Development of New Formulation Dry Powder for Pulmonary Delivery Using Amino Acids to Improve Stability

Abstract

Cationic polymers are being studied as non-viral gene delivery vectors. Poly{N-[N-(2-aminoethyl)-2-aminoethyl]aspartamide} (PAsp(DET)) and their block copolymers with poly(ethylene glycol), PEG-PAsp(DET), have been reported as efficient biodegradable non-viral vectors which form a polyplex with plasmid DNA (pDNA). However, the polyplexes are not stable because PAsp(DET) and PEG-PAsp(DET) are easily subjected to hydrolysis; therefore, they need to be prepared on site. In this study, using the biodegradable polycations as non-viral vectors, PAsp(DET) and PEG-PAsp(DET), we investigated the effects of L-leucine (Leu) on the polyplex. We prepared solutions and dry powders with and without Leu. Both dry powders had large and porous particles and Leu acted as a dispersing agent. The transfection activity of the sample solutions decreased within a month. However, the decrease in the transfection activity was partially suppressed by the dry powder with Leu at 5 and 25°C at 3 months. Furthermore, transfection experiments revealed that Leu exhibited a pDNA-stabilizing effect in the solution and dry powder. Similar results were observed for pDNA integrity, where a polyplex was formed in the dry powder. The results suggest that Leu is a candidate stabilizer to protect pDNA from degradation.

Biodegradable Poly(Amino Ester) with Aromatic Backbone as Efficient Nonviral Gene Delivery Vectors

“…Safe and efficient delivery of genetic material into target cells remains a challenging aspect of gene therapy [ 3 ]. Although viral vectors are considered to have high gene transfer efficiency, their clinical applications have been greatly restricted by a number of key issues such as immunogenicity, insertional mutagenesis, oncogenic effects, and toxicity [ 4 , 5 ]. As safer alternatives, non-viral gene vectors have received increasing attention for their easy preparation and modification [ 6 , 7 , 8 ].…”

Inhalation delivery technology for genome-editing of respiratory diseases

“…On the other hand, while the potential of preparing pDNA dry powder by SFD has been briefly explored by Kuo et al in 2004 [114], it was not until several years later when Mohri et al attempted to develop and optimise the formulations of chitosanconjugated pDNA [115]. Subsequent investigations focused on studying the effect of bovine serum albumin and the amino acid leucine as lyoprotectant during freeze drying [116,117], as well as the use of alternative biodegradable polycation as the transfection vector [118]. Liang et al compared the performances of inhalable dry powders of pDNA conjugated to cationic pH-responsive peptides prepared by SD and SFD, with a more thorough evaluation on their aerodynamic properties [119].…”

Naked pDNA Inhalation Powder Composed of Hyaluronic Acid Exhibits High Gene Expression in the Lungs

“…The addition of Leu to the HA powders may promote favorable inhalation characteristics and an accompanying improvement in gene expression. Referring to our previous studies, 8,23 we administered 10 μg of pDNA to mice. The mass of the powder administered to a mouse was 0.5 mg because of the limitation of the size of the administration apparatus.…”

“…20,21 However, gene inhalation solution for nebulizers has some pharmaceutical issues; it is necessary to prepare them just before use, because the therapeutic gene is generally labile in a solution. DPIs have attracted attention to solve these problems inherent to gene inhalation solution, because a gene in a dry formulation is more stable than that in a solution 22,23 Morphology and particle size are important factors for the development of inhalation powder. 5,24 Previously, we reported a stable gene inhalation powder prepared by precipitation in supercritical carbon dioxide and spray-freeze-drying (SFD); 8,9 it exhibited gene expression in the lungs of mice following pulmonary administration.…”

“…Nebulizer formulations containing siRNA have been used in a clinical trial of gene therapy. , However, gene inhalation solution for nebulizers has some pharmaceutical issues; it is necessary to prepare them just before use, because the therapeutic gene is generally labile in a solution. DPIs have attracted attention to solve these problems inherent to gene inhalation solution, because a gene in a dry formulation is more stable than that in a solution , …”