RESUMO
The Mycobacterium abscessus drug development pipeline is poorly populated, with particularly few validated target-lead couples to initiate de novo drug discovery. Trimethoprim, an inhibitor of dihydrofolate reductase (DHFR) used for the treatment of a range of bacterial infections, is not active against M. abscessus. Thus, evidence that M. abscessus DHFR is vulnerable to pharmacological intervention with a small molecule inhibitor is lacking. Here, we show that the pyrrolo-quinazoline PQD-1, previously identified as a DHFR inhibitor active against Mycobacterium tuberculosis, exerts whole cell activity against M. abscessus. Enzyme inhibition studies showed that PQD-1, in contrast to trimethoprim, is a potent inhibitor of M. abscessus DHFR and over-expression of DHFR causes resistance to PQD-1, providing biochemical and genetic evidence that DHFR is a vulnerable target and mediates PQD-1's growth inhibitory activity in M. abscessus. As observed in M. tuberculosis, PQD-1 resistant mutations mapped to the folate pathway enzyme thymidylate synthase (TYMS) ThyA. Like trimethoprim in other bacteria, PQD-1 synergizes with the dihydropteroate synthase (DHPS) inhibitor sulfamethoxazole (SMX), offering an opportunity to exploit the successful dual inhibition of the folate pathway and develop similarly potent combinations against M. abscessus. PQD-1 is active against subspecies of M. abscessus and a panel of clinical isolates, providing epidemiological validation of the target-lead couple. Leveraging a series of PQD-1 analogs, we have demonstrated a dynamic structure-activity relationship (SAR). Collectively, the results identify M. abscessus DHFR as an attractive target and PQD-1 as a chemical starting point for the discovery of novel drugs and drug combinations that target the folate pathway in M. abscessus.
Assuntos
Antagonistas do Ácido Fólico , Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Mycobacterium tuberculosis , Humanos , Mycobacterium abscessus/genética , Mycobacterium abscessus/metabolismo , Tetra-Hidrofolato Desidrogenase/genética , Tetra-Hidrofolato Desidrogenase/metabolismo , Antagonistas do Ácido Fólico/farmacologia , Trimetoprima/farmacologia , Mycobacterium tuberculosis/metabolismo , Inibidores Enzimáticos/farmacologia , Ácido Fólico , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológicoRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) research and antiviral discovery are hampered by the lack of a cell-based virus replication system that can be readily adopted without biosafety level 3 (BSL-3) restrictions. Here, the construction of a noninfectious SARS-CoV-2 reporter replicon and its application in deciphering viral replication mechanisms and evaluating SARS-CoV-2 inhibitors are presented. The replicon genome is replication competent but does not produce progeny virions. Its replication can be inhibited by RdRp mutations or by known SARS-CoV-2 antiviral compounds. Using this system, a high-throughput antiviral assay has also been developed. Significant differences in potencies of several SARS-CoV-2 inhibitors in different cell lines were observed, which highlight the challenges of discovering antivirals capable of inhibiting viral replication in vivo and the importance of testing compounds in multiple cell culture models. The generation of a SARS-CoV-2 replicon provides a powerful platform to expand the global research effort to combat COVID-19.
Assuntos
Antivirais/farmacologia , COVID-19/virologia , Ensaios de Triagem em Larga Escala/métodos , Replicon/efeitos dos fármacos , SARS-CoV-2/efeitos dos fármacos , Células A549 , Animais , Chlorocebus aethiops , RNA-Polimerase RNA-Dependente de Coronavírus/genética , Células HEK293 , Humanos , Replicon/genética , SARS-CoV-2/genética , Células Vero , Replicação Viral/efeitos dos fármacosRESUMO
Since the initial report of the novel Coronavirus Disease 2019 (COVID-19) emanating from Wuhan, China, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has spread globally. While the effects of SARS-CoV-2 infection are not completely understood, there appears to be a wide spectrum of disease ranging from mild symptoms to severe respiratory distress, hospitalization, and mortality. There are no Food and Drug Administration (FDA)-approved treatments for COVID-19 aside from remdesivir; early efforts to identify efficacious therapeutics for COVID-19 have mainly focused on drug repurposing screens to identify compounds with antiviral activity against SARS-CoV-2 in cellular infection systems. These screens have yielded intriguing hits, but the use of nonhuman immortalized cell lines derived from non-pulmonary or gastrointestinal origins poses any number of questions in predicting the physiological and pathological relevance of these potential interventions. While our knowledge of this novel virus continues to evolve, our current understanding of the key molecular and cellular interactions involved in SARS-CoV-2 infection is discussed in order to provide a framework for developing the most appropriate in vitro toolbox to support current and future drug discovery efforts.
Assuntos
Descoberta de Drogas , SARS-CoV-2/fisiologia , Tropismo Viral , Internalização do Vírus , Replicação Viral , COVID-19/virologia , Catepsinas , Linhagem Celular , Desenvolvimento de Medicamentos , Endocitose , Furina , Humanos , SARS-CoV-2/efeitos dos fármacos , Serina Endopeptidases , Tratamento Farmacológico da COVID-19RESUMO
Tricyclic pyrrolopyrimidines (TPPs) are a new class of antibacterials inhibiting the ATPase of DNA gyrase. TPP8, a representative of this class, is active against Mycobacterium abscessus in vitro. Spontaneous TPP8 resistance mutations mapped to the ATPase domain of M. abscessus DNA gyrase, and the compound inhibited DNA supercoiling activity of recombinant M. abscessus enzyme. Further profiling of TPP8 in macrophage and mouse infection studies demonstrated proof-of-concept activity against M. abscessus ex vivo and in vivo.
Assuntos
Infecções por Mycobacterium não Tuberculosas , Mycobacterium abscessus , Adenosina Trifosfatases , Animais , Antibacterianos/farmacologia , DNA Girase/genética , Camundongos , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium não Tuberculosas/tratamento farmacológico , Infecções por Mycobacterium não Tuberculosas/microbiologia , Micobactérias não Tuberculosas , Pirimidinas , PirróisRESUMO
Novel bacterial topoisomerase inhibitors (NBTIs) are the newest members of gyrase inhibitor broad-spectrum antibacterial agents, represented by the most advanced member, gepotidacin, a 4-amino-piperidine linked NBTI, which is undergoing phase III clinical trials for treatment of urinary tract infections (UTI). We have extensively reported studies on oxabicyclooctane linked NBTIs, including AM-8722. The present study summarizes structure activity relationship (SAR) of AM-8722 leading to identification of 7-fluoro-1-cyanomethyl-1,5-naphthyridin-2-one based NBTI (16, AM-8888) with improved potency and spectrum (MIC values of 0.016-4 µg/mL), with Pseudomonas aeruginosa being the least sensitive strain (MIC 4 µg/mL).
Assuntos
Antibacterianos , Inibidores da Topoisomerase , Antibacterianos/química , Antibacterianos/farmacologia , DNA Girase/metabolismo , DNA Topoisomerase IV , Testes de Sensibilidade Microbiana , Staphylococcus aureus/metabolismo , Relação Estrutura-Atividade , Tioinosina/análogos & derivados , Inibidores da Topoisomerase II/química , Inibidores da Topoisomerase II/farmacologia , Inibidores da Topoisomerase/química , Inibidores da Topoisomerase/farmacologiaRESUMO
The methylotrophic yeast Pichia pastoris (reclassified as Komagataella phaffii) is a versatile protein expression system, yet many commonly used promoters have attributes undesirable for fermentation or its optimization. Hence, the copper-inducible CUP1 gene promoter from the related yeast Saccharomyces cerevisiae was used to express human gelatin. Multimerization of a potential copper response element in the CUP1 promoter, a S. cerevisiae Ace1p binding site, significantly increased gelatin expression. Expression was induced by copper in a dose-dependent fashion and was not dependent on cell density. Gelatin was additionally induced in standard copper-containing fermentation basal salts media. Removal of a S. cerevisiae heat shock factor (Hsf1p) binding site reduced copper-dependent gelatin induction suggesting that a similar protein may regulate this promoter in P. pastoris. This engineered copper inducible promoter expands the yeast recombinant protein production tool kit.
Assuntos
Gelatina/genética , Expressão Gênica , Metalotioneína/genética , Regiões Promotoras Genéticas , Saccharomyces cerevisiae/genética , Saccharomycetales/genética , Cobre/metabolismo , Meios de Cultura , Fermentação , Regulação Fúngica da Expressão Gênica , Humanos , Saccharomyces cerevisiae/metabolismo , Saccharomycetales/efeitos dos fármacos , Saccharomycetales/metabolismoRESUMO
BACKGROUND: Intrathecal opioids are routinely administered during spinal anesthesia for postcesarean analgesia. The effectiveness of intrathecal morphine for postcesarean analgesia is well established, and the use of intrathecal hydromorphone is growing. No prospective studies have compared the effectiveness of equipotent doses of intrathecal morphine versus intrathecal hydromorphone as part of a multimodal analgesic regimen for postcesarean analgesia. The authors hypothesized that intrathecal morphine would result in superior analgesia compared with intrathecal hydromorphone 24 h after delivery. METHODS: In this single-center, double-blinded, randomized trial, 138 parturients undergoing scheduled cesarean delivery were randomized to receive 150 µg of intrathecal morphine or 75 µg of intrathecal hydromorphone as part of a primary spinal anesthetic and multimodal analgesic regimen; 134 parturients were included in the analysis. The primary outcome was the numerical rating scale score for pain with movement 24 h after delivery. Static and dynamic pain scores, nausea, pruritus, degree of sedation, and patient satisfaction were assessed every 6 h for 36 h postpartum. Total opioid consumption was recorded. RESULTS: There was no significant difference in pain scores with movement at 24 h (intrathecal hydromorphone median [25th, 75th] 4 [3, 5] and intrathecal morphine 3 [2, 4.5]) or at any time point (estimated difference, 0.5; 95% CI, 0 to 1; P = 0.139). Opioid received in the first 24 h did not differ between groups (median [25th, 75th] oral morphine milligram equivalents for intrathecal hydromorphone 30 [7.5, 45.06] vs. intrathecal morphine 22.5 [14.0, 37.5], P = 0.769). From Kaplan-Meier analysis, the median time to first opioid request was 5.4 h for hydromorphone and 12.1 h for morphine (log-rank test P = 0.200). CONCLUSIONS: Although the hypothesis was that intrathecal morphine would provide superior analgesia to intrathecal hydromorphone, the results did not confirm this. At the doses studied, both intrathecal morphine and intrathecal hydromorphone provide effective postcesarean analgesia when combined with a multimodal analgesia regimen.
Assuntos
Analgesia Epidural/métodos , Analgesia Obstétrica/métodos , Cesárea , Hidromorfona/uso terapêutico , Morfina/uso terapêutico , Dor Pós-Operatória/tratamento farmacológico , Adulto , Analgésicos Opioides/administração & dosagem , Analgésicos Opioides/uso terapêutico , Método Duplo-Cego , Feminino , Humanos , Hidromorfona/administração & dosagem , Masculino , Morfina/administração & dosagem , Resultado do TratamentoRESUMO
Objective: To compare the effect of a conventional to an intensive blood pressure monitoring regimen on blood pressure in hypertensive patients in the general practice setting. Design: Randomized controlled parallel group trial with 12-month follow-up. Setting: One hundred and ten general practices in all regions of Denmark. Participants: One thousand forty-eight patients with essential hypertension. Intervention: Conventional blood pressure monitoring ('usual group') continued usual ad hoc blood pressure monitoring by office blood pressure measurements, while intensive blood pressure monitoring ('intensive group') supplemented this with frequent home blood pressure monitoring and 24-hour ambulatory blood pressure monitoring. Primary outcome measures: Mean day- and night-time systolic and diastolic 24-hour ambulatory blood pressure. Secondary outcome measures: Change in systolic and diastolic office blood pressure and change in cardiovascular risk profile. Results: Of the patients, 515 (49%) were allocated to the usual group, and 533 (51%) to the intensive group. The reductions in day- and night-time 24-hour ambulatory blood pressure were similar (usual group: 4.6 ± 13.5/2.8 ± 82 mmHg; intensive group: 5.6 ± 13.0/3.5 ± 8.2 mmHg; P = 0.27/P = 0.20). Cardiovascular risk scores were reduced in both groups at follow-up, but more so in the intensive than in the usual group (P = 0.02). Conclusion: An intensive blood pressure monitoring strategy led to a similar blood pressure reduction to conventional monitoring. However, the intensive strategy appeared to improve patients' cardiovascular risk profile through other effects than a reduction of blood pressure. Trial registration: Clinical Trials NCT00244660.
Assuntos
Monitorização Ambulatorial da Pressão Arterial/métodos , Medicina Geral , Serviços de Assistência Domiciliar , Hipertensão/terapia , Anti-Hipertensivos/uso terapêutico , Dinamarca , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de RiscoRESUMO
BACKGROUND: De Barsy syndrome is a rare, autosomal recessive syndrome characterized by cutis laxa, progeroid appearance, ophthalmic opacification, skeletal malformations, growth delays, and intellectual disability. AIMS: The aim of this case series is to identify the anesthetic considerations in the clinical management of patients with de Barsy syndrome. METHODS: A retrospective case review from 1968 to 2016 was performed at a single tertiary medical center to identify patients with de Barsy syndrome who underwent anesthesia for diagnostic and surgical procedures. We collected and analyzed the perioperative records and following data: age, sex, American Society of Anesthesiologists physical status, relevant comorbidities, surgical procedures, anesthesia management, and observed complications. RESULTS: Three patients underwent 64 unique anesthetics for a diverse collection of diagnostic and surgical procedures. An array of anesthetics and techniques were successfully used. Observations of the perioperative period found 7 episodes of intraoperative hyperthermia (>38.3°), a single difficult airway requiring fiberoptic bronchoscopic-guided intubation, and repeatedly difficult intravenous access. CONCLUSION: This expanded case series suggests that providers caring for patients with de Barsy syndrome should be aware of potential challenges with airway management, vascular access, and temperature monitoring.
Assuntos
Anestesia , Opacidade da Córnea/terapia , Cútis Laxa/terapia , Deficiência Intelectual/terapia , Anestésicos Inalatórios , Anestésicos Intravenosos , Criança , Feminino , Febre/epidemiologia , Humanos , Complicações Intraoperatórias/epidemiologia , Complicações Intraoperatórias/terapia , Masculino , Estudos RetrospectivosRESUMO
Oxabicyclooctane-linked novel bacterial topoisomerase inhibitors (NBTIs) represent a new class of recently described antibacterial agents with broad-spectrum activity. NBTIs dually inhibit the clinically validated bacterial targets DNA gyrase and topoisomerase IV and have been shown to bind distinctly from known classes of antibacterial agents directed against these targets. Herein we report the molecular, cellular, and in vivo characterization of AM-8722 as a representative N-alkylated-1,5-naphthyridone left-hand-side-substituted NBTI. Consistent with its mode of action, macromolecular labeling studies revealed a specific effect of AM-8722 to dose dependently inhibit bacterial DNA synthesis. AM-8722 displayed greater intrinsic enzymatic potency than levofloxacin versus both DNA gyrase and topoisomerase IV from Staphylococcus aureus and Escherichia coli and displayed selectivity against human topoisomerase II. AM-8722 was rapidly bactericidal and exhibited whole-cell activity versus a range of Gram-negative and Gram-positive organisms, with no whole-cell potency shift due to the presence of DNA or human serum. Frequency-of-resistance studies demonstrated an acceptable rate of resistance emergence in vitro at concentrations 16- to 32-fold the MIC. AM-8722 displayed acceptable pharmacokinetic properties and was shown to be efficacious in mouse models of bacterial septicemia. Overall, AM-8722 is a selective and potent NBTI that displays broad-spectrum antimicrobial activity in vitro and in vivo.
Assuntos
Antibacterianos/farmacologia , Ciclo-Octanos/farmacologia , DNA Girase/metabolismo , DNA Topoisomerase IV/antagonistas & inibidores , DNA Topoisomerases Tipo II/metabolismo , Inibidores da Topoisomerase II/farmacologia , Animais , Linhagem Celular , DNA Bacteriano/genética , Cães , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Infecções por Escherichia coli/tratamento farmacológico , Humanos , Camundongos , Testes de Sensibilidade Microbiana , Ratos , Ratos Wistar , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genéticaRESUMO
PURPOSE: To evaluate and model the pharmacokinetic and pharmacodynamic behavior in rats of FG-3019, a human monoclonal antibody targeting connective tissue growth factor (CTGF). METHODS: FG-3019, human CTGF (rhCTGF), or the N-terminal domain of rhCTGF were administered intravenously to rats and concentrations of these proteins as well as endogenous CTGF were determined by immunoassays. FG-3019, or (125)I-labeled FG-3019, and human CTGF (rhCTGF) were co-administered to assess the impact of CTGF on the elimination rate and tissue localization of FG-3019, which was further characterized by immunohistochemical analysis. A PK/PD model for target-mediated elimination of FG-3019 was developed to fit the kinetic data. RESULTS: FG-3019 exhibited non-linear pharmacokinetics in rats. Circulating concentrations of the N-terminal half of CTGF increased after dosing with FG-3019, reached maximal levels after 1-5 days, and returned toward baseline levels as FG-3019 cleared from the circulation, whereas the concentration of intact CTGF was unaffected by administration of FG-3019. Co-administration of rhCTGF dramatically enhanced the rate of FG-3019 elimination, redistributing the majority of (125)I-labeled FG-3019 from the blood to the liver, kidney, spleen and adrenal gland. FG-3019 co-administered with CTGF was found along the sinusoids of the liver and adrenal glands, the capillaries of the kidney glomeruli and in the spleen. A pharmacokinetic model for target-mediated elimination of FG-3019 was used to fit the time courses of FG-3019 and endogenous CTGF plasma concentrations, as well as time courses of rhCTGF and rhCTGF N-fragment after intravenous administration of these species. CONCLUSIONS: FG-3019 is subject to target mediated elimination in rats.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Fator de Crescimento do Tecido Conjuntivo/administração & dosagem , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Administração Intravenosa , Animais , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais Humanizados , Relação Dose-Resposta a Droga , Interações Medicamentosas/fisiologia , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/metabolismo , Distribuição Tecidual/efeitos dos fármacos , Distribuição Tecidual/fisiologiaRESUMO
Aspartyl proteases are important pharmacological targets. Historically aspartyl proteases have been commonly targeted with transition state derived peptidomimetics. The strategy to develop aspartyl protease inhibitors has undertaken a dramatic paradigm shift in the last 10 years. The pharmaceutical industry in 2005 disclosed several scaffolds or "head groups" that prompted the field to move beyond peptidomimetic derived inhibitors. Since the discovery of the first amino heterocycle aspartyl protease inhibitor, the amino hydantoin, industry and academia have positioned themselves for a foothold on the new molecular space, designing a variety of related "head groups". Both the design and synthetic efforts involved in constructing these scaffolds are varied and complex. Here we highlight the synthetic strategies used to access these amino heterocycle scaffolds.
Assuntos
Ácido Aspártico Endopeptidases/antagonistas & inibidores , Técnicas de Química Sintética/métodos , Compostos Heterocíclicos/síntese química , Compostos Heterocíclicos/farmacologia , Inibidores de Proteases/síntese química , Inibidores de Proteases/farmacologia , Desenho de Fármacos , Compostos Heterocíclicos/química , Inibidores de Proteases/químicaRESUMO
Distinguishing between the two enantiomers of a molecule is a challenging task due to their nearly identical physical properties. Time-consuming chromatography methods are typically required for this task, which greatly limits the throughput of analysis. Here we describe a fluorescence-based method for the rapid and high-throughput analysis of both small-molecule enantiopurity and concentration. Our approach relies on selective molecular recognition of one enantiomer of the target molecule using a DNA aptamer, and the ability of aptamer-based biosensors to transduce the presence of a target molecule into a dose-dependent fluorescence signal. The key novel aspect of our approach is the implementation of enantiomeric DNA biosensors, which are synthesized from D- and L-DNA, but labeled with orthogonal fluorophores. According to the principle of reciprocal chiral substrate specificity, these biosensors will bind to opposite enantiomers of the target with equal affinity and selectivity, enabling simultaneous quantification of both enantiomers of the target. Using the previously reported DNA biosensor for L-tyrosinamide (L-Tym), we demonstrate the ability to rapidly and accurately measure both enantiopurity and concentration for mixtures of L- and D-Tym. We also apply our enantiomeric biosensors to the optimization of reaction conditions for the synthesis of D-Tym and provide mathematical modeling to suggest that DNA biosensors having only modest binding selectivity can also be used for fluorescence-based enantiopurity measurement. This research provides a generalizable method for high-throughput analysis of reaction mixtures, which is anticipated to significantly accelerate reaction optimization for the synthesis of high-value chiral small molecules.
Assuntos
Aptâmeros de Nucleotídeos/química , Corantes Fluorescentes/química , Tirosina/análogos & derivados , Sequência de Bases , Técnicas Biossensoriais/métodos , DNA/química , Ensaios de Triagem em Larga Escala/métodos , Conformação de Ácido Nucleico , Estereoisomerismo , Tirosina/análise , Tirosina/isolamento & purificaçãoRESUMO
Oxabicyclooctane linked novel bacterial topoisomerase inhibitors (NBTIs) are new class of recently reported broad-spectrum antibacterial agents. They target bacterial DNA gyrase and topoisomerase IV and bind to a site different than quinolones. They show no cross-resistance to known antibiotics and provide opportunity to combat drug-resistant bacteria. A structure activity relationship of the C-2 substituted ether analogs of 1,5-naphthyridine oxabicyclooctane-linked NBTIs are described. Synthesis and antibacterial activities of a total of 63 analogs have been summarized representing alkyl, cyclo alkyl, fluoro alkyl, hydroxy alkyl, amino alkyl, and carboxyl alkyl ethers. All compounds were tested against three key strains each of Gram-positive and Gram-negative bacteria as well as for hERG binding activities. Many key compounds were also tested for the functional hERG activity. Six compounds were evaluated for efficacy in a murine bacteremia model of Staphylococcus aureus infection. Significant tolerance for the ether substitution (including polar groups such as amino and carboxyl) at C-2 was observed for S. aureus activity however the same was not true for Enterococcus faecium and Gram-negative strains. Reduced clogD generally showed reduced hERG activity and improved in vivo efficacy but was generally associated with decreased overall potency. One of the best compounds was hydroxy propyl ether (16), which mainly retained the potency, spectrum and in vivo efficacy of AM8085 associated with the decreased hERG activity and improved physical property.
Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Naftiridinas/química , Relação Estrutura-Atividade , Animais , Antibacterianos/síntese química , Antibacterianos/farmacocinética , Técnicas de Química Sintética , Ciclo-Octanos/química , DNA Girase/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Canal de Potássio ERG1 , Enterococcus faecium/efeitos dos fármacos , Canais de Potássio Éter-A-Go-Go/metabolismo , Camundongos Endogâmicos C57BL , Testes de Sensibilidade Microbiana , Ratos Sprague-Dawley , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Inibidores da Topoisomerase II/química , Inibidores da Topoisomerase II/farmacologiaRESUMO
Bacterial resistance is rapidly growing, necessitating the need to discover new agents. Novel bacterial topoisomerase inhibitors (NBTIs) are new class of broad-spectrum antibacterial agents targeting bacterial DNA gyrase and topoisomerase IV. This class of inhibitors binds to an alternative binding site relative to fluoroquinolones and shows no cross-resistance to quinolones. NBTIs consist of three structural motifs. A structure activity relationship of the left hand motif 1,5-naphthyridine of oxabicyclooctane-linked NBTIs is described. Fifty five compounds were evaluated against a panel of key Gram-positive and Gram-negative strains of bacteria, as well as for hERG activity and five compounds were tested for in vivo efficacy in murine model of Staphylococcus aureus infection. These studies suggest that only a narrow range (activating and deactivating) of substitutions at C-2 and C-7 are tolerated for optimal antibacterial activity and spectrum. An alkoxy (methoxy) and CN at C-2, and a halogen and hydroxyl at C-7, appeared to be preferred in this series. Substitutions on the other three carbons generally have detrimental effect on the activity. No clear hERG activity SAR emerged from these substitutions.
Assuntos
DNA Topoisomerases/metabolismo , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Inibidores da Topoisomerase/química , Inibidores da Topoisomerase/farmacologia , Animais , Camundongos , Estrutura Molecular , Infecções Estafilocócicas/microbiologia , Relação Estrutura-AtividadeRESUMO
Novel bacterial topoisomerase inhibitors (NBTIs) are a new class of broad-spectrum antibacterial agents targeting bacterial Gyrase A and ParC and have potential utility in combating antibiotic resistance. (R)-Hydroxy-1,5-naphthyridinone left-hand side (LHS) oxabicyclooctane linked pyridoxazinone right-hand side (RHS) containing NBTIs showed a potent Gram-positive antibacterial profile. SAR around the RHS moiety, including substitutions around pyridooxazinone, pyridodioxane, and phenyl propenoids has been described. A fluoro substituted pyridoxazinone showed an MIC against Staphylococcus aureus of 0.5 µg/mL with reduced functional hERG activity (IC50 333 µM) and good in vivo efficacy [ED90 12 mg/kg, intravenous (iv) and 15 mg/kg, oral (p.o.)]. A pyridodioxane-containing NBTI showed a S. aureus MIC of 0.5 µg/mL, significantly improved hERG IC50 764 µM and strong efficacy of 11 mg/kg (iv) and 5 mg/kg (p.o.). A phenyl propenoid series of compounds showed potent antibacterial activity, but also showed potent hERG binding activity. Many of the compounds in the hydroxy-tricyclic series showed strong activity against Acinetobacter baumannii, but reduced activity against Escherichia coli and Pseudomonas aeruginosa. Bicyclic heterocycles appeared to be the best RHS moiety for the hydroxy-tricyclic oxabicyclooctane linked NBTIs.
Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Naftiridinas/química , Inibidores da Topoisomerase/química , Inibidores da Topoisomerase/farmacologia , Acinetobacter baumannii/efeitos dos fármacos , Antibacterianos/síntese química , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/metabolismo , DNA Girase/química , DNA Girase/metabolismo , Escherichia coli/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Oxazóis/química , Pseudomonas aeruginosa/efeitos dos fármacos , Staphylococcus aureus/efeitos dos fármacos , Relação Estrutura-Atividade , Inibidores da Topoisomerase/síntese químicaRESUMO
Oxabicyclooctane linked 1,5-naphthyridinyl-pyridoxazinones are novel broad-spectrum bacterial topoisomerase inhibitors (NBTIs) targeting bacterial DNA gyrase and topoisomerase IV at a site different than quinolones. Due to lack of cross-resistance to known antibiotics they present excellent opportunity to combat drug-resistant bacteria. A structure activity relationship of the pyridoxazinone moiety is described in this Letter. Chemical synthesis and activities of NBTIs with substitutions at C-3, C-4 and C-7 of the pyridoxazinone moiety with halogens, alkyl groups and methoxy group has been described. In addition, substitutions of the linker NH proton and its transformation into amide analogs of AM-8085 and AM-8191 have been reported. Fluoro, chloro, and methyl groups at C-3 of the pyridoxazinone moiety retained the potency and spectrum. In addition, a C-3 fluoro analog showed 4-fold better oral efficacy (ED50 3.9 mg/kg) as compared to the parent AM-8085 in a murine bacteremia model of infection of Staphylococcus aureus. Even modest polarity (e.g., methoxy) is not tolerated at C-3 of the pyridoxazinone unit. The basicity and NH group of the linker is important for the activity when CH2 is at the linker position-8. However, amides (with linker position-8 ketone) with a position-7 NH or N-methyl group retained potency and spectrum suggesting that neither basicity nor hydrogen-donor properties of the linker amide NH is essential for the activity. This would suggest likely an altered binding mode of the linker position-7,8 amide containing compounds. The amides showed highly improved hERG (functional IC50 >30 µM) profile.
Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Ciclo-Octanos/química , Avaliação Pré-Clínica de Medicamentos/métodos , Relação Estrutura-Atividade , Inibidores da Topoisomerase/química , Administração Oral , Animais , Antibacterianos/administração & dosagem , Técnicas de Química Sintética , DNA Topoisomerase IV/antagonistas & inibidores , Canal de Potássio ERG1 , Canais de Potássio Éter-A-Go-Go/metabolismo , Compostos Heterocíclicos com 2 Anéis/química , Compostos Heterocíclicos com 2 Anéis/farmacologia , Camundongos , Testes de Sensibilidade Microbiana , Naftiridinas/química , Naftiridinas/farmacologia , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/patogenicidade , Inibidores da Topoisomerase/farmacologiaRESUMO
Novel bacterial topoisomerase inhibitors (NBTIs) represent a new class of broad-spectrum antibacterial agents targeting bacterial Gyrase A and ParC and have potential utility in combating antibiotic resistance. A series of novel oxabicyclooctane-linked NBTIs with new tricyclic-1,5-naphthyridinone left hand side moieties have been described. Compounds with a (R)-hydroxy-1,5-naphthyridinone moiety (7) showed potent antibacterial activity (e.g., Staphylococcus aureus MIC 0.25 µg/mL), acceptable Gram-positive and Gram-negative spectrum with rapidly bactericidal activity. The compound 7 showed intravenous and oral efficacy (ED50) at 3.2 and 27 mg/kg doses, respectively, in a murine model of bacteremia. Most importantly they showed significant attenuation of functional hERG activity (IC50 >170 µM). In general, lower logD attenuated hERG activity but also reduced Gram-negative activity. The co-crystal structure of a hydroxy-tricyclic NBTI bound to a DNA-gyrase complex exhibited a binding mode that show enantiomeric preference for R isomer and explains the activity and SAR. The discovery, synthesis, SAR and X-ray crystal structure of the left-hand-side tricyclic 1,5-naphthyridinone based oxabicyclooctane linked NBTIs are described.
Assuntos
Antibacterianos/farmacologia , Ciclo-Octanos/farmacologia , DNA Topoisomerases Tipo II/metabolismo , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Naftiridinas/farmacologia , Inibidores da Topoisomerase II/farmacologia , Antibacterianos/síntese química , Antibacterianos/química , Ciclo-Octanos/síntese química , Ciclo-Octanos/química , Relação Dose-Resposta a Droga , Bactérias Gram-Negativas/enzimologia , Bactérias Gram-Positivas/enzimologia , Testes de Sensibilidade Microbiana , Modelos Moleculares , Estrutura Molecular , Naftiridinas/síntese química , Naftiridinas/química , Relação Estrutura-Atividade , Inibidores da Topoisomerase II/síntese química , Inibidores da Topoisomerase II/químicaRESUMO
Clostridium difficile is the causative agent of C. difficile-associated diarrhea (CDAD), with increased risk in elderly populations. Kibdelomycin, a novel natural-product inhibitor of type II topoisomerase enzymes, was evaluated for activity against C. difficile and gastrointestinal anaerobic organisms. Toxigenic C. difficile isolates (n=168) from U.S. hospitals and anaerobic Gram-positive and Gram-negative organisms (n=598) from Chicago-area hospitals were tested. Kibdelomycin showed potent activity against toxigenic C. difficile (MIC90=0.25 µg/ml) and most Gram-positive aerobic organisms but had little activity against Bacteroides species (MIC50>32 µg/ml; n=270). Potent anti-C. difficile activity was also observed in the hamster model of C. difficile colitis. Dosing at 1.6 mg/kg (twice-daily oral dose) resulted in protection from a lethal infection and a 2-log reduction in C. difficile cecal counts. A 6.25-mg/kg twice-daily oral dose completely eliminated detectable C. difficile counts in cecal contents. A single 6.25-mg/kg oral dose showed that cecal contents were exposed to the drug at >2 µM (eightfold higher than the MIC), with no significant plasma exposure. These findings support further exploration of kibdelomycin for development of an anti-C. difficile agent.
Assuntos
Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Clostridioides difficile/efeitos dos fármacos , Infecções por Clostridium/tratamento farmacológico , Animais , Antibacterianos/farmacocinética , Cricetinae , Masculino , Camundongos , Testes de Sensibilidade MicrobianaRESUMO
Hepatitis C virus (HCV) infection is a leading cause of liver transplantation and there is an urgent need to develop therapies to reduce rates of HCV infection of transplanted livers. Approved therapeutics for HCV are poorly tolerated and are of limited efficacy in this patient population. Human monoclonal antibody HCV1 recognizes a highly-conserved linear epitope of the HCV E2 envelope glycoprotein (amino acids 412-423) and neutralizes a broad range of HCV genotypes. In a chimpanzee model, a single dose of 250 mg/kg HCV1 delivered 30 minutes prior to infusion with genotype 1a H77 HCV provided complete protection from HCV infection, whereas a dose of 50 mg/kg HCV1 did not protect. In addition, an acutely-infected chimpanzee given 250 mg/kg HCV1 42 days following exposure to virus had a rapid reduction in viral load to below the limit of detection before rebounding 14 days later. The emergent virus displayed an E2 mutation (N415K/D) conferring resistance to HCV1 neutralization. Finally, three chronically HCV-infected chimpanzees were treated with a single dose of 40 mg/kg HCV1 and viral load was reduced to below the limit of detection for 21 days in one chimpanzee with rebounding virus displaying a resistance mutation (N417S). The other two chimpanzees had 0.5-1.0 log(10) reductions in viral load without evidence of viral resistance to HCV1. In vitro testing using HCV pseudovirus (HCVpp) demonstrated that the sera from the poorly-responding chimpanzees inhibited the ability of HCV1 to neutralize HCVpp. Measurement of antibody responses in the chronically-infected chimpanzees implicated endogenous antibody to E2 and interference with HCV1 neutralization although other factors may also be responsible. These data suggest that human monoclonal antibody HCV1 may be an effective therapeutic for the prevention of graft infection in HCV-infected patients undergoing liver transplantation.