title: Prospective screening of novel antibacterial inhibitors of dihydrofolate reductase for mutational resistance.
abstract: Resistance to trimethoprim (TMP) resulting from point mutations in the enzyme drug target dihydrofolate reductase (DHFR) drives the development of new antifolate inhibitors effective against methicillin-resistant Staphylococcus aureus (MRSA). For the past several years we have used structure-based design to create propargyl-linked antifolates that are highly potent antibacterial agents. In order to focus priority on the development of lead compounds with a low propensity to induce resistance, we prospectively evaluated resistance profiles for two of these inhibitors in an MRSA strain. By selection with the lead inhibitors, we generated resistant strains that contain single point mutations F98Y and H30N associated with TMP resistance and one novel mutation, F98I, in DHFR. Encouragingly, the pyridyl propargyl-linked inhibitor selects mutants at low frequency (6.85 × 10(-10) to 1.65 × 10(-9)) and maintains a low MIC (2.5 μg/ml) and a low mutant prevention concentration (1.25 μg/ml), strongly supporting its position as a lead compound. Results from this prospective screening method inform the continued design of antifolates effective against mutations at the Phe 98 position. Furthermore, the method can be used broadly to incorporate ideas for overcoming resistance early in the development process.
authors.affiliation: Department of Pharmaceutical Sciences, University of Connecticut, Storrs, Connecticut, USA.
authors.forename: Kathleen M
authors.initials: KM
authors.lastname: Frey
authors.forename: Kishore
authors.initials: K
authors.lastname: Viswanathan
authors.forename: Dennis L
authors.initials: DL
authors.lastname: Wright
authors.forename: Amy C
authors.initials: AC
authors.lastname: Anderson
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D005493:Folic Acid Antagonists / Q000494:pharmacology; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects / Q000235:genetics; D008826:Microbial Sensitivity Tests; D009154:Mutation; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D014295:Trimethoprim / Q000494:pharmacology; D014296:Trimethoprim Resistance / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D005493:Folic Acid Antagonists; D014295:Trimethoprim; D013762:Tetrahydrofolate Dehydrogenase