1,721,054 research outputs found
AmpC beta-lactamase in complex with 3-(3-nitro-phenylsulfamoyl)-thiophene-2-carboxylic acid
No full textBacterial expression of beta-lactamases is the most widespread resistance mechanism to beta-lactam antibiotics, such as penicillins and cephalosporins. There is a pressing need for novel, non-beta-lactam inhibitors of these enzymes. One previously discovered novel inhibitor of the beta-lactamase AmpC, compound 1, has several favorable properties: it is chemically dissimilar to beta-lactams and is a noncovalent, competitive inhibitor of the enzyme. However, at 26 microM its activity is modest. Using the X-ray structure of the AmpC/1 complex as a template, 14 analogues were designed and synthesized. The most active of these, compound 10, had a K(i) of 1 microM, 26-fold better than the lead. To understand the origins of this improved activity, the structures of AmpC in complex with compound 10 and an analogue, compound 11, were determined by X-ray crystallography to 1.97 and 1.96 A, respectively. Compound 10 was active in cell culture, reversing resistance to the third generation cephalosporin ceftazidime in bacterial pathogens expressing AmpC. In contrast to beta-lactam-based inhibitors clavulanate and cefoxitin, compound 10 did not up-regulate beta-lactamase expression in cell culture but simply inhibited the enzyme expressed by the resistant bacteria. Its escape from this resistance mechanism derives from its dissimilarity to beta-lactam antibiotics
CTM-M-9 in complex with the broad spectrum inhibitor 3-(2- carboxyvinyl)benzo(b)thiophene-2-boronic acid
No full textProduction of β-lactamases (BLs) is the most widespread resistance mechanism adopted by bacteria to fight β-lactam antibiotics. The substrate spectrum of BLs has become increasingly broad, posing a serious health problem. Thus, there is an urgent need for novel BL inhibitors. Boronic acid transition-state analogues are able to reverse the resistance conferred by class A and C BLs. We describe a boronic acid analogue possessing interesting and potent broad-spectrum activity vs class A and C serine-based BLs. Starting from benzo(b)thiophene-2-boronic acid (BZBTH2B), a nanomolar non-β-lactam inhibitor of AmpC that can potentiate the activity of a third-generation cephalosporin against AmpC-producing resistant bacteria, we designed a novel broad-spectrum nanomolar inhibitor of class A and C BLs. Structure-based drug design (SBDD), synthesis, enzymology data, and X-ray crystallography results are discussed. We clarified the inhibitor binding geometry responsible for broad-spectrum activity vs serine-active BLs using double mutant thermodynamic cycle studies
LexA e la risposta SOS nei batteri: design e ottimizzazione di inibitori contrastanti lo sviluppo della resistenza batterica.
No full textProgetto FAR 2017 UNIMORE PI DOnatella Tondi
Abstract
La diffusione di batteri resistenti agli antimicrobici rappresenta una riconosciuta emergenza mondiale che limita sempre più il trattamento delle infezioni batteriche: l’OMS considera la resistenza batterica agli antibiotici fra i principali rischi per la salute umana.
Il progetto ha come oggetto di studio il repressore trascrizionale LexA coinvolto nella risposta SOS, un importante sistema di riparazione del DNA presente nei batteri. In seguito a stress esterno (es. trattamento antibiotico) nei batteri viene attivata l’auto-proteolisi di LexA e come conseguenza la risposta SOS: viene così indotta l'espressione di oltre 60 geni coinvolti, per esempio, nella riparazione e ricombinazione del DNA.
E’ stato dimostrato che il blocco della risposta SOS riduce de facto l'acquisizione e l'integrazione di nuovi geni di resistenza, limitando così le capacità dei batteri di evolvere in cellule resistenti.
Il progetto, avvalendosi di un approccio di Structure-Based, propone l’identificazione di inibitori di LexA e della loro successiva ottimizzazione sintetica hit-to-lead. Le molecole candidate verranno validate in vitro vs LexA e le migliori avviate a studi ex vivo vs i batteri per valutarne la capacità di bloccare la risposta SOS. Per le molecole più promettenti saranno condotti studi di cristallizzazione a raggi-x.
Presso il proponente sono stati avviati studi di validazione di LexA come proteina target: è stato sviluppato un modello cinetico di analisi, la produzione della proteina transgenica è stata messa a punto ed un saggio cellulare è già disponibile.
L’obiettivo finale che il progetto si prefigge è ritardare lo sviluppo di batteri resistenti intervenendo a livello del trasferimento orizzontale e verticale dei geni di resistenza. In questo contesto il blocco dell’attività di LexA e della risposta SOS consentirebbe di riabilitare in terapia numerosi antibiotici ormai abbandonati e di prolungare la vita di quelli di nuovo sviluppo
AmpC beta-lactamase in complex with a p.carboxyphenylboronic acid
No full textA small set of boronic acids acting as low nanomolar inhibitors of AmpC beta-lactamase were designed and synthesized in the effort to improve affinity, pharmacokinetic properties, and to provide a valid lead compound. X-ray crystallography revealed the binary complex of the best inhibitor bound to the enzyme, highlighting possibilities for its further rational derivatization and chemical optimization
Designing novel inhibitors for Metallo beta-lactamases.
No full textThe extensive use of beta-lactam antibiotics has created major resistance problems leading to increased morbidity, mortality and health-care costs. Resistance is most often mediated by beta-lactamases (BLs), which have emerged in both Gram-positive and Gram-negative bacteria [1,2]. Medicinal chemists have introduced beta-lactam-based molecules that inhibit or are stable to their action. These molecules are themselves beta-lactams, making it easier for bacteria to respond by adapting previously evolved mechanisms. Many bacteria are now resistant to these anti-resistance compounds. Thus there is a growing need for new broad-spectrum beta-lactamases inhibitors in general and especially against Metallo BLs (MBLs)[3,4]. Their substrate promiscuity, their resistance to available drugs, the easiness of variants appearance and transferability make MBLs the most worrisome BLs [2].
Focusing on the de novo, non beta-lactam like derivatives we identified, through a structure-based in silico screening of commercially available library using FLAP, several promising candidates active against class B beta-lactamases (MBL). The binding affinities of the high scoring hits were measured in vitro revealing, for some of them, low micromolar affinity towards BLs.
To investigate the potential of these compounds to reverse antibiotic resistance, we are undertaking antimicrobial activity studies in bacterial cell culture. The ability of novel compounds to synergize antibiotics against pathogenic resistant bacteria, as well as their ability to evade those mechanisms normally involved in resistance to beta-lactam-based inhibitors are now under evaluation. Moreover, since our inhibitors are novel, non beta-lactam based, we expect them to do not up-regulate beta-lactamase expression in cell culture.
X-ray crystallography studies are now in progress to confirm our docking prediction and to deeply investigate the structural requirement necessary for proficuous hit to lead generation
AmpC beta-lactamase in complex with 5-diformylaminomethyl-benzo[b]thiophen-2-boronic acid
No full textBenzo[b]thiophene-2-ylboronic acid, 1, is a 27 nM inhibitor of the class C beta-lactamase AmpC and potentiates the activity of beta-lactam antibiotics in bacteria that express this and related enzymes. As is often true, the potency of compound 1 against the enzymes is much attenuated in cell culture against Gram negative bacteria, where the minimum inhibitor concentration of compound 1 is in the mid-micromolar range. Here, we modulated the properties of this lead to enhance its ability to cross the membrane, using a combination of X-ray crystallography, structure-based design, and application of physical models of outer membrane crossing. This strategy led us to derivatives with substantially improved permeability. Also, the greater solubility of these compounds allowed us to measure their efficacy at higher concentrations than with the lead 1, leading to higher maximum potentiation of the antibiotic effect of ceftazidime on resistant bacteria
Structure-based discovery and combinatorial optimization of novel competitive inhibitors of thymidylate synthase
No full textabstract presentazione oral
Enhancement of the drug discovery process by integration of Structure-based drug design and combinatorial chemistry
No full textThis text traces developments in rational drug discovery and combinatorial library design with contributions from 50 leading scientists in academia and industry who offer coverage of basic principles, design strategies, methodologies, software tools and algorithms, and applications. It outlines the fundamentals of pharmacophore modelling and 3D Quantitative Structure-Activity Relationships (QSAR), classical QSAR, and target protein structure-based design methods
Structure-based inhibitor design vs. beta-lactamase
No full textFragment-based screens test multiple low-molecular weight molecules for binding to a target. Fragments often bind with low affinities but typically have better ligand efficiencies (DeltaG(bind)/heavy atom count) than traditional screening hits. This efficiency, combined with accompanying atomic-resolution structures, has made fragments popular starting points for drug discovery programs. Fragment-based design adopts a constructive strategy: affinity is enhanced either by cycles of functional-group addition or by joining two independent fragments together. The final inhibitor is expected to adopt the same geometry as the original fragment hit. Here we consider whether the inverse, deconstructive logic also applies--can one always parse a higher-affinity inhibitor into fragments that recapitulate the binding geometry of the larger molecule? Cocrystal structures of fragments deconstructed from a known beta-lactamase inhibitor suggest that this is not always the case
Phenylboronic acid derivative inhibitors of ?-lactamases, their preparation, pharmaceutical compositions, and therapeutic use.
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