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1.
Nat Microbiol ; 9(5): 1244-1255, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38649414

RESUMO

Carbapenem-resistant Acinetobacter baumannii infections have limited treatment options. Synthesis, transport and placement of lipopolysaccharide or lipooligosaccharide (LOS) in the outer membrane of Gram-negative bacteria are important for bacterial virulence and survival. Here we describe the cerastecins, inhibitors of the A. baumannii transporter MsbA, an LOS flippase. These molecules are potent and bactericidal against A. baumannii, including clinical carbapenem-resistant Acinetobacter baumannii isolates. Using cryo-electron microscopy and biochemical analysis, we show that the cerastecins adopt a serpentine configuration in the central vault of the MsbA dimer, stalling the enzyme and uncoupling ATP hydrolysis from substrate flipping. A derivative with optimized potency and pharmacokinetic properties showed efficacy in murine models of bloodstream or pulmonary A. baumannii infection. While resistance development is inevitable, targeting a clinically unexploited mechanism avoids existing antibiotic resistance mechanisms. Although clinical validation of LOS transport remains undetermined, the cerastecins may open a path to narrow-spectrum treatment modalities for important nosocomial infections.


Assuntos
Infecções por Acinetobacter , Acinetobacter baumannii , Antibacterianos , Proteínas de Bactérias , Lipopolissacarídeos , Acinetobacter baumannii/efeitos dos fármacos , Acinetobacter baumannii/metabolismo , Lipopolissacarídeos/metabolismo , Animais , Infecções por Acinetobacter/microbiologia , Infecções por Acinetobacter/tratamento farmacológico , Camundongos , Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Transporte Biológico , Testes de Sensibilidade Microbiana , Humanos , Microscopia Crioeletrônica , Carbapenêmicos/farmacologia , Carbapenêmicos/metabolismo , Modelos Animais de Doenças , Feminino , Transportadores de Cassetes de Ligação de ATP
2.
J Med Chem ; 67(5): 3400-3418, 2024 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-38387069

RESUMO

The use of ß-lactam (BL) and ß-lactamase inhibitor combination to overcome BL antibiotic resistance has been validated through clinically approved drug products. However, unmet medical needs still exist for the treatment of infections caused by Gram-negative (GN) bacteria expressing metallo-ß-lactamases. Previously, we reported our effort to discover pan inhibitors of three main families in this class: IMP, VIM, and NDM. Herein, we describe our work to improve the GN coverage spectrum in combination with imipenem and relebactam. This was achieved through structure- and property-based optimization to tackle the GN cell penetration and efflux challenges. A significant discovery was made that inhibition of both VIM alleles, VIM-1 and VIM-2, is essential for broad GN coverage, especially against VIM-producing P. aeruginosa. In addition, pharmacokinetics and nonclinical safety profiles were investigated for select compounds. Key findings from this drug discovery campaign laid the foundation for further lead optimization toward identification of preclinical candidates.


Assuntos
Antibacterianos , Inibidores de beta-Lactamases , Humanos , Inibidores de beta-Lactamases/farmacologia , Inibidores de beta-Lactamases/uso terapêutico , Inibidores de beta-Lactamases/química , Antibacterianos/química , Imipenem/farmacologia , beta-Lactamases , Bactérias Gram-Negativas , Testes de Sensibilidade Microbiana
3.
J Med Chem ; 65(24): 16234-16251, 2022 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-36475645

RESUMO

With the emergence and rapid spreading of NDM-1 and existence of clinically relevant VIM-1 and IMP-1, discovery of pan inhibitors targeting metallo-beta-lactamases (MBLs) became critical in our battle against bacterial infection. Concurrent with our fragment and high-throughput screenings, we performed a knowledge-based search of known metallo-beta-lactamase inhibitors (MBLIs) to identify starting points for early engagement of medicinal chemistry. A class of compounds exemplified by 11, discovered earlier as B. fragilis metallo-beta-lactamase inhibitors, was selected for in silico virtual screening. From these efforts, compound 12 was identified with activity against NDM-1 only. Initial exploration on metal binding design followed by structure-guided optimization led to the discovery of a series of compounds represented by 23 with a pan MBL inhibition profile. In in vivo studies, compound 23 in combination with imipenem (IPM) robustly lowered the bacterial burden in a murine infection model and became the lead for the invention of MBLI clinical candidates.


Assuntos
Infecções Bacterianas , Inibidores de beta-Lactamases , Animais , Camundongos , Inibidores de beta-Lactamases/farmacologia , Inibidores de beta-Lactamases/uso terapêutico , Inibidores de beta-Lactamases/química , Imipenem/farmacologia , Imipenem/uso terapêutico , beta-Lactamases/metabolismo , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Antibacterianos/química , Testes de Sensibilidade Microbiana
4.
J Clin Tuberc Other Mycobact Dis ; 25: 100285, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34816020

RESUMO

The clinical development and regulatory approval of bedaquiline, delamanid and pretomanid over the last decade brought about significant progress in the management of drug-resistant tuberculosis, providing all-oral regimens with favorable safety profiles. The Nix-TB and ZeNix trials of a bedaquiline - pretomanid - linezolid regimen demonstrated for the first time that certain forms of drug-resistant tuberculosis can be cured in the majority of patients within 6 months. Ongoing Phase 3 studies containing these drugs may further advance optimized regimen compositions. Investigational drugs in clinical development that target clinically validated mechanisms, such as second generation oxazolidinones (sutezolid, delpazolid, TBI-223) and diarylquinolines (TBAJ-876 and TBAJ-587) promise improved potency and/or safety compared to the first-in-class drugs. Compounds with novel targets involved in diverse bacterial functions such as cell wall synthesis (DrpE1, MmpL3), electron transport, DNA synthesis (GyrB), cholesterol metabolism and transcriptional regulation of ethionamide bioactivation pathways have advanced to early clinical studies with the potential to enhance antibacterial activity when added to new or established anti-TB drug regimens. Clinical validation of preclinical in vitro and animal model predictions of new anti-TB regimens may further improve the translational value of these models to identify optimal anti-TB therapies.

5.
Drug Discov Today ; 26(9): 2152-2158, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33798647

RESUMO

The outer membrane (OM) of Gram-negative bacteria confers a significant barrier to many antibacterial agents targeting periplasmic and cytosolic functions. 'Synergist' approaches to disrupt the OM have been hampered by poor specificity and accompanying toxicities. The OM contains proteins required for optimal growth and pathogenesis, including lipopolysaccharide (LPS) and capsular polysaccharide (CPS) transport, porins for uptake of macromolecules, and transporters for essential elements (such as iron). Does the external proximity of these proteins offer an enhanced potential to identify effective therapies? Here, we review recent experiences in exploiting Gram-negative OM proteins (OMPs) to address the calamity of exploding antimicrobial resistance. Teaser: Multidrug-resistant (MDR) Gram-negative bacteria are a growing crisis. Few new antimicrobial chemotypes or targets have been identified after decades of screening. Are OMP targets a solution to MDR Gram-negative bacteria?


Assuntos
Antibacterianos/farmacologia , Proteínas da Membrana Bacteriana Externa/metabolismo , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Animais , Antibacterianos/uso terapêutico , Bactérias Gram-Negativas/metabolismo , Infecções por Bactérias Gram-Negativas/tratamento farmacológico , Humanos
6.
Proc Natl Acad Sci U S A ; 116(43): 21748-21757, 2019 10 22.
Artigo em Inglês | MEDLINE | ID: mdl-31591200

RESUMO

The development of new antimicrobial drugs is a priority to combat the increasing spread of multidrug-resistant bacteria. This development is especially problematic in gram-negative bacteria due to the outer membrane (OM) permeability barrier and multidrug efflux pumps. Therefore, we screened for compounds that target essential, nonredundant, surface-exposed processes in gram-negative bacteria. We identified a compound, MRL-494, that inhibits assembly of OM proteins (OMPs) by the ß-barrel assembly machine (BAM complex). The BAM complex contains one essential surface-exposed protein, BamA. We constructed a bamA mutagenesis library, screened for resistance to MRL-494, and identified the mutation bamAE470K BamAE470K restores OMP biogenesis in the presence of MRL-494. The mutant protein has both altered conformation and activity, suggesting it could either inhibit MRL-494 binding or allow BamA to function in the presence of MRL-494. By cellular thermal shift assay (CETSA), we determined that MRL-494 stabilizes BamA and BamAE470K from thermally induced aggregation, indicating direct or proximal binding to both BamA and BamAE470K Thus, it is the altered activity of BamAE470K responsible for resistance to MRL-494. Strikingly, MRL-494 possesses a second mechanism of action that kills gram-positive organisms. In microbes lacking an OM, MRL-494 lethally disrupts the cytoplasmic membrane. We suggest that the compound cannot disrupt the cytoplasmic membrane of gram-negative bacteria because it cannot penetrate the OM. Instead, MRL-494 inhibits OMP biogenesis from outside the OM by targeting BamA. The identification of a small molecule that inhibits OMP biogenesis at the cell surface represents a distinct class of antibacterial agents.


Assuntos
Antibacterianos/farmacologia , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Escherichia coli/antagonistas & inibidores , Escherichia coli/efeitos dos fármacos , Multimerização Proteica/efeitos dos fármacos , Triazinas/farmacologia , Proteínas da Membrana Bacteriana Externa/antagonistas & inibidores , Proteínas da Membrana Bacteriana Externa/genética , Transporte Biológico/fisiologia , Membrana Celular/efeitos dos fármacos , Permeabilidade da Membrana Celular/fisiologia , Avaliação Pré-Clínica de Medicamentos , Farmacorresistência Bacteriana/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Testes de Sensibilidade Microbiana
7.
Antivir Ther ; 23(7): 593-603, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30038064

RESUMO

BACKGROUND: In HCV-infected people who fail to achieve sustained virological response after receiving a direct-acting antiviral regimen, virological failure is almost always accompanied by the presence of resistance-associated substitutions (RASs) in the target protein(s). The aim of this long-term observational study was to evaluate the persistence of NS3/4A and NS5A RASs in participants with genotype (GT) 1 infection who relapsed following treatment with a grazoprevir-containing treatment regimen. METHODS: RASs were evaluated at baseline (that is, pre-dose on day 1 of the original treatment), at the time of virological failure, and up to follow-up week 96. A total of 58 participants were included. RESULTS: In participants treated with elbasvir/grazoprevir ± ribavirin, observed baseline NS3 RASs included 56F, 80K/L, 122N and 170V/I, and observed treatment-emergent NS3 RASs included 36M, 56F/H, 122G, 132I, 156G/I/L/P/T, 168A/E/G/V/Y and 170T. Observed baseline NS5A RASs included 28M/T/V, 30H/R, 31M/V and 93H/N, and treatment-emergent NS5A RASs included 28A/G/S/T, 30H/R, 31M/V and 93H/N/S. Baseline NS3 and NS5A RASs present at time of failure tended to persist during follow-up, and most were detectable for more than 2 years following virological failure. Treatment-emergent NS5A RASs present at time of failure also tended to persist for more than 2 years following virological failure (93%). By contrast, >80% of treatment-emergent NS3 RASs detected at failure had been supplanted by wild type by week 36. CONCLUSIONS: Treatment-emergent NS5A RASs can persist for extended periods of time. Retreatment strategies should take account of the presence of these RASs.


Assuntos
Antivirais/uso terapêutico , Benzofuranos/uso terapêutico , Hepacivirus/efeitos dos fármacos , Hepatite C Crônica/tratamento farmacológico , Imidazóis/uso terapêutico , Quinoxalinas/uso terapêutico , Serina Proteases/genética , Proteínas não Estruturais Virais/genética , Adulto , Substituição de Aminoácidos , Esquema de Medicação , Combinação de Medicamentos , Farmacorresistência Viral/genética , Feminino , Seguimentos , Expressão Gênica , Genótipo , Hepacivirus/classificação , Hepacivirus/genética , Hepatite C Crônica/etnologia , Hepatite C Crônica/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo Genético , Recidiva , Ribavirina/uso terapêutico , Falha de Tratamento , Carga Viral/efeitos dos fármacos
8.
Proc Natl Acad Sci U S A ; 115(28): E6614-E6621, 2018 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-29941590

RESUMO

The outer membrane (OM) of Gram-negative bacteria forms a robust permeability barrier that blocks entry of toxins and antibiotics. Most OM proteins (OMPs) assume a ß-barrel fold, and some form aqueous channels for nutrient uptake and efflux of intracellular toxins. The Bam machine catalyzes rapid folding and assembly of OMPs. Fidelity of OMP biogenesis is monitored by the σE stress response. When OMP folding defects arise, the proteases DegS and RseP act sequentially to liberate σE into the cytosol, enabling it to activate transcription of the stress regulon. Here, we identify batimastat as a selective inhibitor of RseP that causes a lethal decrease in σE activity in Escherichia coli, and we further identify RseP mutants that are insensitive to inhibition and confer resistance. Remarkably, batimastat treatment allows the capture of elusive intermediates in the OMP biogenesis pathway and offers opportunities to better understand the underlying basis for σE essentiality.


Assuntos
Proteínas da Membrana Bacteriana Externa , Endopeptidases , Proteínas de Escherichia coli , Escherichia coli , Proteínas de Membrana , Desdobramento de Proteína , Fatores de Transcrição , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Endopeptidases/genética , Endopeptidases/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Fatores de Transcrição/metabolismo
9.
PLoS One ; 12(7): e0180965, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28700746

RESUMO

To combat the threat of antibiotic-resistant Gram-negative bacteria, novel agents that circumvent established resistance mechanisms are urgently needed. Our approach was to focus first on identifying bioactive small molecules followed by chemical lead prioritization and target identification. Within this annotated library of bioactives, we identified a small molecule with activity against efflux-deficient Escherichia coli and other sensitized Gram-negatives. Further studies suggested that this compound inhibited DNA replication and selection for resistance identified mutations in a subunit of E. coli DNA gyrase, a type II topoisomerase. Our initial compound demonstrated weak inhibition of DNA gyrase activity while optimized compounds demonstrated significantly improved inhibition of E. coli and Pseudomonas aeruginosa DNA gyrase and caused cleaved complex stabilization, a hallmark of certain bactericidal DNA gyrase inhibitors. Amino acid substitutions conferring resistance to this new class of DNA gyrase inhibitors reside exclusively in the TOPRIM domain of GyrB and are not associated with resistance to the fluoroquinolones, suggesting a novel binding site for a gyrase inhibitor.


Assuntos
Antibacterianos/farmacologia , DNA Girase/metabolismo , Inibidores da Topoisomerase II/farmacologia , Farmacorresistência Bacteriana/genética , Escherichia coli/efeitos dos fármacos , Escherichia coli/enzimologia , Fluoroquinolonas/farmacologia , Testes de Sensibilidade Microbiana , Domínios Proteicos , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/enzimologia
10.
Antimicrob Agents Chemother ; 60(11): 6471-6482, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27527088

RESUMO

Clostridium difficile causes infections of the colon in susceptible patients. Specifically, gut dysbiosis induced by treatment with broad-spectrum antibiotics facilitates germination of ingested C. difficile spores, expansion of vegetative cells, and production of symptom-causing toxins TcdA and TcdB. The current standard of care for C. difficile infections (CDI) consists of administration of antibiotics such as vancomycin that target the bacterium but also perpetuate gut dysbiosis, often leading to disease recurrence. The monoclonal antitoxin antibodies actoxumab (anti-TcdA) and bezlotoxumab (anti-TcdB) are currently in development for the prevention of recurrent CDI. In this study, the effects of vancomycin or actoxumab/bezlotoxumab treatment on progression and resolution of CDI were assessed in mice and hamsters. Rodent models of CDI are characterized by an early severe phase of symptomatic disease, associated with high rates of morbidity and mortality; high intestinal C. difficile burden; and a disrupted intestinal microbiota. This is followed in surviving animals by gradual recovery of the gut microbiota, associated with clearance of C. difficile and resolution of disease symptoms over time. Treatment with vancomycin prevents disease initially by inhibiting outgrowth of C. difficile but also delays microbiota recovery, leading to disease relapse following discontinuation of therapy. In contrast, actoxumab/bezlotoxumab treatment does not impact the C. difficile burden but rather prevents the appearance of toxin-dependent symptoms during the early severe phase of disease, effectively preventing disease until the microbiota (the body's natural defense against C. difficile) has fully recovered. These data provide insight into the mechanism of recurrence following vancomycin administration and into the mechanism of recurrence prevention observed clinically with actoxumab/bezlotoxumab.


Assuntos
Antibacterianos/efeitos adversos , Anticorpos Monoclonais/farmacologia , Anticorpos Neutralizantes/farmacologia , Antitoxinas/farmacologia , Infecções por Clostridium/tratamento farmacológico , Vancomicina/efeitos adversos , Animais , Antibacterianos/administração & dosagem , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/biossíntese , Toxinas Bacterianas/antagonistas & inibidores , Toxinas Bacterianas/biossíntese , Anticorpos Amplamente Neutralizantes , Clostridioides difficile/efeitos dos fármacos , Clostridioides difficile/crescimento & desenvolvimento , Clostridioides difficile/patogenicidade , Infecções por Clostridium/imunologia , Infecções por Clostridium/microbiologia , Infecções por Clostridium/mortalidade , Convalescença , Cricetulus , Modelos Animais de Doenças , Progressão da Doença , Enterotoxinas/antagonistas & inibidores , Enterotoxinas/biossíntese , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/imunologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Análise de Sobrevida , Vancomicina/administração & dosagem
11.
Antimicrob Agents Chemother ; 60(5): 2954-64, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-26926625

RESUMO

The selection of resistance-associated variants (RAVs) against single agents administered to patients chronically infected with hepatitis C virus (HCV) necessitates that direct-acting antiviral agents (DAAs) targeting multiple viral proteins be developed to overcome failure resulting from emergence of resistance. The combination of grazoprevir (formerly MK-5172), an NS3/4A protease inhibitor, and elbasvir (formerly MK-8742), an NS5A inhibitor, was therefore studied in genotype 1a (GT1a) replicon cells. Both compounds were independently highly potent in GT1a wild-type replicon cells, with 90% effective concentration (EC90) values of 0.9 nM and 0.006 nM for grazoprevir and elbasvir, respectively. No cross-resistance was observed when clinically relevant NS5A and NS3 RAVs were profiled against grazoprevir and elbasvir, respectively. Kinetic analyses of HCV RNA reduction over 14 days showed that grazoprevir and elbasvir inhibited prototypic NS5A Y93H and NS3 R155K RAVs, respectively, with kinetics comparable to those for the wild-type GT1a replicon. In combination, grazoprevir and elbasvir interacted additively in GT1a replicon cells. Colony formation assays with a 10-fold multiple of the EC90 values of the grazoprevir-elbasvir inhibitor combination suppressed emergence of resistant colonies, compared to a 100-fold multiple for the independent agents. The selected resistant colonies with the combination harbored RAVs that required two or more nucleotide changes in the codons. Mutations in the cognate gene caused greater potency losses for elbasvir than for grazoprevir. Replicons bearing RAVs identified from resistant colonies showed reduced fitness for several cell lines and may contribute to the activity of the combination. These studies demonstrate that the combination of grazoprevir and elbasvir exerts a potent effect on HCV RNA replication and presents a high genetic barrier to resistance. The combination of grazoprevir and elbasvir is currently approved for chronic HCV infection.


Assuntos
Antivirais/farmacologia , Inibidores de Proteases/farmacologia , Quinoxalinas/farmacologia , Amidas , Benzofuranos/farmacologia , Carbamatos , Ciclopropanos , Quimioterapia Combinada , Genótipo , Hepacivirus/efeitos dos fármacos , Imidazóis/farmacologia , Mutação/genética , Replicon/efeitos dos fármacos , Replicon/genética , Ribavirina/farmacologia , Sulfonamidas
12.
Bioorg Med Chem Lett ; 25(12): 2473-8, 2015 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-25978963

RESUMO

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 química
13.
Bioorg Med Chem Lett ; 25(9): 1831-5, 2015 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-25851938

RESUMO

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/química
14.
ACS Chem Biol ; 8(11): 2442-51, 2013 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-23957438

RESUMO

Modern medicine is founded on the discovery of penicillin and subsequent small molecules that inhibit bacterial peptidoglycan (PG) and cell wall synthesis. However, the discovery of new chemically and mechanistically distinct classes of PG inhibitors has become exceedingly rare, prompting speculation that intracellular enzymes involved in PG precursor synthesis are not 'druggable' targets. Here, we describe a ß-lactam potentiation screen to identify small molecules that augment the activity of ß-lactams against methicillin-resistant Staphylococcus aureus (MRSA) and mechanistically characterize a compound resulting from this screen, which we have named murgocil. We provide extensive genetic, biochemical, and structural modeling data demonstrating both in vitro and in whole cells that murgocil specifically inhibits the intracellular membrane-associated glycosyltransferase, MurG, which synthesizes the lipid II PG substrate that penicillin binding proteins (PBPs) polymerize and cross-link into the cell wall. Further, we demonstrate that the chemical synergy and cidality achieved between murgocil and the ß-lactam imipenem is mediated through MurG dependent localization of PBP2 to the division septum. Collectively, these data validate our approach to rationally identify new target-specific bioactive ß-lactam potentiation agents and demonstrate that murgocil now serves as a highly selective and potent chemical probe to assist our understanding of PG biosynthesis and cell wall biogenesis across Staphylococcal species.


Assuntos
Antibacterianos/farmacologia , Proteínas da Membrana Bacteriana Externa/antagonistas & inibidores , N-Acetilglucosaminiltransferases/antagonistas & inibidores , Peptidoglicano Glicosiltransferase/metabolismo , Pirazóis/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Esteróis/farmacologia , Simulação por Computador , Farmacorresistência Bacteriana , Inibidores Enzimáticos/farmacologia , Humanos , Microscopia de Fluorescência , Modelos Moleculares , Pirazóis/química , Staphylococcus aureus/enzimologia , Esteróis/química
15.
Chem Biol ; 20(2): 272-84, 2013 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-23438756

RESUMO

Innovative strategies are needed to combat drug resistance associated with methicillin-resistant Staphylococcus aureus (MRSA). Here, we investigate the potential of wall teichoic acid (WTA) biosynthesis inhibitors as combination agents to restore ß-lactam efficacy against MRSA. Performing a whole-cell pathway-based screen, we identified a series of WTA inhibitors (WTAIs) targeting the WTA transporter protein, TarG. Whole-genome sequencing of WTAI-resistant isolates across two methicillin-resistant Staphylococci spp. revealed TarG as their common target, as well as a broad assortment of drug-resistant bypass mutants mapping to earlier steps of WTA biosynthesis. Extensive in vitro microbiological analysis and animal infection studies provide strong genetic and pharmacological evidence of the potential effectiveness of WTAIs as anti-MRSA ß-lactam combination agents. This work also highlights the emerging role of whole-genome sequencing in antibiotic mode-of-action and resistance studies.


Assuntos
Antibacterianos/farmacologia , Parede Celular/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Ácidos Teicoicos/biossíntese , beta-Lactamas/metabolismo , Substituição de Aminoácidos , Antibacterianos/química , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Avaliação Pré-Clínica de Medicamentos , Farmacorresistência Bacteriana , Genoma Bacteriano , Staphylococcus aureus Resistente à Meticilina/genética , Testes de Sensibilidade Microbiana , Concentração Osmolar , Fenótipo , Análise de Sequência de DNA , Ácidos Teicoicos/química , Temperatura , beta-Lactamas/química
16.
J Clin Virol ; 55(2): 134-9, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22824230

RESUMO

BACKGROUND: Vicriviroc (VCV), a small-molecule antagonist of the C-C chemokine receptor 5 (CCR5), blocks HIV's entry into CD4+ cells. Small studies have suggested that resistance to CCR5 antagonists is slow to develop. OBJECTIVES: To examine resistance to VCV in isolates from treatment experienced patients who experienced virologic failure in two phase 3 trials. STUDY DESIGN: Genotypic and phenotypic susceptibility to VCV, and other antiretroviral drugs were evaluated at baseline and at defined intervals during the study. In a post hoc analysis, viral tropism at baseline was evaluated using the Trofile-ES assay. Only subjects with R5-tropic virus were included in the analysis. Viral envelope sequencing was performed on samples from subjects with emergent VCV resistance defined using a relative MPI cutoff. RESULTS: 71/486 subjects treated with VCV for 48 weeks met the protocol-defined virologic failure criteria. 7/71 (10%) had DM/X4 virus at the time of virologic failure; VCV resistance was identified in 4/486 treated subjects (1%). No control subject had detectable DM/X4 virus or VCV resistance at virologic failure. Clonal analysis of envelope sequences from VCV-resistant virus identified 2-5 amino acid substitutions at or near the crown of the V3 loop; however, no signature V3 mutations were identified. Changes outside the V3 loop were also observed in resistant clones; no consistent variant pattern was observed. CONCLUSIONS: In these trials, use of a sensitive tropism assay and potent antiretroviral drug combinations contributed to the infrequent detection of X4-tropic virus and VCV resistance. Substitutions in the V3 loop were associated with VCV resistance, however, no specific pattern of amino acid changes were sufficient to reliably predict VCV susceptibility.


Assuntos
Fármacos Anti-HIV/administração & dosagem , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Piperazinas/administração & dosagem , Pirimidinas/administração & dosagem , Tropismo Viral , Ensaios Clínicos Fase III como Assunto , Genótipo , HIV-1/isolamento & purificação , Humanos , Testes de Sensibilidade Microbiana , Fenótipo , RNA Viral/genética , Análise de Sequência de DNA , Falha de Tratamento , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética
18.
Bioorg Med Chem Lett ; 22(14): 4896-9, 2012 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-22687744

RESUMO

The structure-activity relationship studies of a novel sulfonylurea series of piperazine pyridazine-based small molecule glucan synthase inhibitors is described. The optimization of PK profiles within the series led to the discovery of several compounds with improved pharmacokinetic profiles which demonstrated in vitro potency against clinically relevant strains. However, the advancement of compounds from this series into a non-lethal systemic fungal infection model failed to show in vivo efficacy.


Assuntos
Antifúngicos/química , Inibidores Enzimáticos/química , Glucosiltransferases/antagonistas & inibidores , Chumbo/química , Piperazinas/química , Piridazinas/química , Compostos de Sulfonilureia/química , Animais , Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Linhagem Celular , Inibidores Enzimáticos/farmacologia , Humanos , Estrutura Molecular , Piperazina , Piridazinas/farmacologia , Ratos , Relação Estrutura-Atividade , Compostos de Sulfonilureia/farmacologia
19.
Antimicrob Agents Chemother ; 55(11): 5099-106, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21844320

RESUMO

The echinocandins are a class of semisynthetic natural products that target ß-1,3-glucan synthase (GS). Their proven clinical efficacy combined with minimal safety issues has made the echinocandins an important asset in the management of fungal infection in a variety of patient populations. However, the echinocandins are delivered only parenterally. A screen for antifungal bioactivities combined with mechanism-of-action studies identified a class of piperazinyl-pyridazinones that target GS. The compounds exhibited in vitro activity comparable, and in some cases superior, to that of the echinocandins. The compounds inhibit GS in vitro, and there was a strong correlation between enzyme inhibition and in vitro antifungal activity. In addition, like the echinocandins, the compounds caused a leakage of cytoplasmic contents from yeast and produced a morphological response in molds characteristic of GS inhibitors. Spontaneous mutants of Saccharomyces cerevisiae with reduced susceptibility to the piperazinyl-pyridazinones had substitutions in FKS1. The sites of these substitutions were distinct from those conferring resistance to echinocandins; likewise, echinocandin-resistant isolates remained susceptible to the test compounds. Finally, we present efficacy and pharmacokinetic data on an example of the piperazinyl-pyridazinone compounds that demonstrated efficacy in a murine model of Candida glabrata infection.


Assuntos
Antifúngicos/farmacologia , Glucosiltransferases/antagonistas & inibidores , Animais , Antifúngicos/química , Candida glabrata/efeitos dos fármacos , Candida glabrata/enzimologia , Candida glabrata/patogenicidade , Candidíase/tratamento farmacológico , Masculino , Camundongos , Estrutura Molecular , Piperazinas/química , Piperazinas/farmacologia , Piridazinas/química , Piridazinas/farmacologia , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/enzimologia
20.
Bioorg Med Chem Lett ; 21(10): 2890-3, 2011 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-21489787

RESUMO

A novel series of pyridazinone analogs has been developed as potent ß-1,3-glucan synthase inhibitors through structure-activity relationship study of the lead 5-[4-(benzylsulfonyl)piperazin-1-yl]-4-morpholino-2-phenyl-pyridazin-3(2H)-one (1). The effect of changes to the core structure is described in detail. Optimization of the sulfonamide moiety led to the identification of important compounds with much improved systematic exposure while retaining good antifungal activity against the fungal strains Candida glabrata and Candida albicans.


Assuntos
Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Glucosiltransferases/antagonistas & inibidores , Piridazinas/síntese química , Piridazinas/farmacologia , Antifúngicos/síntese química , Antifúngicos/química , Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Candida glabrata/efeitos dos fármacos , Inibidores Enzimáticos/química , Estrutura Molecular , Piridazinas/química , Relação Estrutura-Atividade
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