RESUMO
Through regulation of the epigenome, the bromodomain and extra terminal (BET) family of proteins represent important therapeutic targets for the treatment of human disease. Through mimicking the endogenous N-acetyl-lysine group and disrupting the protein-protein interaction between histone tails and the bromodomain, several small molecule pan-BET inhibitors have progressed to oncology clinical trials. This work describes the medicinal chemistry strategy and execution to deliver an orally bioavailable tetrahydroquinoline (THQ) pan-BET candidate. Critical to the success of this endeavor was a potency agnostic analysis of a data set of 1999 THQ BET inhibitors within the GSK collection which enabled identification of appropriate lipophilicity space to deliver compounds with a higher probability of desired oral candidate quality properties. SAR knowledge was leveraged via Free-Wilson analysis within this design space to identify a small group of targets which ultimately delivered I-BET567 (27), a pan-BET candidate inhibitor that demonstrated efficacy in mouse models of oncology and inflammation.
Assuntos
Aminoquinolinas/química , Desenho de Fármacos , Proteínas/metabolismo , Administração Oral , Aminoquinolinas/metabolismo , Aminoquinolinas/farmacocinética , Aminoquinolinas/uso terapêutico , Animais , Benzoatos/química , Benzoatos/metabolismo , Sítios de Ligação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cristalografia por Raios X , Cães , Meia-Vida , Humanos , Masculino , Camundongos , Conformação Molecular , Simulação de Dinâmica Molecular , Neoplasias/tratamento farmacológico , Proteínas/antagonistas & inibidores , Ratos , Relação Estrutura-AtividadeRESUMO
The two tandem bromodomains of the BET (bromodomain and extraterminal domain) proteins enable chromatin binding to facilitate transcription. Drugs that inhibit both bromodomains equally have shown efficacy in certain malignant and inflammatory conditions. To explore the individual functional contributions of the first (BD1) and second (BD2) bromodomains in biology and therapy, we developed selective BD1 and BD2 inhibitors. We found that steady-state gene expression primarily requires BD1, whereas the rapid increase of gene expression induced by inflammatory stimuli requires both BD1 and BD2 of all BET proteins. BD1 inhibitors phenocopied the effects of pan-BET inhibitors in cancer models, whereas BD2 inhibitors were predominantly effective in models of inflammatory and autoimmune disease. These insights into the differential requirement of BD1 and BD2 for the maintenance and induction of gene expression may guide future BET-targeted therapies.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Antineoplásicos/farmacologia , Proteínas de Ciclo Celular/antagonistas & inibidores , Histona Acetiltransferases/antagonistas & inibidores , Fatores Imunológicos/farmacologia , Terapia de Alvo Molecular , Fatores de Transcrição/antagonistas & inibidores , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/uso terapêutico , Antineoplásicos/uso terapêutico , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Descoberta de Drogas , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Histona Acetiltransferases/química , Histona Acetiltransferases/genética , Humanos , Doenças do Sistema Imunitário/tratamento farmacológico , Fatores Imunológicos/química , Fatores Imunológicos/uso terapêutico , Inflamação/tratamento farmacológico , Neoplasias/tratamento farmacológico , Domínios Proteicos/efeitos dos fármacos , Fatores de Transcrição/química , Fatores de Transcrição/genéticaRESUMO
OBJECTIVE: Caffeine reduces toxic Ca2+ signals in pancreatic acinar cells via inhibition of inositol 1,4,5-trisphosphate receptor (IP3R)-mediated signalling, but effects of other xanthines have not been evaluated, nor effects of xanthines on experimental acute pancreatitis (AP). We have determined effects of caffeine and its xanthine metabolites on pancreatic acinar IP3R-mediated Ca2+ signalling and experimental AP. DESIGN: Isolated pancreatic acinar cells were exposed to secretagogues, uncaged IP3 or toxins that induce AP and effects of xanthines, non-xanthine phosphodiesterase (PDE) inhibitors and cyclic adenosine monophosphate and cyclic guanosine monophosphate (cAMP/cGMP) determined. The intracellular cytosolic calcium concentration ([Ca2+]C), mitochondrial depolarisation and necrosis were assessed by confocal microscopy. Effects of xanthines were evaluated in caerulein-induced AP (CER-AP), taurolithocholic acid 3-sulfate-induced AP (TLCS-AP) or palmitoleic acid plus ethanol-induced AP (fatty acid ethyl ester AP (FAEE-AP)). Serum xanthines were measured by liquid chromatography-mass spectrometry. RESULTS: Caffeine, dimethylxanthines and non-xanthine PDE inhibitors blocked IP3-mediated Ca2+ oscillations, while monomethylxanthines had little effect. Caffeine and dimethylxanthines inhibited uncaged IP3-induced Ca2+ rises, toxin-induced Ca2+ release, mitochondrial depolarisation and necrotic cell death pathway activation; cAMP/cGMP did not inhibit toxin-induced Ca2+ rises. Caffeine significantly ameliorated CER-AP with most effect at 25â mg/kg (seven injections hourly); paraxanthine or theophylline did not. Caffeine at 25â mg/kg significantly ameliorated TLCS-AP and FAEE-AP. Mean total serum levels of dimethylxanthines and trimethylxanthines peaked at >2â mM with 25â mg/kg caffeine but at <100â µM with 25â mg/kg paraxanthine or theophylline. CONCLUSIONS: Caffeine and its dimethylxanthine metabolites reduced pathological IP3R-mediated pancreatic acinar Ca2+ signals but only caffeine ameliorated experimental AP. Caffeine is a suitable starting point for medicinal chemistry.
Assuntos
Células Acinares/efeitos dos fármacos , Cafeína/farmacologia , Cálcio/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/antagonistas & inibidores , Pâncreas/patologia , Pancreatite/prevenção & controle , Inibidores de Fosfodiesterase/farmacologia , Células Acinares/metabolismo , Animais , Cafeína/uso terapêutico , Morte Celular/efeitos dos fármacos , Células Cultivadas , Ceruletídeo , AMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Citosol/metabolismo , Etanol , Ácidos Graxos Monoinsaturados , Inositol 1,4,5-Trifosfato/metabolismo , Masculino , Camundongos , Microscopia Confocal , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Necrose/diagnóstico por imagem , Pancreatite/sangue , Pancreatite/induzido quimicamente , Inibidores de Fosfodiesterase/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Ácido Taurolitocólico/análogos & derivados , Xantinas/sangue , Xantinas/farmacologiaRESUMO
The bromodomain and extraterminal (BET) protein BRD2-4 inhibitors hold therapeutic promise in preclinical models of hematologic malignancies. However, translation of these data to molecules suitable for clinical development has yet to be accomplished. Herein we expand the mechanistic understanding of BET inhibitors in multiple myeloma by using the chemical probe molecule I-BET151. I-BET151 induces apoptosis and exerts strong antiproliferative effect in vitro and in vivo. This is associated with contrasting effects on oncogenic MYC and HEXIM1, an inhibitor of the transcriptional activator P-TEFb. I-BET151 causes transcriptional repression of MYC and MYC-dependent programs by abrogating recruitment to the chromatin of the P-TEFb component CDK9 in a BRD2-4-dependent manner. In contrast, transcriptional upregulation of HEXIM1 is BRD2-4 independent. Finally, preclinical studies show that I-BET762 has a favorable pharmacologic profile as an oral agent and that it inhibits myeloma cell proliferation, resulting in survival advantage in a systemic myeloma xenograft model. These data provide a strong rationale for extending the clinical testing of the novel antimyeloma agent I-BET762 and reveal insights into biologic pathways required for myeloma cell proliferation.
Assuntos
Antineoplásicos/uso terapêutico , Benzodiazepinas/uso terapêutico , Compostos Heterocíclicos de 4 ou mais Anéis/uso terapêutico , Mieloma Múltiplo/tratamento farmacológico , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzodiazepinas/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Humanos , Camundongos , Mieloma Múltiplo/genética , Mieloma Múltiplo/patologia , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas de Ligação a RNA/genética , Fatores de Transcrição , Ativação Transcricional/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
The bromo and extra C-terminal domain (BET) family of bromodomains are involved in binding epigenetic marks on histone proteins, more specifically acetylated lysine residues. This paper describes the discovery and structure-activity relationships (SAR) of potent benzodiazepine inhibitors that disrupt the function of the BET family of bromodomains (BRD2, BRD3, and BRD4). This work has yielded a potent, selective compound I-BET762 that is now under evaluation in a phase I/II clinical trial for nuclear protein in testis (NUT) midline carcinoma and other cancers.