Sequence-Specific Peptide Nucleic Acid-Based Antisense Inhibitors of TEM-1 β-Lactamase and Mechanism of Adaptive Resistance

The recent surge of drug-resistant superbugs and shrinking antibiotic pipeline are serious challenges to global health. In particular, the emergence of beta-lactamases has caused extensive resistance against the most frequently prescribed class of beta-lactam antibiotics. Here, we develop novel synthetic peptide nucleic acid-based antisense inhibitors that target the start codon and ribosomal binding site of the TEM-1 beta-lactamase transcript and act via translation inhibition mechanism. We show that these antisense inhibitors are capable of resensitizing drug-resistant Escherichia coli to beta-lactam antibiotics exhibiting 10-fold reduction in the minimum inhibitory concentration (MIC). To study the mechanism of resistance, we adapted E. coli at MIC levels of the beta-lactam/antisense inhibitor combination and observed a nonmutational, bet-hedging based adaptive antibiotic resistance response as evidenced by phenotypic heterogeneity as well as heterogeneous expression of key stress response genes. Our data show that both the development of new antimicrobials and an understanding of cellular response during the development of tolerance could aid in mitigating the impending antibiotic crisis.