Either of these approaches could be used to reduce expression of particular genes to enhance antibiotic killing. PNAs have demonstrated effective antimicrobial activity when conjugated to cell-penetrating-peptide (CPP) motifs and targeted to knockdown gene expression 13, 14, 15, 16, 17. Another tool for sequence-specific gene targeting are peptide nucleic acids (PNAs), single stranded DNA mimics that bind tightly to the corresponding antisense mRNAs 12. CRISPR has also been employed to interfere with the expression of target genes (CRISPRi) and has been explored for antibacterial applications 11. This includes transcriptome editing based on clustered regularly interspaced short palindromic repeats (CRISPR)-associated endonucleases (such as Cas9), which has been employed as an antimicrobial to selectively degrade particular genetic elements 9, 10. The design of such therapies has been made possible with recent advances in synthetic biology and the corresponding development of novel tools with sequence-specific targeting capabilities. In doing so, these treatments should theoretically minimize inherent selective pressure for bacteria to evolve escape strategies that negate their impact. Crucially, such therapies must be designed to have minimal direct impact on bacterial fitness when administered independently. Rather than focusing on the development of novel drugs utilizing alternative mechanisms for killing bacteria, this approach aims to devise co-therapies whose sole function is to potentiate the efficacy of existing antibiotic treatment.
Here we seek to highlight a potential therapeutic approach to tackling this fundamental problem in combating antibiotic resistance. Without addressing this central law governing antibiotic resistance, the antibiotic arms race seems all but perpetual. The remarkable capacity for microbes to evolve resistance is due to a combination of relatively quick doubling times and large population sizes, allowing bacteria to rapidly explore a diverse array of genetic possibilities and increasing the likelihood for a fortuitously beneficial variant to emerge. In this model, while scientists continue to discover and bring to market novel drugs, it is almost universally accepted that bacteria will continue to evolve resistance and thus inexorably diminish a therapy’s efficacy over time 8. This has led to what is often referred to as the antibiotic arms race 6, 7. This problem is likely to be exacerbated as multidrug-resistant (MDR) bacteria continue to emerge, necessitating the pursuit of alternative antimicrobial strategies.Ĭurrent antibiotic research has focused largely on developing new drugs that exhibit bactericidal activity through novel mechanisms. Both the World Economic Forum 3 and the World Health Organization 4 warn of calamitous economic and health outcomes if current trends continue unabated while approximately 1 million people die from such infectious yearly, annual deaths attributable to antibiotic resistant infections are estimated to reach 10 million by 2050 5.
Estimates for the yearly cost imposed by antibiotic resistance reaches as high as $55 billion in the United States 1 and €1.5 billion across Europe 2. Our results highlight a promising approach for extending the utility of current antibiotics.Īntibiotic resistance is one of the foremost problems facing humanity. coli showed three cases of re-sensitization with minimal fitness impacts. PNA combined with sub-minimal inhibitory concentrations of trimethoprim against two isolates of Klebsiella pneumoniae and E. Finally, these results informed the design of four antisense peptide nucleic acid (PNA) co-therapies, csgD, fnr, recA and acrA, against four MDR, clinically isolated bacteria. These fitness neutral gene perturbations worked as co-therapies in reducing a Salmonella enterica intracellular infection in HeLa. Identified gene targets were subsequently tested for antibiotic synergy on the transcriptomic level via multiplexed CRISPR-dCas9 and showed successful sensitization of E. We systematically explored 270 gene knockout-antibiotic combinations in Escherichia coli, identifying 90 synergistic interactions. Here we propose using fitness neutral gene expression perturbations to potentiate antibiotics. Proliferation of multidrug-resistant (MDR) bacteria poses a threat to human health, requiring new strategies.
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