RESUMEN
Toxoplasma gondii is a widely distributed apicomplexan parasite causing toxoplasmosis, a critical health issue for immunocompromised individuals and for congenitally infected foetuses. Current treatment options are limited in number and associated with severe side effects. Thus, novel anti-toxoplasma agents need to be identified and developed. 1-Deoxy-d-xylulose 5-phosphate reductoisomerase (DXR) is considered the rate-limiting enzyme in the non-mevalonate pathway for the biosynthesis of the isoprenoid precursors isopentenyl pyrophosphate and dimethylallyl pyrophosphate in the parasite, and has been previously investigated for its key role as a novel drug target in some species, encompassing Plasmodia, Mycobacteria and Escherichia coli. In this study, we present the first crystal structure of T. gondii DXR (TgDXR) in a tertiary complex with the inhibitor fosmidomycin and the cofactor NADPH in dimeric conformation at 2.5â Å resolution revealing the inhibitor binding mode. In addition, we biologically characterize reverse α-phenyl-ß-thia and ß-oxa fosmidomycin analogues and show that some derivatives are strong inhibitors of TgDXR which also, in contrast with fosmidomycin, inhibit the growth of T. gondii in vitro. Here, ((3,4-dichlorophenyl)((2-(hydroxy(methyl)amino)-2-oxoethyl)thio)methyl)phosphonic acid was identified as the most potent anti T. gondii compound. These findings will enable the future design and development of more potent anti-toxoplasma DXR inhibitors.
Asunto(s)
Isomerasas Aldosa-Cetosa , Fosfomicina , Complejos Multienzimáticos , Toxoplasma , Toxoplasma/enzimología , Toxoplasma/efectos de los fármacos , Isomerasas Aldosa-Cetosa/antagonistas & inhibidores , Isomerasas Aldosa-Cetosa/química , Isomerasas Aldosa-Cetosa/metabolismo , Isomerasas Aldosa-Cetosa/genética , Fosfomicina/farmacología , Fosfomicina/análogos & derivados , Fosfomicina/química , Cristalografía por Rayos X , Complejos Multienzimáticos/antagonistas & inhibidores , Complejos Multienzimáticos/química , Complejos Multienzimáticos/metabolismo , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Proteínas Protozoarias/antagonistas & inhibidores , Proteínas Protozoarias/química , Proteínas Protozoarias/metabolismo , Proteínas Protozoarias/genética , NADP/metabolismo , NADP/química , Humanos , Modelos Moleculares , Oxidorreductasas/antagonistas & inhibidores , Oxidorreductasas/química , Oxidorreductasas/metabolismoRESUMEN
The practical and theoretical significance of the resolvability parameter makes it an important factor, particularly in the context of network analysis. Its significance is seen in various applications and consequences: Network security, efficient routing, social network analysis, facility location, and site selection. This article finds the double resolvability parameters of the fosmidomycin anti-malaria drug. Resolvability parameters like double metric, double edge metric, and double mixed metric dimensions of fosmidomycin anti-malaria drug also hold exchange properties in the molecular graph of fosmidomycin. We convert the molecular structures of fosmidomycin into molecular graphs and then find some resolvability parameters.
RESUMEN
Reverse analogs of the phosphonohydroxamic acid antibiotic fosmidomycin are potent inhibitors of the nonmevalonate isoprenoid biosynthesis enzyme 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR, IspC) of Plasmodium falciparum. Some novel analogs with large phenylalkyl substituents at the hydroxamic acid nitrogen exhibit nanomolar PfDXR inhibition and potent in vitro growth inhibition of P. falciparum parasites coupled with good parasite selectivity. X-ray crystallographic studies demonstrated that the N-phenylpropyl substituent of the newly developed lead compound 13e is accommodated in a subpocket within the DXR catalytic domain but does not reach the NADPH binding pocket of the N-terminal domain. As shown for reverse carba and thia analogs, PfDXR selectively binds the S-enantiomer of the new lead compound. In addition, some representatives of the novel inhibitor subclass are nanomolar Escherichia coli DXR inhibitors, whereas the inhibition of Mycobacterium tuberculosis DXR is considerably weaker.
Asunto(s)
Isomerasas Aldosa-Cetosa , Antimaláricos , Fosfomicina , Ácidos Hidroxámicos , Complejos Multienzimáticos , Plasmodium falciparum , Fosfomicina/farmacología , Fosfomicina/análogos & derivados , Fosfomicina/química , Isomerasas Aldosa-Cetosa/antagonistas & inhibidores , Isomerasas Aldosa-Cetosa/metabolismo , Isomerasas Aldosa-Cetosa/química , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/enzimología , Ácidos Hidroxámicos/farmacología , Ácidos Hidroxámicos/química , Antimaláricos/farmacología , Antimaláricos/química , Complejos Multienzimáticos/antagonistas & inhibidores , Complejos Multienzimáticos/metabolismo , Complejos Multienzimáticos/química , Cristalografía por Rayos X , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/síntesis química , Relación Estructura-Actividad , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Escherichia coli/enzimología , Modelos Moleculares , Mycobacterium tuberculosis/efectos de los fármacos , Mycobacterium tuberculosis/enzimología , Dominio Catalítico , Oxidorreductasas/antagonistas & inhibidores , Oxidorreductasas/metabolismoRESUMEN
Fosmidomycin (FOS) is a natural product inhibiting the DXR enzyme in the MEP pathway and has stimulated interest for finding more suitable FOS analogues. Herein, two series of FOS analogue hydroxamate-containing bisphosphonates as proherbicides were designed, with bisphosphonate replacing the phosphonic unit in FOS while retaining the hydroxamate (BPF series) or replacing it with retro-hydroxamate (BPRF series). The BPF series were synthesized through a three-step reaction sequence including Michael addition of vinylidenebisphosphonate, N-acylation, and deprotection, and the BPRF series were synthesized with a retro-Claisen condensation incorporated into the reaction sequence. Evaluation on model plants demonstrated several compounds having considerable herbicidal activities, and in particular, compound 8m exhibited multifold activity enhancement as compared to the control FOS. The proherbicide properties were comparatively validated. Furthermore, DXR enzyme assay, dimethylallyl pyrophosphate rescue, and molecular docking verified 8m to be a promising proherbicide candidate targeting the DXR enzyme. In addition, 8m also displayed good antimalarial activities.
Asunto(s)
Isomerasas Aldosa-Cetosa , Antimaláricos , Fosfomicina , Fosfomicina/análogos & derivados , Difosfonatos , Simulación del Acoplamiento Molecular , Fosfomicina/farmacología , Isomerasas Aldosa-Cetosa/metabolismoRESUMEN
BACKGROUND: The enzymes involved in the 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway are attractive targets of a new mode of action for developing anti-infective drugs and herbicides, and inhibitors against 1-deoxy-d-xylulose 5-phosphate reductoisomerase (IspC), the second key enzyme in the pathway, have been intensively investigated; however, few works are reported regarding IspC inhibitors designed for new herbicide discovery. RESULTS: A series of fosmidomycin (FOS) analogs were designed with nitrogen-containing linkers replacing the trimethylene linker between the two active substructures of FOS, phosphonic acid and hydroxamic acid. Synthesis followed a facile three-step route of sequential aza-Michael addition of α-amino acids to dibenzyl vinylphosphonate, amidation of the amino acid carboxyl with O-benzyl hydroxylamine, and simultaneous removal of the benzyl protective groups. Biological activity evaluation of IspC and model plants revealed that some compounds had moderate enzyme and model plant growth inhibition effects. In particular, compound 10g, which has a N-(4-fluorophenylethyl) nitrogen-containing linker, exhibited the best plant inhibition activities, superior to the control FOS against the model plants Arabidopsis thaliana, Brassica napus L., Amaranthus retroflexus and Echinochloa crus-galli. A dimethylallyl pyrophosphate rescue assay on A. thaliana confirmed that both 10g and FOS exert their herbicidal activity by blocking the MEP pathway. This result consistent with molecular docking, which confirmed 10g and FOS binding to the IspC active site in a similar way. CONCLUSION: Compound 10g has excellent herbicidal activity and represents the first herbicide lead structure of a new mode of action that targets IspC enzyme in the MEP pathway. © 2023 Society of Chemical Industry.
Asunto(s)
Eritritol/análogos & derivados , Fosfomicina , Herbicidas , Fosfatos de Azúcar , Simulación del Acoplamiento Molecular , Fosfomicina/farmacología , Herbicidas/química , NitrógenoRESUMEN
Fosmidomycin is a natural antibiotic with potent IspC (DXR, 1-deoxy-d-xylulose-5-phosphate reductoisomerase) inhibitory activity. This enzyme catalyzes the first committed step of the non-mevalonate isoprenoid biosynthesis pathway, which is essential in most bacteria, including A. baumanii and M. tuberculosis, and apicomplexan parasites, including Plasmodium parasites. Mainly as a result of its high polarity, fosmidomycin displays suboptimal pharmacokinetic properties. Furthermore, fosmidomycin is inactive against A. baumannii and M. tuberculosis as a result of its inability to penetrate the bacterial cell wall. Temporarily masking the phosphonate moiety as a prodrug has the potential to solve both issues. We report on the expansion of the acyloxymethyl and alkoxycarbonyloxymethyl phosphonate ester prodrug series of a fosmidomycin surrogate. Prodrug promoieties were designed based on electronic, lipophilic and siderophoric properties. This investigation led to the discovery of derivatives with two-digit nanomolar and submicromolar IC50-values against P. falciparum and A. baumanii, respectively.
Asunto(s)
Antimaláricos , Antagonistas del Ácido Fólico , Malaria Falciparum , Mycobacterium tuberculosis , Organofosfonatos , Profármacos , Tuberculosis , Humanos , Antimaláricos/farmacología , Antibacterianos/farmacología , Profármacos/farmacologíaRESUMEN
Plasmodium falciparum is the etiological agent of human malaria, one of the most widespread diseases in tropical and subtropical regions. Drug resistance is one of the biggest problems in controlling the disease, which leads to the need to discover new antimalarial compounds. One of the most promissory drugs purposed is fosmidomycin, an inhibitor of the biosynthesis of isoprene units by the methylerythritol 4-phosphate (MEP) pathway, which in some cases failed in clinical studies. Once formed, isoprene units are condensed to form longer structures such as farnesyl and geranylgeranyl pyrophosphate, which are necessary for Heme O and A formation, ubiquinone, and dolichyl phosphate biosynthesis as well as for protein isoprenylation. Even though the natural substrates of polyprenyl transferases and synthases are polyprenyl pyrophosphates, it was already demonstrated that isoprenoid alcohols (polyprenols) such as farnesol (FOH) and geranylgeraniol (GGOH) can rescue parasites from fosmidomycin. This study better investigated how this rescue phenomenon occurs by performing drug-rescue assays. Similarly, to FOH and GGOH, it was observed that phytol (POH), a 20-carbon plant isoprenoid, as well as unsaponifiable lipid extracts from foods rescue parasites from the antimalarial effect of fosmidomycin. Contrarily, neither dolichols nor nonaprenol rescue parasites from fosmidomycin. Considering this, here we characterized the transport of FOH, GGOH, and POH. Once incorporated, it was observed that these substances are phosphorylated, condensed into longer isoprenoid alcohols, and incorporated into proteins and dolichyl phosphates. Through proteomic and radiolabelling approaches, it was found that prenylated proteins are naturally attached to several isoprenoids, derived from GGOH, dolichol, and POH if exogenously added. Furthermore, the results suggest the presence of at least two promiscuous protein prenyltransferases in the parasite: one enzyme which can use FPP among other unidentified substrates and another enzyme that can use GGPP, phytyl pyrophosphate (PPP), and dolichols, among other substrates not identified here. Thus, further evidence was obtained for dolichols and other isoprenoid products attached to proteins. This study helps to better understand the apicoplast-targeting antimalarial mechanism of action and a novel post-translational modification of proteins in P. falciparum.
RESUMEN
Isoprenoids are the output of the polymerization of five-carbon, branched isoprenic chains derived from isopentenyl pyrophosphate (IPP) and its isomer, dimethylallyl pyrophosphate (DMAPP). Isoprene units are consecutively condensed to form longer structures such as farnesyl and geranylgeranyl pyrophosphate (FPP and GGPP, respectively), necessary for the biosynthesis of several metabolites. Polyprenyl transferases and synthases use polyprenyl pyrophosphates as their natural substrates; however, it is known that free polyprenols, such as farnesol (FOH), and geranylgeraniol (GGOH) can be incorporated into prenylated proteins, ubiquinone, cholesterol, and dolichols. Furthermore, FOH and GGOH have been shown to block the effects of isoprenoid biosynthesis inhibitors such as fosmidomycin, bisphosphonates, or statins in several organisms. This phenomenon is the consequence of a short pathway, which was observed for the first time more than 25 years ago: the polyprenol salvage pathway, which works via the phosphorylation of FOH and GGOH. Biochemical studies in bacteria, animals, and plants suggest that this pathway can be carried out by two enzymes: a polyprenol kinase and a polyprenyl-phosphate kinase. However, to date, only a few genes have been unequivocally identified to encode these enzymes in photosynthetic organisms. Nevertheless, pieces of evidence for the importance of this pathway abound in studies related to infectious diseases, cancer, dyslipidemias, and nutrition, and to the mitigation of the secondary effects of several drugs. Furthermore, nowadays it is known that both FOH and GGOH can be incorporated via dietary sources that produce various biological effects. This review presents, in a simplified but comprehensive manner, the most important data on the FOH and GGOH salvage pathway, stressing its biomedical importance The main objective of this review is to bring to light the need to discover and characterize the kinases associated with the isoprenoid salvage pathway in animals and pathogens.
Asunto(s)
Diterpenos , Inhibidores de Hidroximetilglutaril-CoA Reductasas , Animales , Farnesol/farmacología , Diterpenos/farmacología , Diterpenos/metabolismo , Terpenos/farmacología , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacologíaRESUMEN
To address the continued rise of multi-drug-resistant microorganisms, the development of novel drugs with new modes of action is urgently required. While humans biosynthesize the essential isoprenoid precursors isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) via the established mevalonate pathway, pathogenic protozoa and certain pathogenic eubacteria use the less well-known methylerythritol phosphate pathway for this purpose. Important pathogens using the MEP pathway are, for example, Plasmodium falciparum, Mycobacterium tuberculosis, Pseudomonas aeruginosa and Escherichia coli. The enzymes of that pathway are targets for antiinfective drugs that are exempt from target-related toxicity. 2C-Methyl-D-erythritol 4-phosphate (MEP), the second enzyme of the non-mevalonate pathway, has been established as the molecular target of fosmidomycin, an antibiotic that has so far failed to be approved as an anti-infective drug. This review describes the development and anti-infective properties of a wide range of fosmidomycin derivatives synthesized over the last four decades. Here we discuss the DXR inhibitor pharmacophore, which comprises a metal-binding group, a phosphate or phosphonate moiety and a connecting linker. Furthermore, non-fosmidomycin-based DXRi, bisubstrate inhibitors and several prodrug concepts are described. A comprehensive structure-activity relationship (SAR) of nearly all inhibitor types is presented and some novel opportunities for further drug development of DXR inhibitors are discussed.
RESUMEN
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.
Asunto(s)
Antibacterianos/farmacología , Fosfomicina/análogos & derivados , Ácidos Hidroxámicos/farmacología , Isomerasas Aldosa-Cetosa/antagonistas & inhibidores , Isomerasas Aldosa-Cetosa/metabolismo , Farmacorresistencia Bacteriana/efectos de los fármacos , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Escherichia coli/efectos de los fármacos , Escherichia coli/crecimiento & desarrollo , Fosfomicina/síntesis química , Fosfomicina/química , Fosfomicina/farmacología , Pruebas de Sensibilidad MicrobianaRESUMEN
MAIN CONCLUSION: Greening of in vitro callus cultures and andrographolide over-accumulation was achieved by manipulating light exposure and media composition, when the biosynthetic cascade was channeled through the DXP pathway. Andrographolide, the primary biologically active compound of Andrographis paniculata, is produced through coordinated action of two pathways, the classical cytosolic mevalonate pathway and the alternative plastidial non-mevalonate pathway (Deoxy-xylulose Phosphate pathway). In vitro callus cultures of A. paniculata are useful sources of production, as well as, manipulation of andrographolide, and the present study was designed to explore the strategy of pathway inhibition for its overproduction. When the cytosolic mevalonate pathway blocker, lovastatin, was applied to callus cultures of A. paniculata, andrographolide production was enhanced in comparison to untreated control. In contrast, treatment of the callus tissue with the DXP-pathway blocker, fosmidomycin, led to depletion in andrographolide production. The present study also showed that silver nitrate, a potent elicitor of andrographolide production in in vitro callus culture, when added in combination with the pathway inhibitors resulted in alterations in andrographolide production. The highest andrographolide production was obtained in callus treated with a combination of silver nitrate and lovastatin, indicating a predominant role of the plastidial DXP pathway in andrographolide biosynthesis. A positive co-relation with chlorophyll content and andrographolide production in in vitro callus cultures (untreated and treated) observed also supported the above assumption. It could be inferred from this study that greening of callus tissue through organellar organization was a potent strategy for enhancing andrographolide accumulation in callus tissues of A. paniculata.
Asunto(s)
Andrographis , Diterpenos , Extractos VegetalesRESUMEN
BACKGROUND: We report a method to estimate carbon assimilation based on isotope ratio-mass spectrometry (IRMS) of 13CO2 labeled plant tissue. Photosynthetic carbon assimilation is the principal experimental observable which integrates important aspects of primary plant metabolism. It is traditionally measured through gas exchange. Despite its centrality in plant research, gas exchange performs poorly with rosette growth habits typical of Arabidopsis thaliana, mutant lines with limited biomass, and accounts poorly for leaf shading. RESULTS: IRMS-based carbon assimilation values from plants labeled at different light intensities were compared to those obtained by gas exchange, and the two methods yielded similar values. Using this method, we observed a strong correlation between 13C content and labeling time (R2 = 0.999) for 158 wild-type plants labeled for 6 to 42 min. Plants cultivated under different light regimes showed a linear response with respect to carbon assimilation, varying from 7.38 nmol 13C mg-1 leaf tissue min-1 at 80 PAR to 19.27 nmol 13C mg-1 leaf tissue min-1 at 500 PAR. We applied this method to examine the link between inhibition of the 2C-methyl-D-erythritol-4-phosphate (MEP) pathway and suppression of photosynthesis. A significant decrease in carbon assimilation was observed when metabolic activity in the MEP pathway was compromised by mutation or herbicides targeting the MEP pathway. Mutants affected in MEP pathway genes 1-DEOXY-D-XYLULOSE 5-PHOSPHATE SYNTHASE (DXS) or 1-HYDROXY-2-METHYL-2-(E)-BUTENYL 4-DIPHOSPHATE SYNTHASE (HDS) showed assimilation rates 36% and 61% lower than wild type. Similarly, wild type plants treated with the MEP pathway inhibitors clomazone or fosmidomycin showed reductions of 52% and 43%, respectively, while inhibition of the analogous mevalonic acid pathway, which supplies the same isoprenoid intermediates in the cytosol, did not, suggesting inhibition of photosynthesis was specific to disruption of the MEP pathway. CONCLUSIONS: This method provides an alternative to gas exchange that offers several advantages: resilience to differences in leaf overlap, measurements based on tissue mass rather than leaf surface area, and compatibility with mutant Arabidopsis lines which are not amenable to gas exchange measurements due to low biomass and limited leaf surface area. It is suitable for screening large numbers of replicates simultaneously as well as post-hoc analysis of previously labeled plant tissue and is complementary to downstream detection of isotopic label in targeted metabolite pools.
RESUMEN
The Gram-negative bacterium Pseudomonas aeruginosa is an opportunistic pathogen, responsible for many hospital-acquired infections. The bacterium is quite resistant toward many antibiotics, in particular because of the fine-tuned permeability of its outer membrane (OM). General diffusion outer membrane pores are quite rare in this organism. Instead, its OM contains many substrate-specific porins. Their expression is varying according to growth conditions and virulence. Phosphate limitations, as well as pathogenicity factors, result in the induction of the two mono- and polyphosphate-specific porins, OprP and OprO, respectively, together with an inner membrane uptake mechanism and a periplasmic binding protein. These outer membrane channels could serve as outer membrane pathways for the uptake of phosphonates. Among them are not only herbicides, but also potent antibiotics, such as fosfomycin and fosmidomycin. In this study, we investigated the interaction between OprP and OprO and fosmidomycin in detail. We could demonstrate that fosmidomycin is able to bind to the phosphate-specific binding site inside the two porins. The inhibition of chloride conductance of OprP and OprO by fosmidomycin is considerably less than that of phosphate or diphosphate, but it can be measured in titration experiments of chloride conductance and also in single-channel experiments. The results suggest that fosmidomycin transport across the OM of P. aeruginosa occurs through OprP and OprO. Our data with the ones already known in the literature show that phosphonic acid-containing antibiotics are in general good candidates to treat the infections of P. aeruginosa at the very beginning through a favorable OM transport system.
Asunto(s)
Antibacterianos/metabolismo , Proteínas Bacterianas/metabolismo , Fosfomicina/análogos & derivados , Transporte Iónico/fisiología , Porinas/metabolismo , Pseudomonas aeruginosa/metabolismo , Proteínas de la Membrana Bacteriana Externa/metabolismo , Proteínas Bacterianas/genética , Sitios de Unión/fisiología , Cloruros/metabolismo , Farmacorresistencia Bacteriana Múltiple/genética , Fosfomicina/metabolismo , Ácidos Fosforosos/metabolismo , Porinas/genética , Pseudomonas aeruginosa/efectos de los fármacos , Pseudomonas aeruginosa/genéticaRESUMEN
The malaria parasite Plasmodium falciparum contains the apicoplast organelle that synthesizes isoprenoids, which are metabolites necessary for posttranslational modification of Plasmodium proteins. We used fosmidomycin, an antibiotic that inhibits isoprenoid biosynthesis, to identify mechanisms that underlie the development of the parasite's adaptation to the drug at sublethal concentrations. We first determined a concentration of fosmidomycin that reduced parasite growth by â¼50% over one intraerythrocytic developmental cycle (IDC). At this dose, we maintained synchronous parasite cultures for one full IDC and collected metabolomic and transcriptomic data at multiple time points to capture global and stage-specific alterations. We integrated the data with a genome-scale metabolic model of P. falciparum to characterize the metabolic adaptations of the parasite in response to fosmidomycin treatment. Our simulations showed that, in treated parasites, the synthesis of purine-based nucleotides increased, whereas the synthesis of phosphatidylcholine during the trophozoite and schizont stages decreased. Specifically, the increased polyamine synthesis led to increased nucleotide synthesis, while the reduced methyl-group cycling led to reduced phospholipid synthesis and methyltransferase activities. These results indicate that fosmidomycin-treated parasites compensate for the loss of prenylation modifications by directly altering processes that affect nucleotide synthesis and ribosomal biogenesis to control the rate of RNA translation during the IDC. This also suggests that combination therapies with antibiotics that target the compensatory response of the parasite, such as nucleotide synthesis or ribosomal biogenesis, may be more effective than treating the parasite with fosmidomycin alone.
Asunto(s)
Antimaláricos , Apicoplastos , Fosfomicina , Malaria Falciparum , Antimaláricos/uso terapéutico , Fosfomicina/análogos & derivados , Fosfomicina/farmacología , Fosfomicina/uso terapéutico , Humanos , Malaria Falciparum/tratamiento farmacológico , Plasmodium falciparum/genéticaRESUMEN
A series of N-benzylated phosphoramidate esters, containing a 3,4-dihydroxyphenyl Mg2+-chelating group, has been synthesised in five steps as analogues of fosmidomycin, a Plasmodium falciparum 1-deoxy-1-d-xylulose-5-phosphate reductoisomerase (PfDXR) inhibitor. The 3,4-dihydroxyphenyl group effectively replaces the Mg2+-chelating hydroxamic acid group in fosmidomycin. The compounds showed very encouraging anti-parasitic activity with IC50 values of 5.6-16.4 µM against Plasmodium falciparum parasites and IC50 values of 5.2 - 10.2 µM against Trypanosoma brucei brucei (T.b.brucei). Data obtained from in silico docking of the ligands in the PfDXR receptor cavity (3AU9)5 support their potential as PfDXR inhibitors.
Asunto(s)
Amidas/síntesis química , Antimaláricos/síntesis química , Complejos de Coordinación/síntesis química , Magnesio/química , Ácidos Fosfóricos/síntesis química , Plasmodium falciparum/efectos de los fármacos , Antimaláricos/farmacología , Complejos de Coordinación/farmacología , Relación Dosis-Respuesta a Droga , Diseño de Fármacos , Fosfomicina/análogos & derivados , Fosfomicina/farmacología , Células HeLa , Humanos , Ligandos , Simulación del Acoplamiento Molecular , Trypanosoma brucei brucei/efectos de los fármacosRESUMEN
Malaria, despite many efforts, remains among the most problematic infectious diseases worldwide, mainly due to the development of drug resistance by Plasmodium falciparum. The antibiotic fosmidomycin (FSM) is also known for its antimalarial activity by targeting the non-mevalonate isoprenoid synthesis pathway, which is essential for the malaria parasites but is absent in mammalians. In this study, we synthesized and evaluated against the chloroquine-resistant P. falciparum FcB1/Colombia strain, a series of FSM analogs, derivatives, and conjugates with other antimalarial agents, such as artemisinin (ART) and aminochloroquinoline (ACQ). The biological evaluation revealed four new compounds with higher antimalarial activity than FSM: two FSM-ACQ derivatives and two FSM-ART conjugates, with 3.5-5.4 and 41.5-23.1 times more potent activities than FSM, respectively.
Asunto(s)
Antimaláricos/farmacología , Artemisininas/farmacología , Fosfomicina/análogos & derivados , Plasmodium falciparum/efectos de los fármacos , Quinolinas/farmacología , Antimaláricos/síntesis química , Antimaláricos/química , Artemisininas/química , Fosfomicina/síntesis química , Fosfomicina/química , Fosfomicina/farmacología , Estructura Molecular , Pruebas de Sensibilidad Parasitaria , Quinolinas/químicaRESUMEN
Tropical forests absorb large amounts of atmospheric CO2 through photosynthesis, but high surface temperatures suppress this absorption while promoting isoprene emissions. While mechanistic isoprene emission models predict a tight coupling to photosynthetic electron transport (ETR) as a function of temperature, direct field observations of this phenomenon are lacking in the tropics and are necessary to assess the impact of a warming climate on global isoprene emissions. Here we demonstrate that in the early successional species Vismia guianensis in the central Amazon, ETR rates increased with temperature in concert with isoprene emissions, even as stomatal conductance (gs ) and net photosynthetic carbon fixation (Pn ) declined. We observed the highest temperatures of continually increasing isoprene emissions yet reported (50°C). While Pn showed an optimum value of 32.6 ± 0.4°C, isoprene emissions, ETR, and the oxidation state of PSII reaction centers (qL ) increased with leaf temperature with strong linear correlations for ETR (Æ¿ = 0.98) and qL (Æ¿ = 0.99) with leaf isoprene emissions. In contrast, other photoprotective mechanisms, such as non-photochemical quenching, were not activated at elevated temperatures. Inhibition of isoprenoid biosynthesis repressed Pn at high temperatures through a mechanism that was independent of stomatal closure. While extreme warming will decrease gs and Pn in tropical species, our observations support a thermal tolerance mechanism where the maintenance of high photosynthetic capacity under extreme warming is assisted by the simultaneous stimulation of ETR and metabolic pathways that consume the direct products of ETR including photorespiration and the biosynthesis of thermoprotective isoprenoids. Our results confirm that models which link isoprene emissions to the rate of ETR hold true in tropical species and provide necessary "ground-truthing" for simulations of the large predicted increases in tropical isoprene emissions with climate warming.
Asunto(s)
Butadienos , Hemiterpenos , Dióxido de Carbono , Transporte de Electrón , Fotosíntesis , Hojas de la PlantaRESUMEN
Babesia microti, the main pathogen causing human babesiosis, has been reported to exhibit resistance to the traditional treatment of azithromycin + atovaquone and clindamycin + quinine, suggesting the necessity of developing new drugs. The methylerythritol 4-phosphate (MEP) pathway, a unique pathway in apicomplexan parasites, was shown to play a crucial function in the growth of Plasmodium falciparum. In the MEP pathway, 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR) is a rate-limiting enzyme and fosmidomycin (FSM) is a reported inhibitor for this enzyme. DXR has been shown as an antimalarial drug target, but no report is available on B. microti DXR (BmDXR). Here BmDXR was cloned, sequenced, analyzed by bioinformatics, and evaluated as a potential drug target for inhibiting the growth of B. micorti in vitro. Drug assay was performed by adding different concentrations of FSM in B. microti in vitro culture. Rescue experiment was done by supplementing 200 µM isopentenyl pyrophosphate (IPP) or 5 µM geranylgeraniol (GG-ol) in the culture medium together with 5 µM FSM or 10 µM diminazene aceturate. The results indicated that FSM can inhibit the growth of B. microti in in vitro culture with an IC50 of 4.63 ± 0.12 µM, and growth can be restored by both IPP and GG-ol. Additionally, FSM is shown to inhibit the growth of parasites by suppressing the DXR activity, which agreed with the reported results of other apicomplexan parasites. Our results suggest the potential of DXR as a drug target for controlling B. microti and that FSM can inhibit the growth of B. microti in vitro.
RESUMEN
Drug resistance is a major healthcare challenge, resulting in a continuous need to develop new inhibitors. The development of these inhibitors requires an understanding of the mechanisms of resistance for a critical mass of occurrences. Recent genome editing technologies based on high-throughput DNA synthesis and sequencing may help to predict mutations resulting in resistance by testing large mutagenesis libraries. Here we describe the rationale of this approach, with examples and relevance to drug development and resistance in malaria.
Asunto(s)
Isomerasas Aldosa-Cetosa/química , Evolución Molecular Dirigida/métodos , Resistencia a Medicamentos/genética , Malaria/tratamiento farmacológico , Mutagénesis , Isomerasas Aldosa-Cetosa/antagonistas & inhibidores , Isomerasas Aldosa-Cetosa/metabolismo , Antibacterianos/farmacología , Escherichia coli/genética , Escherichia coli/metabolismo , Fosfomicina/análogos & derivados , Fosfomicina/farmacología , Biblioteca de Genes , Mutación , Plasmodium falciparum/genética , Plasmodium falciparum/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismoRESUMEN
Members of the haloacid dehalogenase (HAD) family of metabolite phosphatases play an important role in regulating multiple pathways in Plasmodium falciparum central carbon metabolism. We show that the P. falciparum HAD protein, phosphoglycolate phosphatase (PGP), regulates glycolysis and pentose pathway flux in asexual blood stages via detoxifying the damaged metabolite 4-phosphoerythronate (4-PE). Disruption of the P. falciparumpgp gene caused accumulation of two previously uncharacterized metabolites, 2-phospholactate and 4-PE. 4-PE is a putative side product of the glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase, and its accumulation inhibits the pentose phosphate pathway enzyme, 6-phosphogluconate dehydrogenase (6-PGD). Inhibition of 6-PGD by 4-PE leads to an unexpected feedback response that includes increased flux into the pentose phosphate pathway as a result of partial inhibition of upper glycolysis, with concomitant increased sensitivity to antimalarials that target pathways downstream of glycolysis. These results highlight the role of metabolite detoxification in regulating central carbon metabolism and drug sensitivity of the malaria parasite.IMPORTANCE The malaria parasite has a voracious appetite, requiring large amounts of glucose and nutrients for its rapid growth and proliferation inside human red blood cells. The host cell is resource rich, but this is a double-edged sword; nutrient excess can lead to undesirable metabolic reactions and harmful by-products. Here, we demonstrate that the parasite possesses a metabolite repair enzyme (PGP) that suppresses harmful metabolic by-products (via substrate dephosphorylation) and allows the parasite to maintain central carbon metabolism. Loss of PGP leads to the accumulation of two damaged metabolites and causes a domino effect of metabolic dysregulation. Accumulation of one damaged metabolite inhibits an essential enzyme in the pentose phosphate pathway, leading to substrate accumulation and secondary inhibition of glycolysis. This work highlights how the parasite coordinates metabolic flux by eliminating harmful metabolic by-products to ensure rapid proliferation in its resource-rich niche.