Inhalation Properties and Stability of Nebulized Naked siRNA Solution for Pulmonary Therapy

Abstract

The use of naked unmodified small interfering RNA (N-siRNA) without vector has previously been investigated as a pulmonary therapy. However, little is known regarding stabilities and aerodynamic particle sizes of N-siRNA-containing droplets; nebulizers have not yet been optimized for N-siRNA solutions. Thus, in this study, we investigated the feasibility of inhaled N-siRNA solutions for pulmonary therapy using nebulization. Various nebulizers and N-siRNA concentrations were assessed in terms of siRNA integrity after nebulization, and inhalation properties including aerodynamic particle size were examined. In comparison with ultrasonic-, air-jet-, and vibrating-mesh nebulizers, N-siRNA integrity was not affected by nebulization. Thus, in further experiments, performances of N-siRNA aerosols with different nebulizers and N-siRNA concentrations were evaluated and screened using an aerodynamic particle sizer (APS) which employed the time-of-flight principle or a cascade impactor. Mean mass aerodynamic diameters of N-siRNA-containing droplets from vibrating-mesh nebulizers tended to decrease with increasing N-siRNA concentrations, reflecting the influence of N-siRNA solutions on surface tension, as indicated by contact angles. These data indicate the utility of APS instruments for investigating the nebulized characteristics of expensive drugs including siRNAs and may facilitate the development of N-siRNA inhalation formulations.Key words small interfering RNA (siRNA); vibrating-mesh nebulizer; aerodynamic particle size; surface tension; contact angle Small interfering RNA (siRNA) has a great therapeutic potential as a tool for the post-transcriptional silencing of target gene expression. 1) Recently, the field of siRNA delivery has rapidly progressed, and the local pulmonary delivery of siRNA has been investigated for various lung diseases. 2,3)Inhalation is the most popular noninvasive method for delivering therapeutic siRNA agents to the lungs 4); local siRNA inhalation therapy reduced virus titers in the lung and attenuated local pulmonary chemokine production after acute lung injury and infection.5,6) However, naked (unmodified) siRNA (N-siRNA) is susceptible to nuclease degradation; siRNA delivery systems facilitating gene silencing efficiency, including those using cationic materials, can have cytotoxic effects. N-siRNA has been delivered with some success to the lung, although systemic delivery of N-siRNA generally fails to produce significant gene silencing effects.3,7) N-siRNA lacks delivery vectors that produce toxicity and inflammation; thus, N-siRNA formulation for pulmonary delivery offers advantages for safety and simplicity. However, few developments of N-siRNA formulations have been reported. Direct pulmonary N-siRNA delivery has been achieved in humans following the inhalation of aerosols generated by inhalers or nebulizers. Three types of inhalation devices are currently available for pulmonary drug delivery, including metered dose inhalers, dry powder inhalers, and nebulizers. Liquid nebulization e...

Stability of Naked Nucleic Acids under Physical Treatment and Powder Formation: Suitability for Development as Dry Powder Formulations for Inhalation

“…In contrast, the structural and functional integrities of siRNA were preserved after all the physical treatments and powder formation. The high stability of siRNA observed in the present study is consistent with previous findings [ 21 , 22 , 23 , 24 , 35 , 36 ]. The physical stresses caused by SD in the present study may have been weaker than those reported by Wu et al, who showed that the functional integrity of siRNA decreased in SD powders with increases in the inlet temperature (80 to 200 °C) and atomizing airflow (473 to 742 L/h) [ 24 ].…”

siRNA Therapeutics against Respiratory Viral Infections—What Have We Learned for Potential COVID‐19 Therapies?

“…[ 66 ] Moreover, naked siRNA lends itself easily to a simple inhaler‐based application as siRNA solutions can be aerosolized without integrity loss by various types of nebulizers. [ 67 ] Nanoparticle suspensions of siRNA, on the other hand, are more prone to aggregation and other physical changes when exposed to shear forces during the aerosolization process. [ 68 ] While chemically modified siRNAs have increased stability in vivo, they still cannot necessarily pass through intact cell membranes and may activate an immune response.…”

Oligonucleotide therapeutics in neurodegenerative diseases

“…Here we provide a brief discussion on delivery approaches. Naked delivery of siRNAs has been demonstrated to be effective for localized treatments, including in the eye for age-related macular degeneration [24,25], by inhalation for pulmonary disease [7], and transdermally for various skin conditions [26]. Antisense oligonucleotides are also employed without carriers.…”