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Cited by 289RNA Interference and Gene DeliveryImmunotherapy and Immune ResponsesSARS-CoV-2 and COVID-19 Research

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An Update on Self-Amplifying mRNA Vaccine Development

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Blakney, Anna K., Ip, Shell, Geall, Andrew J.·2021·Vaccines
DOI 10.3390/vaccines9020097PMC PMC7911542

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Abstract

This review will explore the four major pillars required for design and development of an saRNA vaccine: Antigen design, vector design, non-viral delivery systems, and manufacturing (both saRNA and lipid nanoparticles (LNP)). We report on the major innovations, preclinical and clinical data reported in the last five years and will discuss future prospects.

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Figure 1 A comparison of vaccine platforms including vaccines derived from the virus itself and are formulated as a part or whole modified version of the virus (left) and nucleic acid vaccines, such as self-amplifying RNA vaccines (right). Nucleic acid vaccines are derived from knowledge of the viral genome, where glycoproteins are encoded into nucleic acids and delivered with either a synthetic carrier such as a lipid nanoparticle or an inert viral delivery system such as adenoviruses. The encoded antigen sequences are then expressed by the host cells.
Figure 1 A comparison of vaccine platforms including vaccines derived from the virus itself and are formulated as a part or whole modified version of the virus (left) and nucleic acid vaccines, such as self-amplifying RNA vaccines (right). Nucleic acid vaccines are derived from knowledge of the viral genome, where glycoproteins are encoded into nucleic acids and delivered with either a synthetic carrier such as a lipid nanoparticle or an inert viral delivery system such as adenoviruses. The encoded antigen sequences are then expressed by the host cells.CC BY · PubMed Central
Figure 2 The Four Pillars of successful saRNA vaccine development. The antigens, vectors, delivery and manufacturing each represent modular components that need to be combined to make a successful drug product. Each pillar has its set of design and development considerations and associated technologies that are explored in this review.
Figure 2 The Four Pillars of successful saRNA vaccine development. The antigens, vectors, delivery and manufacturing each represent modular components that need to be combined to make a successful drug product. Each pillar has its set of design and development considerations and associated technologies that are explored in this review.CC BY · PubMed Central
Figure 3 A timeline of innovations that have contributed to the development of saRNA vaccines and associated technologies. These include advances in technologies associated with each of the Four Pillars of a successful saRNA vaccine [27,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56].
Figure 3 A timeline of innovations that have contributed to the development of saRNA vaccines and associated technologies. These include advances in technologies associated with each of the Four Pillars of a successful saRNA vaccine [27,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56].CC BY · PubMed Central
Figure 4 A comparison of mRNA vectors. Both conventional (A) and self-amplifying (B) mRNAs share basic elements including a cap, 5′ UTR, 3’ UTR, and poly(A) tail of variable length. Self-amplifying RNA (saRNA) also encode four non-structural proteins (nsP1–4) and a subgenomic promoter derived from the genome of the alphavirus. nsP1–4 encode a replicase responsible for amplification of the saRNA that enable lower doses than non-replicating mRNA.
Figure 4 A comparison of mRNA vectors. Both conventional (A) and self-amplifying (B) mRNAs share basic elements including a cap, 5′ UTR, 3’ UTR, and poly(A) tail of variable length. Self-amplifying RNA (saRNA) also encode four non-structural proteins (nsP1–4) and a subgenomic promoter derived from the genome of the alphavirus. nsP1–4 encode a replicase responsible for amplification of the saRNA that enable lower doses than non-replicating mRNA.CC BY · PubMed Central
Figure 5 Mechanism of self-amplifying mRNA. (1) Following delivery to the cytoplasm, translation of the saRNA produces the non-structural proteins 1–4 (nsP 1–4) that form the (RDRP). (2) RDRP is responsible for replication of the saRNA producing copies of the saRNA. Multiple copies of the subgenomic RNA (3) are hence produced from each saRNA originally delivered. This leads to translation of many more copies of the antigen (4) when compared to a non-amplifying RNA (5).
Figure 5 Mechanism of self-amplifying mRNA. (1) Following delivery to the cytoplasm, translation of the saRNA produces the non-structural proteins 1–4 (nsP 1–4) that form the (RDRP). (2) RDRP is responsible for replication of the saRNA producing copies of the saRNA. Multiple copies of the subgenomic RNA (3) are hence produced from each saRNA originally delivered. This leads to translation of many more copies of the antigen (4) when compared to a non-amplifying RNA (5).CC BY · PubMed Central
Figure 6 Non-viral saRNA delivery systems. Lipid-, polymer-, and emulsion-based delivery systems all use cationic groups to mediate condensation of the anionic RNA as well as delivery across the cell membrane. LNP systems, which have been found to be the most potent vaccine formulatinos, utilize a pH-sensitive ionizable cationic lipids and are taken up in cells through receptor-mediated endocytosis. In the endosome, the lower pH environment ionizes the cationic lipids, which then interacts electrostatically with anionic lipids in the endosomal membrane. These ion pairs cause a phase transition into a porous hexagonal phase (HII) that disrupts the endosome and facilitates release of the RNA into the cytoplasm.
Figure 6 Non-viral saRNA delivery systems. Lipid-, polymer-, and emulsion-based delivery systems all use cationic groups to mediate condensation of the anionic RNA as well as delivery across the cell membrane. LNP systems, which have been found to be the most potent vaccine formulatinos, utilize a pH-sensitive ionizable cationic lipids and are taken up in cells through receptor-mediated endocytosis. In the endosome, the lower pH environment ionizes the cationic lipids, which then interacts electrostatically with anionic lipids in the endosomal membrane. These ion pairs cause a phase transition into a porous hexagonal phase (HII) that disrupts the endosome and facilitates release of the RNA into the cytoplasm.CC BY · PubMed Central
Figure 7 A comparison of vaccine drug product manufacturing processes for egg- and cell-based manufacturing of conventional vaccines, as well as vaccines produced from viral genome sequence information such as the RNA, protein subunit, and viral vectored DNA vaccines against SARS-CoV-2 from Moderna, Novavax, and Johnson & Johnson respectively [130,131,132,133,134,135,136,137,138,139,140,141,142,143]. RNA vaccines offer a cell-free manufacturing process that is responsible for many advantages of the platform, allowing facile and rapid vaccine manufacturing. Moderna’s mRNA vaccine against SARS-CoV-2 (mRNA-1273) began clinical trials just 63 days following the publication of the SARS-CoV-2 genome. * For comparative purposes, we have included historical timelines for the flu pandemic vaccines for egg and cell culture production, but it should be noted that large efficacy trials are not required for these vaccines since they are licensed based on a correlate of protection (hemagglutination inhibition (HI) antibody responses).
Figure 7 A comparison of vaccine drug product manufacturing processes for egg- and cell-based manufacturing of conventional vaccines, as well as vaccines produced from viral genome sequence information such as the RNA, protein subunit, and viral vectored DNA vaccines against SARS-CoV-2 from Moderna, Novavax, and Johnson & Johnson respectively [130,131,132,133,134,135,136,137,138,139,140,141,142,143]. RNA vaccines offer a cell-free manufacturing process that is responsible for many advantages of the platform, allowing facile and rapid vaccine manufacturing. Moderna’s mRNA vaccine against SARS-CoV-2 (mRNA-1273) began clinical trials just 63 days following the publication of the SARS-CoV-2 genome. * For comparative purposes, we have included historical timelines for the flu pandemic vaccines for egg and cell culture production, but it should be noted that large efficacy trials are not required for these vaccines since they are licensed based on a correlate of protection (hemagglutination inhibition (HI) antibody responses).CC BY · PubMed Central
Figure 8 Schematic diagram of the manufacturing process for the RNA drug substance. The process involves a cell-free enzymatic in-vitro transcription reaction followed by purification to remove the DNA template, followed by tangential flow filtration (TFF) for buffer exchange and concentration, followed by sterile filtration through a 0.2 µm filter.
Figure 8 Schematic diagram of the manufacturing process for the RNA drug substance. The process involves a cell-free enzymatic in-vitro transcription reaction followed by purification to remove the DNA template, followed by tangential flow filtration (TFF) for buffer exchange and concentration, followed by sterile filtration through a 0.2 µm filter.CC BY · PubMed Central

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Research works printed but not linked: 3

  1. Mulligan, 2020, Phase I/II study of COVID-19 RNA vaccine BNT162b1 in adults, Nature, 586, 589, 10.1038/s41586-020-2639-4 · DOI · Paper page
  2. Walsh, 2020, Safety and Immunogenicity of Two RNA-Based Covid-19 Vaccine Candidates, N. Engl. J. Med., 383, 2439, 10.1056/NEJMoa2027906 · DOI · Paper page
  3. Alnylam Pharmaceuticals Press (2018). Alnylam Announces First-Ever FDA Approval of an RNAi Therapeutic, ONPATTRO (Patisiran) for the Treatment of the Polyneuropathy of Hereditary Transthyretin-Mediated Amyloidosis in Adults, Alnylam Pharmaceuticals Press.

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Metadata

Authors
Blakney, A. K., Ip, S., & Geall, A. J.
Affiliations
  • University of British Columbia (CA)
  • Precision Nanosystems (Canada) (CA)
Corresponding author
Blakney, Anna K.
ORCID
Venue
Vaccines
Publisher
MDPI AG
Published
2021-01-28
Volume
9
Issue
2
Pages
97
ISSN
2076-393X (ISSN-L)
DOI
10.3390/vaccines9020097
PMC
PMC7911542
OpenAlex
W3112388253
Cited by
288
References
170
Citations by year
2026: 48 · 2025: 55 · 2024: 53 · 2023: 49 · 2022: 50 · 2021: 33
FWCI
4.85
Type
journal article
Language
en
Open access
Licence
Locations
  • Vaccines (publishedVersion, cc-by)
  • Preprints.org (acceptedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • cIRcle (University of British Columbia) (submittedVersion, cc-by)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • Open Collections
Funders
H2020 Marie Skłodowska-Curie Actions (794059)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
  • Life Sciences › Immunology and Microbiology › Immunology › Immunotherapy and Immune Responses
  • Health Sciences › Medicine › Infectious Diseases › SARS-CoV-2 and COVID-19 Research
Keywords
self-amplifying RNA vaccine, antigen design, vector design, lipid nanoparticles, vaccine production

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