Development of an alcohol dilution-lyophilization method for preparing lipid nanoparticles containing encapsulated siRNA
Open accessAbstract
Systems for delivering nucleic acids are now fundamental technologies for realizing personalized medicine. Among the various nucleic acid delivery systems that are currently available, lipid-nanoparticles (LNPs) that contain short interfering RNA (siRNA) have been extensively investigated for clinical applications. LNPs are generally prepared by an alcohol dilution method. In this method, it is necessary to remove the alcohol and then concentrate the LNP sample before they can be used. In this study, we report on the development of an “alcohol dilution–lyophilization method” for preparing siRNA-encapsulating LNPs. This method involves the use of a freeze-drying (lyophilization) method to remove the residual alcohol and to simultaneously concentrate the preparation. At first, the compositions of cryoprotectants and polyethylene glycol (PEG)-lipids that were used were optimized from the point of view of particle stabilization. A combination of sucrose and 1-(monomethoxy polyethyleneglycol5000)-2,3-dimyristoylglycerol (DMG-PEG5000) was found to have the most efficient cryoprotective activity for the LNPs. The knockdown efficiency of the LNP prepared by the alcohol dilution–lyophilization method was comparable to that of an LNP prepared by the conventional ultrafiltration method.
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Shirane et al. found no difference between the gene knockdown efficiency of freshly prepared and reconstituted freeze-dried siRNA-LNP formulations in vivo.
“Shirane et al. freeze dried the ethanol-containing siRNA-LNP dispersion obtained immediately after LNP formation and found no difference between the gene knockdown efficiency of the freshly prepared (ethanol removed by ultrafiltration) and the reconstituted freeze-dried formulations in vivo (Shirane et al., 2018).”
From mRNA-lipid nanoparticle COVID-19 vaccines: structure and stability (2021) · cites this paper as Shirane et al., 2018
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- Belliveau NM, Huft J, Lin PJ, Chen S, Leung AK, Leaver TJ, Wild AW, Lee JB, Taylor RJ, Tam YK, Hansen CL, Cullis PR. Microflu- idic synthesis of highly potent limit-size lipid nanoparticles for in vivo delivery of siRNA. Molecular Therapy Nucleic Acids, 1, e37 (2012). · DOI · Paper page
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- Authors
- Shirane, D., Tanaka, H., Nakai, Y., Yoshioka, H., & Akita, H.
- Affiliations
- Chiba University (JP)
- NOF Corporation (Japan) (JP)
- Venue
- Biological and Pharmaceutical Bulletin
- Publisher
- Pharmaceutical Society of Japan
- Published
- 2018-08-01
- Volume
- 41
- Issue
- 8
- Pages
- 1291-1294
- ISSN
- 0918-6158 (ISSN-L)
- 1347-5215
- OpenAlex
- W2886823346
- Cited by
- 30
- References
- 14
- Citations by year
- 2026: 5 · 2025: 3 · 2024: 2 · 2023: 12 · 2022: 5 · 2021: 2 · 2020: 1
- FWCI
- 0.27
- Type
- journal article
- Language
- en
- Open access
- diamond
- doi.org
- Locations
- Biological and Pharmaceutical Bulletin (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Asahi Glass Foundation
- Chiba University
- Japan Society for the Promotion of Science (17K19473, CREST, 17H06558, 18K18377)
- Core Research for Evolutional Science and Technology (JPMJCR17H1)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Physical Sciences › Materials Science › Biomaterials › Nanoparticle-Based Drug Delivery
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmaceutical Science › Advancements in Transdermal Drug Delivery
- Keywords
- lipid nanoparticles, siRNA delivery, freeze-drying, cryoprotectants, sucrose, particle stabilization, knockdown efficiency, nucleic acid delivery
- MeSH terms
- Animals, Cholesterol, Cryoprotective Agents, Drug Compounding, Factor VII, Freeze Drying, Liver, Male, Mice, Inbred ICR, Phosphatidylcholines, Polyethylene Glycols, Sucrose, Triglycerides, Myristic Acid, 1-Butanol, alpha-Tocopherol, RNA, Small Interfering, Nanoparticles
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