 1 as a possible culprit. Through the installation of a metabolic soft spot distal to this moiety, potent renin inhibitors with improved CYP profile were identified.
Assuntos
Inibidores do Citocromo P-450 CYP3A , Inibidores Enzimáticos/farmacologia , Propionatos/química , Renina/antagonistas & inibidores , Amidas/química , Citocromo P-450 CYP3A , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacocinética , Humanos , Concentração Inibidora 50 , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/enzimologiaRESUMO
The discovery and SAR of a new series of substituted amino propanamide renin inhibitors are herein described. This work has led to the preparation of compounds with in vitro and in vivo profiles suitable for further development. Specifically, challenges pertaining to oral bioavailability, covalent binding and time-dependent CYP 3A4 inhibition were overcome thereby culminating in the identification of compound 50 as an optimized renin inhibitor with good efficacy in the hypertensive double-transgenic rat model.
Assuntos
Anti-Hipertensivos/química , Anti-Hipertensivos/uso terapêutico , Hipertensão/tratamento farmacológico , Renina/antagonistas & inibidores , Renina/metabolismo , Animais , Anti-Hipertensivos/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Cristalografia por Raios X , Cães , Humanos , Modelos Moleculares , Ligação Proteica , Ratos , Ratos Sprague-Dawley , Renina/química , Relação Estrutura-AtividadeRESUMO
The discovery of novel and selective inhibitors of human 5-lipoxygenase (5-LO) is described. These compounds are potent, orally bioavailable, and active at inhibiting leukotriene biosynthesis in vivo in a dog PK/PD model. A major focus of the optimization process was to reduce affinity for the human ether-a-go-go gene potassium channel while preserving inhibitory potency on 5-LO. These efforts led to the identification of inhibitor (S)-16 (MK-0633, setileuton), a compound selected for clinical development for the treatment of respiratory diseases.
RESUMO
Caspase-3 is a cysteinyl protease that mediates apoptotic cell death. Its inhibition may have an important impact on the treatment of several degenerative diseases. Here we report the synthesis of reversible inhibitors via a solid-support palladium-catalyzed amination of 3-bromopyrazinones and the discovery of a pan-caspase reversible inhibitor.
Assuntos
Inibidores de Caspase , Paládio/química , Inibidores de Proteases/síntese química , Inibidores de Proteases/farmacologia , Pirazinas/síntese química , Pirazinas/farmacologia , Aminação , Catálise , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Humanos , Indicadores e Reagentes , Espectrometria de Massas , Proteínas Recombinantes/química , Relação Estrutura-AtividadeRESUMO
Leukotriene biosynthesis inhibitors have potential as therapeutic agents for asthma and inflammatory diseases. A novel series of substituted coumarin derivatives has been synthesized and the structure-activity relationship was evaluated with respect to their ability to inhibit the formation of leukotrienes via the human 5-lipoxygenase enzyme.
Assuntos
Cumarínicos/farmacologia , Inibidores Enzimáticos/farmacologia , Inibidores de Lipoxigenase , Cumarínicos/síntese química , Cumarínicos/química , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
A rapid and quantitative method for measuring the activity and fractional inhibition of enzymes within their natural cellular environment remains an unmet need in drug discovery. We describe the use of a nonradioactive quantitative enzyme-linked immunosorbent assay (ELISA) for measuring intracellular caspase activity that is amenable to robotic automation. The ELISA specifically detects active-caspase-3 and was used to correlate the in-cell activity of caspase-3 with the progress of caspase-3-mediated events under varying concentrations of caspase-3 inhibitors in NT2 cells. We examined the cleavage of endogenous substrates (poly(ADP-ribose)polymerase and alphaII-spectrin), the extent of DNA fragmentation, and the autocatalytic removal of the caspase-3 prodomain as markers of caspase-3 activity. To impart inhibition of the downstream markers, a greater level of caspase-3 inhibition was required. Although the functional markers were found not to accurately predict intracellular caspase-3 activity, we found that the inhibition of intracellular caspase-3 was highly correlated (R(2) = 0.96) to the inhibition of DNA fragmentation. Also, by comparing the potency of the different inhibitors against the intracellular enzyme versus the purified enzyme, the effects of inhibitor functional groups on whole-cell activity were addressed.
Assuntos
Inibidores de Caspase , Caspases/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Apoptose , Biotina/análogos & derivados , Biotina/farmacologia , Caspase 3 , Linhagem Celular , Cumarínicos/farmacologia , Inibidores de Cisteína Proteinase/farmacologia , Fragmentação do DNA/efeitos dos fármacos , Dipeptídeos/farmacologia , Eletroforese em Gel de Poliacrilamida , Humanos , Concentração Inibidora 50 , Oligopeptídeos/farmacologia , Inibidores de Poli(ADP-Ribose) Polimerases , Sensibilidade e Especificidade , Espectrina/antagonistas & inibidoresRESUMO
Caspase 3 is a cysteinyl protease that mediates apoptotic cell death. Its inhibition may have an important impact in the treatment of several degenerative diseases. The P1 aspartic acid residue is a required element of recognition for this enzyme that was maintained constant along with the adjacent natural valine as the P2 group. The thiobenzylmethylketone warhead on the aspartate was conveniently handled through solid-phase synthesis allowing modification in the P3 region that eventually led to simpler derivatives with increased potency against caspase 3. The key to such an effect is the introduction of hydroxyl group alpha to the P3 carbonyl.
Assuntos
Inibidores de Caspase , Dipeptídeos/síntese química , Cetonas/síntese química , Ácido Aspártico , Caspase 3 , Técnicas de Química Combinatória , Dipeptídeos/farmacologia , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Humanos , Ligação de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Concentração Inibidora 50 , Cetonas/farmacologia , Proteínas Recombinantes , Relação Estrutura-Atividade , ValinaRESUMO
A series of potent and selective inhibitors of the inducible microsomal PGE2 synthase (mPGES-1) has been developed based on the indole FLAP inhibitor MK-886. Compounds 23 and 30 inhibit mPGES-1 with potencies in the low nanomolar range and with selectivities of at least 100-fold compared to their inhibition of mPGES-2, thromboxane synthase and binding affinity to FLAP. They also block the production of PGE2 in cell based assays but with a decreased potency and more limited selectivity compared to the enzyme assays.
Assuntos
Inibidores de Ciclo-Oxigenase/síntese química , Inibidores de Ciclo-Oxigenase/farmacologia , Indóis/síntese química , Indóis/farmacologia , Oxirredutases Intramoleculares/antagonistas & inibidores , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Inibidores de Ciclo-Oxigenase/química , Humanos , Indóis/química , Microssomos/enzimologia , Prostaglandina-E Sintases , Relação Estrutura-AtividadeRESUMO
Caspase-3-deficient mice of the 129S1/SvImJ (129) strain show severe brain development defects resulting in brain overgrowth and perinatal lethality, whereas on the C57BL/6J (B6) background, these mice develop normally. We therefore sought to identify the strain-dependent ameliorating gene. We biochemically isolated caspase-7 from B6-caspase-3-null (Casp3-/-) tissues as being the enzyme with caspase-3-like properties and capability of performing a caspase-3 surrogate function, apoptotic DNA fragmentation. Moreover, we show that, in contrast to the human enzymes, mouse caspase-7 is as efficient as caspase-3 at cleaving and thus inactivating ICAD (inhibitor of caspase-activated DNase), the inhibitor of apoptotic DNA fragmentation. Low levels of caspase-7 expression and activation correlate with lack of DNA fragmentation in 129-Casp3-/- apoptotic precursor neurons, whereas B6-Casp3-/- cells, which can fragment their DNA, show higher levels of caspase-7 expression and activation. The amount of caspase-7 activation in apoptotic precursor neurons is independent of the presence of caspase-3. Together, our findings demonstrate for the first time a strong correlation between caspase-7 activity, normal brain development, and apoptotic DNA fragmentation in Casp3-/- mice.
Assuntos
Encéfalo/anormalidades , Caspases/deficiência , Caspases/fisiologia , Cisteína Endopeptidases/fisiologia , Animais , Apoptose/genética , Proteínas Reguladoras de Apoptose , Encéfalo/embriologia , Caspase 3 , Caspase 7 , Inibidores de Caspase , Caspases/genética , Caspases/metabolismo , Cisteína Endopeptidases/metabolismo , Fragmentação do DNA/genética , Ativação Enzimática/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Proteínas/metabolismo , Especificidade da EspécieRESUMO
Caspases are cysteine proteases that specifically cleave Asp-Xxx bonds. They are key agents in inflammation and apoptosis and are attractive targets for therapy against inflammation, neurodegeneration, ischemia, and cancer. Many caspase structures are known, but most involve either peptide or protein inhibitors, unattractive candidates for drug development. We present seven crystal structures of inhibited caspase-3 that illustrate several approaches to reducing the peptidyl characteristics of the inhibitors while maintaining their potency and selectivity. The inhibitors reduce the peptidyl nature of inhibitors while preserving binding potency by (1). exploiting a hydrophobic binding site C-terminal to the cleavage site, (2). replacing the negatively charged aspartyl residue at P4 with neutral groups, and (3). using a peptidomimetic 5,6,7-tricyclic system or a pyrazinone at P2-P3. In addition, we have found that two nicotinic acid aldehydes induce a significant conformational change in the S2 and S3 subsites of caspase-3, revealing an unexpected binding mode. These results advance the search for caspase-directed drugs by revealing how unacceptable molecular features can be removed without loss of potency.
Assuntos
Caspases/química , Inibidores Enzimáticos/química , Peptídeos/química , Caspase 3 , Inibidores de Caspase , Cristalografia por Raios X , Compostos Heterocíclicos com 3 Anéis/química , Ligantes , Modelos Moleculares , Mimetismo Molecular , Estrutura Molecular , Niacina/análogos & derivados , Niacina/química , Oligopeptídeos/química , Conformação Proteica , Pirazinas/química , Relação Estrutura-AtividadeRESUMO
A robust method for the solid phase synthesis of a series of selective caspase-3 peptide inhibitors is described. The inhibitors can be obtained after cleavage from the solid support without further purification.
Assuntos
Inibidores de Caspase , Oligopeptídeos/síntese química , Oligopeptídeos/farmacologia , Caspase 3 , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Química Combinatória , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Humanos , Concentração Inibidora 50RESUMO
The discovery of a series of potent, selective and reversible dipeptidyl caspase-3 inhibitors are reported. The iterative discovery process of using combinatorial chemistry, parallel synthesis, moleculare modelling and structural biology will be discussed.
Assuntos
Ácido Aspártico/química , Inibidores de Caspase , Dipeptídeos , Inibidores Enzimáticos , Cetonas , Sítios de Ligação , Caspase 3 , Células Cultivadas , Técnicas de Química Combinatória , Dipeptídeos/síntese química , Dipeptídeos/farmacologia , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Humanos , Cetonas/síntese química , Cetonas/farmacologia , Modelos Moleculares , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
Caspase-3 is a cysteinyl protease that mediates apoptotic cell death. Its inhibition may have an important impact in the treatment of several degenerative diseases. Since P(1) aspartic acid is a required element of recognition for this enzyme, a library of capped aspartyl aldehydes was synthesized using solid-phase chemistry. The 5-bromonicotinamide derivative of the aspartic acid aldehyde was identified to be an inhibitor of caspase-3. Substitution at the 5-position of the pyridine ring and conversion of the aldehyde to ketones led to a series of potent inhibitors of caspase-3.
Assuntos
Inibidores de Caspase , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Cetonas/síntese química , Cetonas/farmacologia , Aldeídos/síntese química , Aldeídos/farmacologia , Ácido Aspártico/química , Caspase 3 , Linhagem Celular , Fragmentação do DNA/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Humanos , Indicadores e Reagentes , Biblioteca de Peptídeos , Piridinas/químicaRESUMO
The molecular mechanisms mediating degeneration of midbrain dopamine neurons in Parkinson's disease (PD) are poorly understood. Here, we provide evidence to support a role for the involvement of the calcium-dependent proteases, calpains, in the loss of dopamine neurons in a mouse model of PD. We show that administration of N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) evokes an increase in calpain-mediated proteolysis in nigral dopamine neurons in vivo. Inhibition of calpain proteolysis using either a calpain inhibitor (MDL-28170) or adenovirus-mediated overexpression of the endogenous calpain inhibitor protein, calpastatin, significantly attenuated MPTP-induced loss of nigral dopamine neurons. Commensurate with this neuroprotection, MPTP-induced locomotor deficits were abolished, and markers of striatal postsynaptic activity were normalized in calpain inhibitor-treated mice. However, behavioral improvements in MPTP-treated, calpain inhibited mice did not correlate with restored levels of striatal dopamine. These results suggest that protection against nigral neuron degeneration in PD may be sufficient to facilitate normalized locomotor activity without necessitating striatal reinnervation. Immunohistochemical analyses of postmortem midbrain tissues from human PD cases also displayed evidence of increased calpain-related proteolytic activity that was not evident in age-matched control subjects. Taken together, our findings provide a potentially novel correlation between calpain proteolytic activity in an MPTP model of PD and the etiology of neuronal loss in PD in humans.