Co-delivery of free vancomycin and transcription factor decoy-nanostructured lipid carriers can enhance inhibition of methicillin resistant Staphylococcus aureus (MRSA).

Alan Hibbitts,Jesús M De La Fuente,Inés Serrano-Sevilla,Laura De Matteis,Ainhoa Lucía,Michael Mcarthur,Fabrice Navarro,José A Aínsa,Prabagaran Narayanasamy

doi:10.1371/journal.pone.0220684

Abstract

Bacterial resistance to antibiotics is widely regarded as a major public health concern with last resort MRSA treatments like vancomycin now encountering resistant strains. TFDs (Transcription Factor Decoys) are oligonucleotide copies of the DNA-binding sites for transcription factors. They bind to and sequester the targeted transcription factor, thus inhibiting transcription of many genes. By developing TFDs with sequences aimed at inhibiting transcription factors controlling the expression of highly conserved bacterial cell wall proteins, TFDs present as a potential method for inhibiting microbial growth without encountering typical resistance mechanisms. However, the efficient protection and delivery of the TFDs inside the bacterial cells is a critical step for the success of this technology. Therefore, in our study, specific TFDs against S. aureus were complexed with two different types of nanocarriers: cationic nanostructured lipid carriers (cNLCs) and chitosan-based nanoparticles (CS-NCs). These TFD-carrier nanocomplexes were characterized for size, zeta potential and TFD complexation or loading efficiency in a variety of buffers. In vitro activity of the nanocomplexes was examined alone and in combination with vancomycin, first in methicillin susceptible strains of S. aureus with the lead candidate advancing to tests against MRSA cultures. Results found that both cNLCs and chitosan-based carriers were adept at complexing and protecting TFDs in a range of physiological and microbiological buffers up to 72 hours. From initial testing, chitosan-TFD particles demonstrated no visible improvements in effect when co-administered with vancomycin. However, co-delivery of cNLC-TFD with vancomycin reduced the MIC of vancomycin by over 50% in MSSA and resulted in significant decreases in viability compared with vancomycin alone in MRSA cultures. Furthermore, these TFD-loaded particles demonstrated very low levels of cytotoxicity and haemolysis in vitro. To our knowledge, this is the first attempt at a combined antibiotic/oligonucleotide-TFD approach to combatting MRSA and, as such, highlights a new avenue of MRSA treatment combining traditional small molecules drugs and bacterial gene inhibition.

Highlights

Antimicrobial resistance to conventional antibiotics is an increasingly serious threat to global public health that requires urgent action
CNLC-transcription factor decoys (TFDs) nanocomplexes dispersed in phosphate buffered saline (PBS) were the only samples that retained a cationic surface charge throughout the 72 hours of testing
All the cell and microbiology culture media displayed roughly the same anionic surface charges (-3.6 mV to -4.1 mV) immediately after dispersion. cationic nanostructured lipid carriers (cNLCs)-TFD nanocomplexes dispersed in culture media demonstrated further decreases at 24-hours incubation with the lowest zeta potential recorded by nanocomplexes dispersed in A549 medium showing -10.2 mV

Summary

Introduction

Antimicrobial resistance to conventional antibiotics is an increasingly serious threat to global public health that requires urgent action. Antimicrobial-resistant infections carry higher incidents of mortality and present a considerable economic burden of over 20 billion dollars per year in the US alone [1] This translates to approximately 23,000 deaths in the US and 33,000 in the EU annually as a direct result of an antimicrobial-resistant infection [1, 2] with global mortality expected to rise to up to 300 million deaths by 2050 [1]. The emergence of strains resistant to methicillin and other antimicrobial agents has added to the cost and length of treatment resulting in growing concern among medical professionals [5] This is especially so in the hospital environment due to the higher mortality rates arising from systemic methicillin-resistant S. aureus (MRSA) infections [6, 7]. Patients infected with MRSA are estimated to be 64% more likely to die than those infected with a non-resistant form of S. aureus

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