Production, Characterization, and Assessment of Permanently Cationic and Ionizable Lipid Nanoparticles for Use in the Delivery of Self-Amplifying RNA Vaccines
Open accessAbstract
Africa bears the highest burden of infectious diseases, yet the continent is heavily reliant on First World countries for the development and supply of life-saving vaccines. The COVID-19 pandemic was a stark reminder of Africa’s vaccine dependence and since then great interest has been generated in establishing mRNA vaccine manufacturing capabilities on the African continent. Herein, we explore alphavirus-based self-amplifying RNAs (saRNAs) delivered by lipid nanoparticles (LNPs) as an alternative to the conventional mRNA vaccine platform. The approach is intended to produce dose-sparing vaccines which could assist resource-constrained countries to achieve vaccine independence. Protocols to synthesize high-quality saRNAs were optimized and in vitro expression of reporter proteins encoded by saRNAs was achieved at low doses and observed for an extended period. Permanently cationic or ionizable LNPs (cLNPs and iLNPs, respectively) were successfully produced, incorporating saRNAs either exteriorly (saRNA-Ext-LNPs) or interiorly (saRNA-Int-LNPs). DOTAP and DOTMA saRNA-Ext-cLNPs performed best and were generally below 200 nm with good PDIs (<0.3). DOTAP and DDA saRNA-Int-cLNPs performed optimally, allowing for saRNA amplification. These were slightly larger, with higher PDIs as a result of the method used, which will require further optimization. In both cases, the N:P ratio and lipid molar ratio had a distinct effect on saRNA expression kinetics, and RNA was encapsulated at high percentages of >90%. These LNPs allow the delivery of saRNA with no significant toxicity. The optimization of saRNA production and identification of potential LNP candidates will facilitate saRNA vaccine and therapeutic development. The dose-sparing properties, versatility, and manufacturing simplicity of the saRNA platform will facilitate a rapid response to future pandemics.
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The current understanding of the field is that incorporating modified NTPs into saRNA abrogates downstream efficacy.
“However, the current understanding of the field is that incorporating modified NTPs into saRNA abrogates downstream efficacy, as reported by multiple independent groups5,19–28.”
From Complete substitution with modified nucleotides in self-amplifying RNA suppresses the interferon response and increases potency (2025) · cites this paper as [27]
The Discussion reports strong reporter-protein expression from saRNA made without modifications used in conventional mRNA, but does not say what happens when modified NTPs are incorporated into saRNA. The Results discuss NTP concentration and capping, not a comparison of modified and unmodified NTPs. Searches for modified NTPs, modified nucleotides, and effects on replication or efficacy found no passage addressing the claim. (LLM, conf=0.75)
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References 28 · Cited by 17
- Authors
- Kairuz, D., Samudh, N., Ely, A., Arbuthnot, P., & Bloom, K.
- Affiliations
- South African Medical Research Council (ZA)
- Corresponding author
- Bloom, Kristie
- Venue
- Pharmaceutics
- Publisher
- MDPI AG
- Published
- 2023-04-07
- Volume
- 15
- Issue
- 4
- Pages
- 1173
- ISSN
- 1999-4923 (ISSN-L)
- PMC
- PMC10143526
- OpenAlex
- W4362699318
- Cited by
- 17
- References
- 28
- Citations by year
- 2026: 3 · 2025: 9 · 2024: 3 · 2023: 2
- FWCI
- 1.51
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- Pharmaceutics (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Funders
- South African National Research Foundation (120383, 21/74, 20/46, KBLM022)
- Poliomyelitis Research Foundation (120383, 21/74, 20/46, KBLM022)
- South African Medical Research Council (120383, 21/74, 20/46, KBLM022)
- Medicines Patent Pool (120383, 21/74, 20/46, KBLM022)
- 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
- self-amplifying RNA vaccine, lipid nanoparticles, ionizable lipids, N:P ratio
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