Disease relevance of ENT-26398

• Previous studies have shown that 100 nM 5-fluoroorotate (5-FO) is sufficient to block the in vitro proliferation of Plasmodium falciparum without causing toxicity to mammalian cells [1].
• This second wave of parasitemia cleared without additional 5-fluoroorotate treatment [2].
• On the basis of the pharmacokinetics profile, mice infected with Plasmodium yoelii were treated with 5-fluoroorotate at a dose of 0.2 or 5 mg/kg (body weight) every 4 h for 3 days [2].
• 5-Fluoroorotate-resistant mutants of Salmonella typhimurium [3].

High impact information on ENT-26398

• Using Ty1-URA3 reporters and negative selection with 5-fluoroorotic acid, it is shown that Ty1 genes can undergo transcriptional cosuppression that is independent of DNA methylation and polycomb-mediated repression [4].
• Basal transcription was monitored by the use of a GAL1 promoter-URA3 reporter construct whose uninduced activity can be detected through cellular sensitivity to the drug, 5-fluoroorotic acid [5].
• Liver rough endoplasmic reticulum (RER) membranes were isolated from rats given [3H]orotic acid for 48 h (ribosomal RNA [rRNA] label) or for 3 h along with 5-fluoroorotate; this latter procedure permits the labeling of cytoplasmic messenger RNAs (mRNAs) in the absence of rRNA labeling [6].
• To test this hypothesis, different Plasmodium falciparum clones having varying numbers of preexisting resistance mechanisms were treated with two new antimalarial agents: 5-fluoroorotate and atovaquone [7].
• The engineered Saccharomyces cerevisiae cells used in this study express two interacting proteins after induction with galactose whose interaction results in cell death in the presence of 5-fluoroorotic acid (inducible reverse two-hybrid assay) [8].

Biological context of ENT-26398

• A promoter containing 25 repeats allows expression of a URA3 reporter gene and yields sensitivity to the drug 5-fluoroorotic acid [9].
• Excision of the vector sequences and one copy of the duplicated globin sequences by homologous recombination is required for cell survival when exposed to the selective agent 5-fluoroorotic acid, which is toxic to ura+ yeast cells [10].
• By utilizing a 5-fluoroorotic acid-mediated plasmid shuffle in the EcMetAP2 yeast strain, a yeast strain can be generated whose growth is dependent on MetAP2 from any organism [11].
• These Ura- segregants were selected on 5-fluoroorotic acid-containing medium and used in the next round of mutagenesis [12].
• Compared with malarial parasites, the mammalian cells were relatively tolerant of 5-fluoroorotic acid (50% inhibitory concentration, 0.9 to 10 microM) [13].

Anatomical context of ENT-26398

• Two drugs tested, methotrexate-gamma-aspartate and 5-fluoroorotate, show increased cytotoxicity on various cultured cell lines following encapsulation in liposomes and can be considered liposome-dependent agents [14].
• We have selected 143 independent Nicotiana plumbaginifolia cell lines that survive in the presence of 5-fluoroorotic acid [15].
• Protoplasts prepared from different yeasts were fused to the protoplasts of the ade2-1, ura3-52, kar1-1, rho0 strain of S. cerevisiae and were selected for respiring cybrids on plates containing 5-fluoroorotic acid and a non-fermentable carbon source [16].
• On the contrary, lapacol and 5-fluoroorotate, inhibitors of DHODH besides LFM, suppressed the biogenesis of mitochondria during the cell cycle progression [17].

Associations of ENT-26398 with other chemical compounds

• In contrast to wild-type cell lines, the fluoroorotic acid-selected cell lines did not respond to thymine or to other biochemicals that affect thymine levels [15].
• Incubation of malarial parasites with 3 to 10 microM radioactive 5-fluoroorotic acid for 48 h resulted in significant incorporation of radioactivity into the RNA fraction of P. falciparum; approximately 9% of the uridine residues were substituted with 5-fluorouridine [18].
• Clonal viability measurements on Plasmodium falciparum to assess in vitro schizonticidal activity of leupeptin, chloroquine, and 5-fluoroorotate [19].
• Consistent with a proposed mechanism for the potent antimalarial activity of 5-fluoroorotate, 5-fluoro-2'-deoxyuridylate inhibited Plasmodium falciparum thymidylate synthase with a Ki of 2 nM [20].
• The ability of conjugates of peptides and 5-fluorocytosine or 5-fluoroorotic acid to enter Candida albicans was investigated [21].

Gene context of ENT-26398

• Using this PDR5 promoter-URA3 recombinant, we further demonstrate a simple extension of the method that yields regulatory mutants via 5-fluoroorotic acid selection [22].
• Several strategies commonly used in Saccharomyces cerevisiae and other eukaryotes were unsuccessful, due to the frequent occurrence of ectopic integration, linear plasmid formation, and spontaneous resistance to 5-fluoroorotic acid, which is a selective agent for URA5 gene inactivation [23].
• Plasmid shuffling experiment with Saccharomyces cerevisiae GTase subunit gene CEG1 null mutant demonstrated that deletion mutants 211-567 and 211-585 were able to support cell viability in the presence of 5-fluoroorotic acid, whereas 211-537 and 211-560 were not [24].
• Dihydrofolate reductase activity, which resides on the same bifunctional protein as thymidylate synthase, was not affected by 5-fluoroorotate treatment [18].
• Using the toxicity of the DctA substrate fluoroorotic acid as a selective agent in an iterated selection procedure, four independent S. meliloti dctA mutants were isolated that retained some ability to transport dicarboxylates [25].

Analytical, diagnostic and therapeutic context of ENT-26398

• When plasmid DNA containing the cloned WdURA5 gene was introduced by electroporation into a wdura5 auxotrophic recipient strain derived previously by selection with 5-fluoroorotic acid (5-FOA), an apparent gene repair event occurred at high frequency without any evidence of integration of the plasmid DNA [26].
• Dihydroorotic acid hydrazide (DHOH) did not inhibit the replication of potatovirus X (PVX) in leaf disks of Nicotiana tabacum 'Samsun'. In contrast, 5-fluoroorotic acid (5-FOA) completely inhibited the replication of PVX, as demonstrated by a serological virus assay as well as a local lesion bioassay using Gomphrena globosa as the test plant [27].

References