The overuse of antibiotics plays a major role in the emergence and spread of multidrug-resistant bacteria. A molecularly targeted, specific treatment regimen that acts in narrow spectrum would provide a great advantage by reducing the selective pressure during bacterial growth. Herein, we introduce a gene therapeutic approach based on nonviral delivery of genome editing material for the treatment of the multidrug-resistant infections and preventing the spread of these pathogens. We apply the CRISPR-Cas9 system, which has been recognized as an innovative tool for highly specific and efficient genome engineering in different organisms, as the therapeutic cargo. Since most gene editing strategies using CRISPR-Cas9 have been based on viral vectors and thus limited in applicability for therapeutic purposes, we have developed a nonviral system utilizing a nanocomplex of chemically modified Cas9 endonuclease and single-guide RNA. We show that CRISPR nanocomplexes are successfully formed, by characterization using DLS and TEM analyses, while maintaining the functional activity for double-strand DNA cleavage. We also demonstrate that nanocomplexes designed to target mecA- the major gene involved in methicillin resistance- can be efficiently delivered into Methicillin-resistant <i>Staphylococcus</i> <i>aureus </i>(MRSA), and are also able to edit the bacterial genome by targeting mecA. The present study shows the potential applicability of the CRISPR nanocomplex in the clinic as a target specific antimicrobial, and ultimately contribute to control of antibacterial resistance and global health.

Nonviral genome editing based on a CRISPR nanocomplex for target-specific treatment of multidrug-resistant bacterial infections

253rd National Meeting (2017)

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