Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0024530 (malaria)
 44,886 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cambodia is located in an area of resistance to multiple antimalarials and has been the first country to implement the systematic use of an artesunate-mefloquine combination as first-line treatment for Plasmodium falciparum malaria. Little is known, however, about the prevalence of resistance mutations within the natural parasite populations, impeding rational drug policy in this context. Using direct sequencing of PCR products, we have analyzed sequence polymorphism of the dihydrofolate reductase-thymidylate synthase, dihydropteroate synthetase, and multidrug resistance 1 genes in a large number of clinical P. falciparum isolates collected in various areas of Cambodia. This highlighted a 100% prevalence of haplotypes with multiple mutations in the target genes of antifolates after more than a decade without use of antifolates for malaria therapy. A high prevalence of mutations in Pfmdr1, including mutations associated with decreased in vitro susceptibility to mefloquine and quinine, was also observed. In addition, novel, low-frequency mutations were detected in Pfmdr1. Our findings show an alarming rate of multilocus resistance genotypes in Cambodia, requiring diligent surveillance and imposing limitations on possible future drug combinations.
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PMID:Countrywide survey shows very high prevalence of Plasmodium falciparum multilocus resistance genotypes in Cambodia. 1604 16

Inhibitors of dihydrofolate reductase (DHFR) have been mainstays in the treatment of falciparum malaria. Resistance to one of these antifolates, pyrimethamine, is now common in Plasmodium falciparum populations. Antifolates have not traditionally been recommended for treatment of vivax malaria. However, recent studies have suggested that a third-generation antifolate, WR99210, is remarkably effective even against highly pyrimethamine-resistant parasites from both species. Two methods were used to identify a compound that is effective against quadruple mutant alleles from P. falciparum (N51I/C59R/S108N/I164L) and from Plasmodium vivax (57L/111L/117T/173F). The first was simple yeast system used to screen a panel of WR99210 analogs. The biguanide prodrug, JPC-2056, of the 2-chloro-4-trifluoromethoxy analog of WR99210 was effective against both the P. falciparum and P. vivax enzymes, and has been selected for further development. The second method compared the analogs in silico by docking them in the known structure of the P. falciparum DHFR-thymidylate synthase. The program reproduced well the position of the triazine ring, but the calculated energies of ligand binding were very similar for different compounds and therefore did not reproduce the observed trends in biological activity. The WR99210 family of molecules is flexible due to a long bridge between the triazine ring and the substituted benzene. During docking, multiple conformations were observed for the benzene ring part of the molecules in the DHFR active site, making computer-based predictions of binding energy less informative than for more rigid ligands. This flexibility is a key factor in their effectiveness against the highly mutant forms of DHFR.
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PMID:Identification of the optimal third generation antifolate against P. falciparum and P. vivax. 1618 88

The rodent malaria Plasmodium yoelii is a useful model to study protective immunity to pre-erythrocytic stages of infection, pathogenesis of erythrocytic stages, and vaccine development. However, the utility of the P. yoelii model system has not been fully realized because transfection and genetic manipulation methodologies for this rodent species are less developed than that of another rodent species Plasmodium berghei. Here we report improved transfection efficiency using the AMAXA nucleofector system compared to conventional transfection methodologies. We also show that heterologous promoters from P. berghei can be used to drive expression of a green fluorescent protein (GFP) reporter protein in P. yoelii. In an effort to develop additional selectable markers for this parasite, we also tested positive selectable markers that have been used successfully in P. falciparum and P. berghei. Human dihydrofolate reductase (hdhfr) and Toxoplasma gondii dihydrofolate reductase-thymidylate synthase (Tgdhfr-ts) conferred drug resistance to WR99210 and pyrimethamine, respectively, when introduced as episomes. These improvements should make genetic manipulation of P. yoelii more amenable and facilitate further studies of host-parasite interactions using this attractive rodent model.
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PMID:Improved transfection and new selectable markers for the rodent malaria parasite Plasmodium yoelii. 1645 71

Pyrimethamine analogs were examined as potential agents against vivax malaria using a bacterial surrogate system carrying Plasmodium vivax dihydrofolate reductase-thymidylate synthase (PvDHFR-TS), in which the PvDHFR complemented chemically knocked out host dihydrofolate reductase. The system was initially tested with P. falciparum dihydrofolate reductase-thymidylate synthase and was found to have good correlation with the parasite-based system. The 50% inhibitory concentrations derived from PvDHFR-TS-dependent bacteria were correlated with their corresponding inhibition constants (Ki) from an enzyme inhibition assay, pointing to the likelihood that the potent enzyme inhibitors will also have potent antimalarial activities. Active compounds against both wild-type and S58R S117N (SP21) double-mutant P. vivax include analogs with structures which can avert a steric clash with the asparagine (S117N) side chain of the mutant, similar to those found for homologous Plasmodium falciparum mutants, raising the possibility that the same compounds can be developed against both types of antifolate-resistant malaria. This rapid and convenient drug screening system should be useful for development of new antifolates against P. vivax, for which a continuous culture system is not yet available.
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PMID:Evaluation of the activities of pyrimethamine analogs against Plasmodium vivax and Plasmodium falciparum dihydrofolate reductase-thymidylate synthase using in vitro enzyme inhibition and bacterial complementation assays. 1695 16

The introduction of avian malaria (Plasmodium relictum) to Hawaii has provided a model system for studying the influence of exotic disease on naive host populations. Little is known, however, about the origin or the genetic variation of Hawaii's malaria and traditional classification methods have confounded attempts to place the parasite within a global ecological and evolutionary context. Using fragments of the parasite mitochondrial gene cytochrome b and the nuclear gene dihydrofolate reductase-thymidylate synthase obtained from a global survey of greater than 13000 avian samples, we show that Hawaii's avian malaria, which can cause high mortality and is a major limiting factor for many species of native passerines, represents just one of the numerous lineages composing the morphological parasite species. The single parasite lineage detected in Hawaii exhibits a broad host distribution worldwide and is dominant on several other remote oceanic islands, including Bermuda and Moorea, French Polynesia. The rarity of this lineage in the continental New World and the restriction of closely related lineages to the Old World suggest limitations to the transmission of reproductively isolated parasite groups within the morphological species.
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PMID:Global phylogeographic limits of Hawaii's avian malaria. 1701 60

The dihydrofolate reductase (DHFR, EC 1.5.1.3) domain of Plasmodium falciparum bifunctional dihydrofolate reductase-thymidylate synthase (DHFR-TS) is an attractive target of two important antifolate antimalarials: pyrimethamine (Pyr) and cycloguanil (Cyc). Over recent years, knowledge of malarial DHFR and mechanism(s) of antifolate resistance have increased substantially. These observations have provided an important framework for better understanding the molecular basis of antifolate resistance in malaria. This article provides a brief review and update on molecular aspects relevant to antifolate resistance in malaria.
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PMID:Dihydrofolate reductase and antifolate resistance in malaria. 1709 21

Plasmodium falciparum, the protozoan that causes the most lethal form of human malaria, has been controlled principally by two safe, affordable drugs, chloroquine and sulfadoxine-pyrimethamine (SP). Studies in the laboratory and in the field have demonstrated that resistance to SP depends on non-synonymous point mutations in the dihydrofolate reductase (DHFR), and dihydropteroate synthase (DHPS) coding regions. Parasites that carry dhfr genes with 3 or 4 point mutations (51I/59R/108N triple mutation or 51I/59R/108N/164L quadruple mutation) are resistant to pyrimethamine in vitro and patients infected with these parasites respond poorly to SP treatment. The wide spread of these pyrimethamine-resistant alleles demonstrates the increased fitness over drug-sensitive alleles in the presence of the drug. However, it is not clear whether these alleles might reduce the fitness of parasites in the absence of drug pressure. As a first step, we compared the kinetic properties of the wild type, and three mutant alleles to determine whether the native DHFR-thymidylate synthase form of the mutant proteins showed compromised activity in vitro. The mutant enzymes had K(m) values for their substrate, dihydrofolate that were significantly lower than the wild type, k(cat) values in the same range as the wild type enzyme, and k(cat)/K(m) values higher than wild type. In contrast, the K(m) values for the NADPH cofactor were higher than wild type for the mutant enzymes. These observations suggest that the fitness of these parasites may not be compromised relative to those that carry the wild type allele, even without sustained SP drug pressure.
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PMID:Pyrimethamine-resistant dihydrofolate reductase enzymes of Plasmodium falciparum are not enzymatically compromised in vitro. 1745 22

With the emergence of drug-resistant vivax malaria, in vitro studies are urgently needed to examine resistance mechanisms and for drug development. Currently, Plasmodium vivax culturing is inadequate for addressing these needs; therefore, surrogate biological systems have been developed. Although these systems are informative, they do not address Plasmodium species-specific mechanisms, such as drug delivery through erythrocytes and parasite membranes. Here, we demonstrate that P. falciparum is an excellent biological system for expression of P. vivax dhfr-ts alleles to assess dihydrofolate reductase (DHFR)-thymidylate synthase interactions with antifolates. Our results show that the P. vivax dhfr-ts quadruple-mutant allele AMRU1, expressed in P. falciparum, provides significant protection against pyrimethamine, cycloguanil, and clocicguanil. Moreover, the PvDHFR quadruple mutant confers greater resistance to cycloguanil, clociguanil, and WR99210 than the PfDHFR quadruple mutant. Modeling of both P. vivax and P. falciparum DHFR quadruple mutants suggests that mutations unique to P. vivax DHFR are responsible for differences seen in parasite susceptibility to antifolates.
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PMID:A novel Plasmodium falciparum expression system for assessing antifolate resistance caused by mutant P. vivax dihydrofolate reductase-thymidylate synthase. 1759 62

Folate metabolism of the malaria parasites provides two targets for current antimalarials: dihydrofolate reductase and dihydropteroate synthase. Dihydrofolate reductase inhibitors have been used as antimalarials over the past few decades, often in combination with dihydropteroate synthase inhibitors. Resistance to these antifolate drugs developed through mutations in both target enzymes. However, limited mutation possibilities gave opportunities for the development of new drugs. Furthermore, other enzymes in the folate and related pathways are potential new targets that remain to be exploited. These include thymidylate synthase, an enzyme fused with dihydrofolate reductase in the same protein chain, serine hydroxymethyltransferase, methylene tetrahydrofolate dehydrogenase, methionine synthase and enzymes in the glycine cleavage pathway.
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PMID:Folate metabolism as a source of molecular targets for antimalarials. 1766 90

Plasmodium malariae, the parasite responsible for quartan malaria, is transmitted in most areas of malaria endemicity and is associated with significant morbidity. The sequence of the gene coding for the enzyme dihydrofolate reductase-thymidylate synthase (DHFR-TS) was obtained from field isolates of P. malariae and from the closely related simian parasite Plasmodium brasilianum. The two sequences were nearly 100% homologous, adding weight to the notion that they represent genetically distinct lines of the same species. A survey of polymorphisms of the dhfr sequences in 35 isolates of P. malariae collected from five countries in Asia and Africa revealed a low number of nonsynonymous mutations in five codons. In five of the isolates collected from southeast Asia, a nonsynonymous mutation was found at one of the three positions known to be associated with antifolate resistance in other Plasmodium species. Five isolates with the wild-type DHFR could be assayed for drug susceptibility in vitro and were found to be sensitive to pyrimethamine (mean 50% inhibitory concentration, 2.24 ng/ml [95% confidence interval, 0.4 to 3.1]).
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PMID:Genetic analysis of the dihydrofolate reductase-thymidylate synthase gene from geographically diverse isolates of Plasmodium malariae. 1768 97


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