RESUMO
In this work, we implemented for the first time the cycloSaligenyl prodrug strategy to increase the bioavailability of fosmidomycin phosphate analogs in bacteria. Here, we report the synthesis of 34 cycloSaligenyl prodrugs of fosfoxacin and its derivatives. Among them, fifteen double prodrugs efficiently prevented the growth of the non-pathogenic, fast-growing Mycobacterium smegmatis.
Assuntos
Pró-Fármacos , Monofosfato de Citidina , Mycobacterium smegmatis , FosfatosRESUMO
Isoprenoids, a diverse class of natural products, are present in all living organisms. Their two universal building blocks are synthesized via two independent pathways: the mevalonate pathway and the 2-C-methyl-ᴠ-erythritol 4-phosphate (MEP) pathway. The presence of the latter in pathogenic bacteria and its absence in humans make all its enzymes suitable targets for the development of novel antibacterial drugs. (E)-4-Hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP), the last intermediate of this pathway, is a natural ligand for the human Vγ9Vδ2 T cells and the most potent natural phosphoantigen known to date. Moreover, 5-hydroxypentane-2,3-dione, a metabolite produced by Escherichia coli 1-deoxy-ᴠ-xylulose 5-phosphate synthase (DXS), the first enzyme of the MEP pathway, structurally resembles (S)-4,5-dihydroxy-2,3-pentanedione, a signal molecule implied in bacterial cell communication. In this review, we shed light on the diversity of potential uses of the MEP pathway in antibacterial therapies, starting with an overview of the antibacterials developed for each of its enzymes. Then, we provide insight into HMBPP, its synthetic analogs, and their prodrugs. Finally, we discuss the potential contribution of the MEP pathway to quorum sensing mechanisms. The MEP pathway, providing simultaneously antibacterial drug targets and potent immunostimulants, coupled with its potential role in bacterial cell-cell communication, opens new therapeutic perspectives.
Assuntos
Fosfatos Açúcares , Humanos , Fosfatos Açúcares/metabolismo , Terpenos/farmacologia , Terpenos/metabolismo , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Eritritol/metabolismoRESUMO
Three α,α-difluorophosphonate derivatives of fosmidomycin were synthesized from diethyl 1,1-difluorobut-3-enylphosphonate and were evaluated on Escherichia coli. Two of them are among the best 1-deoxy-d-xylulose 5-phosphate reductoisomerase inhibitors, with IC50 in the nM range, much better than fosmidomycin, the reference compound. They also showed an enhanced antimicrobial activity against E. coli on Petri dishes in comparison with the corresponding phosphates and the non-fluorinated phosphonate.
Assuntos
Antibacterianos/farmacologia , Fosfomicina/análogos & derivados , Ácidos Hidroxâmicos/farmacologia , Aldose-Cetose Isomerases/antagonistas & inibidores , Aldose-Cetose Isomerases/metabolismo , Farmacorresistência Bacteriana/efeitos dos fármacos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Fosfomicina/síntese química , Fosfomicina/química , Fosfomicina/farmacologia , Testes de Sensibilidade MicrobianaRESUMO
Aryl phosphoramidate prodrugs of fosfoxacin derivatives 15a-b and 8a-b were synthesized and investigated for their ability to target bacteria. No growth inhibition was observed neither for Mycobacterium smegmatis nor for Escherichia coli on solid medium, demonstrating the absence of release of the active compounds in the bacterial cells. Investigation of the stability of the prodrugs and their multienzymatic cleavage in abiotic and biotic conditions showed that the use of aryl phosphoramidate prodrug approach to deliver non-nucleotides compounds is not obvious and might not be appropriate for an antimicrobial drug.
Assuntos
Amidas/síntese química , Monofosfato de Citidina/análogos & derivados , Ácidos Fosfóricos/síntese química , Pró-Fármacos/síntese química , Amidas/química , Monofosfato de Citidina/síntese química , Monofosfato de Citidina/química , Estrutura Molecular , Ácidos Fosfóricos/química , Pró-Fármacos/químicaRESUMO
Hydroxamate analogs of fosfoxacin, the phosphate homolog of fosmidomycin, have been synthesized and their activity tested on Escherichia coli and Mycobacterium smegmatis DXRs. Except for compound 4b, the IC50 values of phosphate derivatives are approximately 10-fold higher than those of the corresponding phosphonates. Although their inhibitory activity on Escherichia coli DXR is less efficient than their phosphonate analogs, we report the ability of phosphate compounds to inhibit the growth of Escherichia coli. This work points out that the uptake of fosfoxacin and its analogs is taking place via the GlpT and UhpT transporters. As expected, these compounds are inefficient to inhibit the growth of M. smegmatis growth inhibition probably due to a lack of uptake.
Assuntos
Aldose-Cetose Isomerases/antagonistas & inibidores , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Escherichia coli/enzimologia , Fosfomicina/análogos & derivados , Mycobacterium smegmatis/enzimologia , Fosfatos/farmacologia , Aldose-Cetose Isomerases/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Fosfomicina/síntese química , Fosfomicina/química , Fosfomicina/farmacologia , Estrutura Molecular , Fosfatos/química , Relação Estrutura-AtividadeRESUMO
Flavonoids, due to their physical and chemical properties (among them hydrophobicity and metal chelation abilities), are potential inhibitors of the 1-deoxyxylulose 5-phosphate reductoisomerase and most of the tested flavonoids effectively inhibited its activity with encouraging IC50 values in the micromolar range. The addition of 0.01% Triton X100 in the assays led however, to a dramatic decrease of the inhibition revealing that a non-specific inhibition probably takes place. Our study highlights the possibility of erroneous conclusions regarding the inhibition of enzymes by flavonoids that are able to produce aggregates in micromolar range. Therefore, the addition of a detergent in the assays prevents possible false positive hits in high throughput screenings.
Assuntos
Aldose-Cetose Isomerases/antagonistas & inibidores , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Escherichia coli/enzimologia , Flavonoides/química , Flavonoides/farmacologia , Aldose-Cetose Isomerases/metabolismo , Escherichia coli/efeitos dos fármacos , Infecções por Escherichia coli/microbiologia , HumanosRESUMO
To develop more effective inhibitors than fosmidomycin, a natural compound which inhibits the deoxyxylulose 5-phosphate reductoisomerase (DXR), the second enzyme of the MEP pathway, we designed molecules possessing on the one hand a catechol that is able to chelate the magnesium dication and on the other hand a group able to occupy the NADPH recognition site. Catechol-rhodanine derivatives (1-6) were synthesized and their potential inhibition was tested on the DXR of Escherichia coli. For the inhibitors 1 and 2, the presence of detergent in the enzymatic assays led to a dramatic decrease of the inhibition suggesting, that these compounds are rather promiscuous inhibitors. The compounds 4 and 5 kept their inhibition capacity in the presence of Triton X100 and could be considered as specific inhibitors of DXR. Compound 4 showed antimicrobial activity against Escherichia coli. The only partial protection of NADPH against the inhibition suggested that the catechol-rhodanine derivatives did not settle in the coenzyme binding site. This paper points out the necessity to include a detergent in the DXR enzymatic assays to avoid false positive when putative hydrophobic inhibitors are tested and especially when the IC50, are in the micromolar range.
Assuntos
Aldose-Cetose Isomerases/antagonistas & inibidores , Antibacterianos/farmacologia , Catecóis/farmacologia , Inibidores Enzimáticos/farmacologia , Escherichia coli/efeitos dos fármacos , Rodanina/farmacologia , Aldose-Cetose Isomerases/metabolismo , Antibacterianos/síntese química , Antibacterianos/química , Catecóis/química , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Escherichia coli/enzimologia , Escherichia coli/crescimento & desenvolvimento , Testes de Sensibilidade Microbiana , Estrutura Molecular , Rodanina/química , Relação Estrutura-AtividadeRESUMO
This work is focused on the design of new antimicrobial drugs and on the development of lipophilic inhibitors of the DXR, the second enzyme of the MEP pathway for the biosynthesis of isoprene units in most bacteria, by replacing the phosphonate group of fosmidomycin derivatives by a tetrazoyl moiety capable of multiple hydrogen bonding. The N- and C-substituted tetrazole analogues of phosphonohydroxamate inhibitors were synthesized and tested on the DXR of Escherichia coli. This work points out the hypothesis that the phosphonate/phosphate recognition site might be too rigid to accommodate other functional groups.
Assuntos
Anti-Infecciosos/síntese química , Ácidos Hidroxâmicos/química , Complexos Multienzimáticos/antagonistas & inibidores , Tetrazóis/química , Anti-Infecciosos/química , Anti-Infecciosos/metabolismo , Domínio Catalítico , Escherichia coli/metabolismo , Proteínas de Escherichia coli/antagonistas & inibidores , Proteínas de Escherichia coli/metabolismo , Fosfomicina/análogos & derivados , Fosfomicina/química , Fosfomicina/metabolismo , Ligação de Hidrogênio , Ácidos Hidroxâmicos/metabolismo , Complexos Multienzimáticos/metabolismo , Ligação Proteica , Tetrazóis/síntese química , Tetrazóis/metabolismoRESUMO
Fosmidomycin derivatives in which the hydroxamic acid group has been replaced by several bidentate chelators as potential hydroxamic alternatives were prepared and tested against the DXR from Escherichia coli. These results illustrate the predominant role of the hydroxamate functional group as the most effective metal binding group in DXR inhibitors.
Assuntos
Aldose-Cetose Isomerases/química , Quelantes/química , Inibidores Enzimáticos/química , Fosfomicina/análogos & derivados , Ácidos Hidroxâmicos/química , Aldose-Cetose Isomerases/antagonistas & inibidores , Quelantes/farmacologia , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Escherichia coli/enzimologia , Fosfomicina/química , Fosfomicina/farmacologia , Concentração Inibidora 50 , Metaloproteínas/química , Estrutura Molecular , Ligação Proteica/efeitos dos fármacosRESUMO
Since Mycobacterium tuberculosis sets up several multiple anti-tuberculosis drug resistance mechanisms, development of new drugs with innovative target is urgent. The methylerythritol phosphate pathway (MEP) involved in the biosynthesis of essential metabolites for the survival of mycobacteria, represents such a target. Fosmidomycin 1a and FR900098 1b, two inhibitors of DXR, do not affect the viability of M. tuberculosis cells, due to a lack of uptake. To overcome the absence of the mycobacterial cell wall crossing of these compounds, we synthesized and tested the inhibition potency of acyloxymethyl phosphonate esters as prodrugs of fosmidomycin 1a, FR900098 1b and their analogs 2a and 2b on Mycobacterium smegmatis. Only the prodrugs 4b-6b inhibit the bacterial growth and could be effective anti-mycobacterial agents.
Assuntos
Aldose-Cetose Isomerases/antagonistas & inibidores , Antibacterianos/farmacologia , Inibidores Enzimáticos/metabolismo , Complexos Multienzimáticos/antagonistas & inibidores , Mycobacterium smegmatis/efeitos dos fármacos , Mycobacterium smegmatis/crescimento & desenvolvimento , Oxirredutases/antagonistas & inibidores , Pró-Fármacos/farmacologia , Aldose-Cetose Isomerases/metabolismo , Antibacterianos/síntese química , Antibacterianos/química , Antibacterianos/metabolismo , Fosfomicina/análogos & derivados , Fosfomicina/metabolismo , Interações Hidrofóbicas e Hidrofílicas , Cinética , Viabilidade Microbiana/efeitos dos fármacos , Complexos Multienzimáticos/metabolismo , Mycobacterium smegmatis/enzimologia , Mycobacterium smegmatis/metabolismo , Organofosfonatos/síntese química , Organofosfonatos/química , Organofosfonatos/metabolismo , Organofosfonatos/farmacologia , Oxazinas/metabolismo , Oxirredutases/metabolismo , Pró-Fármacos/síntese química , Pró-Fármacos/química , Pró-Fármacos/metabolismo , Xantenos/metabolismoRESUMO
Fosmidomycin and its analogue FR-900098 are potent inhibitors of 1-deoxy-d-xylulose 5-phosphate reducto-isomerase (DXR), the second enzyme of the MEP pathway for the biosynthesis of isoprenoids. This paper describes the synthesis of analogues of the two reverse phosphonohydroxamic acids 3 and 4, in which the length of the carbon spacer is modified, the N-methyl group of 3 is replaced by an ethyl group, and the phosphate group is replaced by potential isosteric moieties, i.e., sulfonate or carboxylate functionalities. The potential of the synthesized analogues to inhibit the E. coli DXR was evaluated.
Assuntos
Aldose-Cetose Isomerases/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Eritritol/análogos & derivados , Fosfomicina/análogos & derivados , Complexos Multienzimáticos/antagonistas & inibidores , Organofosfonatos/química , Oxirredutases/antagonistas & inibidores , Fosfatos Açúcares/metabolismo , Terpenos/farmacologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Eritritol/química , Eritritol/metabolismo , Fosfomicina/química , Estrutura Molecular , Relação Estrutura-Atividade , Fosfatos Açúcares/química , Terpenos/química , Terpenos/metabolismoRESUMO
A novel, mild, and efficient method was described to introduce a dibenzyl phosphate by ring opening of benzylglycidol mediated by Lewis acids. This methodology was used as a key step for synthesizing the dihydroxyacetone phosphate (DHAP) in only three steps with an overall yield of 74% from the commercially available racemic benzylglycidol.
RESUMO
Isoprenoid biosynthesis via the methylerythritol phosphate pathway is a target against pathogenic bacteria and the malaria parasite Plasmodium falciparum. 4-(Hydroxyamino)-4-oxobutylphosphonic acid and 4-[hydroxy(methyl)amino]-4-oxobutyl phosphonic acid, two novel inhibitors of DXR (1-deoxy-D-xylulose 5-phosphate reducto-isomerase), the second enzyme of the pathway, have been synthesized and compared with fosmidomycin, the best known inhibitor of this enzyme. The latter phosphonohydroxamic acid showed a high inhibitory activity towards DXR, much like fosmidomycin, as well as significant antibacterial activity against Escherichia coli in tests on Petri dishes.
Assuntos
Aldose-Cetose Isomerases/antagonistas & inibidores , Antibacterianos/farmacologia , Antimaláricos/farmacologia , Inibidores Enzimáticos/farmacologia , Proteínas de Escherichia coli/antagonistas & inibidores , Fosfomicina/análogos & derivados , Ácidos Hidroxâmicos/farmacologia , Complexos Multienzimáticos/antagonistas & inibidores , Organofosfonatos/farmacologia , Oxirredutases/antagonistas & inibidores , Terpenos/metabolismo , Antibacterianos/síntese química , Antimaláricos/síntese química , Butiratos/síntese química , Butiratos/farmacologia , Desenho de Fármacos , Farmacorresistência Bacteriana Múltipla , Inibidores Enzimáticos/síntese química , Escherichia coli/efeitos dos fármacos , Escherichia coli/enzimologia , Fosfomicina/farmacologia , Ácidos Hidroxâmicos/síntese química , Testes de Sensibilidade Microbiana , NADP/metabolismo , Organofosfonatos/síntese químicaRESUMO
The mevalonate-independent methylerythritol phosphate pathway is a long overlooked metabolic pathway for isoprenoid biosynthesis. It is present in most bacteria, including pathogens and opportunistic pathogens, in some unicellular eukaryotes, including the parasite responsible for malaria, and in the chloroplasts of all phototrophic organisms. It represents an alternative to the mevalonate pathway, which is only present in animals, fungi, the plant cytoplasm, archaebacteria and some eubacteria. This biosynthetic pathway is thus a potential target for antibacterial and antiparasitic drugs. An isopentenyl diphosphate isomerase that differs from the previously known isopentenyl diphosphate isomerase found in all other organisms, including animals, was discovered in several Gram-positive bacteria possessing the mevalonate pathway, adding another target related to isoprenoid biosynthesis.
Assuntos
Antibacterianos/administração & dosagem , Antiparasitários/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Terpenos/metabolismo , Animais , Humanos , Terpenos/antagonistas & inibidoresRESUMO
(3,4)-3,4-Dihydroxy-5-oxohexylphosphonic acid, an isosteric analogue of 1-deoxy-D-xylulose 5-phosphate (DXP), was obtained in enantiomerically pure form from (+)-2,3--benzylidene--threitol by a seven-step sequence. This phosphonate did not affect the growth of. It did not inhibit the 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR), but was converted by this enzyme into (3,4)-3,4,5-trihydroxy-3-methylpentylphosphonic acid, an isosteric analogue of 2-C-methyl-D-erythritol 4-phosphate. The enzyme was, however, less efficient with the methylene phosphonate analogue than with the natural substrate.
Assuntos
Aldose-Cetose Isomerases/metabolismo , Eritritol/análogos & derivados , Eritritol/metabolismo , Complexos Multienzimáticos/metabolismo , Compostos Organofosforados/síntese química , Oxirredutases/metabolismo , Pentosefosfatos/síntese química , Fosfatos de Poli-Isoprenil/biossíntese , Fosfatos Açúcares/metabolismo , Aldose-Cetose Isomerases/química , Eritritol/química , Escherichia coli/metabolismo , Cinética , Espectroscopia de Ressonância Magnética , Complexos Multienzimáticos/química , Compostos Organofosforados/química , Oxirredutases/química , Pentosefosfatos/química , Pentosefosfatos/metabolismo , Fosfatos de Poli-Isoprenil/química , Fosfatos Açúcares/químicaRESUMO
In plants, two pathways are utilized for the synthesis of isopentenyl diphosphate, the universal precursor for isoprenoid biosynthesis. The key enzyme of the cytoplasmic mevalonic acid (MVA) pathway is 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR). Treatment of Tobacco Bright Yellow-2 (TBY-2) cells by the HMGR-specific inhibitor mevinolin led to growth reduction and induction of apparent HMGR activity, in parallel to an increase in protein representing two HMGR isozymes. Maximum induction was observed at 24 h. 1-Deoxy-d-xylulose (DX), the dephosphorylated first precursor of the plastidial 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway, complemented growth inhibition by mevinolin in the low millimolar concentration range. Furthermore, DX partially re-established feedback repression of mevinolin-induced HMGR activity. Incorporation studies with [1,1,1,4-2H4]DX showed that sterols, normally derived from MVA, in the presence of mevinolin are synthesized via the MEP pathway. Fosmidomycin, an inhibitor of 1-deoxy-d-xylulose-5-phosphate reductoisomerase, the second enzyme of the MEP pathway, was utilized to study the reverse complementation. Growth inhibition by fosmidomycin of TBY-2 cells could be partially overcome by MVA. Chemical complementation was further substantiated by incorporation of [2-13C]MVA into plastoquinone, representative of plastidial isoprenoids. Best rates of incorporation of exogenous stably labeled precursors were observed in the presence of both inhibitors, thereby avoiding internal isotope dilution.
Assuntos
Eritritol/análogos & derivados , Eritritol/metabolismo , Fosfomicina/análogos & derivados , Ácido Mevalônico/metabolismo , Nicotiana/metabolismo , Fosfatos Açúcares/metabolismo , Xilulose/análogos & derivados , Citosol/metabolismo , Fosfomicina/farmacologia , Hidroximetilglutaril-CoA Redutases/metabolismo , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Lovastatina/farmacologia , Fitosteróis/biossíntese , Plastídeos/metabolismo , Plastoquinona/metabolismo , Transdução de Sinais , Nicotiana/citologia , Nicotiana/efeitos dos fármacos , Xilulose/farmacologiaRESUMO
The 1-deoxyxylulose 5-phosphate reductoisomerase (DXR, EC 1.1.1.267) catalyzes the conversion of 1-deoxy-d-xylulose 5-phosphate (DXP) into 2-C-methyl-d-erythritol 4-phosphate (MEP). This transformation is a two-step process involving a rearrangement of DXP into the putative intermediate 2-C-methyl-d-erythrose 4-phosphate followed by a NADPH-dependent reduction of the latter aldehyde. By using [1-(13)C]DXP as a substrate, the rearrangement of DXP into [5-(13)C]2-C-methyl-d-erythrose 4-phosphate was shown to be NADPH dependent, although it does not involve areduction step. The putative aldehyde intermediate, obtained by chemical synthesis, was converted into MEP by the DXR in the presence of NADPH and into DXP in the presence of NADP(+), indicating the reversibility of the reaction catalyzed by the DXR. This reversibility was confirmed by the conversion of MEP into DXP in the presence of NADP(+). The equilibrium was, however, largely displaced in favour of the formation of MEP. The reduction step required the presence of a divalent cation such as Mg(2+) or Mn(2+).
Assuntos
Aldose-Cetose Isomerases/metabolismo , Eritritol/análogos & derivados , Eritritol/metabolismo , Complexos Multienzimáticos/metabolismo , Oxirredutases/metabolismo , Pentosefosfatos/metabolismo , Fosfatos Açúcares/metabolismo , Eritritol/química , Escherichia coli/metabolismo , Cinética , NADP/metabolismo , Pentosefosfatos/química , Fosfatos Açúcares/químicaRESUMO
In the bacterium Escherichia coli, the mevalonic-acid (MVA)-independent 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway is characterized by two branches leading separately to isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). The signature of this branching is the retention of deuterium in DMAPP and the deuterium loss in IPP after incorporation of 1-[4-(2)H]deoxy-d-xylulose ([4-(2)H]DX). Feeding tobacco BY-2 cell-suspension cultures with [4-(2)H]DX resulted in deuterium retention in the isoprene units derived from DMAPP, as well as from IPP in the plastidial isoprenoids, phytoene and plastoquinone, synthesized via the MEP pathway. This labelling pattern represents direct evidence for the presence of the DMAPP branch of the MEP pathway in a higher plant, and shows that IPP can be synthesized from DMAPP in plant plastids, most probably via a plastidial IPP isomerase.