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1.
Adv Exp Med Biol ; 1313: 155-177, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34661895

RESUMO

Leprosy is an ancient insidious disease caused by Mycobacterium leprae, where the skin and peripheral nerves undergo chronic granulomatous infections, leading to sensory and motor impairment with characteristic deformities. Susceptibility to leprosy and its disease state are determined by the manifestation of innate immune resistance mediated by cells of monocyte lineage. Due to insufficient innate resistance, granulomatous infection is established, influencing the specific cellular immunity. The clinical presentation of leprosy ranges between two stable polar forms (tuberculoid to lepromatous) and three unstable borderline forms. The tuberculoid form involves Th1 response, characterized by a well demarcated granuloma, infiltrated by CD4+ T lymphocytes, containing epitheloid and multinucleated giant cells. In the lepromatous leprosy, there is no characteristic granuloma but only unstructured accumulation of ineffective macrophages containing engulfed pathogens. Th1 response, characterised by IFN-γ and IL-2 production, activates macrophages in order to kill intracellular pathogens. Conversely, a Th2 response, characterized by the production of IL-4, IL-5 and IL-10, helps in antibody production and consequently downregulates the cell-mediated immunity induced by the Th1 response. M. lepare has a long generation time and its inability to grow in culture under laboratory conditions makes its study challenging. The nine-banded armadillo still remains the best clinical and immunological model to study host-pathogen interaction in leprosy. In this chapter, we present cellular morphology and the genomic uniqueness of M. leprae, and how the pathogen shows tropism for Schwann cells, macrophages and dendritic cells.


Assuntos
Hanseníase , Humanos , Imunidade Celular , Mycobacterium leprae , Pele , Linfócitos T
2.
Adv Exp Med Biol ; 1313: 217-240, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34661897

RESUMO

Tuberculosis (TB) is a highly contagious disease caused by Mycobacterium tuberculosis (Mtb) and is the major cause of morbidity and mortality across the globe. The clinical outcome of TB infection and susceptibility varies among individuals and even among different populations, contributed by host genetic factors such as polymorphism in the human leukocyte antigen (HLA) alleles as well as in cytokine genes, nutritional differences between populations, immunometabolism, and other environmental factors. Till now, BCG is the only vaccine available to prevent TB but the protection rendered by BCG against pulmonary TB is not uniform. To deliver a vaccine which can give consistent protection against TB is a great challenge with rising burden of drug-resistant TB. Thus, expectations are quite high with new generation vaccines that will improve the efficiency of BCG without showing any discordance for all forms of TB, effective for individual of all ages in all parts of the world. In order to enhance or improve the efficacy of BCG, different strategies are being implemented by considering the immunogenicity of various Mtb virulence factors as well as of the recombinant strains, co-administration with adjuvants and use of appropriate vehicle for delivery. This chapter discusses several such pre-clinical attempts to boost BCG with subunit vaccines tested in murine models and also highlights various recombinant TB vaccines undergoing clinical trials. Promising candidates include new generation of live recombinant BCG (rBCG) vaccines, VPM1002, which are deleted in one or two virulence genes. They encode for the mycobacteria-infected macrophage-inhibitor proteins of host macrophage apoptosis and autophagy, key events in killing and eradication of Mtb. These vaccines are rBCG- ΔureC::hly HMR, and rBCG-ΔureC::hly ΔnuoG. The former vaccine has passed phase IIb in clinical trials involving South African infants and adults. Thus, with an aim of elimination of TB by 2050, all these cumulative efforts to develop a better TB vaccine possibly is new hope for positive outcomes.


Assuntos
Mycobacterium tuberculosis , Vacinas contra a Tuberculose , Adulto , Animais , Vacina BCG , Humanos , Lactente , Camundongos , Mycobacterium tuberculosis/genética , Vacinação
3.
Antibiotics (Basel) ; 10(8)2021 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-34438951

RESUMO

The bulbs of Allium species are a known source of antibacterial phytochemicals. Anti-infective, efflux pump and biofilm inhibitory activities of bulb extracts of selected Ghanaian shallots Allium cepa var aggregatum were evaluated using the HT-SPOTi assay and other whole-cell phenotypic screening techniques to determine their possible mechanisms of action. Ethanol and aqueous extracts of white A. cepa inhibited the growth of Mycobacterium smegmatis mc2 155 and Escherichia coli, respectively. The majority of the Allium extracts significantly (p < 0.05) exhibited efflux pump inhibitory activity against all the acid-fast, Gram-positive and Gram-negative strains used. Hexane and chloroform extract of the pink A. cepa and the aqueous extract of the white A. cepa significantly inhibited M. smegmatis biofilm formation. For Pseudomonas aeruginosa, the inhibition was observed at 250 µg/mL for the aqueous extract (~77.34%) and 125 µg/mL for the hexane extract (~76.51%). The results suggest that Ghanaian shallots could potentially be useful when further developed to tackle antimicrobial resistance, particularly in tuberculosis (TB).

5.
JAC Antimicrob Resist ; 3(1): dlab028, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-34223102

RESUMO

Objectives: Identification and validation of novel therapeutic targets is imperative to tackle the rise of drug resistance in tuberculosis. An essential Mur ligase-like gene (Rv3712), expected to be involved in cell-wall peptidoglycan (PG) biogenesis and conserved across mycobacteria, including the genetically depleted Mycobacterium leprae, was the primary focus of this study. Methods: Biochemical analysis of Rv3712 was performed using inorganic phosphate release assays. The operon structure was identified using reverse-transcriptase PCR and a transcription/translation fusion vector. In vivo mycobacterial protein fragment complementation assays helped generate the interactome. Results: Rv3712 was found to be an ATPase. Characterization of its operon revealed a mycobacteria-specific promoter driving the co-transcription of Rv3712 and Rv3713. The two gene products were found to interact with each other in vivo. Sequence-based functional assignments reveal that Rv3712 and Rv3713 are likely to be the mycobacterial PG precursor-modifying enzymes MurT and GatD, respectively. An in vivo network involving Mtb-MurT, regulatory proteins and cell division proteins was also identified. Conclusions: Understanding the role of the enzyme complex in the context of PG metabolism and cell division, and the implications for antimicrobial resistance and host immune responses will facilitate the design of therapeutics that are targeted specifically to M. tuberculosis.

6.
Pharmaceuticals (Basel) ; 14(4)2021 Apr 14.
Artigo em Inglês | MEDLINE | ID: mdl-33919737

RESUMO

The emergence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis (TB) has reinforced the need for the development of new anti-TB drugs. The first line drug isoniazid inhibits InhA. This is a prodrug requiring activation by the enzyme KatG. Mutations in KatG have largely contributed to clinical isoniazid resistance. We aimed to design new 'direct' InhA inhibitors that obviate the need for activation by KatG, circumventing pre-existing resistance. In silico molecular modelling was used as part of a rational structure-based drug-design approach involving inspection of protein crystal structures of InhA:inhibitor complexes, including the broad spectrum antibiotic triclosan (TCS). One crystal structure exhibited the unusual presence of two triclosan molecules within the Mycobacterium tuberculosis InhA binding site. This became the basis of a strategy for the synthesis of novel inhibitors. A series of new, flexible ligands were designed and synthesised, expanding on the triclosan structure. Low Minimum Inhibitory Concentrations (MICs) were obtained for benzylphenyl compounds (12, 43 and 44) and di-triclosan derivative (39), against Mycobacterium bovis BCG although these may also be inhibiting other enzymes. The ether linked di-triclosan derivative (38) displayed excellent in vitro isolated enzyme inhibition results comparable with triclosan, but at a higher MIC (125 µg mL-1). These compounds offer good opportunities as leads for further optimisation.

7.
Antibiotics (Basel) ; 10(1)2021 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-33477812

RESUMO

Tuberculosis (TB) remains a global health emergency, with an estimated 2 billion people infected across the world, and 1.4 million people dying to this disease every year. Many aspects of the causative agent, Mycobacterium tuberculosis, make this disease difficult for healthcare and laboratory researchers to fight against, such as unique pathophysiology, latent infection and long and complex treatment regimens, thus causing patient non-compliance with the treatment. Development of new drugs is critical for tackling these problems. Repurposing drugs is a promising strategy for generating an effective drug treatment whilst circumventing many of the challenges of conventional drug development. In this regard, the incorporation of immunomodulatory drugs into the standard regimen to potentiate frontline drugs is found to be highly appealing. Drugs of diverse chemical classes and drug categories are increasingly being evidenced to possess antitubercular activity, both in vitro and in vivo. This article explores and discusses the molecular entities that have shown promise in being repurposed for use in anti-TB adjunctive therapy and aims to provide the most up-to-date picture of their progress.

8.
Antibiotics (Basel) ; 10(2)2021 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-33498280

RESUMO

Tuberculosis (TB) is still a leading cause of death worldwide. Treatments remain unsatisfactory due to an incomplete understanding of the underlying host-pathogen interactions during infection. In the present study, weighted gene co-expression network analysis (WGCNA) was conducted to identify key macrophage modules and hub genes associated with mycobacterial infection. WGCNA was performed combining our own transcriptomic results using Mycobacterium aurum-infected human monocytic macrophages (THP1) with publicly accessible datasets obtained from three types of macrophages infected with seven different mycobacterial strains in various one-to-one combinations. A hierarchical clustering tree of 11,533 genes was built from 198 samples, and 47 distinct modules were revealed. We identified a module, consisting of 226 genes, which represented the common response of host macrophages to different mycobacterial infections that showed significant enrichment in innate immune stimulation, bacterial pattern recognition, and leukocyte chemotaxis. Moreover, by network analysis applied to the 74 genes with the best correlation with mycobacteria infection, we identified the top 10 hub-connecting genes: NAMPT, IRAK2, SOCS3, PTGS2, CCL20, IL1B, ZC3H12A, ABTB2, GFPT2, and ELOVL7. Interestingly, apart from the well-known Toll-like receptor and inflammation-associated genes, other genes may serve as novel TB diagnosis markers and potential therapeutic targets.

9.
J Med Chem ; 64(1): 71-100, 2021 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-33372516

RESUMO

The alarming rise in drug-resistant clinical cases of tuberculosis (TB) has necessitated the rapid development of newer chemotherapeutic agents with novel mechanisms of action. The mycobactin biosynthesis pathway, conserved only among the mycolata family of actinobacteria, a group of intracellularly surviving bacterial pathogens that includes Mycobacterium tuberculosis, generates a salicyl-capped peptide mycobactin under iron-stress conditions in host macrophages to support the iron demands of the pathogen. This in vivo essentiality makes this less explored mycobactin biosynthesis pathway a promising endogenous target for novel lead-compounds discovery. In this Perspective, we have provided an up-to-date account of drug discovery efforts targeting selected enzymes (MbtI, MbtA, MbtM, and PPTase) from the mbt gene cluster (mbtA-mbtN). Furthermore, a succinct discussion on non-specific mycobactin biosynthesis inhibitors and the Trojan horse approach adopted to impair iron metabolism in mycobacteria has also been included in this Perspective.


Assuntos
Antituberculosos/farmacologia , Mycobacterium tuberculosis/metabolismo , Oxazóis/metabolismo , Tuberculose/tratamento farmacológico , Proteínas de Bactérias/metabolismo , Vias Biossintéticas , Descoberta de Drogas , Farmacorresistência Bacteriana/efeitos dos fármacos , Humanos , Ferro/metabolismo , Mycobacterium tuberculosis/efeitos dos fármacos
10.
Antibiotics (Basel) ; 9(12)2020 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-33339285

RESUMO

The present study aimed to screen plants for bioactive compounds with potential antibacterial activities. In our efforts to evaluate plants from Borneo, we isolated and elucidated the structures of four natural products from the bioactive fraction of a chloroform extract of Goniothalamus longistipetes using various chromatographic and spectroscopic techniques. The bioactive compounds were identified as a known styryllactone, (+)-altholactone ((2S,3R,3aS,7aS)-3-hydroxy-2-phenyl-2,3,3a,7a-tetrahydrobenzo-5(4H)-5-one) (1), a new styryllactone, (2S,3R,3aS,7aS)-3-hydroxy-2-phenyl-2,3,3a,7a-tetrahydrobenzo-5(4H)-5-one) (2) as well as a new alkaloid, 2,6-dimethoxyisonicotinaldehyde (3) and a new alkenyl-5-hydroxyl-phenyl benzoic acid (4). 1 and 4 showed broad-spectrum anti-bacterial activities against Gram-positive and Gram-negative bacteria as well as acid-fast model selected for this study. Compound 2 only demonstrated activities against Gram-positive bacteria whilst 3 displayed selective inhibitory activities against Gram-positive bacterial strains. Additionally, their mechanisms of anti-bacterial action were also investigated. Using Mycobacterium smegmatis as a fast-growing model of tubercle bacilli, compounds 1, 2 and 4 demonstrated inhibitory activities against whole-cell drug efflux and biofilm formation; two key intrinsic mechanisms of antibiotic resistance. Interestingly, the amphiphilic compound 4 exhibited inhibitory activity against the conjugation of plasmid pKM101 in Escherichia coli using a plate conjugation assay. Plasmid conjugation is a mechanism by which Gram-positive and Gram-negative-bacteria acquire drug resistance and virulence. These results indicated that bioactive compounds isolated from Goniothalamus longistipetes can be potential candidates as 'hits' for further optimisation.

11.
Access Microbiol ; 2(4): acmi000105, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33005869

RESUMO

With antimicrobial resistance creating a major public health crisis, the designing of novel antimicrobial compounds that effectively combat bacterial infection is becoming increasingly critical. Interdisciplinary approaches integrate the best features of whole-cell phenotypic evaluation to validate novel therapeutic targets and discover new leads to combat antimicrobial resistance. In this project, whole-cell phenotypic evaluation such as testing inhibitors on bacterial growth, viability, efflux pump, biofilm formation and their interaction with other drugs were performed on a panel of Gram-positive, Gram-negative and acid-fast group of bacterial species. This enabled additional antimicrobial activities of compounds belonging to the flavonoid family including ketones, chalcones and stilbenes, to be identified. Flavonoids have received renewed attention in literature over the past decade, and a variety of beneficial effects of these compounds have been illuminated, including anti-cancer, anti-inflammatory, anti-tumour as well as anti-fungal and anti-bacterial. However, their mechanisms of action are yet to be identified. In this paper, we found that the compounds belonging to the flavonoid family exerted a range of anti-infective properties being identified as novel efflux pump inhibitors, whilst offering the opportunity to be used in combination therapy. The compound 2-phenylacetophenone displayed broad-spectrum efflux pump inhibition activity, whilst trans-chalcone, displayed potent activity against Gram-negative and mycobacterial efflux pumps causing inhibition higher than known potent efflux pump inhibitors, verapamil and chlorpromazine. Drug-drug interaction studies also highlighted that 2-phenylacetophenone not only has the potential to work additively with known antibacterial agents that affect the cell-wall and DNA replication but also trans-chalcone has the potential to work synergistically with anti-tubercular agents. Overall, this paper shows how whole-cell phenotypic analysis allows for the discovery of new antimicrobial agents and their consequent mode of action whilst offering the opportunity for compounds to be repurposed, in order to contribute in the fight against antimicrobial resistance.

12.
J Antimicrob Chemother ; 75(11): 3194-3201, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32790867

RESUMO

BACKGROUND: The rise of antimicrobial drug resistance in Mycobacterium tuberculosis coupled with the shortage of new antibiotics has elevated TB to a major global health priority. Repurposing drugs developed or used for other conditions has gained special attention in the current scenario of accelerated drug development for several global infectious diseases. In a similar effort, previous studies revealed that carprofen, a non-steroidal anti-inflammatory drug, selectively inhibited the growth of replicating, non-replicating and MDR clinical isolates of M. tuberculosis. OBJECTIVES: We aimed to reveal the whole-cell phenotypic and transcriptomic effects of carprofen in mycobacteria. METHODS: Integrative molecular and microbiological approaches such as resazurin microtitre plate assay, high-throughput spot-culture growth inhibition assay, whole-cell efflux inhibition, biofilm inhibition and microarray analyses were performed. Analogues of carprofen were also synthesized and assessed for their antimycobacterial activity. RESULTS: Carprofen was found to be a bactericidal drug that inhibited mycobacterial drug efflux mechanisms. It also restricted mycobacterial biofilm growth. Transcriptome profiling revealed that carprofen likely acts by targeting respiration through the disruption of membrane potential. The pleiotropic nature of carprofen's anti-TB action may explain why spontaneous drug-resistant mutants could not be isolated in practice. CONCLUSIONS: This immunomodulatory drug and its chemical analogues have the potential to reverse TB antimicrobial drug resistance, offering a swift path to clinical trials of novel TB drug combinations.


Assuntos
Mycobacterium tuberculosis , Tuberculose Resistente a Múltiplos Medicamentos , Tuberculose , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Carbazóis , Resistência Microbiana a Medicamentos , Humanos , Tuberculose/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico
13.
Antibiotics (Basel) ; 9(7)2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32630175

RESUMO

In this study, the inhibitory potential of 3-(5-nitrofuran-2-yl)prop-2-en-1-one derivatives was evaluated against a panel of bacteria, as well as mammalian cell lines to determine their therapeutic index. In addition, we investigated the mechanism of antibiotic action of the derivatives to identify their therapeutic target. We discovered compound 2 to be an extremely potent inhibitor of Mycobacterium tuberculosis H37Rv growth (MIC: 0.031 mg/L) in vitro, performing better than the currently used first-line antituberculosis drugs such as isoniazid, rifampicin, ethambutol, and pretomanid in vitro. Furthermore, compound 3 was equipotent to pretomanid against a multidrug-resistant M. tuberculosis clinical isolate. The derivatives were selective and bactericidal towards slow-growing mycobacteria. They showed low cytotoxicity towards murine RAW 264.7 and human THP-1 cell lines, with high selectivity indices. Compound 1 effectively eliminated the intracellular mycobacteria in a mycobacteria-infected macrophage model. The derivatives were assessed for their potential to inhibit mycobacterial arylamine N-acetyltransferase (NAT) and were identified as good inhibitors of recombinant mycobacterial NAT, a novel target essential for the intracellular survival of M. tuberculosis. This study provided hits for designing new potent and selective antituberculosis leads, having mycobacterial NAT inhibition as their possible endogenous mechanisms of action.

14.
Antibiotics (Basel) ; 9(7)2020 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-32650510

RESUMO

Carvotacetones (1-7) isolated from Sphaeranthus africanus were screened for their antimycobacterial and efflux pump (EP) inhibitory potential against the mycobacterial model strains Mycobacterium smegmatis mc2 155, Mycobacterium aurum ATCC 23366, and Mycobacterium bovis BCG ATCC 35734. The minimum inhibitory concentrations (MICs) of the carvotacetones were detected through high-throughput spot culture growth inhibition (HT-SPOTi) and microbroth dilution assays. In order to assess the potential of the compounds 1 and 6 to accumulate ethidium bromide (EtBr) in M. smegmatis and M. aurum, a microtiter plate-based fluorometric assay was used to determine efflux activity. Compounds 1 and 6 were analyzed for their modulating effects on the MIC of EtBr and the antibiotic rifampicin (RIF) against M. smegmatis. Carvotacetones 1 and 6 had potent antibacterial effects on M. aurum and M. bovis BCG (MIC ≤ 31.25 mg/L) and could successfully enhance EtBr activity against M. smegmatis. Compound 1 appeared as the most efficient agent for impairing the efflux mechanism in M. smegmatis. Both compounds 1 and 6 were highly effective against M. aurum and M. bovis BCG. In particular, compound 1 was identified as a valuable candidate for inhibiting mycobacterial efflux mechanisms and as a promising adjuvant in the therapy of tuberculosis or other non-tubercular mycobacterial infections.

15.
ACS Med Chem Lett ; 11(5): 638-644, 2020 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-32435364

RESUMO

A series of N-phenyl-2,5-dimethylpyrrole derivatives, designed as hybrids of the antitubercular agents BM212 and SQ109, have been synthesized and evaluated against susceptible and drug-resistant mycobacteria strains. Compound 5d, bearing a cyclohexylmethylene side chain, showed high potency against M. tuberculosis including MDR-TB strains at submicromolar concentrations. The new compound shows bacteriostatic activity and low toxicity and proved to be effective against intracellular mycobacteria too, showing an activity profile similar to isoniazid.

16.
Immunobiology ; 225(3): 151951, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32423788

RESUMO

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb) infection, remains a major global health emergency. It is estimated that one third of global population are affected, predominantly with latent granuloma form of the disease. Mtb co-evolved with humans, for its obligatory intra-macrophage phagosome habitat and slow replication, balanced against unique mycobacterial innate immunity, which appears to be highly complex. TB is transmitted via cough aerosol Mtb inhalation. Bovine TB attenuated Bacillus Calmette Guerin (BCG) live vaccine has been in practice for protection of young children from severe disseminated Mtb infection, but not sufficiently for their lungs, as obtained by trials in TB endemic community. To augment BCG vaccine-driven innate and adaptive immunity for neonates and better protection against adult pulmonary TB, a number of BCG pre-vaccination based, subset vaccine candidates have been tested via animal preclinical, followed by safe clinical trials. BCG also enhances innate macrophage trained immunity and memory, through primordial intracellular Toll-like receptors (TLRs) 7 and 9, which recognise distinct mycobacterial molecular pattern signature. This signature is transmitted by TLR signalling via nuclear factor-κB, for activating innate immune transcription and expression of gene profiling in a mycobacterial signature-specific manner. These are epigenetically imprinted in reprogramming of distinct chromatin areas for innate immune memory, to be recalled following lung reinfection. Unique TB innate immunity and its trained memory are considered independent from adaptive immune B and T cells. On the other hand, adaptive immunity is crucial in Mtb containment in granulomatous latency, supported by innate immune cell infiltration. In nearly 5-10 % of susceptible people, latent TB may be activated due to immune evasion by Mtb from intracellular phagosome within macrophage, perpetrating TB. However, BCG and new recombinant BCG vaccines have the capacity, as indicated in pre- and clinical trials, to overcome such Mtb evasion. Various strategies include pro-inflammatory-bactericidal type 1 polarisation (M1) phenotype of the infected macrophage, involving thrombospondin-TLR pathway. Saprophytic M. smegmatis-based recombinant vaccines are also promising candidates against TB. BCG vaccination of neonates/infants in TB endemic countries also reduced their pneumonia caused by various microbes independent of TB immunity. Here, we discuss host immune response against Mtb, its immune evasion strategies, and the important role innate immunity plays in the development of protection against TB.


Assuntos
Imunidade Adaptativa , Resistência à Doença/imunologia , Interações Hospedeiro-Patógeno/imunologia , Imunidade Inata , Mycobacterium tuberculosis/imunologia , Tuberculose/imunologia , Vacina BCG/imunologia , Biomarcadores , Humanos , Evasão da Resposta Imune , Tuberculose/metabolismo , Tuberculose/microbiologia , Vacinas contra a Tuberculose/imunologia
17.
Molecules ; 25(5)2020 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-32164166

RESUMO

Here we sought to evaluate the contribution of the PBD unit to the biological activity of PBD-conjugates and, to this end, an adenosine nucleoside was attached to the PBD A-ring C8 position. A convergent approach was successfully adopted for the synthesis of a novel C8-linked pyrrolo(2,1-c)(1,4)benzodiazepine(PBD)-adenosine(ADN) hybrid. The PBD and adenosine (ADN) moieties were synthesized separately and then linked through a pentynyl linker. To our knowledge, this is the first report of a PBD connected to a nucleoside. Surprisingly, the compound showed no cytotoxicity against murine cells and was inactive against Mycobacterium aurum and M. bovis strains and did not bind to guanine-containing DNA sequences, as shown by DNase I footprinting experiments. Molecular dynamics simulations revealed that the PBD-ADN conjugate was poorly accommodated in the DNA minor groove of two DNA sequences containing the AGA-PBD binding motif, with the adenosine moiety of the ligand preventing the covalent binding of the PBD unit to the guanine amino group of the DNA duplex. These interesting findings shed further light on the ability of the substituents attached at the C8 position of PBDs to affect and modulate the biological and biophysical properties of PBD hybrids.


Assuntos
Adenosina/química , Dioxinas/química , Animais , Sequência de Bases , Linhagem Celular , DNA/química , Guanina/química , Ligantes , Camundongos , Simulação de Dinâmica Molecular
18.
Molecules ; 25(3)2020 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-32046221

RESUMO

Therapeutic treatment options for opportunistic non-tuberculous mycobacterial (NTM) infection and/or serious mycobacterial infections such as tuberculosis (TB) and leprosy are limited due to the spread of antimicrobial resistance mechanism. Plant-derived natural compounds as prospective efflux pump inhibitors may present a promising adjunct to conventional chemotherapy by enhancing mycobacterial susceptibility to antibiotics. This study served to evaluate the antimicrobial and resistance-modifying profile of a range of plant-derived flavonoids against the mycobacterial model strains: M. smegmatis, M. aurum, and M. bovis BCG. The minimum inhibitory concentrations (MICs) of the compounds against the mycobacterial strains were determined using both agar dilution and broth dilution assays, while their efflux inhibitory activity was investigated via an ethidium bromide-based fluorometric assay. All compounds were screened for their synergistic effects with ethidium bromide (EtBr) and rifampicin (RIF) against M. smegmatis. Skullcapflavone II (5,2'-dihydroxy-6,7,8,6'-tetramethoxyflavone, 1) exerted potent antimicrobial activity against M. aurum and M. bovis BCG and considerably increased the susceptibility of M. smegmatis to EtBr and RIF. Nobiletin (5,6,7,8,3',4'-hexamethoxyflavone, 2) was determined to be the most potent efflux-inhibitor in M. aurum and M. smegmatis. However, a connection between strong modulatory and putative efflux activity of the compounds could not be observed. Nevertheless, the results highlight two polymethoxyflavones, skullcapflavone II and nobiletin, with potent antimycobacterial and antibiotic resistance modulating activities as valuable adjuvants in anti-mycobacterial therapies.


Assuntos
Anti-Infecciosos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Flavonoides/farmacologia , Mycobacterium/efeitos dos fármacos , Proteínas de Bactérias/metabolismo , Transporte Biológico/efeitos dos fármacos , Etídio/química , Proteínas de Membrana Transportadoras/metabolismo , Testes de Sensibilidade Microbiana/métodos , Mycobacterium/metabolismo , Infecções por Mycobacterium/tratamento farmacológico , Rifampina/farmacologia
19.
Chem Biol Drug Des ; 95(1): 192-199, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31560814

RESUMO

We report the biological evaluation of 5-(5-nitrothiophen-2-yl)-4,5-dihydro-1H-pyrazole derivatives against bacteria, eukaryotic cell lines and the assessment of their mechanisms of action to determine their prospects of being developed into potent antituberculosis agents. The compounds were evaluated for their antibacterial property against Mycobacterium tuberculosis H37Rv, multidrug-resistant M. tuberculosis, Mycobacterium bovis BCG, Mycobacterium aurum, Escherichia coli, and Staphylococcus aureus using high-throughput spot-culture growth inhibition assay. They were found to be selective toward slow-growing mycobacteria and Gram-positive bacteria. In M. bovis BCG, they exhibited a bactericidal mode of action. Cytotoxicity was assessed in human THP-1 and murine RAW 264.7 cell lines, and the compounds showed a lower cytotoxicity potential when compared with their antibacterial activity. They were found to be excellent whole-cell efflux pump inhibitors of the mycobacterial surrogate M. aurum, performing better than known efflux pump inhibitor verapamil. The 5-nitrothiophene moiety was identified for the first time as a prospective inhibitor scaffold of mycobacterial arylamine N-acetyltransferase enzyme, which is the key enzyme in metabolizing isoniazid, a first-line antituberculosis drug. The two aforementioned findings make the compounds potential hits in the development of adjunctive tuberculosis therapy.


Assuntos
Antibacterianos/química , Arilamina N-Acetiltransferase/antagonistas & inibidores , Proteínas de Bactérias/antagonistas & inibidores , Inibidores Enzimáticos/química , Pirazóis/química , Animais , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Ensaios de Triagem em Larga Escala , Humanos , Camundongos , Pirazóis/farmacologia
20.
ChemMedChem ; 14(19): 1735-1741, 2019 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-31454170

RESUMO

Iron is essential for the pathogenicity and virulence of Mycobacterium tuberculosis, which synthesises salicyl-capped siderophores (mycobactins) to acquire this element from the host. MbtA is the adenylating enzyme that catalyses the initial reaction of mycobactin biosynthesis and is solely expressed by mycobacteria. A 3200-member library comprised of lead-like, structurally diverse compounds was screened against M. tuberculosis for whole-cell inhibitory activity. A set of 846 compounds that inhibited the tubercle bacilli growth were then tested for their ability to bind to MbtA using a fluorescence-based thermal shift assay and NMR-based Water-LOGSY and saturation transfer difference (STD) experiments. We identified an attractive hit molecule, 5-hydroxyindol-3-ethylamino-(2-nitro-4-trifluoromethyl)benzene (5), that bound with high affinity to MbtA and produced a MIC90 value of 13 µm. The ligand was docked into the MbtA crystal structure and displayed an excellent fit within the MbtA active pocket, adopting a binding mode different from that of the established MbtA inhibitor Sal-AMS.


Assuntos
Antituberculosos/química , Ligases/metabolismo , Mycobacterium tuberculosis/enzimologia , Bibliotecas de Moléculas Pequenas/química , Adenosina/química , Antituberculosos/farmacologia , Domínio Catalítico , Sobrevivência Celular/efeitos dos fármacos , Células Hep G2 , Humanos , Ferro/química , Ligantes , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Estrutura Molecular , Oxazóis/química , Sideróforos/química , Bibliotecas de Moléculas Pequenas/farmacologia , Relação Estrutura-Atividade
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