Highly compacted biodegradable DNA nanoparticles capable of overcoming the mucus barrier for inhaled lung gene therapy

Abstract

Gene therapy has emerged as an alternative for the treatment of diseases refractory to conventional therapeutics. Synthetic nanoparticle-based gene delivery systems offer highly tunable platforms for the delivery of therapeutic genes. However, the inability to achieve sustained, high-level transgene expression in vivo presents a significant hurdle. The respiratory system, although readily accessible, remains a challenging target, as effective gene therapy mandates colloidal stability in physiological fluids and the ability to overcome biological barriers found in the lung. We formulated highly stable DNA nanoparticles based on state-of-theart biodegradable polymers, poly(β-amino esters) (PBAEs), possessing a dense corona of polyethylene glycol. We found that these nanoparticles efficiently penetrated the nanoporous and highly adhesive human mucus gel layer that constitutes a primary barrier to reaching the underlying epithelium. We also discovered that these PBAE-based mucus-penetrating DNA nanoparticles (PBAE-MPPs) provided uniform and high-level transgene expression throughout the mouse lungs, superior to several gold standard gene delivery systems. PBAE-MPPs achieved robust transgene expression over at least 4 mo following a single administration, and their transfection efficiency was not attenuated by repeated administrations, underscoring their clinical relevance. Importantly, PBAE-MPPs demonstrated a favorable safety profile with no signs of toxicity following intratracheal administration.lung gene therapy | mucus-penetrating particles | nanotechnology | biodegradable polymer | nonviral gene delivery

Protein nanocages that penetrate airway mucus and tumor tissue

“…We have previously demonstrated that muco-inert nanoparticles capable of efficiently penetrating airway mucus provide widespread in vivo distribution throughout the mouse airways following inhalation, whereas muco-adhesive nanoparticles were unable to do so (28,29). We thus investigated the airway distribution of Cy5-labeled FTn, with or without PEG surface coatings, following intranasal administration in mice.…”

“…The sections were stained with Prolong Gold antifade with DAPI (Life Technologies), and fluorescence images of the sections were obtained using a Zeiss confocal microscope. To quantify the particle distribution, the acquired images were analyzed by following an image-based analysis method that we have previously reported (28). Briefly, at least 10 fluorescence images at 10× magnification were taken of the lungs harvested from individual animals.…”

Lessons learned from lung and liver in-vivo gene therapy: implications for the future

“…Furthermore, mucus buildup is often associated with inflammation, which can lead to reduced efficacy of viral vectors when they are taken up by an increased number of resident lung macrophages, attracted by a boosted immune system. Both these issues can possibly be overcome by mucus-penetrating nanoparticles, or the co-administration of mucolytic drugs [45]. …”

The Mucus Barrier to Inhaled Gene Therapy

“…18,49 Most nonviral gene vectors are formulated with positivelycharged carrier materials, including cationic polymers and lipids, that condense nucleic acids into NP with net positively-charged surfaces. [79][80][81][82] Sanders et al, showed the diffusion of cationic 1,2-dioleoyl-3-trimethylammoniumpropane (DOTAP) lipoplexes was strongly hindered in CF mucus. 79 This effect was also observed for some polymer-based gene delivery systems, including those made using polyethylenimine, 80 polyamidoamine dendrimer, 81 and poly-β-amino ester, 82 each of which also carry positively-charged surfaces (>20 mV ζ-potential).…”