(228bp) Sequence Specific Antisense Inhibitors of Non-Traditional Antibiotic Pathways for Eliminating Multidrug-Resistant Bacteria

Multidrug-resistant (MDR) infections are a pressing concern to global health which is made worse by the lack of new antibiotics being introduced. The overuse and misuse of current antibiotics has created pan-drug resistant bacteria to which there are no longer efficacious antibiotics. These current antibiotics are typically limited to three main pathways in bacteria: DNA replication and cell growth, protein biosynthesis, and cell wall biosynthesis. Developing antisense therapeutics, which enable sequence specific targeting of genes, allows us to take advantage of the many non-traditional antibiotic pathways to both uncover novel antibiotic targets and elucidate potential combination therapies which capitalize on the unexplored bacterial pathways. Using Escherichia coli as a model organism we created peptide nucleic acid based antisense molecules which target essential genes in non-traditional antibiotic pathways including metabolism, cell signaling, stress response, and gene regulation via antisense inhibition. Although the antisense molecules were designed against essential genes in E. coli, these molecules demonstrate therapeutic potential against multiple pathogenic bacteria including in clinical isolates of E. coli, Klebsiella pneumoniae, and Salmonella typhimurium, thus highlighting the potential to create broad-spectrum yet gene specific antibiotics. The clinical isolates used in this study are highly resistant to most classes of antibiotics, yet when the antisense molecules are used in combination with traditional antibiotics we observe a strong synergistic effect that significantly inhibits cell growth greater than either mono-therapy. We are able to re-sensitize the clinical isolates to Clinical Laboratory Standards Institute (CLSI) levels of antibiotics using the antisense inhibitors of non-traditional antibiotic pathways. Our findings highlight the potential utility of both targeting novel targets in non-traditional antibiotic pathways as well as the promise of combination therapies taking advantage of the novel targets in bacteria.