Inhibition of pyrimidine biosynthesis de novo in Plasmodium falciparum by 2-(4-t-butylcyclohexyl)-3-hydroxy-1,4-naphthoquinone in vitro

1985 ◽  
Vol 14 (1) ◽  
pp. 97-109 ◽  
Author(s):  
D HAMMOND ◽  
J BURCHELL ◽  
M PUDNEY
Keyword(s):  
Plasmodium Falciparum ◽  
De Novo ◽  
Pyrimidine Biosynthesis

scholarly journals Localization of ferrochelatase in Plasmodium falciparum

2004 ◽  
Vol 384 (2) ◽  
pp. 429-436 ◽  
Author(s):  
Sundaramurthy VARADHARAJAN ◽  
B. K. Chandrashekar SAGAR ◽  
Pundi N. RANGARAJAN ◽  
Govindarajan PADMANABAN
Keyword(s):  
Plasmodium Falciparum ◽  
De Novo ◽  
Enzymic Activity ◽  
Malarial Parasite ◽  
Parasite Genome ◽  
Marker Proteins ◽  
Parasite Plasmodium ◽  
Theoretical Predictions ◽  
Parasite Plasmodium Falciparum

Our previous studies have demonstrated de novo haem biosynthesis in the malarial parasite (Plasmodium falciparum and P. berghei). It has also been shown that the first enzyme of the pathway is the parasite genome-coded ALA (δ-aminolaevulinate) synthase localized in the parasite mitochondrion, whereas the second enzyme, ALAD (ALA dehydratase), is accounted for by two species: one species imported from the host red blood cell into the parasite cytosol and another parasite genome-coded species in the apicoplast. In the present study, specific antibodies have been raised to PfFC (parasite genome-coded ferrochelatase), the terminal enzyme of the haem-biosynthetic pathway, using recombinant truncated protein. With the use of these antibodies as well as those against the hFC (host red cell ferrochelatase) and other marker proteins, immunofluorescence studies were performed. The results reveal that P. falciparum in culture manifests a broad distribution of hFC and a localized distribution of PfFC in the parasite. However, PfFC is not localized to the parasite mitochondrion. Immunoelectron-microscopy studies reveal that PfFC is indeed localized to the apicoplast, whereas hFC is distributed in the parasite cytoplasm. These results on the localization of PfFC are unexpected and are at variance with theoretical predictions based on leader sequence analysis. Biochemical studies using the parasite cytosolic and organellar fractions reveal that the cytosol containing hFC accounts for 80% of FC enzymic activity, whereas the organellar fraction containing PfFC accounts for the remaining 20%. Interestingly, both the isolated cytosolic and organellar fractions are capable of independent haem synthesis in vitro from [4-14C]ALA, with the cytosol being three times more efficient compared with the organellar fraction. With [2-14C]glycine, most of the haem is synthesized in the organellar fraction. Thus haem is synthesized in two independent compartments: in the cytosol, using the imported host enzymes, and in the organellar fractions, using the parasite genome-coded enzymes.

scholarly journals Emergence and clonal expansion of in vitro artemisinin-resistant Plasmodium falciparum kelch13 R561H mutant parasites in Rwanda

2020 ◽  
Vol 26 (10) ◽  
pp. 1602-1608 ◽  
Author(s):  
Aline Uwimana ◽  
Eric Legrand ◽  
Barbara H. Stokes ◽  
Jean-Louis Mangala Ndikumana ◽  
Marian Warsame ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Plasmodium Falciparum ◽  
Gene Editing ◽  
De Novo ◽  
Artemisinin Resistance ◽  
Clonal Expansion ◽  
Efficacy Trials ◽  
Cure Rates ◽  
Antimalarial Chemotherapy

Abstract Artemisinin resistance (delayed P. falciparum clearance following artemisinin-based combination therapy), is widespread across Southeast Asia but to date has not been reported in Africa1–4. Here we genotyped the P. falciparum K13 (Pfkelch13) propeller domain, mutations in which can mediate artemisinin resistance5,6, in pretreatment samples collected from recent dihydroarteminisin-piperaquine and artemether-lumefantrine efficacy trials in Rwanda7. While cure rates were >95% in both treatment arms, the Pfkelch13 R561H mutation was identified in 19 of 257 (7.4%) patients at Masaka. Phylogenetic analysis revealed the expansion of an indigenous R561H lineage. Gene editing confirmed that this mutation can drive artemisinin resistance in vitro. This study provides evidence for the de novo emergence of Pfkelch13-mediated artemisinin resistance in Rwanda, potentially compromising the continued success of antimalarial chemotherapy in Africa.

scholarly journals MCT4 regulates de novo pyrimidine biosynthesis in GBM in a lactate-independent manner

2020 ◽  
Vol 2 (1) ◽  
Author(s):  
Raffaella Spina ◽  
Dillon M Voss ◽  
Xiaohua Yang ◽  
Jason W Sohn ◽  
Robert Vinkler ◽  
...  
Keyword(s):  
De Novo ◽  
Apoptotic Cell ◽  
Pyrimidine Biosynthesis ◽  
Monocarboxylate Transporter ◽  
Independent Manner ◽  
Pyrimidine Nucleosides ◽  
Significant Prolongation ◽  
Orthotopic Xenografts

Abstract Background Necrotic foci with surrounding hypoxic cellular pseudopalisades and microvascular hyperplasia are histological features found in glioblastoma (GBM). We have previously shown that monocarboxylate transporter 4 (MCT4) is highly expressed in necrotic/hypoxic regions in GBM and that increased levels of MCT4 are associated with worse clinical outcomes. Methods A combined transcriptomics and metabolomics analysis was performed to study the effects of MCT4 depletion in hypoxic GBM neurospheres. Stable and inducible MCT4-depletion systems were used to evaluate the effects of and underlining mechanisms associated with MCT4 depletion in vitro and in vivo, alone and in combination with radiation. Results This study establishes that conditional depletion of MCT4 profoundly impairs self-renewal and reduces the frequency and tumorigenicity of aggressive, therapy-resistant, glioblastoma stem cells. Mechanistically, we observed that MCT4 depletion induces anaplerotic glutaminolysis and abrogates de novo pyrimidine biosynthesis. The latter results in a dramatic increase in DNA damage and apoptotic cell death, phenotypes that were readily rescued by pyrimidine nucleosides supplementation. Consequently, we found that MCT4 depletion promoted a significant prolongation of survival of animals bearing established orthotopic xenografts, an effect that was extended by adjuvant treatment with focused radiation. Conclusions Our findings establish a novel role for MCT4 as a critical regulator of cellular deoxyribonucleotide levels and provide a new therapeutic direction related to MCT4 depletion in GBM.

scholarly journals Biosynthesis of GDP-fucose and Other Sugar Nucleotides in the Blood Stages of Plasmodium falciparum

2013 ◽  
Vol 288 (23) ◽  
pp. 16506-16517 ◽  
Author(s):  
Sílvia Sanz ◽  
Giulia Bandini ◽  
Diego Ospina ◽  
Maria Bernabeu ◽  
Karina Mariño ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
De Novo ◽  
Cell Communication ◽  
Developmental Cycle ◽  
Metabolic Labeling ◽  
Sugar Nucleotides ◽  
Liquid Chromatography Tandem Mass ◽  
Intraerythrocytic Developmental Cycle

Carbohydrate structures play important roles in many biological processes, including cell adhesion, cell-cell communication, and host-pathogen interactions. Sugar nucleotides are activated forms of sugars used by the cell as donors for most glycosylation reactions. Using a liquid chromatography-tandem mass spectrometry-based method, we identified and quantified the pools of UDP-glucose, UDP-galactose, UDP-N-acetylglucosamine, GDP-mannose, and GDP-fucose in Plasmodium falciparum intraerythrocytic life stages. We assembled these data with the in silico functional reconstruction of the parasite metabolic pathways obtained from the P. falciparum annotated genome, exposing new active biosynthetic routes crucial for further glycosylation reactions. Fucose is a sugar present in glycoconjugates often associated with recognition and adhesion events. Thus, the GDP-fucose precursor is essential in a wide variety of organisms. P. falciparum presents homologues of GDP-mannose 4,6-dehydratase and GDP-l-fucose synthase enzymes that are active in vitro, indicating that most GDP-fucose is formed by a de novo pathway that involves the bioconversion of GDP-mannose. Homologues for enzymes involved in a fucose salvage pathway are apparently absent in the P. falciparum genome. This is in agreement with in vivo metabolic labeling experiments showing that fucose is not significantly incorporated by the parasite. Fluorescence microscopy of epitope-tagged versions of P. falciparum GDP-mannose 4,6-dehydratase and GDP-l-fucose synthase expressed in transgenic 3D7 parasites shows that these enzymes localize in the cytoplasm of P. falciparum during the intraerythrocytic developmental cycle. Although the function of fucose in the parasite is not known, the presence of GDP-fucose suggests that the metabolite may be used for further fucosylation reactions.

Cytotoxic Effects of Inhibitors of de Novo Pyrimidine Biosynthesis upon Plasmodium falciparum

Biochemistry ◽  
1994 ◽  
Vol 33 (17) ◽  
pp. 5268-5274 ◽  
Author(s):  
Kristen K. Seymour ◽  
Stephen D. Lyons ◽  
Leonidas Phillips ◽  
Karl H. Rieckmann ◽  
Richard I. Christopherson
Keyword(s):  
Plasmodium Falciparum ◽  
De Novo ◽  
Pyrimidine Biosynthesis ◽  
Cytotoxic Effects

scholarly journals Potent and Selective Activity of a Combination of Thymidine and 1843U89, a Folate-Based Thymidylate Synthase Inhibitor, against Plasmodium falciparum

2000 ◽  
Vol 44 (4) ◽  
pp. 1047-1050 ◽  
Author(s):  
Lei Jiang ◽  
Pei-Chieh Lee ◽  
John White ◽  
Pradipsinh K. Rathod
Keyword(s):  
Plasmodium Falciparum ◽  
Thymidylate Synthase ◽  
Mammalian Cells ◽  
De Novo ◽  
Antimalarial Activity ◽  
Malarial Parasite ◽  
Drug Resistant ◽  
Additional Tool ◽  
Synthase Inhibitor

ABSTRACT Unlike mammalian cells, malarial parasites are completely dependent on the de novo pyrimidine pathway and lack the enzymes to salvage preformed pyrimidines. In the present study, first, it is shown that 1843U89, even without polyglutamylation, is a potent folate-based inhibitor of purified malarial parasite thymidylate synthase. The binding was noncompetitive with respect to methylenetetrahydrofolate, and 1843U89 had a Ki of 1 nM. The compound also had potent antimalarial activity in vitro. Plasmodium falciparum cells in culture were inhibited by 1843U89, with a 50% inhibitory concentration of about 70 nM. The compound was effective against drug-sensitive as well as drug-resistant clones ofP. falciparum. As predicted by the biochemistry of the parasite, the potent inhibition of parasite proliferation by 1843U89 could not be reversed with 10 μM thymidine. In contrast, in the presence of 10 μM thymidine, mammalian cells were unaffected by 1843U89 even at concentrations as high as 0.1 mM, thus offering a selectivity window of more than 1,000-fold. On this basis, folate-based thymidylate synthase inhibitors may represent a powerful additional tool that can be used to combat drug-resistant malaria.

scholarly journals A De novo Peptide from a High Throughput Peptide Library Blocks Myosin A -MTIP Complex Formation in Plasmodium falciparum

2020 ◽  
Vol 21 (17) ◽  
pp. 6158
Author(s):  
Zill e Anam ◽  
Nishant Joshi ◽  
Sakshi Gupta ◽  
Preeti Yadav ◽  
Ayushi Chaurasiya ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
De Novo ◽  
Peptide Library ◽  
Tail Domain ◽  
Interacting Protein ◽  
Membrane Complex ◽  
Inner Membrane Complex ◽  
De Novo Peptide

Apicomplexan parasites, through their motor machinery, produce the required propulsive force critical for host cell-entry. The conserved components of this so-called glideosome machinery are myosin A and myosin A Tail Interacting Protein (MTIP). MTIP tethers myosin A to the inner membrane complex of the parasite through 20 amino acid-long C-terminal end of myosin A that makes direct contacts with MTIP, allowing the invasion of Plasmodium falciparum in erythrocytes. Here, we discovered through screening a peptide library, a de-novo peptide ZA1 that binds the myosin A tail domain. We demonstrated that ZA1 bound strongly to myosin A tail and was able to disrupt the native myosin A tail MTIP complex both in vitro and in vivo. We then showed that a shortened peptide derived from ZA1, named ZA1S, was able to bind myosin A and block parasite invasion. Overall, our study identified a novel anti-malarial peptide that could be used in combination with other antimalarials for blocking the invasion of Plasmodium falciparum.

scholarly journals Identification and Deconvolution of Cross-Resistance Signals from Antimalarial Compounds Using Multidrug-Resistant Plasmodium falciparum Strains

2014 ◽  
Vol 59 (2) ◽  
pp. 1110-1118 ◽  
Author(s):  
Monika Chugh ◽  
Christian Scheurer ◽  
Sibylle Sax ◽  
Elizabeth Bilsland ◽  
Donelly A. van Schalkwyk ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Mode Of Action ◽  
De Novo ◽  
Real Life ◽  
Multidrug Resistant ◽  
Chloroquine Resistance ◽  
Cross Resistance ◽  
Content Type ◽  
Antimalarial Compounds

ABSTRACTPlasmodium falciparum, the most deadly agent of malaria, displays a wide variety of resistance mechanisms in the field. The ability of antimalarial compounds in development to overcome these must therefore be carefully evaluated to ensure uncompromised activity against real-life parasites. We report here on the selection and phenotypic as well as genotypic characterization of a panel of sensitive and multidrug-resistantP. falciparumstrains that can be used to optimally identify and deconvolute the cross-resistance signals from an extended panel of investigational antimalarials. As a case study, the effectiveness of the selected panel of strains was demonstrated using the 1,2,4-oxadiazole series, a newly identified antimalarial series of compounds within vitroactivity againstP. falciparumat nanomolar concentrations. This series of compounds was to be found inactive against several multidrug-resistant strains, and the deconvolution of this signal implicatedpfcrt, the genetic determinant of chloroquine resistance. Targeted mode-of-action studies further suggested that this new chemical series might act as falcipain 2 inhibitors, substantiating the suggestion that these compounds have a site of action similar to that of chloroquine but a distinct mode of action. New antimalarials must overcome existing resistance and, ideally, prevent itsde novoappearance. The panel of strains reported here, which includes recently collected as well as standard laboratory-adapted field isolates, is able to efficiently detect and precisely characterize cross-resistance and, as such, can contribute to the faster development of new, effective antimalarial drugs.

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