Abstract
Nowadays, the downregulation of genes involved in the pathogenesis of severe lung diseases through local siRNA delivery appears an interesting therapeutic approach. In this study, we propose novel hybrid lipid-polymer nanoparticles (hNPs) consisting of poly(lactic-co-glycolic) acid (PLGA) and dipalmitoyl phosphatidylcholine (DPPC) as siRNA inhalation system. A panel of DPPC/PLGA hNPs was prepared by emulsion/solvent diffusion and fully characterized. A combination of model siRNAs against the sodium transepithelial channel (ENaC) was entrapped in optimized hNPs comprising or not poly(ethylenimine) (PEI) as third component. siRNA-loaded hNPs were characterized for encapsulation efficiency, release kinetics, aerodynamic properties, and stability in artificial mucus (AM). The fate and cytotoxicity of hNPs upon aerosolization on a triple cell co-culture model (TCCC) mimicking human epithelial airway barrier were assessed. Finally, the effect of siRNA-loaded hNPs on ENaC protein expression at 72 hours was evaluated in A549 cells. Optimized muco-inert hNPs encapsulating model siRNA with high efficiency were produced. The developed hNPs displayed a hydrodynamic diameter of ∼150 nm, a low polydispersity index, a negative ζ potential close to -25 mV, and a peculiar triphasic siRNA release lasting for 5 days, which slowed down in the presence of PEI. siRNA formulations showed optimal in vitro aerosol performance after delivery with a vibrating mesh nebulizer. Furthermore, small-angle X-ray scattering analyses highlighted an excellent stability upon incubation with AM, confirming the potential of hNPs for direct aerosolization on mucus-lined airways. Studies in TCCC confirmed that fluorescent hNPs are internalized inside airway epithelial cells and do not exert any cytotoxic or acute proinflammatory effect. Finally, a prolonged inhibition of ENaC protein expression was observed in A549 cells upon treatment with siRNA-loaded hNPs. Results demonstrate the great potential of hNPs as carriers for pulmonary delivery of siRNA, prompting toward investigation of their therapeutic effectiveness in severe lung diseases.
Highlights
RNA interference refers to the inhibition of gene expression by small, double-stranded RNA molecules that direct the mRNA machinery to degrade a specific mRNA. siRNA therapeutics have many advantages in terms of clinical applications, the first of which is their virtual potential to silence any gene underlying more-or-less complex pathologies.[1,2] In particular, severe lung diseases, such as lung cancer or cystic fibrosis (CF), nowadays represent a very important area of application for siRNA-based therapies.[3]
In-depth formulation studies were performed on empty hybrid lipid-polymer nanoparticles (hNPs) to establish the optimal ratio between the selected components, that is, poly(lactic-co-glycolic) acid (PLGA), dipalmitoyl phosphatidylcholine (DPPC), and PEI
Independent of the amount of DPPC added to the organic phase and the presence of PEI in the internal water phase, the adopted formulation conditions lead to the formation of hNPs with DH ranging between *135 and *169 nm, low polydispersity index (PI) (
Summary
RNA interference refers to the inhibition of gene expression by small, double-stranded RNA molecules (small interference RNA or siRNA) that direct the mRNA machinery to degrade a specific mRNA. siRNA therapeutics have many advantages in terms of clinical applications, the first of which is their virtual potential to silence any gene underlying more-or-less complex pathologies.[1,2] In particular, severe lung diseases, such as lung cancer or cystic fibrosis (CF), nowadays represent a very important area of application for siRNA-based therapies.[3]. Amid advanced drug delivery systems, the encapsulation of siRNA into colloidal carriers is considered a very promising formulation approach for inhaled treatment of severe lung diseases.[4,5] In this context, biodegradable poly(lactic-co-glycolic) acid (PLGA) nanoparticles (NPs) are gaining considerable interest since they may provide protection of the therapeutic cargo from enzymatic degradation, prolonged release (i.e., decreased number of administrations), and improved retention in the lungs.[6,7] particle engineering with excipients aimed to overcome the mucus-lined human lung epithelial barrier is essential.[8,9]. Conclusions: Results demonstrate the great potential of hNPs as carriers for pulmonary delivery of siRNA, prompting toward investigation of their therapeutic effectiveness in severe lung diseases
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
