RESUMO
With resistance to current agricultural fungicides rising, a great need has emerged for new antifungals with unexploited targets. In response, we report a novel series of diazaborines with potent activity against representative fungal plant pathogens. To identify their mode of action, we selected for resistant isolates using the model fungus Saccharomyces cerevisiae. Whole-genome sequencing of independent diazaborine-resistant lineages identified a recurring mutation in ERG25, which encodes a C-4 methyl sterol oxidase required for ergosterol biosynthesis in fungi. Haploinsufficiency and allele-swap experiments provided additional genetic evidence for Erg25 as the most biologically relevant target of our diazaborines. Confirming Erg25 as putative target, sterol profiling of compound-treated yeast revealed marked accumulation of the Erg25 substrate, 4,4-dimethylzymosterol and depletion of both its immediate product, zymosterol, as well as ergosterol. Encouraged by these mechanistic insights, the potential utility of targeting Erg25 with a diazaborine was demonstrated in soybean-rust and grape-rot models of fungal plant disease.
Assuntos
Ergosterol , Oxigenases de Função Mista , Antifúngicos/farmacologia , Oxigenases de Função Mista/genética , Saccharomyces cerevisiae/genética , EsteróisRESUMO
Several boron-containing small molecules have been approved by the US FDA to treat human diseases. We explored potential applications of boron-containing compounds in modern agriculture by pursuing multiple research and development programs. Here, we report a novel series of multi-substitution benzoxaboroles (1-36), a compound class that we recently reported as targeting geranylgeranyl transferase I (GGTase I) and thereby inhibiting protein prenylation (Kim et al., 2020). These compounds were designed, synthesized, and tested against the agriculturally important fungal pathogens Mycosphaerella fijiensis and Colletotrichum sublineolum in a structure-activity relationship (SAR) study. Compounds 13, 28, 30, 34 and 36 were identified as active leads with excellent antifungal MIC95 values in the range of 1.56-3.13 ppm against M. fijiensis and 0.78-3.13 ppm against C. sublineolum.
Assuntos
Antifúngicos/farmacologia , Ascomicetos/efeitos dos fármacos , Compostos de Boro/farmacologia , Colletotrichum/efeitos dos fármacos , Fungicidas Industriais/farmacologia , Agricultura , Alquil e Aril Transferases/antagonistas & inibidores , Alquil e Aril Transferases/metabolismo , Antifúngicos/síntese química , Antifúngicos/química , Ascomicetos/metabolismo , Compostos de Boro/síntese química , Compostos de Boro/química , Colletotrichum/metabolismo , Relação Dose-Resposta a Droga , Fungicidas Industriais/síntese química , Fungicidas Industriais/química , Testes de Sensibilidade Microbiana , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
Fungal pathogens pose an increasing threat to global food security through devastating effects on staple crops and contamination of food supplies with carcinogenic toxins. Widespread deployment of agricultural fungicides has increased crop yields but is driving increasingly frequent resistance to available agents and creating environmental reservoirs of drug-resistant fungi that can also infect susceptible human populations. To uncover non-cross-resistant modes of antifungal action, we leveraged the unique chemical properties of boron chemistry to synthesize novel 6-thiocarbamate benzoxaboroles with broad spectrum activity against diverse fungal plant pathogens. Through whole genome sequencing of Saccharomyces cerevisiae isolates selected for stable resistance to these compounds, we identified mutations in the protein prenylation-related genes, CDC43 and ERG20. Allele-swapping experiments confirmed that point mutations in CDC43, which encodes an essential catalytic subunit within geranylgeranyl transferase I (GGTase I) complex, were sufficient to confer resistance to the benzoxaboroles. Mutations in ERG20, which encodes an upstream farnesyl pyrophosphate synthase in the geranylgeranylation pathway, also conferred resistance. Consistent with impairment of protein prenylation, the compounds disrupted membrane localization of the classical geranylgeranylation substrate Cdc42. Guided by molecular docking predictions, which favored Cdc43 as the most likely direct target, we overexpressed and purified functional GGTase I complex to demonstrate direct binding of benzoxaboroles to it and concentration-dependent inhibition of its transferase activity. Further development of the boron-containing scaffold described here offers a promising path to the development of GGTase I inhibitors as a mechanistically distinct broad spectrum fungicide class with reduced potential for cross-resistance to antifungals in current use.
Assuntos
Antifúngicos/farmacologia , Compostos de Boro/farmacologia , Prenilação de Proteína/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/metabolismo , Tiocarbamatos/farmacologia , Alquil e Aril Transferases/antagonistas & inibidores , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Antifúngicos/síntese química , Antifúngicos/metabolismo , Compostos de Boro/síntese química , Compostos de Boro/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Membrana Celular/efeitos dos fármacos , Dimetilaliltranstransferase/genética , Dimetilaliltranstransferase/metabolismo , Farmacorresistência Fúngica/genética , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Fungos/efeitos dos fármacos , Fungos/genética , Simulação de Acoplamento Molecular , Mutação Puntual , Ligação Proteica , Proteínas de Saccharomyces cerevisiae/genética , Tiocarbamatos/síntese química , Tiocarbamatos/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismoRESUMO
AN12855 is a direct, cofactor-independent inhibitor of InhA in Mycobacterium tuberculosis In the C3HeB/FeJ mouse model with caseous necrotic lung lesions, AN12855 proved efficacious with a significantly lower resistance frequency than isoniazid. AN12855 drug levels were better retained in necrotic lesions and caseum where the majority of hard to treat, extracellular bacilli reside. Owing to these combined attributes, AN12855 represents a promising alternative to the frontline antituberculosis agent isoniazid.
Assuntos
Antituberculosos/farmacologia , Compostos Aza/farmacologia , Compostos de Boro/farmacologia , Hidrocarbonetos Fluorados/farmacologia , Inibinas/antagonistas & inibidores , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose Pulmonar/tratamento farmacológico , Animais , Carga Bacteriana/efeitos dos fármacos , Modelos Animais de Doenças , Desenvolvimento de Medicamentos , Feminino , Isoniazida/farmacologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C3H , Testes de Sensibilidade Microbiana , Tuberculose Pulmonar/microbiologiaRESUMO
New antitubercular agents are needed to combat the spread of multidrug- and extensively drug-resistant strains of Mycobacterium tuberculosis. The frontline antitubercular drug isoniazid (INH) targets the mycobacterial enoyl-ACP reductase, InhA. Resistance to INH is predominantly through mutations affecting the prodrug-activating enzyme KatG. Here, we report the identification of the diazaborines as a new class of direct InhA inhibitors. The lead compound, AN12855, exhibited in vitro bactericidal activity against replicating bacteria and was active against several drug-resistant clinical isolates. Biophysical and structural investigations revealed that AN12855 binds to and inhibits the substrate-binding site of InhA in a cofactor-independent manner. AN12855 showed good drug exposure after i.v. and oral delivery, with 53% oral bioavailability. Delivered orally, AN12855 exhibited dose-dependent efficacy in both an acute and chronic murine model of tuberculosis infection that was comparable with INH. Combined, AN12855 is a promising candidate for the development of new antitubercular agents.
RESUMO
We identified a series of novel 7-phenyl benzoxaborole compounds with activity against Mycobacterium tuberculosis. Compounds had a range of activity with inhibitory concentrations (IC90) as low as 5.1 µM and no cytotoxicity against eukaryotic cells (IC50 > 50 µM). Compounds were active against intracellular mycobacteria cultured in THP-1 macrophages. We isolated and characterized resistant mutants with mutations in NADH dehydrogenase (Ndh) or the regulatory protein Mce3R. Mutations suggest that Ndh may be the target of this series.
Assuntos
Compostos de Boro/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Compostos de Boro/química , Compostos de Boro/toxicidade , Compostos Bicíclicos Heterocíclicos com Pontes/química , Compostos Bicíclicos Heterocíclicos com Pontes/toxicidade , Relação Dose-Resposta a Droga , Farmacorresistência Bacteriana , Humanos , Testes de Sensibilidade Microbiana , Mutação , Mycobacterium tuberculosis/enzimologia , Mycobacterium tuberculosis/genética , NADH Desidrogenase/antagonistas & inibidores , NADH Desidrogenase/genética , NADH Desidrogenase/metabolismo , Células THP-1RESUMO
There is an urgent need to develop new and safer antitubercular agents that possess a novel mode of action. We synthesized and evaluated a novel series of 3-aminomethyl 4-halogen benzoxaboroles as Mycobacterium tuberculosis (Mtb) leucyl-tRNA synthetase (LeuRS) inhibitors. A number of Mtb LeuRS inhibitors were identified that demonstrated good antitubercular activity with high selectivity over human mitochondrial and cytoplasmic LeuRS. Further evaluation of these Mtb LeuRS inhibitors by in vivo pharmacokinetics (PK) and murine tuberculosis (TB) efficacy models led to the discovery of GSK3036656 (abbreviated as GSK656). This molecule shows potent inhibition of Mtb LeuRS (IC50 = 0.20 µM) and in vitro antitubercular activity (Mtb H37Rv MIC = 0.08 µM). Additionally, it is highly selective for the Mtb LeuRS enzyme with IC50 of >300 µM and 132 µM for human mitochondrial LeuRS and human cytoplasmic LeuRS, respectively. In addition, it exhibits remarkable PK profiles and efficacy against Mtb in mouse TB infection models with superior tolerability over initial leads. This compound has been progressed to clinical development for the treatment of tuberculosis.
Assuntos
Antituberculosos/síntese química , Antituberculosos/farmacologia , Compostos de Boro/farmacologia , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Compostos Heterocíclicos com 2 Anéis/farmacologia , Leucina-tRNA Ligase/antagonistas & inibidores , Mycobacterium tuberculosis/efeitos dos fármacos , Animais , Antituberculosos/farmacocinética , Compostos de Boro/síntese química , Compostos de Boro/farmacocinética , Descoberta de Drogas , Inibidores Enzimáticos/farmacocinética , Feminino , Compostos Heterocíclicos com 2 Anéis/síntese química , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis/enzimologia , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
Benzoxaboroles are effective against bacterial, fungal and protozoan pathogens. We report potent activity of the benzoxaborole AN3661 against Plasmodium falciparum laboratory-adapted strains (mean IC50 32 nM), Ugandan field isolates (mean ex vivo IC50 64 nM), and murine P. berghei and P. falciparum infections (day 4 ED90 0.34 and 0.57 mg kg-1, respectively). Multiple P. falciparum lines selected in vitro for resistance to AN3661 harboured point mutations in pfcpsf3, which encodes a homologue of mammalian cleavage and polyadenylation specificity factor subunit 3 (CPSF-73 or CPSF3). CRISPR-Cas9-mediated introduction of pfcpsf3 mutations into parental lines recapitulated AN3661 resistance. PfCPSF3 homology models placed these mutations in the active site, where AN3661 is predicted to bind. Transcripts for three trophozoite-expressed genes were lost in AN3661-treated trophozoites, which was not observed in parasites selected or engineered for AN3661 resistance. Our results identify the pre-mRNA processing factor PfCPSF3 as a promising antimalarial drug target.
Assuntos
Antimaláricos/farmacologia , Compostos de Boro/farmacologia , Fator de Especificidade de Clivagem e Poliadenilação/química , Plasmodium falciparum/efeitos dos fármacos , Proteínas de Protozoários/química , RNA Mensageiro/genética , Sequência de Aminoácidos , Animais , Antimaláricos/síntese química , Compostos de Boro/síntese química , Sistemas CRISPR-Cas , Domínio Catalítico , Fator de Especificidade de Clivagem e Poliadenilação/antagonistas & inibidores , Fator de Especificidade de Clivagem e Poliadenilação/genética , Fator de Especificidade de Clivagem e Poliadenilação/metabolismo , Resistência a Medicamentos/genética , Eritrócitos/efeitos dos fármacos , Eritrócitos/parasitologia , Edição de Genes/métodos , Humanos , Malária/tratamento farmacológico , Malária/parasitologia , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Camundongos , Simulação de Acoplamento Molecular , Mutação , Plasmodium berghei/efeitos dos fármacos , Plasmodium berghei/genética , Plasmodium berghei/crescimento & desenvolvimento , Plasmodium berghei/metabolismo , Plasmodium falciparum/genética , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium falciparum/metabolismo , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Proteínas de Protozoários/antagonistas & inibidores , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Trofozoítos/efeitos dos fármacos , Trofozoítos/genética , Trofozoítos/crescimento & desenvolvimento , Trofozoítos/metabolismoRESUMO
The recent development and spread of extensively drug-resistant and totally drug-resistant resistant (TDR) strains of Mycobacterium tuberculosis highlight the need for new antitubercular drugs. Protein synthesis inhibitors have played an important role in the treatment of tuberculosis (TB) starting with the inclusion of streptomycin in the first combination therapies. Although parenteral aminoglycosides are a key component of therapy for multidrug-resistant TB, the oxazolidinone linezolid is the only orally available protein synthesis inhibitor that is effective against TB. Here, we show that small-molecule inhibitors of aminoacyl-tRNA synthetases (AARSs), which are known to be excellent antibacterial protein synthesis targets, are orally bioavailable and effective against M. tuberculosis in TB mouse infection models. We applied the oxaborole tRNA-trapping (OBORT) mechanism, which was first developed to target fungal cytoplasmic leucyl-tRNA synthetase (LeuRS), to M. tuberculosis LeuRS. X-ray crystallography was used to guide the design of LeuRS inhibitors that have good biochemical potency and excellent whole-cell activity against M. tuberculosis Importantly, their good oral bioavailability translates into in vivo efficacy in both the acute and chronic mouse models of TB with potency comparable to that of the frontline drug isoniazid.
Assuntos
Antituberculosos/farmacologia , Leucina-tRNA Ligase/antagonistas & inibidores , Mycobacterium tuberculosis/efeitos dos fármacos , Inibidores da Síntese de Proteínas/farmacologia , Administração Oral , Animais , Antituberculosos/administração & dosagem , Antituberculosos/química , Antituberculosos/farmacocinética , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos/métodos , Feminino , Humanos , Leucina-tRNA Ligase/química , Leucina-tRNA Ligase/genética , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Testes de Sensibilidade Microbiana , Mycobacterium smegmatis/efeitos dos fármacos , Mycobacterium smegmatis/genética , Mycobacterium tuberculosis/genética , Inibidores da Síntese de Proteínas/administração & dosagem , Inibidores da Síntese de Proteínas/química , Inibidores da Síntese de Proteínas/farmacocinética , Relação Estrutura-Atividade , Tuberculose/tratamento farmacológico , Células VeroRESUMO
Psoriasis and atopic dermatitis are skin diseases affecting millions of patients. Here, we characterize benzoxaborole phosphodiesterase (PDE)-4 inhibitors, a new topical class that has demonstrated therapeutic benefit for psoriasis and atopic dermatitis in phase 2 or phase 3 studies. Crisaborole [AN2728, 4-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)benzonitrile], compd2 [2-ethoxy-6-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)nicotinonitrile], compd3 [6-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)-2-(2-isopropoxyethoxy)nicotinonitrile], and compd4 [5-chloro-6-((1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-5-yl)oxy)-2-((4-oxopentyl)oxy)nicotinonitrile] are potent PDE4 inhibitors with similar affinity for PDE4 isoforms and equivalent inhibition on the catalytic domain and the full-length enzyme. These benzoxaboroles are less active on other PDE isozymes. Compd4 binds to the catalytic domain of PDE4B2 with the oxaborole group chelating the catalytic bimetal and overlapping with the phosphate in cAMP during substrate hydrolysis, and the interaction extends into the adenine pocket. In cell culture, benzoxaborole PDE4 inhibitors suppress the release of tumor necrosis factor-α, interleukin (IL)-23, IL-17, interferon-γ, IL-4, IL-5, IL-13, and IL-22, and these cytokines contribute to the pathologic changes in skin structure and barrier functions as well as immune dysregulation in atopic dermatitis and psoriasis. Treatment with compd3 or N(6),2'-O-dibutyryladenosine 3',5'-cyclic monophosphate increases cAMP response element binding protein phosphorylation in human monocytes and decreases extracellular signal-regulated kinase phosphorylation in human T cells; these changes lead to reduced cytokine production and are among the mechanisms by which compd3 blocks cytokine release. Topical compd3 penetrates the skin and suppresses phorbol myristate acetate-induced IL-13, IL-22, IL-17F, and IL-23 transcription and calcipotriol-induced thymic stromal lymphopoietin expression in mouse skin. Skin thinning is a major dose-limiting side effect of glucocorticoids. By contrast, repeated application of compd3 did not thin mouse skin. These findings show the potential benefits and safety of benzoxaborole PDE4 inhibitors for the treatment of psoriasis and atopic dermatitis.
Assuntos
Compostos de Boro/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Dermatite Atópica/tratamento farmacológico , Inibidores da Fosfodiesterase 4/farmacologia , Psoríase/tratamento farmacológico , Pele/efeitos dos fármacos , Pele/patologia , Administração Tópica , Animais , Compostos de Boro/administração & dosagem , Compostos de Boro/uso terapêutico , Compostos Bicíclicos Heterocíclicos com Pontes/administração & dosagem , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Domínio Catalítico , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/química , Citocinas/metabolismo , Dermatite Atópica/metabolismo , Dermatite Atópica/patologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Modelos Moleculares , Inibidores da Fosfodiesterase 4/administração & dosagem , Inibidores da Fosfodiesterase 4/uso terapêutico , Fosforilação/efeitos dos fármacos , Psoríase/metabolismo , Psoríase/patologia , Pele/metabolismo , Linfopoietina do Estroma do TimoRESUMO
Novel isoxazoline amide benzoxaboroles were designed and synthesized to optimize the ectoparasiticide activity of this chemistry series against ticks and fleas. The study identified an orally bioavailable molecule, (S)-N-((1-hydroxy-3,3-dimethyl-1,3-dihydrobenzo[c][1,2]oxaborol-6-yl)methyl)-2-methyl-4-(5-(3,4,5-trichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)benzamide (23), with a favorable pharmacodynamics profile in dogs (Cmax=7.42ng/mL; Tmax=26.0h; terminal half-life t1/2=127h). Compound 23, a development candidate, demonstrated 100% therapeutic effectiveness within 24h of treatment, with residual efficacy of 97% against American dog ticks (Dermacentor variabilis) on day 30 and 98% against cat fleas (Ctenocephalides felis) on day 32 after a single oral dose at 25mg/kg in dogs.
Assuntos
Amidas/farmacologia , Antiparasitários/farmacologia , Compostos de Boro/farmacologia , Ctenocephalides/efeitos dos fármacos , Dermacentor/efeitos dos fármacos , Ectoparasitoses/tratamento farmacológico , Isoxazóis/farmacologia , Administração Oral , Amidas/administração & dosagem , Amidas/química , Animais , Antiparasitários/administração & dosagem , Antiparasitários/química , Compostos de Boro/administração & dosagem , Compostos de Boro/química , Gatos , Cães , Relação Dose-Resposta a Droga , Ectoparasitoses/parasitologia , Isoxazóis/administração & dosagem , Isoxazóis/química , Estrutura Molecular , Testes de Sensibilidade Parasitária , Relação Estrutura-AtividadeRESUMO
A novel series of isoxazoline benzoxaborole small molecules was designed and synthesized for a structure-activity relationship (SAR) investigation to assess the ectoparasiticide activity against ticks and fleas. The study identified an orally bioavailable molecule, (S)-3,3-dimethyl-5-(5-(3,4,5-trichlorophenyl)-5-(trifluoromethyl)-4,5-dihydroisoxazol-3-yl)benzo[c][1,2]oxaborol-1(3H)-ol (38, AN8030), which was long lasting in dogs (t1/2=22 days). Compound 38 demonstrated 97.6% therapeutic effectiveness within 24 h of treatment, with residual efficacy of 95.3% against American dog ticks (Dermacentor variabilis) on day 30% and 100% against cat fleas (Ctenocephalides felis) on day 32 after a single oral dose at 50 mg/kg in dogs.
Assuntos
Compostos de Boro/química , Doenças do Cão/tratamento farmacológico , Descoberta de Drogas , Ectoparasitoses/tratamento farmacológico , Isoxazóis/síntese química , Administração Oral , Animais , Compostos de Boro/administração & dosagem , Compostos de Boro/farmacologia , Doenças do Cão/parasitologia , Cães , Isoxazóis/administração & dosagem , Isoxazóis/química , Isoxazóis/farmacologia , Estrutura Molecular , Relação Estrutura-Atividade , Fatores de TempoRESUMO
Utilizing structure-based drug design, a novel dihydropyridopyrimidinone series which exhibited potent Hsp90 inhibition, good pharmacokinetics upon oral administration, and an excellent pharmacokinetic/pharmacodynamic relationship in vivo was developed from a commercial hit. The exploration of this series led to the selection of NVP-HSP990 as a development candidate.
Assuntos
Antineoplásicos/síntese química , Piridonas/síntese química , Pirimidinas/síntese química , Animais , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Camundongos , Piridonas/farmacocinética , Piridonas/farmacologia , Pirimidinas/farmacocinética , Pirimidinas/farmacologia , Relação Estrutura-AtividadeRESUMO
Benzoxaboroles are a novel class of drug-like compounds that have been rich sources of novel inhibitors for various enzymes and of new drugs. While examining benzoxaborole activity in phenotypic screens, our attention was attracted by the (aminomethylphenoxy)benzoxaborole family, which potently inhibited Toll-like receptor-stimulated cytokine secretion from leukocytes. After considering their structure-activity relationships and the central role of kinases in leukocyte biology, we performed a kinome-wide screen to investigate the members of the (aminomethylphenoxy)benzoxaborole family. This technique identified Rho-activated kinase (ROCK) as a target. We showed competitive behavior, with respect to ATP, and then determined the ROCK2-drug cocrystal structure. The drug occupies the ATP site in which the oxaborole moiety provides hydrogen bond donors and acceptors to the hinge, and the aminomethyl group interacts with the magnesium/ATP-interacting aspartic acid common to protein kinases. The series exhibits excellent selectivity against most of the kinome, with greater than 15-fold selectivity against the next best member of the AGC protein kinase subfamily. Medicinal chemistry efforts with structure-based design resulted in a compound with a Ki of 170 nM. Cellular studies revealed strong enzyme inhibition rank correlation with suppression of intracellular phosphorylation of a ROCK substrate. The biochemical potencies of these compounds also translated to functional activity, causing smooth muscle relaxation in rat aorta and guinea pig trachea. The series exhibited oral availability and one member reduced rat blood pressure, consistent with ROCK's role in smooth muscle contraction. Thus, the benzoxaborole moiety represents a novel hinge-binding kinase scaffold that may have potential for therapeutic use.
Assuntos
Compostos de Boro/metabolismo , Inibidores de Proteínas Quinases/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Quinases Associadas a rho/antagonistas & inibidores , Quinases Associadas a rho/metabolismo , Animais , Aorta Torácica/efeitos dos fármacos , Pressão Sanguínea/efeitos dos fármacos , Citocinas/sangue , Humanos , Células Jurkat , Modelos Moleculares , Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Fosforilação , Inibidores de Proteínas Quinases/química , Proteína Fosfatase 1/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Traqueia/efeitos dos fármacos , Quinases Associadas a rho/genéticaRESUMO
Gram-negative bacteria cause approximately 70% of the infections in intensive care units. A growing number of bacterial isolates responsible for these infections are resistant to currently available antibiotics and to many in development. Most agents under development are modifications of existing drug classes, which only partially overcome existing resistance mechanisms. Therefore, new classes of Gram-negative antibacterials with truly novel modes of action are needed to circumvent these existing resistance mechanisms. We have previously identified a new a way to inhibit an aminoacyl-tRNA synthetase, leucyl-tRNA synthetase (LeuRS), in fungi via the oxaborole tRNA trapping (OBORT) mechanism. Herein, we show how we have modified the OBORT mechanism using a structure-guided approach to develop a new boron-based antibiotic class, the aminomethylbenzoxaboroles, which inhibit bacterial leucyl-tRNA synthetase and have activity against Gram-negative bacteria by largely evading the main efflux mechanisms in Escherichia coli and Pseudomonas aeruginosa. The lead analogue, AN3365, is active against Gram-negative bacteria, including Enterobacteriaceae bearing NDM-1 and KPC carbapenemases, as well as P. aeruginosa. This novel boron-based antibacterial, AN3365, has good mouse pharmacokinetics and was efficacious against E. coli and P. aeruginosa in murine thigh infection models, which suggest that this novel class of antibacterials has the potential to address this unmet medical need.
Assuntos
Aminoacil-tRNA Sintetases/antagonistas & inibidores , Antibacterianos/farmacologia , Compostos de Boro/farmacologia , Escherichia coli/efeitos dos fármacos , Infecções por Bactérias Gram-Negativas/tratamento farmacológico , Pseudomonas aeruginosa/efeitos dos fármacos , Aminoacil-tRNA Sintetases/metabolismo , Animais , Antibacterianos/síntese química , Antibacterianos/farmacocinética , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/metabolismo , Compostos de Boro/síntese química , Compostos de Boro/farmacocinética , Cristalografia por Raios X , Descoberta de Drogas , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Escherichia coli/enzimologia , Feminino , Infecções por Bactérias Gram-Negativas/microbiologia , Humanos , Leucina/metabolismo , Camundongos , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Pseudomonas aeruginosa/enzimologia , Relação Estrutura-Atividade , Coxa da Perna/microbiologia , Inibidores de beta-Lactamases , beta-Lactamases/metabolismoRESUMO
We have designed and synthesized a novel class of compounds based on fluoroquinolone antibacterial prototype. The design concept involved the replacement of the 3-carboxylic acid in ciprofloxacin with an oxaborole-fused ring as an acid-mimicking group. The synthetic method employed in this work provides a good example of incorporating boron atom in complex molecules with multiple functional groups. The antibacterial activity of the newly synthesized compounds has been evaluated.
Assuntos
Antibacterianos/síntese química , Antibacterianos/farmacologia , Compostos de Boro/síntese química , Compostos de Boro/farmacologia , Fluoroquinolonas/síntese química , Fluoroquinolonas/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/síntese química , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Ciprofloxacina/química , Ciprofloxacina/farmacologia , Relação Estrutura-AtividadeRESUMO
We have synthesized and evaluated a series of novel HCV NS3 protease inhibitors with various P4 capping groups, which include urea, carbamate, methoxy-carboxamide, cyclic carbamate and amide, pyruvic amide, oxamate, oxalamide and cyanoguanidine. Most of these compounds are remarkably potent, exhibiting single-digit to sub-nanomolar activity in the enzyme assay and cell-based replicon assay. Selected compounds were also evaluated in the protease-inhibitor-resistant mutant transient replicon assay, and they were found to show quite different potency profiles against a panel of HCV protease-inhibitor-resistant mutants.
Assuntos
Antivirais/síntese química , Antivirais/farmacologia , Hepacivirus/efeitos dos fármacos , Inibidores de Serina Proteinase/síntese química , Inibidores de Serina Proteinase/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Amidas/química , Animais , Antivirais/química , Carbamatos/química , Relação Dose-Resposta a Droga , Farmacorresistência Viral/genética , Guanidinas/química , Hepacivirus/enzimologia , Hepacivirus/genética , Testes de Sensibilidade Microbiana , Estrutura Molecular , Ácido Oxâmico/química , Ratos , Inibidores de Serina Proteinase/química , Relação Estrutura-Atividade , Ureia/química , Proteínas não Estruturais Virais/metabolismoRESUMO
We have used boron-based molecules to create novel, competitive, reversible inhibitors of phosphodiesterase 4 (PDE4). The co-crystal structure reveals a binding configuration which is unique compared to classical catechol PDE4 inhibitors, with boron binding to the activated water in the bimetal center. These phenoxybenzoxaboroles can be optimized to generate submicromolar potency enzyme inhibitors, which inhibit TNF-α, IL-2, IFN-γ, IL-5 and IL-10 activities in vitro and show safety and efficacy for topical treatment of human psoriasis. They provide a valuable new route for creating novel potent anti-PDE4 inhibitors.
Assuntos
Compostos de Boro/química , Compostos de Boro/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/química , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Inibidores de Fosfodiesterase/química , Inibidores de Fosfodiesterase/farmacologia , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Ligação Competitiva , Compostos Bicíclicos Heterocíclicos com Pontes/química , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Domínio Catalítico , Cristalografia por Raios X , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/genética , Citocinas/biossíntese , Humanos , Técnicas In Vitro , Isoenzimas/antagonistas & inibidores , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Cinética , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/imunologia , Metais/química , Modelos Moleculares , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismoRESUMO
The macrocyclic urea 2, a byproduct in the synthesis of benzoxaborole 1, was identified to be a novel and potent HCV protease inhibitor. We further explored this motif by synthesizing additional urea-based inhibitors and by characterizing them in replicase HCV protease-resistant mutants assay. Several compounds, exemplified by 12, were found to be more potent in HCV replicon assays than leading second generation inhibitors such as danoprevir and TMC-435350. Additionally, following oral administration, inhibitor 12 was found in rat liver in significantly higher concentrations than those reported for both danoprevir and TMC-435350, suggesting that inhibitor 12 has the combination of anti-HCV and pharmacokinetic properties that warrants further development of this series.
Assuntos
Antivirais/síntese química , Farmacorresistência Viral , Hepacivirus/efeitos dos fármacos , Inibidores de Serina Proteinase/síntese química , Ureia/análogos & derivados , Ureia/síntese química , Proteínas não Estruturais Virais/antagonistas & inibidores , Administração Oral , Animais , Antivirais/farmacocinética , Antivirais/farmacologia , Hepacivirus/enzimologia , Hepacivirus/genética , Interações Hidrofóbicas e Hidrofílicas , Fígado/metabolismo , Mutação , Ratos , Replicon/efeitos dos fármacos , Inibidores de Serina Proteinase/farmacocinética , Inibidores de Serina Proteinase/farmacologia , Relação Estrutura-Atividade , Ureia/farmacocinética , Ureia/farmacologia , Proteínas não Estruturais Virais/genéticaRESUMO
A novel series of P3 oxo-modified macrocyclic hepatitis C virus NS3/4A serine protease inhibitor was designed, synthesized and biologically evaluated. The hydroxy-substituted inhibitor 10 demonstrated high potency in genotype 1a and 1b replicon and in the panel of HCV protease mutants. Interestingly, the t-butyl carbonate analog 9c, while not the most potent one in this series, exhibited a virtually flat potency profile in the panel of HCV protease mutants, thus providing opportunity for further optimization.