Optimized pulmonary gene transfection in mice by spray–freeze dried powder inhalation

Mohri, Kunihiko; Okuda, Tomoyuki; Mori, Asami; Danjo, Kazumi; Okamoto, Hirokazu

doi:10.1016/j.jconrel.2010.02.018

Cited by 75 publications

(48 citation statements)

References 39 publications

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“…RESULTS AND DISCUSSION Table 1 summarizes the formulation of each DPI studied. Similar to a previous report by Okamoto and colleagues 20,28) ICG was also encapsulated in the DPI formulation to confirm delivery into the lung. Particle size and zeta potential were measured to evaluate the physicochemical properties of the DPI formulations.…”

Section: ) Pcmv-luc Was Amplified In the Dh5αmentioning

confidence: 74%

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Bovine Serum Albumin as a Lyoprotectant for Preparation of DNA Dry Powder Formulations Using the Spray-Freeze Drying Method

Tsukamoto

Okuda

Okamoto

et al. 2012

Biological & Pharmaceutical Bulletin

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The development of efficient and selective therapeutic gene delivery methods is a potential medical treatment for intractable diseases. Dry powder inhalers (DPIs) can efficiently deliver drugs locally to the lung. Many reports discuss preparation methods for DPIs. Spray-freeze drying is a method by which highly porous particulates can be prepared. However, altered physical properties after preparation may result in changes in gene expression. In this study, bovine serum albumin (BSA) was added as a lyoprotectant, and 1,2-dioleoyl-3-trimethylammonium propane/cholesterol liposomes/pCMV-Luc DPIs (lipoplex DPIs) were prepared by spray-freeze drying. The mean particle sizes of the lipoplex DPIs prepared without BSA increased by approximately 6.7-fold compared with that of the lipoplexes solution. In contrast, the mean particle sizes of the lipoplex/BSA DPIs increased only slightly. Gene expression was evaluated after the intratracheal administration of the lipoplexes solution, with maximum gene expression observed at 12 h after the administration. In contrast, maximum gene expression of the lipoplex/BSA DPIs occurred at 6 h after administration. The gene expression associated with the lipoplex DPIs was significantly lower compared with that of the lipoplex/BSA DPIs at 6 (p<0.01), 12 (p<0.01), and 24 h (p<0.05). These variances may be due to the difference in mean particle size between the DPI formulations. The results suggest that BSA is a useful lyoprotectant for dry powder formulation preparations of DNA using the spray-freeze drying method, given that the preparation results in minimal variation of physical properties and gene expression.Key words dry powder inhaler; spray-freeze drying; lyoprotectant; bovine serum albumin; pulmonary gene delivery; lipoplex Gene therapy is expected to be a cure for intractable diseases. However, efficient and selective delivery of therapeutic genes to the target cells is necessary for gene therapy to be effective.1) Although viral vectors may show high gene expression, toxicity as a result of immunogenicity is a major problem associated with their use. Therefore, non-viral vectors have received attention because of their relatively low toxicity. 2)Many researchers have demonstrated that cationic liposomes show high gene expression and low toxicity.3-5) Lung diseases such as lung cancer and cystic fibrosis are intractable, and therefore, the realization of efficient gene therapy to treat such conditions is essential. However, the development of delivery methods that enable the local delivery of therapeutic genes to the lung is needed.The aerosolization of gene vectors is an efficient delivery method, and many related studies have been carried out. 6,7)However, it is necessary for aerosols to be sprayed immediately because the stability of gene vectors in aqueous solution is low. 8) Alternatively, dry powder inhalers (DPIs) are widely used and accepted as an effective system for pulmonary drug delivery. DPIs have many advantages, including increased shelf life of drugs including pla...

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Section: ) Pcmv-luc Was Amplified In the Dh5αmentioning

confidence: 74%

“…Highly porous dry powders can be obtained by this method. 19,20) However, with the spray-freeze drying method, gene expression may be decreased due to the increase in particle size of the gene vectors during the process of freeze-drying. Many studies report that it is possible to maintain physical properties during freeze-drying.…”

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confidence: 99%

Bovine Serum Albumin as a Lyoprotectant for Preparation of DNA Dry Powder Formulations Using the Spray-Freeze Drying Method

Tsukamoto

Okuda

Okamoto

et al. 2012

Biological & Pharmaceutical Bulletin

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“…28) As an alternative one-step method for inhalable dry siRNA/chitosan powders, a spray-freeze-drying technique might be applied since a porous dry pDNA/chitosan powder could be stably prepared as shown in our previous report. 44) …”

Section: Discussionmentioning

confidence: 99%

Gene Silencing in a Mouse Lung Metastasis Model by an Inhalable Dry Small Interfering RNA Powder Prepared Using the Supercritical Carbon Dioxide Technique

Okuda

Kito

Oiwa

et al. 2013

Biological & Pharmaceutical Bulletin

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In this study, a novel dry small interfering RNA (siRNA) powder for inhalation, containing chitosan and mannitol, was prepared using the supercritical carbon dioxide (CO 2 ) technique. Although the siRNA/chitosan powder was difficult to disperse because of a long needle-like structure, it could be reduced to fragments of 10-20 µm by manual grinding, which allowed for administration into mice. Electrophoresis revealed that the supercritical CO 2 technique and manual grinding didn't greatly affect the integrity of the siRNA. Furthermore, the siRNA was more stable in the lungs than in blood, suggesting the utility of pulmonary delivery. Biodistribution experiments using Cy5.5-labeled siRNA demonstrated that pulmonary administration of the powder achieved a prolonged exposure of the siRNA/chitosan complex on the lung epithelial surface at a higher concentration. For the evaluation of the in-vivo gene silencing effect of the siRNA/chitosan powder, mice bearing colon26/Luc cells were used. The powder significantly inhibited the increase in luminescence intensity in the lungs, but the siRNA/chitosan solution and a non-specific dry siRNA/chitosan powder didn't, indicating the effective and specific gene silencing against the tumor cells metastasized in the lungs of mice by the siRNA/chitosan powder. These results strongly indicate that inhalable dry siRNA powders have the possibility of effective pulmonary gene silencing and that the supercritical CO 2 technique can be applied to the production.Key words small interfering RNA; inhalable dry powder; supercritical fluid technique; pulmonary gene delivery; chitosan RNA interference (RNAi), first reported by Fire et al., is a unique cellular phenomenon in which double-stranded RNA (dsRNA) regulates gene expression, contributing to cellular defenses against viral infections and transposon expansion.

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“…26) The powders are expected to be applicable for clinical use as inhaled formulations since their characteristic morphology allows for high dispersibility and superior lung delivery. Due to the increased specific surface area of the powders, however, moisture absorption may readily occur, subsequently leading to a decrease in output efficiency and losses of their inhalation potential.…”

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Development of Spray-Freeze-Dried Powders for Inhalation with High Inhalation Performance and Antihygroscopic Property

2016

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Spray-freeze-drying (SFD) is a unique powderization technique to produce highly porous dry powders with a low density. The characteristic morphology can markedly contribute to the superior inhalation performances of SFD powders. Due to the increased specific surface area of the powders, however, moisture adsorption may readily occur, subsequently leading to losses of their inhalation potentials. In this study, hydrophobic amino acids were newly applied as pharmaceutical excipients to obtain SFD powders with both a favorable inhalation performance and antihygroscopic property. SFD powders composed of several hydrophobic amino acids were prepared. The morphology, particle size distribution, and crystallinity of the prepared powders were evaluated by scanning electron micrography, laser diffraction, and X-ray powder diffraction, respectively. The inhalation characteristics of the SFD powders were examined using a twin-stage liquid impinger equipped with an inspiratory pattern simulator and devices. To investigate their antihygroscopicity, moreover, the SFD powders were stored under a humidified condition to assess the morphology, crystallinity, and inhalation performance as described above. It was demonstrated that a SFD powder composed of L-leucine, L-isoleucine, or L-phenylalanine showed a superior inhalation performance, which was sufficiently maintained after storage under the humidified condition, strongly indicating their antihygroscopicity. These results indicated that the hygroscopicity of SFD powders can be effectively improved by the application of hydrophobic amino acids as excipients. Key words dry powder inhaler (DPI); spray-freeze-drying (SFD); hydrophobic amino acid; inhalation performance; antihygroscopic propertyInhalants that can deliver a drug deep into the lungs directly and noninvasively have attracted attention as dosage forms for the therapy of not only local diseases such as asthma, chronic obstructive pulmonary disease, and cystic fibrosis but also systemic diseases such as diabetes. [1][2][3] There are three kinds of inhalant on the market: pressurized metered-dose inhalers (pMDIs), inhalation solutions, and dry powder inhalers (DPIs). DPIs have many advantages compared to the other two forms: (1) no propellant or battery is required, (2) the device is small, and (3) powders are dispersed on inhalation by a patient, minimizing the mismatch between the timing of inhalation and drug dispersal. 4,5) The powders for DPIs should have an appropriate mass median aerodynamic diameter (MMAD) suitable for inhalation and small adhesion cohesiveness. The commonly recognized optimum MMAD is in the range of 1-6 µm.4) However, the adhesion cohesiveness of such small particles is usually high, and they easily aggregate to form large particles that are not suitable for inhalation. 6) To solve this problem, the small particles of an active ingredient are generally mixed with large inert carrier particles (50-100 µm) such as lactose monohydrate to prevent the small particles of an active ingredient from unde...

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Optimized pulmonary gene transfection in mice by spray–freeze dried powder inhalation

Cited by 75 publications

References 39 publications

Bovine Serum Albumin as a Lyoprotectant for Preparation of DNA Dry Powder Formulations Using the Spray-Freeze Drying Method

Bovine Serum Albumin as a Lyoprotectant for Preparation of DNA Dry Powder Formulations Using the Spray-Freeze Drying Method

Gene Silencing in a Mouse Lung Metastasis Model by an Inhalable Dry Small Interfering RNA Powder Prepared Using the Supercritical Carbon Dioxide Technique

Development of Spray-Freeze-Dried Powders for Inhalation with High Inhalation Performance and Antihygroscopic Property

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