Overcoming Target-Mediated Quinolone Resistance in Topoisomerase IV by Introducing Metal-Ion-Independent Drug–Enzyme Interactions

Abstract

Quinolones, which target gyrase and topoisomerase IV, are the most widely prescribed antibacterials worldwide. Unfortunately, their use is threatened by the increasing prevalence of target-mediated drug resistance. Greater than 90% of mutations that confer quinolone resistance act by disrupting enzyme-drug interactions coordinated by a critical water-metal ion bridge. Quinazolinediones are quinolone-like drugs, but lack the skeletal features necessary to support the bridge interaction. These compounds are of clinical interest, however, because they retain activity against the most common quinolone resistance mutations. We utilized a chemical biology approach to determine how quinazolinediones overcome quinolone resistance in Bacillus anthracis topoisomerase IV. Quinazolinediones that retain activity against quinolone-resistant topoisomerase IV do so primarily by establishing novel interactions through the C7 substituent, rather than the drug skeleton. Because some quinolones are highly active against human topoisomerase IIα, we also determined how clinically relevant quinolones discriminate between the bacterial and human enzymes. Clinically relevant quinolones display poor activity against topoisomerase IIα because the human enzyme cannot support drug interactions mediated by the water-metal ion bridge. However, the inclusion of substituents that allow quinazolinediones to overcome topoisomerase IV-mediated quinolone resistance can cause cross-reactivity against topoisomerase IIα. Therefore, a major challenge in designing drugs that overcome quinolone resistance lies in the ability to identify substituents that mediate strong interactions with the bacterial, but not the human, enzymes. Based on our understanding of quinolone-enzyme interactions, we have identified three compounds that display high activity against quinolone-resistant B. anthracis topoisomerase IV but low activity against human topoisomerase IIα.

Fluoroquinolone interactions with Mycobacterium tuberculosis gyrase: Enhancing drug activity against wild-type and resistant gyrase

“…Results indicate that the water-metal ion bridge is partially functional in WT gyrase and that the most common resistance mutations cause a decrease in bridge-mediated drug affinity for the enzyme. In contrast to other species (32,33), quinolone interactions within the gyrase-cleaved DNA complex depend more heavily on substituents at C7 and C8. Based on an analysis of structure-activity relationships at these two positions, we identified fluoroquinolones that display significantly improved activity against WT and resistant M. tuberculosis gyrase compared with moxifloxacin.…”

“…In several other species, quinazolinediones containing a 3′-(aminomethyl) pyrrolidinyl [3′-(AM)P] (or a related) group at C7 maintain activity against enzymes containing mutations that disrupt the water-metal ion bridge by forming binding interactions primarily through the C7 substituent (20,21,32,(38)(39)(40)(41)(42). As seen in Fig.…”

Characterization of the Novel DNA Gyrase Inhibitor AZD0914: Low Resistance Potential and Lack of Cross-Resistance in Neisseria gonorrhoeae

“…3C). The manual superimposition of AZD0914, maintaining the flat planar stacking mode similar to the fluoroquinolones, suggests that unlike the fluoroquinolones and similar to the quinazolinediones, AZD0914 does not have a metal chelating group and as a result is impervious to the substitutions that confer resistance to fluoroquinolones via the displacement of the metal-binding residues (32,33). In addition, the intercalation of AZD0914 in this pocket puts the Lys450 residue within the correct distance to have a favorable polar interaction with the oxygen of the pyrimidinetrione group of AZD0914 (Fig.…”

Bacillus anthracis GrlA V96A Topoisomerase IV, a Quinolone Resistance Mutation That Does Not Affect the Water-Metal Ion Bridge

“…Furthermore, the level of resistance caused by this mutation is lower than that seen in the presence of Ser81 or Glu85 mutations in B. anthracis topoisomerase IV (34)(35)(36). Finally, the only quinazolinedione resistance mutations reported to date occur in the TOPRIM domain of the B subunit of gyrase and topoisomerase IV (46,48).…”

“…This water-metal ion bridge is formed when the C-3/C-4 keto acid of the drug chelates a divalent metal ion, which interacts with the protein through water molecules that are coordinated by the conserved serine and acidic amino acid residues. In Bacillus anthracis topoisomerase IV, mutations in the bridge-anchoring residues (Ser81 and Glu85) cause drug resistance that is specific to quinolones (34)(35)(36). Resistance is caused by a partial or complete loss of bridge function that is accompanied by a marked decrease in quinolone affinity (34,35).…”