Overcoming Resistance to Novel Bacterial Topoisomerase Inhibitors

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PROJECT SUMMARY The rising tide of antimicrobial resistance threatens catastrophic increases in mortality in the coming decades. Methicillin-resistant Staphylococcus aureus (MRSA) remains a leading pathogen. New antibacterial classes are urgently needed to ensure adequate therapeutic options for MRSA and other resistant bacteria. Novel Bacterial Type II Topoisomerase Inhibitors (NBTIs) derive their efficacy by targeting the clinically validated essential enzymes, DNA gyrase and topoisomerase IV (TopoIV). A novel binding mode avoids target-based cross-resistance to fluoroquinolones and establishes NBTIs as a new antibacterial class. A lead, gepotidacin, stands at the threshold of FDA approval, with several completed Phase 2 and ongoing Phase 3 clinical trials. Resistance to gepotidacin has been observed but is very poorly characterized. The transformative potential of the NBTIs will require a better understanding of mechanisms of action/resistance and new medicinal chemistry strategies to deliver highly efficacious successor NBTIs, the areas of focus in the present proposal. To date, we have synthesized >250 highly diverse NBTIs. Our anti-MRSA lead, 147, showed in vivo efficacy in two infection models and a favorable cardiovascular safety profile by rationally designed reductions of basicity and lipophilicity. We have generated NBTIs with improved dual-targeting of gyrase and TopoIV, reduced rates of spontaneous resistance, and greater antibacterial activity over gepotidacin against NBTI- resistant MRSA. In contrast to gepotidacin, several newly synthesized amide-containing NBTIs induced DNA double strand breaks which we will investigate as a new mechanism of action for the NBTI class. Critically, we also propose that studies with our existing and planned NBTIs, coupled with our demonstrated expertise in microbiology, biochemical pharmacology, computational chemistry, and structural biology, will effectively address major unanswered questions regarding the emergence of resistance to NBTIs and strategies to overcome this issue. Overall, our goal is to generate lead compounds as innovative chemical tools and/or clinical candidates for further development. Three integrated specific aims will be pursued by our interdisciplinary team to: 1) Synthesize structurally and mechanistically distinct NBTIs with druglike properties 2) Evaluate new NBTIs for antibacterial activity & identify/characterize key NBTI-resistant S. aureus mutants 3) Elucidate the mechanism(s) of action of and molecular resistance to new lead NBTIs Aim 1 serves as the innovation engine for the proposal. Aims 2 and 3 support Aim 1 through iterative cycles of rigorous assays to provide new lead compounds. New fundamental information concerning the origin, mechanism, and impact/circumvention of acquired resistance to NBTIs will advance this new class of antibacterials as a pathway to promote human health by addressing the crisis in antimicrobial resistance.