fosfomycin and Malaria

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.

Topics: Aldose-Ketose Isomerases; Anti-Bacterial Agents; Directed Molecular Evolution; Drug Resistance; Escherichia coli; Fosfomycin; Gene Library; Malaria; Mutagenesis; Mutation; Plasmodium falciparum; Saccharomyces cerevisiae

With first indications of resistance against artemisinin compounds, the development of novel alternative antimalarials remains an urgent need. One candidate is fosmidomycin (Fos), a phosphonic acid derivative. This PRISMA guideline-adhering and PROSPERO-registered systematic review and meta-analysis provides an overview of the state-of-the-art of the clinical development of Fos as an antimalarial. Pooling six clinical trials of Fos against uncomplicated malaria in African children yielded an overall day 28 cure rate of 85% (95% CI: 71-98%); a parasite clearance time of 39 h; and a fever clearance time of 30 h. In four adult cohorts, the corresponding values were 70% (95% CI: 40-100%), 49 and 42 h, respectively. Data suggest that besides the partner drug, formulation determines efficacy. We advocate further clinical development of Fos-combinations. PROSPERO registration number: CRD42014013688.

Topics: Adult; Antimalarials; Child; Child, Preschool; Clinical Trials as Topic; Drug Therapy, Combination; Fosfomycin; Humans; Malaria; Malaria, Falciparum; Plasmodium falciparum

Approximately 40% of the world population live in areas with the risk of malaria. Each year, 300-500 million people suffer from acute malaria, and 0.5-2.5 million die from the disease. Although malaria has been widely eradicated in many parts of the world, the global number of cases continues to rise. The most important reason for this alarming situation is the rapid spread of malaria parasites that are resistant to antimalarial drugs, especially chloroquine, which is by far the most frequently used. The development of new antimalarial drugs has been neglected since the 1970s owing to the end colonialism, changes in the areas of military engagement, and the restricted market potential. Only in recent years, in part supported by public funding programs, has interest in the development of antimalarial drugs been renewed. New data available from the recently sequenced genome of the malaria parasite Plasmodium falciparum and the application of methods of modern drug design promise to bring significant development in the fight against this disease.

Topics: Aminoquinolines; Animals; Antimalarials; Artemisinins; Artesunate; Atovaquone; Clinical Trials as Topic; Dapsone; Drug Combinations; Drug Design; Drug Resistance; Ethanolamines; Flavonoids; Fluorenes; Fosfomycin; Global Health; Humans; Lumefantrine; Malaria; Naphthoquinones; Naphthyridines; Plasmodium falciparum; Proguanil; Sesquiterpenes

Topics: Amikacin; Anemia, Sickle Cell; Animals; Anti-Bacterial Agents; Azepines; Burkitt Lymphoma; Fosfomycin; Hepatitis, Viral, Human; Herpesvirus 4, Human; Humans; Lassa Fever; Liver; Malaria; Marburg Virus Disease; Oxamniquine; Parasitic Diseases; Penicillanic Acid; Pneumonia, Pneumocystis; Smallpox; Ticarcillin; Trichinellosis; Vaccination; Virus Diseases

Malaria parasites rely on a plastid organelle for survival during the blood stages of infection. However, the entire organelle is dispensable as long as the isoprenoid precursor, isopentenyl pyrophosphate (IPP), is supplemented in the culture medium. We engineered parasites to produce isoprenoid precursors from a mevalonate-dependent pathway, creating a parasite line that replicates normally after the loss of the apicoplast organelle. We show that carbon-labeled mevalonate is specifically incorporated into isoprenoid products, opening new avenues for researching this essential class of metabolites in malaria parasites. We also show that essential apicoplast proteins, such as the enzyme target of the drug fosmidomycin, can be deleted in this mevalonate bypass parasite line, providing a new method to determine the roles of other important apicoplast-resident proteins. Several antibacterial drugs kill malaria parasites by targeting basic processes, such as transcription, in the organelle. We used metabolomic and transcriptomic methods to characterize parasite metabolism after azithromycin treatment triggered loss of the apicoplast and found that parasite metabolism and the production of apicoplast proteins is largely unaltered. These results provide insight into the effects of apicoplast-disrupting drugs, several of which have been used to treat malaria infections in humans. Overall, the mevalonate bypass system provides a way to probe essential aspects of apicoplast biology and study the effects of drugs that target apicoplast processes.

Topics: Animals; Anti-Bacterial Agents; Apicoplasts; Azithromycin; Fosfomycin; Hemiterpenes; Humans; Malaria; Mevalonic Acid; Organophosphorus Compounds; Parasites; Plasmodium falciparum; Plastids; Protozoan Proteins

Fosmidomycin inhibits IspC (Dxr, 1-deoxy-d-xylulose 5-phosphate reductoisomerase), a key enzyme in nonmevalonate isoprenoid biosynthesis that is essential in Plasmodium falciparum. The drug has been used successfully to treat malaria patients in clinical studies, thus validating IspC as an antimalarial target. However, improvement of the drug's pharmacodynamics and pharmacokinetics is desirable. Here, we show that the conversion of the phosphonate moiety into acyloxymethyl and alkoxycarbonyloxymethyl groups can increase the in vitro activity against asexual blood stages of P. falciparum by more than 1 order of magnitude. We also synthesized double prodrugs by additional esterification of the hydroxamate moiety. Prodrugs with modified hydroxamate moieties are subject to bioactivation in vitro. All prodrugs demonstrated improved antiplasmodial in vitro activity. Selected prodrugs and parent compounds were also tested for their cytotoxicity toward HeLa cells and in vivo in a Plasmodium berghei malaria model as well as in the SCID mouse P. falciparum model.

Topics: Animals; Antimalarials; Cell Survival; Disease Models, Animal; Dose-Response Relationship, Drug; Fosfomycin; HeLa Cells; Humans; Malaria; Mice; Mice, SCID; Molecular Structure; Plasmodium berghei; Plasmodium falciparum; Prodrugs; Structure-Activity Relationship

Previous studies have shown that fosmidomycin, risedronate, and nerolidol exert antimalarial activity in vitro. We included squalestatin, an inhibitor of the isoprenoid metabolism in Erwinia uredovora, and found that combinations of compounds which act on different targets of the plasmodial isoprenoid pathway possess important supra-additivity effects.

Topics: Antimalarials; Biosynthetic Pathways; Bridged Bicyclo Compounds, Heterocyclic; Drug Interactions; Fosfomycin; Malaria; Parasitic Sensitivity Tests; Plasmodium falciparum; Risedronic Acid; Sesquiterpenes; Terpenes; Tricarboxylic Acids

Reverse hydroxamate-based inhibitors of IspC, a key enzyme of the non-mevalonate pathway of isoprenoid biosynthesis and a validated antimalarial target, were synthesized and biologically evaluated. The binding mode of one derivative in complex with EcIspC and a divalent metal ion was clarified by X-ray analysis. Pilot experiments have demonstrated in vivo potential.

Topics: Aldose-Ketose Isomerases; Animals; Antimalarials; Crystallography, X-Ray; Erythrocytes; Fosfomycin; Humans; Malaria; Mice; Models, Molecular; Molecular Structure; Multienzyme Complexes; Oxidoreductases; Parasitic Sensitivity Tests; Plasmodium berghei; Plasmodium falciparum; Structure-Activity Relationship

Plasmodium spp parasites harbor an unusual plastid organelle called the apicoplast. Due to its prokaryotic origin and essential function, the apicoplast is a key target for development of new anti-malarials. Over 500 proteins are predicted to localize to this organelle and several prokaryotic biochemical pathways have been annotated, yet the essential role of the apicoplast during human infection remains a mystery. Previous work showed that treatment with fosmidomycin, an inhibitor of non-mevalonate isoprenoid precursor biosynthesis in the apicoplast, inhibits the growth of blood-stage P. falciparum. Herein, we demonstrate that fosmidomycin inhibition can be chemically rescued by supplementation with isopentenyl pyrophosphate (IPP), the pathway product. Surprisingly, IPP supplementation also completely reverses death following treatment with antibiotics that cause loss of the apicoplast. We show that antibiotic-treated parasites rescued with IPP over multiple cycles specifically lose their apicoplast genome and fail to process or localize organelle proteins, rendering them functionally apicoplast-minus. Despite the loss of this essential organelle, these apicoplast-minus auxotrophs can be grown indefinitely in asexual blood stage culture but are entirely dependent on exogenous IPP for survival. These findings indicate that isoprenoid precursor biosynthesis is the only essential function of the apicoplast during blood-stage growth. Moreover, apicoplast-minus P. falciparum strains will be a powerful tool for further investigation of apicoplast biology as well as drug and vaccine development.

Topics: Animals; Anti-Bacterial Agents; Cell Death; Chloroplasts; Fosfomycin; Genome, Protozoan; Hemiterpenes; Humans; Life Cycle Stages; Malaria; Models, Biological; Organophosphorus Compounds; Parasites; Plasmodium falciparum; Protein Transport; Protozoan Proteins; Terpenes

FR-900098 is a potent chemotherapeutic agent for the treatment of malaria. Here we report the heterologous production of this compound in Escherichia coli by reconstructing the entire biosynthetic pathway using a three-plasmid system. Based on this system, whole-cell feeding assays in combination with in vitro enzymatic activity assays reveal an unusual functional role of nucleotide conjugation and lead to the complete elucidation of the previously unassigned late biosynthetic steps. These studies also suggest a biosynthetic route to a second phosphonate antibiotic, FR-33289. A thorough understanding of the FR-900098 biosynthetic pathway now opens possibilities for metabolic engineering in E. coli to increase production of the antimalarial antibiotic and combinatorial biosynthesis to generate novel derivatives of FR-900098.

Topics: Antimalarials; Biosynthetic Pathways; Escherichia coli; Escherichia coli Proteins; Fosfomycin; Genes, Bacterial; Humans; Malaria

Three alpha-halogenated analogues of 3-(acetylhydroxyamino)propylphosphonic acid (FR900098) have been synthesized from diethyl but-3-enylphosphonate using a previously described method for the alpha-halogenation of alkylphosphonates. These analogues were evaluated for antimalarial potential in vitro against Plasmodium falciparum and in vivo in the P. berghei mouse model. All three analogues showed higher in vitro and/or in vivo potency than the reference compounds.

Topics: Acute Disease; Animals; Antimalarials; Fosfomycin; Malaria; Mice; Plasmodium berghei; Plasmodium falciparum; Structure-Activity Relationship

Fosmidomycin, which acts through inhibition of 1-deoxy-D-xylulose phosphate reductoisomerase (DXR) in the non-mevalonate pathway, represents a valuable recent addition to the armamentarium against uncomplicated malaria. In this paper, we describe the synthesis and biological evaluation of E- and Z-alpha,beta-unsaturated alpha-aryl-substituted analogues of FR900098, a fosmidomycin congener, utilizing a Stille or a Suzuki coupling to introduce the aryl group. In contrast with our expectations based on the promising activity earlier observed for several alpha-substituted fosmidomycin analogues, all synthesized analogues exhibited much lower binding affinity for DXR than fosmidomycin.

Topics: Aldose-Ketose Isomerases; Animals; Antimalarials; Chemistry, Pharmaceutical; Drug Design; Drug Evaluation, Preclinical; Enzyme Inhibitors; Fosfomycin; Inhibitory Concentration 50; Malaria; Models, Chemical; Molecular Structure; Multienzyme Complexes; Oxidoreductases; Plasmodium falciparum; Structure-Activity Relationship

FR900098 represents a derivative of the new antimalarial drug fosmidomycin with enhanced activity. The mechanism of action is the inhibition of the 1-desoxy-D-xylulose 5-phosphate (DOXP) reductoisomerase, an essential enzyme of the mevalonate independent pathway of isoprenoid biosynthesis. Prodrugs with increased oral activity in mice infected with the rodent malaria parasite Plasmodium vinckei were obtained by masking the phosphonate moiety of FR900098 as alkoxycarbonyloxyethyl esters.

Topics: Administration, Oral; Aldose-Ketose Isomerases; Animals; Antimalarials; Biological Availability; Dose-Response Relationship, Drug; Fosfomycin; Malaria; Mice; Mice, Inbred BALB C; Multienzyme Complexes; Oxidoreductases; Plasmodium; Prodrugs; Quantitative Structure-Activity Relationship

FR900098 represents an improved derivative of the new antimalarial drug fosmidomycin and acts through inhibition of the 1-deoxy-D-xylulose 5-phosphate (DOXP) reductoisomerase, an essential enzyme of the mevalonate independent pathway of isoprenoid biosynthesis. Prodrugs with increased activity after oral administration were obtained by chemical modification of the phosphonate moiety to yield acyloxyalkyl esters. The most successful compound demonstrated 2-fold increased activity in mice infected with the rodent malaria parasite Plasmodium vinckei.

Topics: Animals; Antimalarials; Biological Availability; Fosfomycin; Indicators and Reagents; Lysophospholipids; Malaria; Mevalonic Acid; Mice; Mice, Inbred BALB C; Prodrugs

Fosmidomycin acts through inhibition of 1-deoxy-D-xylulose 5-phosphate (DOXP) reductoisomerase, a key enzyme of the nonmevalonate pathway of isoprenoid biosynthesis. It possesses potent antimalarial activity in vitro and in murine malaria. In a recent clinical study, fosmidomycin was effective and well tolerated in the treatment of patients with acute uncomplicated Plasmodium falciparum malaria but resulted in an unacceptably high rate of recrudescence. In order to identify a potential combination partner, the interaction of fosmidomycin with a number of antimalarial drugs in current use was investigated in a series of in vitro experiments. Synergy was observed between fosmidomycin and the lincosamides, lincomycin and clindamycin. The efficacy of a combination of fosmidomycin and clindamycin was subsequently demonstrated in the Plasmodium vinckei mouse model.

Topics: Animals; Anti-Bacterial Agents; Antimalarials; Clindamycin; Dose-Response Relationship, Drug; Drug Synergism; Fosfomycin; Humans; Malaria; Mice; Plasmodium falciparum

The fosmidomycin derivative FR900098 represents an inhibitor of the 1-deoxy-D-xylulose 5-phosphate (DOXP) reductoisomerase with potent antimalarial activity. Prodrugs of FR900098 with increased activity after oral administration were obtained by chemical modification of the phosphonate moiety to yield phosphodiaryl esters. One diaryl ester prodrug demonstrated efficacy in mice infected with the rodent malaria parasite Plasmodium vinckei comparable to i.p. drug administration.

Topics: Aldose-Ketose Isomerases; Animals; Antimalarials; Disease Models, Animal; Fosfomycin; Malaria; Mice; Multienzyme Complexes; Organophosphonates; Oxidoreductases; Plasmodium; Prodrugs; Treatment Outcome

A mevalonate-independent pathway of isoprenoid biosynthesis present in Plasmodium falciparum was shown to represent an effective target for chemotherapy of malaria. This pathway includes 1-deoxy-D-xylulose 5-phosphate (DOXP) as a key metabolite. The presence of two genes encoding the enzymes DOXP synthase and DOXP reductoisomerase suggests that isoprenoid biosynthesis in P. falciparum depends on the DOXP pathway. This pathway is probably located in the apicoplast. The recombinant P. falciparum DOXP reductoisomerase was inhibited by fosmidomycin and its derivative, FR-900098. Both drugs suppressed the in vitro growth of multidrug-resistant P. falciparum strains. After therapy with these drugs, mice infected with the rodent malaria parasite P. vinckei were cured.

Topics: Aldose-Ketose Isomerases; Amino Acid Sequence; Animals; Antimalarials; Cloning, Molecular; Enzyme Inhibitors; Fosfomycin; Genes, Protozoan; Hemiterpenes; Malaria; Malaria, Falciparum; Mevalonic Acid; Mice; Molecular Sequence Data; Multienzyme Complexes; Organelles; Organophosphorus Compounds; Oxidoreductases; Pentosephosphates; Plasmodium falciparum; Recombinant Proteins; Reverse Transcriptase Polymerase Chain Reaction; Terpenes

Topics: Aldose-Ketose Isomerases; Animals; Antimalarials; Drug Design; Enzyme Inhibitors; Fosfomycin; Hemiterpenes; Humans; Malaria; Malaria, Falciparum; Mice; Multienzyme Complexes; Organelles; Organophosphorus Compounds; Oxidoreductases; Plasmodium falciparum; Steroids; Transferases