RESUMO
Malaria is one of the most widespread parasitic diseases, especially in Africa, Southeast Asia and South America. One of the greatest problems for control of the disease is the emergence of drug resistance, which leads to a need for the development of new antimalarial compounds. The biosynthesis of isoprenoids has been investigated as part of a strategy to identify new targets to obtain new antimalarial drugs. Several isoprenoid quinones, including menaquinone-4 (MK-4/vitamin K2), α- and γ-tocopherol and ubiquinone (UQ) homologs UQ-8 and UQ-9, were previously detected in in vitro cultures of Plasmodium falciparum in asexual stages. Herein, we described for the first time the presence of phylloquinone (PK/vitamin K1) in P. falciparum and discuss the possible origins of this prenylquinone. While our results in metabolic labeling experiments suggest a biosynthesis of PK prenylation via phytyl pyrophosphate (phytyl-PP) with phytol being phosphorylated, on the other hand, exogenous PK attenuated atovaquone effects on parasitic growth and respiration, showing that this metabolite can be transported from extracellular environment and that the mitochondrial electron transport system (ETS) of P. falciparum is capable to interact with PK. Although the natural role and origin of PK remains elusive, this work highlights the PK importance in plasmodial metabolism and future studies will be important to elucidate in seeking new targets for antimalarial drugs.
Assuntos
Antimaláricos , Malária Falciparum , Malária , Antimaláricos/farmacologia , Humanos , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Plasmodium falciparum , Vitamina K 1/metabolismo , Vitamina K 1/farmacologiaRESUMO
Human parasitic protozoa cause a large number of diseases worldwide and, for some of these diseases, there are no effective treatments to date, and drug resistance has been observed. For these reasons, the discovery of new etiological treatments is necessary. In this sense, parasitic metabolic pathways that are absent in vertebrate hosts would be interesting research candidates for the identification of new drug targets. Most likely due to the protozoa variability, uncertain phylogenetic origin, endosymbiotic events, and evolutionary pressure for adaptation to adverse environments, a surprising variety of prenylquinones can be found within these organisms. These compounds are involved in essential metabolic reactions in organisms, for example, prevention of lipoperoxidation, participation in the mitochondrial respiratory chain or as enzymatic cofactors. This review will describe several prenylquinones that have been previously characterized in human pathogenic protozoa. Among all existing prenylquinones, this review is focused on ubiquinone, menaquinone, tocopherols, chlorobiumquinone, and thermoplasmaquinone. This review will also discuss the biosynthesis of prenylquinones, starting from the isoprenic side chains to the aromatic head group precursors. The isoprenic side chain biosynthesis maybe come from mevalonate or non-mevalonate pathways as well as leucine dependent pathways for isoprenoid biosynthesis. Finally, the isoprenic chains elongation and prenylquinone aromatic precursors origins from amino acid degradation or the shikimate pathway is reviewed. The phylogenetic distribution and what is known about the biological functions of these compounds among species will be described, as will the therapeutic strategies associated with prenylquinone metabolism in protozoan parasites.
Assuntos
Antineoplásicos/farmacologia , Antiprotozoários/farmacologia , Parasitos/efeitos dos fármacos , Quinonas/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/metabolismo , Antiprotozoários/química , Antiprotozoários/metabolismo , Vias Biossintéticas , Humanos , Estrutura Molecular , Parasitos/metabolismo , Quinonas/química , Quinonas/metabolismo , Simbiose/efeitos dos fármacosRESUMO
Farnesyl diphosphate synthase/geranylgeranyl diphosphate synthase (FPPS/GGPPS) is a key enzyme in the synthesis of isoprenic chains. Risedronate, a bisphosphonate containing nitrogen (N-BP), is a potent inhibitor of blood stage Plasmodium. Here, we show that P. falciparum parasites overexpressing FPPS/GGPPS are more resistant to risedronate, suggesting that this enzyme is an important target, and bisphosphonate analogues can be used as potential antimalarial drugs.
Assuntos
Antimaláricos/farmacologia , Farnesiltranstransferase/biossíntese , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/enzimologia , Ácido Risedrônico/farmacologia , Análise de Variância , Animais , Antimaláricos/análise , Western Blotting , Resistência a Medicamentos , Farnesiltranstransferase/análise , Plasmodium falciparum/crescimento & desenvolvimento , Valores de Referência , Ácido Risedrônico/análiseRESUMO
The development of new drugs is one of the strategies to control malaria. Isoprenoid biosynthesis in Plasmodium falciparum is an essential pathway for parasite survival, and is therefore a potential target for new antimalarial drugs. Indeed, plant-derived secondary metabolites, such as terpenes, exhibit antimalarial activity in vitro by inhibiting isoprenoid biosynthesis in P. falciparum. In this study, the in vitro antiplasmodial activity of perillyl alcohol (POH) was evaluated, along with its in vitro toxicity and its effect on the isoprenylation process. In addition, the efficacy of intranasally administered POH in preventing Plasmodium berghei ANKA-induced experimental cerebral malaria (ECM) was determined. The 50% inhibitory concentrations of POH for 3D7 and K1 P. falciparum were 4.8 µM and 10.4 µM, respectively. POH inhibited farnesylation of 20-37 kDa proteins in P. falciparum (3D7), but no toxic effects in Vero cells were observed. A 500 mg/kg/d dose of POH had no effect on P. berghei ANKA parasitaemia, but showed marked efficacy in preventing ECM development (70% survival compared with 30% for untreated animals). This effect was associated with the downregulation of cerebrovascular inflammation and damage, with marked decreases in brain leucocyte accumulation and the incidence of brain microhaemorrhage. POH also downregulated interleukin (IL)-10, IL-6, tumour necrosis factor-α, interferon-γ, IL-12 and monocyte chemoattractant protein-1 levels in the brain and spleen. In conclusion, POH shows antiplasmodial activity in vitro and, despite there being no evidence of antiplasmodial activity in vivo following intranasal administration, POH prevented cerebrovascular inflammation/damage and expression of pro-inflammatory cytokines.
Assuntos
Antimaláricos/administração & dosagem , Antimaláricos/farmacologia , Malária Cerebral/prevenção & controle , Monoterpenos/administração & dosagem , Monoterpenos/farmacologia , Plasmodium berghei/efeitos dos fármacos , Plasmodium falciparum/efeitos dos fármacos , Administração Intranasal , Animais , Encéfalo/patologia , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Modelos Animais de Doenças , Células Epiteliais/efeitos dos fármacos , Concentração Inibidora 50 , Masculino , Camundongos Endogâmicos C57BL , Testes de Sensibilidade Parasitária , Resultado do Tratamento , Células VeroRESUMO
BACKGROUND: Plasmodium falciparum is sensitive to oxidative stress in vitro and in vivo, and many drugs such as artemisinin, chloroquine and cercosporin interfere in the parasite's redox system. To minimize the damage caused by reactive radicals, antioxidant enzymes and their substrates found in parasites and in erythrocytes must be functionally active. It was shown that P. falciparum synthesizes vitamin E and that usnic acid acts as an inhibitor of its biosynthesis. Vitamin E is a potent antioxidant that protects polyunsaturated fatty acids from lipid peroxidation, and this activity can be measured by detecting its oxidized product and by evaluating reactive oxygen species (ROS) levels. RESULTS: Here, we demonstrated that ROS levels increased in P. falciparum when vitamin E biosynthesis was inhibited by usnic acid treatment and decreased to basal levels if exogenous vitamin E was added. Furthermore, we used metabolic labelling to demonstrate that vitamin E biosynthesized by the parasite acts as an antioxidant since we could detect its radiolabeled oxidized product. The treatment with chloroquine or cercosporin of the parasites increased the ratio between α-tocopherolquinone and α-tocopherol. CONCLUSIONS: Our findings demonstrate that vitamin E produced endogenously by P. falciparum is active as an antioxidant, probably protecting the parasite from the radicals generated by drugs.
Assuntos
Estresse Oxidativo , Plasmodium falciparum/metabolismo , Vitamina E/metabolismo , Animais , Antimaláricos/farmacologia , Benzofuranos/farmacologia , Cloroquina/farmacologia , Eritrócitos/parasitologia , Eritrócitos/ultraestrutura , Humanos , Malária Falciparum/parasitologia , Microscopia de Fluorescência , Perileno/análogos & derivados , Perileno/farmacologia , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/crescimento & desenvolvimento , Proteínas de Protozoários/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Vitamina E/biossínteseRESUMO
Malaria, an infectious disease that kills more than 438,000 people per year worldwide, is a major public health problem. The emergence of strains resistant to conventional therapeutic agents necessitates the discovery of new drugs. We previously demonstrated that various substances, including terpenes, have antimalarial activity in vitro and in vivo. Nerolidol is a sesquiterpene present as an essential oil in several plants that is used in scented products and has been approved by the US Food and Drug Administration as a food-flavouring agent. In this study, the antimalarial activity of nerolidol was investigated in a mouse model of malaria. Mice were infected with Plasmodium berghei ANKA and were treated with 1000 mg/kg/dose nerolidol in two doses delivered by the oral or inhalation route. In mice treated with nerolidol, parasitaemia was inhibited by >99% (oral) and >80% (inhalation) until 14 days after infection (P <0.0001). On Day 30 post-infection, the survival rate of orally treated mice was 90% compared with 16% in controls (P <0.0001). In contrast, inhalation-treated mice showed a survival rate of 50% vs. 42% in controls (P > 0.05). The toxicity of nerolidol administered by either route was not significant, whilst genotoxicity was observed only at the highest dose tested. These results indicate that combined use of nerolidol and other drugs targeting different points of the same isoprenoid pathway may be an effective treatment for malaria.
Assuntos
Antimaláricos/administração & dosagem , Malária/tratamento farmacológico , Plasmodium berghei/efeitos dos fármacos , Sesquiterpenos/administração & dosagem , Administração por Inalação , Administração Oral , Animais , Antimaláricos/efeitos adversos , Antimaláricos/farmacologia , Modelos Animais de Doenças , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Masculino , Camundongos Endogâmicos BALB C , Parasitemia/tratamento farmacológico , Sesquiterpenos/efeitos adversos , Sesquiterpenos/farmacologia , Análise de Sobrevida , Terpenos/farmacologia , Resultado do TratamentoRESUMO
Malaria is a tropical disease with significant morbidity and mortality. A better understanding of the metabolism of its most important etiological agent, Plasmodium falciparum, is paramount to the development of better treatment and other mitigation measures. Farnesyldiphosphate synthase/geranylgeranyldiphosphate synthase (FPPS/GGPPS) is a key enzyme in the synthesis of isoprenic chains present in many essential structures. In P. falciparum, as well as a handful of other organisms, FPPS/GGPPS has been shown to be a bifunctional enzyme. By genetic tagging and microscopy, we observed a changing localization of FPPS/GGPPS in blood stage parasites. Given the great importance of alternative splicing and other transcriptional phenomena in gene regulation and the generation of protein diversity, we have investigated the processing of the FPPS/GGPPS transcript in P. falciparum by high-throughput sequencing methods in four time-points along the intraerythrocytic cycle of P. falciparum. We have identified levels of transcript diversity an order of magnitude higher than previously observed in this organism, as well as a few stage-specific splicing events. Our data suggest that alternative splicing in P. falciparum is an important feature for gene regulation and the generation of protein diversity.
Assuntos
Processamento Alternativo/genética , Geraniltranstransferase/genética , Malária Falciparum/genética , Transcrição Gênica , Animais , Regulação Enzimológica da Expressão Gênica , Variação Genética , Geraniltranstransferase/sangue , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Malária Falciparum/sangue , Malária Falciparum/parasitologia , Plasmodium falciparum/enzimologia , Plasmodium falciparum/genética , Plasmodium falciparum/patogenicidadeRESUMO
The increasing resistance of malaria parasites to almost all available drugs calls for the characterization of novel targets and the identification of new compounds. Carotenoids are polyisoprenoids from plants, algae, and some bacteria, and they are biosynthesized by Plasmodium falciparum but not by mammalian cells. Biochemical and reverse genetics approaches were applied to demonstrate that phytoene synthase (PSY) is a key enzyme for carotenoid biosynthesis in P. falciparum and is essential for intraerythrocytic growth. The known PSY inhibitor squalestatin reduces biosynthesis of phytoene and kills parasites during the intraerythrocytic cycle. PSY-overexpressing parasites showed increased biosynthesis of phytoene and its derived product phytofluene and presented a squalestatin-resistant phenotype, suggesting that this enzyme is the primary target of action of this drug in the parasite.
Assuntos
Antimaláricos/farmacologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Carotenoides/biossíntese , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/metabolismo , Antimaláricos/química , Compostos Bicíclicos Heterocíclicos com Pontes/químicaRESUMO
The 2-C-methyl-D-erythritol-4-phosphate and shikimate pathways were found to be active in Plasmodium falciparum and both can result in vitamin E biosynthesis in plants and algae. This study biochemically confirmed vitamin E biosynthesis in the malaria parasite, which can be inhibited by usnic acid. Furthermore, we found evidence pointing to a role of this vitamin in infected erythrocytes. These findings not only contribute to current understanding of P. falciparum biology but also reveal a pathway that could serve as a chemotherapeutic target.
Assuntos
Eritrócitos/parasitologia , Estágios do Ciclo de Vida , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium falciparum/metabolismo , Vitamina E/biossíntese , Animais , Benzofuranos/farmacologia , Cromatografia Gasosa-Espectrometria de Massas , Estágios do Ciclo de Vida/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/fisiologia , Esquizontes/efeitos dos fármacos , Esquizontes/metabolismo , Vitamina E/análise , alfa-Tocoferol/análise , alfa-Tocoferol/metabolismo , gama-Tocoferol/análise , gama-Tocoferol/metabolismoRESUMO
Herein, we show that intraerythrocytic stages of Plasmodium falciparum have an active pathway for biosynthesis of menaquinone. Kinetic assays confirmed that plasmodial menaquinone acts at least in the electron transport. Similarly to Escherichia coli, we observed increased levels of menaquinone in parasites kept under anaerobic conditions. Additionally, the mycobacterial inhibitor of menaquinone synthesis Ro 48-8071 also suppressed menaquinone biosynthesis and growth of parasites, although off-targets may play a role in this growth-inhibitory effect. Due to its absence in humans, the menaquinone biosynthesis can be considered an important drug target for malaria.
Assuntos
Eritrócitos/parasitologia , Estágios do Ciclo de Vida , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium falciparum/metabolismo , Vitamina K 2/análogos & derivados , Anaerobiose , Animais , Benzofenonas/farmacologia , Elétrons , Malária/tratamento farmacológico , Malária/metabolismo , Terapia de Alvo Molecular , Plasmodium falciparum/efeitos dos fármacos , Vitamina K 2/metabolismoRESUMO
Carotenoids are widespread lipophilic pigments synthesized by all photosynthetic organisms and some nonphotosynthetic fungi and bacteria. All carotenoids are derived from the C40 isoprenoid precursor geranylgeranyl pyrophosphate, and their chemical and physical properties are associated with light absorption, free radical scavenging, and antioxidant activity. Carotenoids are generally synthesized in well defined subcellular organelles, the plastids, which are also present in the phylum Apicomplexa, which comprises a number of important human parasites, such as Plasmodium and Toxoplasma. Recently, it was demonstrated that Toxoplasma gondii synthesizes abscisic acid. We therefore asked if Plasmodium falciparum is also capable of synthesizing carotenoids. Herein, biochemical findings demonstrated the presence of carotenoid biosynthesis in the intraerythrocytic stages of the apicomplexan parasite P. falciparum. Using metabolic labeling with radioisotopes, in vitro inhibition tests with norflurazon, a specific inhibitor of plant carotenoid biosynthesis, the results showed that intraerythrocytic stages of P. falciparum synthesize carotenoid compounds. A plasmodial enzyme that presented phytoene synthase activity was also identified and characterized. These findings not only contribute to the current understanding of P. falciparum evolution but shed light on a pathway that could serve as a chemotherapeutic target.
Assuntos
Carotenoides/metabolismo , Eritrócitos/parasitologia , Plasmodium falciparum/metabolismo , Animais , Clonagem Molecular , Herbicidas/farmacologia , Humanos , Cinética , Malária/terapia , Espectrometria de Massas/métodos , Modelos Químicos , Piridazinas/farmacologia , Terpenos/química , Toxoplasma/metabolismoRESUMO
In Plasmodium falciparum, the formation of isopentenyl diphosphate and dimethylallyl diphosphate, central intermediates in the biosynthesis of isoprenoids, occurs via the methylerythritol phosphate (MEP) pathway. Fosmidomycin is a specific inhibitor of the second enzyme of the MEP pathway, 1-deoxy-D-xylulose-5-phosphate reductoisomerase. We analyzed the effect of fosmidomycin on the levels of each intermediate and its metabolic requirement for the isoprenoid biosynthesis, such as dolichols and ubiquinones, throughout the intraerythrocytic cycle of P. falciparum. The steady-state RNA levels of the MEP pathway-associated genes were quantified by real-time polymerase chain reaction and correlated with the related metabolite levels. Our results indicate that MEP pathway metabolite peak precede maximum transcript abundance during the intraerythrocytic cycle. Fosmidomycin-treatment resulted in a decrease of the intermediate levels in the MEP pathway as well as in ubiquinone and dolichol biosynthesis. The MEP pathway associated transcripts were modestly altered by the drug, indicating that the parasite is not strongly responsive at the transcriptional level. This is the first study that compares the effect of fosmidomycin on the metabolic and transcript profiles in P. falciparum, which has only the MEP pathway for isoprenoid biosynthesis.
Assuntos
Eritritol/análogos & derivados , Eritrócitos/parasitologia , Fosfomicina/análogos & derivados , Plasmodium falciparum/efeitos dos fármacos , Fosfatos Açúcares/metabolismo , Animais , Eritritol/metabolismo , Fosfomicina/farmacologia , Genes de Protozoários , Plasmodium falciparum/genética , Plasmodium falciparum/metabolismo , Reação em Cadeia da PolimeraseRESUMO
In Plasmodium falciparum, the formation of isopentenyl diphosphate and dimethylallyl diphosphate, central intermediates in the biosynthesis of isoprenoids, occurs via the methylerythritol phosphate (MEP) pathway. Fosmidomycin is a specific inhibitor of the second enzyme of the MEP pathway, 1-deoxy-D-xylulose-5-phosphate reductoisomerase. We analyzed the effect of fosmidomycin on the levels of each intermediate and its metabolic requirement for the isoprenoid biosynthesis, such as dolichols and ubiquinones, throughout the intraerythrocytic cycle of P. falciparum. The steady-state RNA levels of the MEP pathway-associated genes were quantified by real-time polymerase chain reaction and correlated with the related metabolite levels. Our results indicate that MEP pathway metabolite peak precede maximum transcript abundance during the intraerythrocytic cycle. Fosmidomycin-treatment resulted in a decrease of the intermediate levels in the MEP pathway as well as in ubiquinone and dolichol biosynthesis. The MEP pathway associated transcripts were modestly altered by the drug, indicating that the parasite is not strongly responsive at the transcriptional level. This is the first study that compares the effect of fosmidomycin on the metabolic and transcript profiles in P. falciparum, which has only the MEP pathway for isoprenoid biosynthesis.
Assuntos
Animais , Eritritol/análogos & derivados , Eritritol/metabolismo , Eritrócitos/parasitologia , Fosfomicina/análogos & derivados , Fosfomicina/farmacologia , Plasmodium falciparum/metabolismo , Fosfatos Açúcares/metabolismo , Genes de Protozoários , Reação em Cadeia da Polimerase , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/genética , Plasmodium falciparum/crescimento & desenvolvimentoRESUMO
Isoprenoids play important roles in all living organisms as components of structural cholesterol, steroid hormones in mammals, carotenoids in plants, and ubiquinones. Significant differences occur in the length of the isoprenic side chains of ubiquinone between different organisms, suggesting that different enzymes are involved in the synthesis of these side chains. Whereas in Plasmodium falciparum the isoprenic side chains of ubiquinone contain 7-9 isoprenic units, 10-unit side chains are found in humans. In a search for the P. falciparum enzyme responsible for the biosynthesis of isoprenic side chains attached to the benzoquinone ring of ubiquinones, we cloned and expressed a putative polyprenyl synthase. Polyclonal antibodies raised against the corresponding recombinant protein confirmed the presence of the native protein in trophozoite and schizont stages of P. falciparum. The recombinant protein, as well as P. falciparum extracts, showed an octaprenyl pyrophosphate synthase activity, with the formation of a polyisoprenoid with eight isoprenic units, as detected by reverse-phase HPLC and reverse-phase TLC, and confirmed by electrospray ionization and tandem MS analysis. The recombinant and native versions of the enzyme had similar Michaelis constants with the substrates isopentenyl pyrophosphate and farnesyl pyrophosphate. The recombinant enzyme could be competitively inhibited in the presence of the terpene nerolidol. This is the first report that directly demonstrates an octaprenyl pyrophosphate synthase activity in parasitic protozoa. Given the rather low similarity of the P. falciparum enzyme to its human counterpart, decaprenyl pyrophosphate synthase, we suggest that the identified enzyme and its recombinant version could be exploited in the screening of novel drugs.
Assuntos
Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica , Plasmodium falciparum/enzimologia , Plasmodium falciparum/crescimento & desenvolvimento , Alquil e Aril Transferases/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Clonagem Molecular , Dados de Sequência Molecular , Plasmodium falciparum/efeitos dos fármacos , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Sesquiterpenos/farmacologia , Especificidade por SubstratoRESUMO
Two genes encoding the enzymes 1-deoxy-D-xylulose-5-phosphate synthase and 1-deoxy-D-xylulose-5-phosphate reductoisomerase have been recently identified, suggesting that isoprenoid biosynthesis in Plasmodium falciparum depends on the methylerythritol phosphate (MEP) pathway, and that fosmidomycin could inhibit the activity of 1-deoxy-D-xylulose-5-phosphate reductoisomerase. The metabolite 1-deoxy-D-xylulose-5-phosphate is not only an intermediate of the MEP pathway for the biosynthesis of isopentenyl diphosphate but is also involved in the biosynthesis of thiamin (vitamin B1) and pyridoxal (vitamin B6) in plants and many microorganisms. Herein we report the first isolation and characterization of most downstream intermediates of the MEP pathway in the three intraerythrocytic stages of P. falciparum. These include, 1-deoxy-D-xylulose-5-phosphate, 2-C-methyl-D-erythritol-4-phosphate, 4-(cytidine-5-diphospho)-2-C-methyl-D-erythritol, 4-(cytidine-5-diphospho)-2-C-methyl-D-erythritol-2-phosphate, and 2-C-methyl-D-erythritol-2,4-cyclodiphosphate. These intermediates were purified by HPLC and structurally characterized via biochemical and electrospray mass spectrometric analyses. We have also investigated the effect of fosmidomycin on the biosynthesis of each intermediate of this pathway and isoprenoid biosynthesis (dolichols and ubiquinones). For the first time, therefore, it is demonstrated that the MEP pathway is functionally active in all intraerythrocytic forms of P. falciparum, and de novo biosynthesis of pyridoxal in a protozoan is reported. Its absence in the human host makes both pathways very attractive as potential new targets for antimalarial drug development.
Assuntos
Eritritol/análogos & derivados , Eritritol/metabolismo , Fosfomicina/análogos & derivados , Plasmodium falciparum/metabolismo , Fosfato de Piridoxal/análogos & derivados , Fosfatos Açúcares/metabolismo , Animais , Antimaláricos/farmacologia , Dolicóis/biossíntese , Eritrócitos/parasitologia , Fosfomicina/farmacologia , Genes de Protozoários , Humanos , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Estrutura Molecular , Pentosefosfatos/biossíntese , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/genética , Plasmodium falciparum/crescimento & desenvolvimento , Fosfato de Piridoxal/biossíntese , Espectrometria de Massas por Ionização por Electrospray , Ubiquinona/biossínteseRESUMO
Development of new drugs is one of the strategies for malaria control. The biosynthesis of several isoprenoids in Plasmodium falciparum was recently described. Interestingly, some intermediates and final products biosynthesized by this pathway in mammals differ from those biosynthesized in P. falciparum. These facts prompted us to evaluate various terpenes, molecules with a similar chemical structure to the intermediates of the isoprenoids pathway, as potential antimalarial drugs. Different terpenes and S-farnesylthiosalicylic acid were tested on cultures of the intraerythrocytic stages of P. falciparum, and the 50% inhibitory concentrations for each one were found: farnesol, 64 microM; nerolidol, 760 nM; limonene, 1.22 mM; linalool, 0.28 mM; and S-farnesylthiosalicylic acid, 14 microM. All the terpenes tested inhibited dolichol biosynthesis in the trophozoite and schizont stages when [1-(n)-(3)H]farnesyl pyrophosphate triammonium salt ([(3)H]FPP) was used as precursor. Farnesol, nerolidol, and linalool showed stronger inhibitory activity on the biosynthesis of the isoprenic side chain of the benzoquinone ring of ubiquinones in the schizont stage. Treatment of schizont stages with S-farnesylthiosalicylic acid led to a decrease in intensity of the band corresponding a p21(ras) protein. The inhibitory effect of terpenes and S-farnesylthiosalicylic acid on the biosynthesis of both dolichol and the isoprenic side chain of ubiquinones and the isoprenylation of proteins in the intraerythrocytic stages of P. falciparum appears to be specific, because overall protein biosynthesis was not affected. Combinations of some terpenes or S-farnesylthiosalicylic acid tested in this work with other antimalarial drugs, like fosmidomycin, could be a new strategy for the treatment of malaria.
Assuntos
Farneseno Álcool/análogos & derivados , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium falciparum/metabolismo , Terpenos/metabolismo , Terpenos/farmacologia , Animais , Cromatografia Líquida de Alta Pressão , Cromatografia em Camada Fina , Depressão Química , Dolicóis/biossíntese , Eletroforese em Gel de Poliacrilamida , Eritrócitos/parasitologia , Farneseno Álcool/farmacologia , Metabolismo dos Lipídeos , Malária Falciparum/parasitologia , Plasmodium falciparum/efeitos dos fármacos , Testes de Precipitina , Salicilatos/farmacologia , Terpenos/isolamento & purificação , Ubiquinona/biossínteseRESUMO
Little is known about isoprenoid biosynthesis in parasitic protozoa. The presence of dolichol and isoprenylated proteins has been detected in Plasmodium falciparum, but no studies are available about the biosynthesis of the isoprenic side chain attached to the benzoquinone ring of coenzyme Q. In the present study, using metabolic labelling with different intermediates, we demonstrated the presence of an active isoprenoid pathway for the biosynthesis of the isoprenic chain of coenzyme Q. Our results show that P. falciparum is able to synthesize different homologs (coenzyme Q(8) and coenzyme Q(9)), depending on the given intermediate. Parasites treated with nerolidol at doses 2.2 times below the IC(50) showed a decreased ability to synthesize the isoprenic chain attached to coenzyme Q at all intraerythrocytic stages. Treatment with nerolidol arrested development of the intraerythrocytic stages of the parasites, indicating that the drug may have an antimalarial potential.
Assuntos
Plasmodium falciparum/enzimologia , Fosfatos de Poli-Isoprenil/química , Fosfatos de Poli-Isoprenil/metabolismo , Prenilação de Proteína , Ubiquinona/biossíntese , Animais , Radioisótopos de Carbono/metabolismo , Eritrócitos/efeitos dos fármacos , Eritrócitos/parasitologia , Humanos , Malária Falciparum/parasitologia , Testes de Sensibilidade Parasitária , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/crescimento & desenvolvimento , Sesquiterpenos/farmacologia , Trítio/metabolismo , Ubiquinona/químicaRESUMO
A critical role has been proposed for the switch from non-cytophilic IgG2 to cytophilic antibodies of IgG1 and IgG3 subclasses observed in the humoral immune responses to Plasmodium falciparum of some Africans. These Africans have acquired clinically immunity naturally, after several years of exposure to holo-endemic malaria. In the present study, the possibility that life-long exposure to low levels of malarial endemicity may be associated with changes in the IgG-subclass composition of antibodies to P. falciparum was investigated in a native Amazonian community. The subjects were 138 malaria-exposed but non-infected Karitiana Indians. In a separate investigation, the concentrations of IgG-subclass antibodies in acutely ill patients with severe malaria (N = 22) were compared with those in age- and sex-matched controls who had uncomplicated malaria (N = 44). Plasma concentrations of IgG against a detergent-soluble extract of P. falciparum schizonts were measured by quantitative ELISA, using indirect standardization. Among the Karitiana, the concentrations of anti-parasite antibodies of all subclasses increased with age, and there was no correlation between age and the proportion of such antibodies which was cytophilic. The predominance of cytophilic IgG1 and non-cytophilic IgG2 antibodies in all age-groups of the Karitiana provides an example of an intermediate pattern of immune responses to P. falciparum which contrasts with those previously described in both clinically immune and non-immune populations. Although mean concentrations of cytophilic IgG1 against P. falciparum were significantly higher in the controls than in the patients with severe malaria, there were no significant differences in other IgG subclasses. Lack of exposure to malaria in the past was associated with disease severity (odds ratio = 4.75; 95% confidence interval = 1.31-17.42), and may explain, at least partially, the occurrence of defective, low-IgG1 antibody responses to P. falciparum in those subjects who had severe malaria.