title: Target-based resistance in Pseudomonas aeruginosa and Escherichia coli to NBTI 5463, a novel bacterial type II topoisomerase inhibitor.
abstract: In a previous report (T. J. Dougherty, A. Nayar, J. V. Newman, S. Hopkins, G. G. Stone, M. Johnstone, A. B. Shapiro, M. Cronin, F. Reck, and D. E. Ehmann, Antimicrob Agents Chemother 58:2657-2664, 2014), a novel bacterial type II topoisomerase inhibitor, NBTI 5463, with activity against Gram-negative pathogens was described. First-step resistance mutations in Pseudomonas aeruginosa arose exclusively in the nfxB gene, a regulator of the MexCD-OprJ efflux pump system. The present report describes further resistance studies with NBTI 5463 in both Pseudomonas aeruginosa and Escherichia coli. Second-step mutations in P. aeruginosa arose at aspartate 82 of the gyrase A subunit and led to 4- to 8-fold increases in the MIC over those seen in the parental strain with a first-step nfxB efflux mutation. A third-step mutant showed additional GyrA changes, with no changes in topoisomerase IV. Despite repeated efforts, resistance mutations could not be selected in E. coli. Genetic introduction of the Asp82 mutations observed in P. aeruginosa did not significantly increase the NBTI MIC in E. coli. However, with the aspartate 82 mutation present, it was possible to select second-step mutations in topoisomerase IV that did lead to MIC increases of 16- and 128-fold. As with the gyrase aspartate 82 mutation, the mutations in topoisomerase IV did not by themselves raise the NBTI MIC in E. coli. Only the presence of mutations in both targets of E. coli led to an increase in NBTI MIC values. This represents a demonstration of the value of balanced dual-target activity in mitigating resistance development.
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, Massachusetts, USA.
authors.forename: Asha S
authors.initials: AS
authors.lastname: Nayar
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, Massachusetts, USA.
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Dougherty
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, Massachusetts, USA.
authors.forename: Folkert
authors.initials: F
authors.lastname: Reck
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, Massachusetts, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Thresher
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, Massachusetts, USA.
authors.forename: Ning
authors.initials: N
authors.lastname: Gao
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, Massachusetts, USA.
authors.forename: Adam B
authors.initials: AB
authors.lastname: Shapiro
authors.affiliation: Infection Innovative Medicines Unit, AstraZeneca R&D Boston, Waltham, Massachusetts, USA david.ehmann@astrazeneca.com.
authors.forename: David E
authors.initials: DE
authors.lastname: Ehmann
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics; D027081:DNA Gyrase / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000187:drug effects* / Q000235:genetics; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics; D008826:Microbial Sensitivity Tests; D009025:Morpholines / Q000494:pharmacology*; D009154:Mutation; D009287:Naphthyridines / Q000494:pharmacology*; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000235:genetics; D059005:Topoisomerase II Inhibitors / Q000494:pharmacology*
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D009025:Morpholines; C000600719:NBTI 5463; D009287:Naphthyridines; D059005:Topoisomerase II Inhibitors; D027081:DNA Gyrase