Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 84
Filtrar
Mais filtros

Base de dados
Tipo de documento
Intervalo de ano de publicação
1.
Nucleic Acids Res ; 51(1): 144-165, 2023 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-36546765

RESUMO

The emergence of drug-resistant Mycobacterium tuberculosis strains highlights the need to discover anti-tuberculosis drugs with novel mechanisms of action. Here we discovered a mycobactericidal strategy based on the prodrug activation of selected chemical derivatives classified as nitronaphthofurans (nNFs) mediated by the coordinated action of the sigH and mrx2 genes. The transcription factor SigH is a key regulator of an extensive transcriptional network that responds to oxidative, nitrosative, and heat stresses in M. tuberculosis. The nNF action induced the SigH stress response which in turn induced the mrx2 overexpression. The nitroreductase Mrx2 was found to activate nNF prodrugs, killing replicating, non-replicating and intracellular forms of M. tuberculosis. Analysis of SigH DNA sequences obtained from spontaneous nNF-resistant M. tuberculosis mutants suggests disruption of SigH binding to the mrx2 promoter site and/or RNA polymerase core, likely promoting the observed loss of transcriptional control over Mrx2. Mutations found in mrx2 lead to structural defects in the thioredoxin fold of the Mrx2 protein, significantly impairing the activity of the Mrx2 enzyme against nNFs. Altogether, our work brings out the SigH/Mrx2 stress response pathway as a promising target for future drug discovery programs.


Assuntos
Antibacterianos , Mycobacterium tuberculosis , Pró-Fármacos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Resposta ao Choque Térmico/genética , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/metabolismo , Pró-Fármacos/farmacologia , Regiões Promotoras Genéticas , Transcrição Gênica , Antibacterianos/farmacologia
2.
Vet Res ; 55(1): 32, 2024 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-38493182

RESUMO

Outbreaks of West Nile virus (WNV) occur periodically, affecting both human and equine populations. There are no vaccines for humans, and those commercialised for horses do not have sufficient coverage. Specific antiviral treatments do not exist. Many drug discovery studies have been conducted, but since rodent or primate cell lines are normally used, results cannot always be transposed to horses. There is thus a need to develop relevant equine cellular models. Here, we used induced pluripotent stem cells to develop a new in vitro model of WNV-infected equine brain cells suitable for microplate assay, and assessed the cytotoxicity and antiviral activity of forty-one chemical compounds. We found that one nucleoside analog, 2'C-methylcytidine, blocked WNV infection in equine brain cells, whereas other compounds were either toxic or ineffective, despite some displaying anti-viral activity in human cell lines. We also revealed an unexpected proviral effect of statins in WNV-infected equine brain cells. Our results thus identify a potential lead for future drug development and underscore the importance of using a tissue- and species-relevant cellular model for assessing the activity of antiviral compounds.


Assuntos
Doenças dos Cavalos , Células-Tronco Pluripotentes Induzidas , Febre do Nilo Ocidental , Vírus do Nilo Ocidental , Animais , Cavalos , Humanos , Febre do Nilo Ocidental/veterinária , Febre do Nilo Ocidental/epidemiologia , Encéfalo , Antivirais/farmacologia , Antivirais/uso terapêutico , Doenças dos Cavalos/tratamento farmacológico
3.
Nucleic Acids Res ; 49(13): 7695-7712, 2021 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-34232992

RESUMO

The multidomain non-structural protein 3 (Nsp3) is the largest protein encoded by coronavirus (CoV) genomes and several regions of this protein are essential for viral replication. Of note, SARS-CoV Nsp3 contains a SARS-Unique Domain (SUD), which can bind Guanine-rich non-canonical nucleic acid structures called G-quadruplexes (G4) and is essential for SARS-CoV replication. We show herein that the SARS-CoV-2 Nsp3 protein also contains a SUD domain that interacts with G4s. Indeed, interactions between SUD proteins and both DNA and RNA G4s were evidenced by G4 pull-down, Surface Plasmon Resonance and Homogenous Time Resolved Fluorescence. These interactions can be disrupted by mutations that prevent oligonucleotides from folding into G4 structures and, interestingly, by molecules known as specific ligands of these G4s. Structural models for these interactions are proposed and reveal significant differences with the crystallographic and modeled 3D structures of the SARS-CoV SUD-NM/G4 interaction. Altogether, our results pave the way for further studies on the role of SUD/G4 interactions during SARS-CoV-2 replication and the use of inhibitors of these interactions as potential antiviral compounds.


Assuntos
COVID-19/virologia , Proteases Semelhantes à Papaína de Coronavírus/metabolismo , Quadruplex G , Domínios e Motivos de Interação entre Proteínas , SARS-CoV-2 , Sequência de Aminoácidos , Proteases Semelhantes à Papaína de Coronavírus/química , Humanos , Ligantes , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Análise Espectral , Relação Estrutura-Atividade , Replicação Viral
4.
Int J Mol Sci ; 24(19)2023 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-37834238

RESUMO

Infection with Ebola virus (EBOV) is responsible for hemorrhagic fever in humans with a high mortality rate. Combined efforts of prevention and therapeutic intervention are required to tackle highly variable RNA viruses, whose infections often lead to outbreaks. Here, we have screened the 2P2I3D chemical library using a nanoluciferase-based protein complementation assay (NPCA) and isolated two compounds that disrupt the interaction of the EBOV protein fragment VP35IID with the N-terminus of the dsRNA-binding proteins PKR and PACT, involved in IFN response and/or intrinsic immunity, respectively. The two compounds inhibited EBOV infection in cell culture as well as infection by measles virus (MV) independently of IFN induction. Consequently, we propose that the compounds are antiviral by restoring intrinsic immunity driven by PACT. Given that PACT is highly conserved across mammals, our data support further testing of the compounds in other species, as well as against other negative-sense RNA viruses.


Assuntos
Ebolavirus , Doença pelo Vírus Ebola , Humanos , Animais , Doença pelo Vírus Ebola/tratamento farmacológico , Doença pelo Vírus Ebola/metabolismo , Ebolavirus/fisiologia , Antivirais/farmacologia , Antivirais/uso terapêutico , Mamíferos
5.
Molecules ; 28(3)2023 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-36770826

RESUMO

The chemokine receptor CXCR4 and its ligand CXCL12 regulate leukocyte trafficking, homeostasis and functions and are potential therapeutic targets in many diseases such as HIV-1 infection and cancers. Here, we identified new CXCR4 ligands in the CERMN chemical library using a FRET-based high-throughput screening assay. These are bis-imidazoline compounds comprising two imidazole rings linked by an alkyl chain. The molecules displace CXCL12 binding with submicromolar potencies, similarly to AMD3100, the only marketed CXCR4 ligand. They also inhibit anti-CXCR4 mAb 12G5 binding, CXCL12-mediated chemotaxis and HIV-1 infection. Further studies with newly synthesized derivatives pointed out to a role of alkyl chain length on the bis-imidazoline properties, with molecules with an even number of carbons equal to 8, 10 or 12 being the most potent. Interestingly, these differ in the functions of CXCR4 that they influence. Site-directed mutagenesis and molecular docking predict that the alkyl chain folds in such a way that the two imidazole groups become lodged in the transmembrane binding cavity of CXCR4. Results also suggest that the alkyl chain length influences how the imidazole rings positions in the cavity. These results may provide a basis for the design of new CXCR4 antagonists targeting specific functions of the receptor.


Assuntos
Imidazolinas , Transdução de Sinais , Ligantes , Simulação de Acoplamento Molecular , Receptores CXCR4 , Imidazóis/farmacologia
6.
Molecules ; 25(12)2020 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-32560578

RESUMO

A series of Mycobacterium tuberculosis TMPK (MtbTMPK) inhibitors based on a reported compound 3 were synthesized and evaluated for their capacity to inhibit MtbTMPK catalytic activity and the growth of a virulent M. tuberculosis strain (H37Rv). Modifications of the scaffold of 3 failed to afford substantial improvements in MtbTMPK inhibitory activity and antimycobacterial activity. Optimization of the substitution pattern of the D ring of 3 resulted in compound 21j with improved MtbTMPK inhibitory potency (three-fold) and H37Rv growth inhibitory activity (two-fold). Moving the 3-chloro substituent of 21j to the para-position afforded isomer 21h, which, despite a 10-fold increase in IC50-value, displayed promising whole cell activity (minimum inhibitory concentration (MIC) = 12.5 µM).


Assuntos
Antituberculosos , Proteínas de Bactérias/antagonistas & inibidores , Inibidores Enzimáticos , Mycobacterium tuberculosis/enzimologia , Núcleosídeo-Fosfato Quinase/antagonistas & inibidores , Timina , Antituberculosos/síntese química , Antituberculosos/química , Antituberculosos/farmacologia , Proteínas de Bactérias/metabolismo , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Humanos , Modelos Moleculares , Estrutura Molecular , Núcleosídeo-Fosfato Quinase/metabolismo , Relação Estrutura-Atividade , Timina/análogos & derivados , Timina/síntese química , Timina/química , Timina/farmacologia
7.
J Enzyme Inhib Med Chem ; 34(1): 1730-1739, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31822127

RESUMO

A series of readily accessible 1-(piperidin-3-yl)thymine amides was designed, synthesised and evaluated as Mycobacterium tuberculosis TMPK (MtbTMPK) inhibitors. In line with the modelling results, most inhibitors showed reasonable MtbTMPK inhibitory activity. Compounds 4b and 4i were slightly more potent than the parent compound 3. Moreover, contrary to the latter, amide analogue 4g was active against the avirulent M. tuberculosis H37Ra strain (MIC50=35 µM). This finding opens avenues for future modifications.


Assuntos
Amidas/farmacologia , Antituberculosos/farmacologia , Inibidores Enzimáticos/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Núcleosídeo-Fosfato Quinase/antagonistas & inibidores , Timina/farmacologia , Amidas/síntese química , Amidas/química , Antituberculosos/síntese química , Antituberculosos/química , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Testes de Sensibilidade Microbiana , Estrutura Molecular , Mycobacterium tuberculosis/enzimologia , Núcleosídeo-Fosfato Quinase/metabolismo , Relação Estrutura-Atividade , Timina/síntese química , Timina/química
8.
Artigo em Inglês | MEDLINE | ID: mdl-29632009

RESUMO

Aspergillus fumigatus can cause pulmonary aspergillosis in immunocompromised patients and is associated with a high mortality rate due to a lack of reliable treatment options. This opportunistic pathogen requires zinc in order to grow and cause disease. Novel compounds that interfere with fungal zinc metabolism may therefore be of therapeutic interest. We screened chemical libraries containing 59,223 small molecules using a resazurin assay that compared their effects on an A. fumigatus wild-type strain grown under zinc-limiting conditions and on a zinc transporter knockout strain grown under zinc-replete conditions to identify compounds affecting zinc metabolism. After a first screen, 116 molecules were selected whose inhibitory effects on fungal growth were further tested by using luminescence assays and hyphal length measurements to confirm their activity, as well as by toxicity assays on HeLa cells and mice. Six compounds were selected following a rescreening, of which two were pyrazolones, two were porphyrins, and two were polyaminocarboxylates. All three groups showed good in vitro activity, but only one of the polyaminocarboxylates was able to significantly improve the survival of immunosuppressed mice suffering from pulmonary aspergillosis. This two-tier screening approach led us to the identification of a novel small molecule with in vivo fungicidal effects and low murine toxicity that may lead to the development of new treatment options for fungal infections by administration of this compound either as a monotherapy or as part of a combination therapy.


Assuntos
Antifúngicos/uso terapêutico , Aspergillus fumigatus/efeitos dos fármacos , Aspergillus fumigatus/patogenicidade , Aspergilose Pulmonar/tratamento farmacológico , Aspergilose Pulmonar/metabolismo , Zinco/metabolismo , Animais , Modelos Animais de Doenças , Medições Luminescentes , Camundongos , Testes de Sensibilidade Microbiana , Pirazolonas/uso terapêutico
9.
Artigo em Inglês | MEDLINE | ID: mdl-28807907

RESUMO

De novo pyrimidine biosynthesis is a key metabolic pathway involved in multiple biosynthetic processes. Here, we identified an original series of 3-(1H-indol-3-yl)-2,3-dihydro-4H-furo[3,2-c]chromen-4-one derivatives as a new class of pyrimidine biosynthesis inhibitors formed by two edge-fused polycyclic moieties. We show that identified compounds exhibit broad-spectrum antiviral activity and immunostimulatory properties, in line with recent reports linking de novo pyrimidine biosynthesis with innate defense mechanisms against viruses. Most importantly, we establish that pyrimidine deprivation can amplify the production of both type I and type III interferons by cells stimulated with retinoic acid-inducible gene 1 (RIG-I) ligands. Altogether, our results further expand the current panel of pyrimidine biosynthesis inhibitors and illustrate how the production of antiviral interferons is tightly coupled to this metabolic pathway. Functional and structural similarities between this new chemical series and dicoumarol, which was reported before to inhibit pyrimidine biosynthesis at the dihydroorotate dehydrogenase (DHODH) step, are discussed.


Assuntos
Antivirais/farmacologia , Vírus Chikungunya/imunologia , Cromonas/farmacologia , Inibidores Enzimáticos/farmacologia , Indóis/farmacologia , Interferon Tipo I/biossíntese , Interferons/biossíntese , Vírus do Sarampo/imunologia , Pirimidinas/biossíntese , Antivirais/química , Linhagem Celular , Cromonas/química , Dicumarol/farmacologia , Di-Hidro-Orotato Desidrogenase , Inibidores Enzimáticos/química , Células HEK293 , Humanos , Imunidade Inata/imunologia , Indóis/química , Interferon Tipo I/imunologia , Interferons/imunologia , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Relação Estrutura-Atividade , Interferon lambda
10.
Antimicrob Agents Chemother ; 60(10): 5631-9, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27401578

RESUMO

Aspergillus fumigatus can infect immunocompromised patients, leading to high mortality rates due to the lack of reliable treatment options. This pathogen requires uptake of zinc from host tissues in order to successfully grow and cause virulence. Reducing the availability of that micronutrient could help treat A. fumigatus infections. In this study, we examined the in vitro effects of seven chelators using a bioluminescent strain of A. fumigatus 1,10-Phenanthroline and N,N,N',N'-tetrakis(2-pyridylmethyl)ethane-1,2-diamine (TPEN) proved to be the chelators most effective at inhibiting fungal growth. Intraperitoneal administration of either phenanthroline or TPEN resulted in a significant improvement in survival and decrease of weight loss and fungal burden for immunosuppressed mice intranasally infected with A. fumigatus In vitro both chelators had an indifferent effect when employed in combination with caspofungin. The use of TPEN in combination with caspofungin also significantly increased survival compared to that when using these drugs individually. Our results suggest that zinc chelation may be a valid strategy for dealing with A. fumigatus infections and that both phenanthroline and TPEN could potentially be used either independently or in combination with caspofungin, indicating that their use in combination with other antifungal treatments might also be applicable.


Assuntos
Antifúngicos/farmacologia , Aspergilose/tratamento farmacológico , Aspergillus fumigatus/efeitos dos fármacos , Quelantes/farmacologia , Equinocandinas/farmacologia , Lipopeptídeos/farmacologia , Animais , Aspergilose/imunologia , Aspergilose/microbiologia , Aspergilose/mortalidade , Aspergillus fumigatus/crescimento & desenvolvimento , Aspergillus fumigatus/patogenicidade , Caspofungina , Clioquinol/farmacologia , Quimioterapia Combinada , Etilaminas/farmacologia , Soros Imunes/farmacologia , Hospedeiro Imunocomprometido , Masculino , Camundongos Endogâmicos BALB C , Fenantrolinas/farmacologia , Pneumonia/microbiologia , Pneumonia/patologia , Piridinas/farmacologia , Resultado do Tratamento , Zinco
12.
Acta Crystallogr D Biol Crystallogr ; 71(Pt 9): 1890-9, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26327379

RESUMO

Inosine-5'-monophosphate dehydrogenases (IMPDHs), which are the rate-limiting enzymes in guanosine-nucleotide biosynthesis, are important therapeutic targets. Despite in-depth functional and structural characterizations of various IMPDHs, the role of the Bateman domain containing two CBS motifs remains controversial. Their involvement in the allosteric regulation of Pseudomonas aeruginosa IMPDH by Mg-ATP has recently been reported. To better understand the function of IMPDH and the importance of the CBS motifs, the structure of a variant devoid of these modules (ΔCBS) was solved at high resolution in the apo form and in complex with IMP. In addition, a single amino-acid substitution variant, D199N, was also structurally characterized: the mutation corresponds to the autosomal dominant mutant D226N of human IMPDH1, which is responsible for the onset of the retinopathy adRP10. These new structures shed light onto the possible mechanism of regulation of the IMPDH enzymatic activity. In particular, three conserved loops seem to be key players in this regulation as they connect the tetramer-tetramer interface with the active site and show significant modification upon substrate binding.


Assuntos
IMP Desidrogenase/química , Pseudomonas aeruginosa/enzimologia , Regulação Alostérica , Sequência de Aminoácidos , Cristalografia por Raios X , Dados de Sequência Molecular , Conformação Proteica , Homologia de Sequência de Aminoácidos
13.
Antimicrob Agents Chemother ; 60(3): 1438-49, 2015 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-26666917

RESUMO

In a search for new antifungal compounds, we screened a library of 4,454 chemicals for toxicity against the human fungal pathogen Aspergillus fumigatus. We identified sr7575, a molecule that inhibits growth of the evolutionary distant fungi A. fumigatus, Cryptococcus neoformans, Candida albicans, and Saccharomyces cerevisiae but lacks acute toxicity for mammalian cells. To gain insight into the mode of inhibition, sr7575 was screened against 4,885 S. cerevisiae mutants from the systematic collection of haploid deletion strains and 977 barcoded haploid DAmP (decreased abundance by mRNA perturbation) strains in which the function of essential genes was perturbed by the introduction of a drug resistance cassette downstream of the coding sequence region. Comparisons with previously published chemogenomic screens revealed that the set of mutants conferring sensitivity to sr7575 was strikingly narrow, affecting components of the endoplasmic reticulum-associated protein degradation (ERAD) stress response and the ER membrane protein complex (EMC). ERAD-deficient mutants were hypersensitive to sr7575 in both S. cerevisiae and A. fumigatus, indicating a conserved mechanism of growth inhibition between yeast and filamentous fungi. Although the unfolded protein response (UPR) is linked to ERAD regulation, sr7575 did not trigger the UPR in A. fumigatus and UPR mutants showed no enhanced sensitivity to the compound. The data from this chemogenomic analysis demonstrate that sr7575 exerts its antifungal activity by disrupting ER protein quality control in a manner that requires ERAD intervention but bypasses the need for the canonical UPR. ER protein quality control is thus a specific vulnerability of fungal organisms that might be exploited for antifungal drug development.


Assuntos
Antifúngicos/farmacologia , Antifúngicos/toxicidade , Aspergillus fumigatus/efeitos dos fármacos , Degradação Associada com o Retículo Endoplasmático/efeitos dos fármacos , Animais , Aspergillus fumigatus/genética , Candida albicans/efeitos dos fármacos , Cryptococcus neoformans/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Degradação Associada com o Retículo Endoplasmático/genética , Células HeLa/efeitos dos fármacos , Humanos , Camundongos Endogâmicos , Testes de Sensibilidade Microbiana , Mutação , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Resposta a Proteínas não Dobradas/efeitos dos fármacos
14.
PLoS Pathog ; 9(10): e1003678, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24098125

RESUMO

Searching for stimulators of the innate antiviral response is an appealing approach to develop novel therapeutics against viral infections. Here, we established a cell-based reporter assay to identify compounds stimulating expression of interferon-inducible antiviral genes. DD264 was selected out of 41,353 compounds for both its immuno-stimulatory and antiviral properties. While searching for its mode of action, we identified DD264 as an inhibitor of pyrimidine biosynthesis pathway. This metabolic pathway was recently identified as a prime target of broad-spectrum antiviral molecules, but our data unraveled a yet unsuspected link with innate immunity. Indeed, we showed that DD264 or brequinar, a well-known inhibitor of pyrimidine biosynthesis pathway, both enhanced the expression of antiviral genes in human cells. Furthermore, antiviral activity of DD264 or brequinar was found strictly dependent on cellular gene transcription, nuclear export machinery, and required IRF1 transcription factor. In conclusion, the antiviral property of pyrimidine biosynthesis inhibitors is not a direct consequence of pyrimidine deprivation on the virus machinery, but rather involves the induction of cellular immune response.


Assuntos
Infecções por Alphavirus/metabolismo , Antivirais/farmacologia , Vírus Chikungunya/metabolismo , Imunidade Inata/efeitos dos fármacos , Pirimidinas/biossíntese , Infecções por Alphavirus/tratamento farmacológico , Infecções por Alphavirus/genética , Infecções por Alphavirus/imunologia , Animais , Antivirais/química , Febre de Chikungunya , Vírus Chikungunya/genética , Vírus Chikungunya/imunologia , Chlorocebus aethiops , Células HeLa , Humanos , Imunidade Inata/imunologia , Fator Regulador 1 de Interferon/genética , Fator Regulador 1 de Interferon/imunologia , Fator Regulador 1 de Interferon/metabolismo , Pirimidinas/imunologia , Células Vero
15.
Med Sci (Paris) ; 31(1): 98-104, 2015 Jan.
Artigo em Francês | MEDLINE | ID: mdl-25658737

RESUMO

RNA viruses are responsible for major human diseases such as flu, bronchitis, dengue, hepatitis C or measles. They also represent an emerging threat because of increased worldwide exchanges and human populations penetrating more and more natural ecosystems. Recent progresses in our understanding of cellular pathways controlling viral replication suggest that compounds targeting host cell functions, rather than the virus itself, could inhibit a large panel of RNA viruses. In particular, several academic laboratories and private companies are now seeking molecules that stimulate the host innate antiviral response. One appealing strategy is to identify molecules that induce the large cluster of antiviral genes known as Interferon-Stimulated Genes (ISGs). To reach this goal, we have developed a phenotypic assay based on human cells transfected with a luciferase reporter gene under control of an interferon-stimulated response element (ISRE). This system was used in a high-throughput screening of chemical libraries comprising around 54,000 compounds. Among validated hits, compound DD264 was shown to boost the innate immune response in cell cultures, and displayed a broad-spectrum antiviral activity. While deciphering its mode of action, DD264 was found to target the fourth enzyme of de novo pyrimidine biosynthesis, namely the dihydroorotate dehydrogenase (DHODH). Thus, our data unraveled a yet unsuspected link between pyrimidine biosynthesis and the innate antiviral response.


Assuntos
Antivirais/farmacologia , Inibidores Enzimáticos/farmacologia , Imunidade Inata/efeitos dos fármacos , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Pirimidinas/biossíntese , Vírus/imunologia , Antivirais/isolamento & purificação , Di-Hidro-Orotato Desidrogenase , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/isolamento & purificação , Ensaios de Triagem em Larga Escala , Humanos , Fenótipo , Bibliotecas de Moléculas Pequenas/análise
16.
Front Pharmacol ; 15: 1329011, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38444943

RESUMO

De novo purine nucleotide biosynthesis (DNPNB) consists of sequential reactions that are majorly conserved in living organisms. Several regulation events take place to maintain physiological concentrations of adenylate and guanylate nucleotides in cells and to fine-tune the production of purine nucleotides in response to changing cellular demands. Recent years have seen a renewed interest in the DNPNB enzymes, with some being highlighted as promising targets for therapeutic molecules. Herein, a review of two newly revealed modes of regulation of the DNPNB pathway has been carried out: i) the unprecedent allosteric regulation of one of the limiting enzymes of the pathway named inosine 5'-monophosphate dehydrogenase (IMPDH), and ii) the supramolecular assembly of DNPNB enzymes. Moreover, recent advances that revealed the therapeutic potential of DNPNB enzymes in bacteria could open the road for the pharmacological development of novel antibiotics.

17.
Eur J Med Chem ; 280: 116920, 2024 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-39369481

RESUMO

Antimicrobial resistance poses a significant threat to global health, necessitating the development of novel therapeutic agents with unique mechanisms of action. Inosine 5'-monophosphate dehydrogenase (IMPDH), an essential enzyme in guanine nucleotide biosynthesis, is a promising target for the discovery of new antimicrobial agents. High-throughput screening studies have previously identified several urea-based leads as potential inhibitors, although many of these are characterised by reduced chemical stability. In this work, we describe the design and synthesis of a series of heteroaryl-susbtituted analogues and the evaluation of their inhibitory potency against IMPDHs. Our screening targets ESKAPEE pathogens, including Pseudomonas aeruginosa, Staphylococcus aureus and Escherichia coli. Several analogues with submicromolar inhibitory potency are identified and show no inhibitory potency on human IMPDH nor cytotoxic effects on human cells. Kinetic studies revealed that these molecules act as noncompetitive inhibitors with respect to the substrates and ligand virtual docking simulations provided insights into the binding interactions at the interface of the NAD+ and IMP binding sites on IMPDH.

18.
J Antimicrob Chemother ; 68(6): 1285-96, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23378416

RESUMO

OBJECTIVES: Candida albicans is the most prevalent fungal pathogen of humans, causing a wide range of infections from harmless superficial to severe systemic infections. Improvement of the antifungal arsenal is needed since existing antifungals can be associated with limited efficacy, toxicity and antifungal resistance. Here we aimed to identify compounds that act synergistically with echinocandin antifungals and that could contribute to a faster reduction of the fungal burden. METHODS: A total of 38 758 compounds were tested for their ability to act synergistically with aminocandin, a ß-1,3-glucan synthase inhibitor of the echinocandin family of antifungals. The synergy between echinocandins and an identified hit was studied with chemogenomic screens and testing of individual Saccharomyces cerevisiae and C. albicans mutant strains. RESULTS: We found that colistin, an antibiotic that targets membranes in Gram-negative bacteria, is synergistic with drugs of the echinocandin family against all Candida species tested. The combination of colistin and aminocandin led to faster and increased permeabilization of C. albicans cells than either colistin or aminocandin alone. Echinocandin susceptibility was a prerequisite to be able to observe the synergy. A large-scale screen for genes involved in natural resistance of yeast cells to low doses of the drugs, alone or in combination, identified efficient sphingolipid and chitin biosynthesis as necessary to protect S. cerevisiae and C. albicans cells against the antifungal combination. CONCLUSIONS: These results suggest that echinocandin-mediated weakening of the cell wall facilitates colistin targeting of fungal membranes, which in turn reinforces the antifungal activity of echinocandins.


Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Colistina/farmacologia , Equinocandinas/farmacologia , Animais , Antifúngicos/uso terapêutico , Candida/genética , Candidíase/tratamento farmacológico , Candidíase/microbiologia , Permeabilidade da Membrana Celular/efeitos dos fármacos , Parede Celular/efeitos dos fármacos , Quitina/biossíntese , Colistina/uso terapêutico , Corantes , Sinergismo Farmacológico , Equinocandinas/uso terapêutico , Biblioteca Gênica , Aptidão Genética , Genótipo , Técnicas de Diluição do Indicador , Lipopeptídeos/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Mutação/genética , Propídio , Saccharomyces cerevisiae/efeitos dos fármacos , Saccharomyces cerevisiae/genética , Esfingolipídeos/biossíntese
19.
Bioorg Med Chem ; 21(1): 257-68, 2013 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-23199481

RESUMO

We report the synthesis of 5'-modified thymidines (16, 18, 21, 23) and 5,5'-bis-substituted 2'-deoxyuridine analogues (30, 47) as inhibitors of thymidine monophosphate kinase of Mycobacterium tuberculosis (TMPKmt). These analogues were evaluated for their capacity to inhibit TMPKmt and solely two 5'-modified thymidines were found to possess moderate inhibitory activity. In addition, a feasibility study of protecting groups for the 5-CH(2)OH moiety of 2'-deoxyuridines is described that enables to introduce the desired 5'-modification.


Assuntos
Antituberculosos/farmacologia , Mycobacterium tuberculosis/enzimologia , Núcleosídeo-Fosfato Quinase/antagonistas & inibidores , Timidina/análogos & derivados , Timidina/farmacologia , Antituberculosos/química , Humanos , Mycobacterium tuberculosis/efeitos dos fármacos , Núcleosídeo-Fosfato Quinase/metabolismo , Timidina/química , Tuberculose/tratamento farmacológico
20.
Nucleic Acids Res ; 39(8): 3458-72, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21149268

RESUMO

Nucleoside Monophosphate Kinases (NMPKs) family are key enzymes in nucleotide metabolism. Bacterial UMPKs depart from the main superfamily of NMPKs. Having no eukaryotic counterparts they represent attractive therapeutic targets. They are regulated by GTP and UTP, while showing different mechanisms in Gram(+), Gram(-) and archaeal bacteria. In this work, we have characterized the mycobacterial UMPK (UMPKmt) combining enzymatic and structural investigations with site-directed mutagenesis. UMPKmt exhibits cooperativity toward ATP and an allosteric regulation by GTP and UTP. The crystal structure of the complex of UMPKmt with GTP solved at 2.5 Å, was merely identical to the modelled apo-form, in agreement with SAXS experiments. Only a small stretch of residues was affected upon nucleotide binding, pointing out the role of macromolecular dynamics rather than major structural changes in the allosteric regulation of bacterial UMPKs. We further probe allosteric regulation by site-directed mutagenesis. In particular, a key residue involved in the allosteric regulation of this enzyme was identified.


Assuntos
Proteínas de Bactérias/química , Mycobacterium tuberculosis/enzimologia , Núcleosídeo-Fosfato Quinase/química , Regulação Alostérica , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Cristalografia por Raios X , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Núcleosídeo-Fosfato Quinase/genética , Núcleosídeo-Fosfato Quinase/metabolismo , Homologia de Sequência de Aminoácidos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA