The effect of moxifloxacin on its target topoisomerases from Escherichia coli and Staphylococcus aureus.

The effect of moxifloxacin on its target enzymes was evaluated by three different approaches: (i) the MICs of moxifloxacin and nine other fluoroquinolones were determined for mutants of Escherichia coli (n = 13) and Staphylococcus aureus (n = 5) carrying different combinations of resistance mutations; (ii) the activity of moxifloxacin on isolated targets was determined as IC50 values for wild-type and mutant type II topoisomerases from E. coli; and (iii) the mutation frequencies were determined for two single-step mutants (MI with a Ser83-->Leu mutation in gyrA and WT-4 with a Ser80-->Ile mutation in parC) and their parent strain (WT). Of the quinolones tested, moxifloxacin was the only one showing an equivalent high activity against both targets. This is reflected by a comparable high susceptibility of the test strains of E. coli and S. aureus and by the IC50 values of moxifloxacin which were 50-90% lower than those of ciprofloxacin, norfloxacin and sparfloxacin for the wild-type and single mutant enzymes of gyrase and topoisomerase IV. However, double mutant GyrA was significantly more sensitive to moxifloxacin than to the other fluoroquinolones tested, while wild-type topoisomerase IV was two-fold more refractory. Mutation rates of WT, MI and WT-4 for ciprofloxacin and moxifloxacin were 5 x 10(-8) vs 4 x 10(-10); <6 x 10(-11) vs <6 x 10(-11); and 2 x 10(-6) vs 5 x 10(-7), respectively. These data indicate an equivalent high inhibitory activity of moxifloxacin on DNA gyrase and topoisomerase IV of E. coli.

Syntheses and biological activities of new N1-aryl substituted quinolone antibacterials.

A series of quinolones with a systematically varied substitution at the phenyl ring at N1 has been synthesized. Three lipophilicity descriptors (log K, log P, Rm) and the pKa values have been determined as well as the microbiological activity: The MIC values for eight different strains of three Gram-positive and three Gram-negative species and the inhibitory concentrations of DNA supercoiling (IC90 and IC100) were determined. From a principal component and a QSAR analysis relationships between antibacterial activity concerning the whole-cell system and electronic properties as well as the length of the substituents at the phenyl rings could be derived. The activity in a cell-free system was governed by the lipophilicity and the width of the substituents. It is speculated that the quinolones take a defined place in the DNA gyrase-DNA complex which is characterized by polar amino acids. This is in agreement with findings from studies of mutant gyrases.

Mutations in the gyrA gene of a highly fluoroquinolone-resistant clinical isolate of Escherichia coli.

We have determined the DNA sequence of the gyrA gene of the fluoroquinolone-resistant Escherichia coli isolate 205096 (MIC of ciprofloxacin, 128 micrograms/ml), which was recently demonstrated to be a gyrA mutant (P. Heisig and B. Wiedemann, Antimicrob. Agents Chemother. 35:2031-2036, 1991). Compared with the gyrA+ gene of E. coli K-12, 55 nucleotide changes were found. Three of these resulted in amino acid exchanges: Ser-83-->Leu, Asp-87-->Gly, and Asp-678-->Glu. A 0.7-kb DNA fragment containing two of these mutations (Ser-83-->Leu and Asp-87-->Gly) was isolated and fused in frame to the residual 3' coding region of gyrA+ in a plasmid to yield a chimeric gyrA gene (gyrA#). After introduction into E. coli 205096, this gyrA# gene does not increase the fluoroquinolone susceptibility of the resulting heterodiploid strain in a dominance test, while the gyrA+ gene does. The ciprofloxacin concentration necessary to inhibit by 90% (IC90) the supercoiling activity of gyrase isolated from E. coli 205096 is above 2,000 micrograms/ml. An identical result was found for gyrase reconstituted in vitro from the gyrB+ gene product and the chimeric gyrA# gene product. This is more than a 4,000-fold increase compared with the IC90 determined for gyrase from E. coli K-12 (gyrA+) (IC90, 0.5 microgram of ciprofloxacin per ml). No indications for the involvement of the gyrB gene or for alterations in quinolone permeation were found.