ABSTRACT
Although targeting TfR1 to deliver oligonucleotides to skeletal muscle has been demonstrated in rodents, effectiveness and pharmacokinetic/pharmacodynamic (PKPD) properties remained unknown in higher species. We developed antibody-oligonucleotide conjugates (AOCs) towards mice or monkeys utilizing anti-TfR1 monoclonal antibodies (αTfR1) conjugated to various classes of oligonucleotides (siRNA, ASOs and PMOs). αTfR1 AOCs delivered oligonucleotides to muscle tissue in both species. In mice, αTfR1 AOCs achieved a > 15-fold higher concentration to muscle tissue than unconjugated siRNA. A single dose of an αTfR1 conjugated to an siRNA against Ssb mRNA produced > 75% Ssb mRNA reduction in mice and monkeys, and mRNA silencing was greatest in skeletal and cardiac (striated) muscle with minimal to no activity in other major organs. In mice the EC50 for Ssb mRNA reduction in skeletal muscle was >75-fold less than in systemic tissues. Oligonucleotides conjugated to control antibodies or cholesterol produced no mRNA reduction or were 10-fold less potent, respectively. Tissue PKPD of AOCs demonstrated mRNA silencing activity primarily driven by receptor-mediated delivery in striated muscle for siRNA oligonucleotides. In mice, we show that AOC-mediated delivery is operable across various oligonucleotide modalities. AOC PKPD properties translated to higher species, providing promise for a new class of oligonucleotide therapeutics.
Subject(s)
Oligonucleotides, Antisense , Oligonucleotides , Mice , Animals , Antibodies/therapeutic use , RNA, Small Interfering/genetics , RNA, Messenger/genetics , Muscle, SkeletalABSTRACT
INTRODUCTION: Prior to the emergence of SARS-CoV-2, the potential use of mRNA vaccines for a rapid pandemic response had been well described in the scientific literature, however during the SARS-CoV-2 outbreak we witnessed the large-scale deployment of the platform in a real pandemic setting. Of the three RNA platforms evaluated in clinical trials, including 1) conventional, non-amplifying mRNA (mRNA), 2) base-modified, non-amplifying mRNA (bmRNA), which incorporate chemically modified nucleotides, and 3) self-amplifying RNA (saRNA), the bmRNA technology emerged with superior clinical efficacy. AREAS COVERED: This review describes the current state of these mRNA vaccine technologies, evaluates their strengths and limitations, and argues that saRNA may have significant advantages if the limitations of stability and complexities of manufacturing can be overcome. EXPERT OPINION: The success of the SARS-CoV-2 mRNA vaccines has been remarkable. However, several challenges remain to be addressed before this technology can successfully be applied broadly to other disease targets. Innovation in the areas of mRNA engineering, novel delivery systems, antigen design, and high-quality manufacturing will be required to achieve the full potential of this disruptive technology.
Subject(s)
COVID-19 , Vaccines , Humans , COVID-19/prevention & control , SARS-CoV-2/genetics , RNA , RNA, Messenger , Vaccines, SyntheticABSTRACT
RNA technology has recently come to the forefront of innovative medicines and is being explored for a wide range of therapies, including prophylactic and therapeutic vaccines, biotherapeutic protein expression and gene therapy. In addition to conventional mRNA platforms now approved for prophylactic SARS-CoV2 vaccines, synthetic self-replicating RNA vaccines are currently being evaluated in the clinic for infectious disease and oncology. The prototypical srRNA vectors in clinical development are derived from alphaviruses, specifically Venezuelan Equine Encephalitis Virus (VEEV). While non-VEEV alphaviral strains have been explored as single cycle viral particles, their use as synthetic vectors largely remains under-utilized in clinical applications. Here we describe the potential commonalities and differences in synthetic alphaviral srRNA vectors in host cell interactions, immunogenicity, cellular delivery, and cargo expression. Thus, unlike the current thinking that VEEV-based srRNA is a one-size-fits-all platform, we argue that a new drug development approach leveraging panels of customizable, synthetic srRNA vectors will be required for clinical success.
Subject(s)
COVID-19 , Vaccines , Viral Vaccines , Animals , Horses/genetics , RNA, Viral , SARS-CoV-2/genetics , Immunotherapy , Viral Vaccines/geneticsABSTRACT
To prevent SARS-CoV-2 infections and generate long-lasting immunity, vaccines need to generate strong viral-specific B and T cell responses. Previous results from our lab and others have shown that immunizations in the presence of an OX40 agonist antibody lead to higher antibody titers and increased numbers of long-lived antigen-specific CD4 and CD8 T cells. Using a similar strategy, we explored the effect of OX40 co-stimulation in a prime and boost vaccination scheme using an adjuvanted SARS-CoV-2 spike protein vaccine in C57BL/6 mice. Our results show that OX40 engagement during vaccination significantly increases long-lived antibody responses to the spike protein. In addition, after immunization spike protein-specific proliferation was greatly increased for both CD4 and CD8 T cells, with enhanced, spike-specific secretion of IFN-γ and IL-2. Booster (3rd injection) immunizations combined with an OX40 agonist (7 months post-prime) further increased vaccine-specific antibody and T cell responses. Initial experiments assessing a self-amplifying mRNA (saRNA) vaccine encoding the spike protein antigen show a robust antigen-specific CD8 T cell response. The saRNA spike-specific CD8 T cells express high levels of GrzmB, IFN-γ and TNF-α which was not observed with protein immunization and this response was further increased by the OX40 agonist. Similar to protein immunizations the OX40 agonist also increased vaccine-specific CD4 T cell responses. In summary, this study compares and contrasts the effects and benefits of both protein and saRNA vaccination and the extent to which an OX40 agonist enhances and sustains the immune response against the SARS-CoV-2 spike protein.
Subject(s)
COVID-19 , Vaccines , Animals , COVID-19/prevention & control , Humans , Interleukin-2 , Mice , Mice, Inbred C57BL , RNA, Messenger , SARS-CoV-2 , Spike Glycoprotein, Coronavirus , Tumor Necrosis Factor-alphaABSTRACT
Self-amplifying RNA (saRNA) is a next-generation vaccine platform, but like all nucleic acids, requires a delivery vehicle to promote cellular uptake and protect the saRNA from degradation. To date, delivery platforms for saRNA have included lipid nanoparticles (LNP), polyplexes and cationic nanoemulsions; of these LNP are the most clinically advanced with the recent FDA approval of COVID-19 based-modified mRNA vaccines. While the effect of RNA on vaccine immunogenicity is well studied, the role of biomaterials in saRNA vaccine effectiveness is under investigated. Here, we tested saRNA formulated with either pABOL, a bioreducible polymer, or LNP, and characterized the protein expression and vaccine immunogenicity of both platforms. We observed that pABOL-formulated saRNA resulted in a higher magnitude of protein expression, but that the LNP formulations were overall more immunogenic. Furthermore, we observed that both the helper phospholipid and route of administration (intramuscular versus intranasal) of LNP impacted the vaccine immunogenicity of two model antigens (influenza hemagglutinin and SARS-CoV-2 spike protein). We observed that LNP administered intramuscularly, but not pABOL or LNP administered intranasally, resulted in increased acute interleukin-6 expression after vaccination. Overall, these results indicate that delivery systems and routes of administration may fulfill different delivery niches within the field of saRNA genetic medicines.
Subject(s)
COVID-19 , Influenza Vaccines , Nanoparticles , Humans , Lipids , Polymers , RNA , SARS-CoV-2 , Spike Glycoprotein, CoronavirusABSTRACT
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.
ABSTRACT
Venezuelan equine encephalitis virus (VEEV) is a known biological defense threat. A live-attenuated investigational vaccine, TC-83, is available, but it has a high non-response rate and can also cause severe reactogenicity. We generated two novel VEE vaccine candidates using self-amplifying mRNA (SAM). LAV-CNE is a live-attenuated VEE SAM vaccine formulated with synthetic cationic nanoemulsion (CNE) and carrying the RNA genome of TC-83. IAV-CNE is an irreversibly-attenuated VEE SAM vaccine formulated with CNE, delivering a TC-83 genome lacking the capsid gene. LAV-CNE launches a TC-83 infection cycle in vaccinated subjects but eliminates the need for live-attenuated vaccine production and potentially reduces manufacturing time and complexity. IAV-CNE produces a single cycle of RNA amplification and antigen expression without generating infectious viruses in subjects, thereby creating a potentially safer alternative to live-attenuated vaccine. Here, we demonstrated that mice vaccinated with LAV-CNE elicited immune responses similar to those of TC-83, providing 100% protection against aerosol VEEV challenge. IAV-CNE was also immunogenic, resulting in significant protection against VEEV challenge. These studies demonstrate the proof of concept for using the SAM platform to streamline the development of effective attenuated vaccines against VEEV and closely related alphavirus pathogens such as western and eastern equine encephalitis and Chikungunya viruses.
Subject(s)
Encephalitis Virus, Venezuelan Equine/immunology , Encephalomyelitis, Venezuelan Equine/drug therapy , Gene Amplification , Immunogenicity, Vaccine , RNA, Messenger/genetics , Vaccines, Attenuated/therapeutic use , Viral Vaccines/therapeutic use , A549 Cells , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Disease Models, Animal , Emulsions/chemistry , Encephalomyelitis, Venezuelan Equine/virology , Female , Humans , Mice , Mice, Inbred BALB C , Transfection , Viral Vaccines/pharmacology , Virus ReplicationABSTRACT
Plasmodium species produce an ortholog of the cytokine macrophage migration inhibitory factor, PMIF, which modulates the host inflammatory response to malaria. Using a novel RNA replicon-based vaccine, we show the impact of PMIF immunoneutralization on the host response and observed improved control of liver and blood-stage Plasmodium infection, and complete protection from re-infection. Vaccination against PMIF delayed blood-stage patency after sporozoite infection, reduced the expression of the Th1-associated inflammatory markers TNF-α, IL-12, and IFN-γ during blood-stage infection, augmented Tfh cell and germinal center responses, increased anti-Plasmodium antibody titers, and enhanced the differentiation of antigen-experienced memory CD4 T cells and liver-resident CD8 T cells. Protection from re-infection was recapitulated by the adoptive transfer of CD8 or CD4 T cells from PMIF RNA immunized hosts. Parasite MIF inhibition may be a useful approach to promote immunity to Plasmodium and potentially other parasite genera that produce MIF orthologous proteins.
Subject(s)
Adaptive Immunity/drug effects , Antibodies, Protozoan/biosynthesis , Macrophage Migration-Inhibitory Factors/antagonists & inhibitors , Malaria Vaccines/administration & dosage , Malaria/prevention & control , Protozoan Proteins/antagonists & inhibitors , Vaccines, DNA/administration & dosage , Adoptive Transfer , Animals , CD4-Positive T-Lymphocytes/drug effects , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/parasitology , CD8-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/parasitology , Female , Gene Expression , Germinal Center/drug effects , Germinal Center/immunology , Germinal Center/parasitology , Immunologic Memory/drug effects , Interferon-gamma/genetics , Interferon-gamma/immunology , Interleukin-12/genetics , Interleukin-12/immunology , Macrophage Migration-Inhibitory Factors/genetics , Macrophage Migration-Inhibitory Factors/immunology , Malaria/immunology , Malaria/parasitology , Malaria Vaccines/biosynthesis , Mice , Mice, Inbred BALB C , Plasmodium berghei/drug effects , Plasmodium berghei/genetics , Plasmodium berghei/immunology , Protein Isoforms/antagonists & inhibitors , Protein Isoforms/genetics , Protein Isoforms/immunology , Protozoan Proteins/genetics , Protozoan Proteins/immunology , RNA, Protozoan/genetics , RNA, Protozoan/immunology , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/immunology , Vaccines, DNA/biosynthesisABSTRACT
RNA-based vaccines have recently emerged as a promising alternative to the use of DNA-based and viral vector vaccines, in part because of the potential to simplify how vaccines are made and facilitate a rapid response to newly emerging infections. SAM vaccines are based on engineered self-amplifying mRNA (SAM) replicons encoding an Ag, and formulated with a synthetic delivery system, and they induce broad-based immune responses in preclinical animal models. In our study, in vivo imaging shows that after the immunization, SAM Ag expression has an initial gradual increase. Gene expression profiling in injection-site tissues from mice immunized with SAM-based vaccine revealed an early and robust induction of type I IFN and IFN-stimulated responses at the site of injection, concurrent with the preliminary reduced SAM Ag expression. This SAM vaccine-induced type I IFN response has the potential to provide an adjuvant effect on vaccine potency, or, conversely, it might establish a temporary state that limits the initial SAM-encoded Ag expression. To determine the role of the early type I IFN response, SAM vaccines were evaluated in IFN receptor knockout mice. Our data indicate that minimizing the early type I IFN responses may be a useful strategy to increase primary SAM expression and the resulting vaccine potency. RNA sequence modification, delivery optimization, or concurrent use of appropriate compounds might be some of the strategies to finalize this aim.
Subject(s)
Drug Design , Interferon Type I/immunology , RNA, Messenger/immunology , Viral Vaccines/immunology , Adjuvants, Immunologic , Animals , Antibodies, Viral , Antigens/immunology , Imaging, Three-Dimensional/methods , Interferon Type I/biosynthesis , Mice , RNA, Messenger/administration & dosage , RNA, Messenger/physiology , RNA, Viral/immunology , Respiratory Syncytial Viruses/chemistry , Respiratory Syncytial Viruses/immunology , Vaccination , Vaccine Potency , Viral Vaccines/geneticsABSTRACT
Nucleic acid vaccines represent an attractive approach to vaccination, combining the positive attributes of both viral vectors and live-attenuated vaccines, without the inherent limitations of each technology. We have developed a novel technology, the Self-Amplifying mRNA (SAM) platform, which is based on the synthesis of self-amplifying mRNA formulated and delivered as a vaccine. SAM vaccines have been shown to stimulate robust innate and adaptive immune responses in small animals and non-human primates against a variety of viral antigens, thus representing a safe and versatile tool against viral infections. To assess whether the SAM technology could be used for a broader range of targets, we investigated the immunogenicity and efficacy of SAM vaccines expressing antigens from Group A (GAS) and Group B (GBS) Streptococci, as models of bacterial pathogens. Two prototype bacterial antigens (the double-mutated GAS Streptolysin-O (SLOdm) and the GBS pilus 2a backbone protein (BP-2a)) were successfully expressed by SAM vectors. Mice immunized with both vaccines produced significant amounts of fully functional serum antibodies. The antibody responses generated by SAM vaccines were capable of conferring consistent protection in murine models of GAS and GBS infections. Inclusion of a eukaryotic secretion signal or boosting with the recombinant protein resulted in higher specific-antibody levels and protection. Our results support the concept of using SAM vaccines as potential solution for a wide range of both viral and bacterial pathogens, due to the versatility of the manufacturing processes and the broad spectrum of elicited protective immune response.
Subject(s)
Antigens, Bacterial/immunology , RNA, Messenger/biosynthesis , Streptococcal Infections/prevention & control , Streptococcal Vaccines/immunology , Streptococcus agalactiae/immunology , Streptococcus pyogenes/immunology , Animals , Antibodies, Bacterial/blood , Antigens, Bacterial/biosynthesis , Antigens, Bacterial/genetics , Disease Models, Animal , Female , Mice , RNA, Messenger/genetics , Streptococcal Vaccines/administration & dosage , Streptococcal Vaccines/genetics , Streptococcus agalactiae/genetics , Streptococcus pyogenes/geneticsABSTRACT
Current hemagglutinin (HA)-based seasonal influenza vaccines induce vaccine strain-specific neutralizing antibodies that usually fail to provide protection against mismatched circulating viruses. Inclusion in the vaccine of highly conserved internal proteins such as the nucleoprotein (NP) and the matrix protein 1 (M1) was shown previously to increase vaccine efficacy by eliciting cross-reactive T-cells. However, appropriate delivery systems are required for efficient priming of T-cell responses. In this study, we demonstrated that administration of novel self-amplifying mRNA (SAM®) vectors expressing influenza NP (SAM(NP)), M1 (SAM(M1)), and NP and M1 (SAM(M1-NP)) delivered with lipid nanoparticles (LNP) induced robust polyfunctional CD4 T helper 1 cells, while NP-containing SAM also induced cytotoxic CD8 T cells. Robust expansions of central memory (TCM) and effector memory (TEM) CD4 and CD8 T cells were also measured. An enhanced recruitment of NP-specific cytotoxic CD8 T cells was observed in the lungs of SAM(NP)-immunized mice after influenza infection that paralleled with reduced lung viral titers and pathology, and increased survival after homologous and heterosubtypic influenza challenge. Finally, we demonstrated for the first time that the co-administration of RNA (SAM(M1-NP)) and protein (monovalent inactivated influenza vaccine (MIIV)) was feasible, induced simultaneously NP-, M1- and HA-specific T cells and HA-specific neutralizing antibodies, and enhanced MIIV efficacy against a heterologous challenge. In conclusion, systemic administration of SAM vectors expressing conserved internal influenza antigens induced protective immune responses in mice, supporting the SAM® platform as another promising strategy for the development of broad-spectrum universal influenza vaccines.
Subject(s)
Antigens, Viral/genetics , Antigens, Viral/immunology , Conserved Sequence , Influenza A virus/immunology , Influenza A virus/physiology , Influenza Vaccines/genetics , Influenza Vaccines/immunology , Animals , Cell Line , Cricetinae , Gene Amplification , Gene Expression , Genetic Vectors/genetics , Influenza A Virus, H1N1 Subtype/immunology , Influenza A Virus, H1N1 Subtype/physiology , Influenza A Virus, H3N2 Subtype/immunology , Influenza A Virus, H3N2 Subtype/physiology , Lung/immunology , Mice , RNA, Messenger/genetics , T-Lymphocytes, Cytotoxic/immunology , Vaccines, Inactivated/genetics , Vaccines, Inactivated/immunology , Viral Core Proteins/genetics , Viral Core Proteins/immunology , Viral Matrix Proteins/genetics , Viral Matrix Proteins/immunologyABSTRACT
Nucleic acid-based vaccines are being developed as a means to combine the positive attributes of both live-attenuated and subunit vaccines. Viral vectors and plasmid DNA vaccines have been extensively evaluated in human clinical trials and have been shown to be safe and immunogenic, although none have yet been licensed for human use. Recently, mRNA based vaccines have emerged as an alternative approach. They promise the flexibility of plasmid DNA vaccines, without the need for electroporation, but with enhanced immunogenicity and safety. In addition, they avoid the limitations of anti-vector immunity seen with viral vectors, and can be dosed repeatedly. This review highlights the key papers published over the past few years and summarizes prospects for the near future.
Subject(s)
RNA, Messenger/genetics , Vaccines, DNA/genetics , Vaccines, DNA/immunology , Animals , HumansABSTRACT
UNLABELLED: Seasonal influenza is a vaccine-preventable disease that remains a major health problem worldwide, especially in immunocompromised populations. The impact of influenza disease is even greater when strains drift, and influenza pandemics can result when animal-derived influenza virus strains combine with seasonal strains. In this study, we used the SAM technology and characterized the immunogenicity and efficacy of a self-amplifying mRNA expressing influenza virus hemagglutinin (HA) antigen [SAM(HA)] formulated with a novel oil-in-water cationic nanoemulsion. We demonstrated that SAM(HA) was immunogenic in ferrets and facilitated containment of viral replication in the upper respiratory tract of influenza virus-infected animals. In mice, SAM(HA) induced potent functional neutralizing antibody and cellular immune responses, characterized by HA-specific CD4 T helper 1 and CD8 cytotoxic T cells. Furthermore, mice immunized with SAM(HA) derived from the influenza A virus A/California/7/2009 (H1N1) strain (Cal) were protected from a lethal challenge with the heterologous mouse-adapted A/PR/8/1934 (H1N1) virus strain (PR8). Sera derived from SAM(H1-Cal)-immunized animals were not cross-reactive with the PR8 virus, whereas cross-reactivity was observed for HA-specific CD4 and CD8 T cells. Finally, depletion of T cells demonstrated that T-cell responses were essential in mediating heterologous protection. If the SAM vaccine platform proves safe, well tolerated, and effective in humans, the fully synthetic SAM vaccine technology could provide a rapid response platform to control pandemic influenza. IMPORTANCE: In this study, we describe protective immune responses in mice and ferrets after vaccination with a novel HA-based influenza vaccine. This novel type of vaccine elicits both humoral and cellular immune responses. Although vaccine-specific antibodies are the key players in mediating protection from homologous influenza virus infections, vaccine-specific T cells contribute to the control of heterologous infections. The rapid production capacity and the synthetic origin of the vaccine antigen make the SAM platform particularly exploitable in case of influenza pandemic.
Subject(s)
Hemagglutinin Glycoproteins, Influenza Virus/genetics , Hemagglutinin Glycoproteins, Influenza Virus/immunology , Influenza Vaccines/immunology , Orthomyxoviridae Infections/prevention & control , RNA, Messenger/genetics , RNA, Messenger/metabolism , Vaccines, DNA/immunology , Animals , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Cross Protection , Disease Models, Animal , Female , Ferrets , Influenza Vaccines/administration & dosage , Influenza Vaccines/genetics , Leukocyte Reduction Procedures , Mice, Inbred BALB C , Orthomyxoviridae Infections/immunology , Respiratory System/virology , Survival Analysis , Treatment Outcome , Vaccines, DNA/administration & dosage , Vaccines, DNA/genetics , Viral LoadABSTRACT
Self-amplifying mRNAs (SAM(®) ) are a novel class of nucleic acid vaccines, delivered by a non-viral delivery system. They are effective at eliciting potent and protective immune responses and are being developed as a platform technology with potential to be used for a broad range of targets. However, their mechanism of action has not been fully elucidated. To date, no evidence of in vivo transduction of professional antigen-presenting cells (APCs) by SAM vector has been reported, while the antigen expression has been shown to occur mostly in the muscle fibres. Here we show that bone-marrow-derived APCs rather than muscle cells are responsible for induction of MHC class-I restricted CD8 T cells in vivo, but direct transfection of APCs by SAM vectors is not required. Based on all our in vivo and in vitro data we propose that upon SAM vaccination the antigen is expressed within muscle cells and then transferred to APCs, suggesting cross-priming as the prevalent mechanism for priming the CD8 T-cell response by SAM vaccines.
Subject(s)
Antigen-Presenting Cells/immunology , Bone Marrow Cells/immunology , CD8-Positive T-Lymphocytes/immunology , Cross-Priming , Influenza A Virus, H1N1 Subtype/immunology , Influenza Vaccines/immunology , Muscle Fibers, Skeletal/immunology , RNA, Messenger/immunology , RNA, Viral/immunology , RNA-Binding Proteins/immunology , Viral Core Proteins/immunology , Animals , Antigen-Presenting Cells/virology , Bone Marrow Cells/virology , Bone Marrow Transplantation , CD8-Positive T-Lymphocytes/virology , Cell Communication , Cell Line , Cricetinae , Female , Histocompatibility Antigens Class I/genetics , Histocompatibility Antigens Class I/immunology , Influenza A Virus, H1N1 Subtype/genetics , Influenza Vaccines/genetics , Mice, Inbred C3H , Mice, Inbred C57BL , Mice, Transgenic , Muscle Fibers, Skeletal/virology , Nucleocapsid Proteins , RNA, Messenger/genetics , RNA, Viral/genetics , RNA-Binding Proteins/genetics , Transfection , Transplantation Chimera , Viral Core Proteins/geneticsABSTRACT
Alphavirus-based replicons are a promising nucleic acid vaccine platform characterized by robust gene expression and immune responses. To further explore their use in vaccination, replicons were engineered to allow conditional control over their gene expression. Riboswitches, comprising a ribozyme actuator and RNA aptamer sensor, were engineered into the replicon 3' UTR. Binding of ligand to aptamer modulates ribozyme activity and, therefore, gene expression. Expression from DNA-launched and VRP-packaged replicons containing riboswitches was successfully regulated, achieving a 47-fold change in expression and modulation of the resulting type I interferon response. Moreover, we developed a novel control architecture where riboswitches were integrated into the 3' and 5' UTR of the subgenomic RNA region of the TC-83 virus, leading to an 1160-fold regulation of viral replication. Our studies demonstrate that the use of riboswitches for control of RNA replicon expression and viral replication holds promise for development of novel and safer vaccination strategies.
Subject(s)
Alphavirus/genetics , Aptamers, Nucleotide/metabolism , Gene Expression Regulation, Viral/drug effects , Molecular Biology/methods , Riboswitch/drug effects , Virology/methods , Alphavirus/physiology , Genetic Engineering/methods , Virus Replication/drug effectsABSTRACT
This chapter provides a brief introduction to nucleic acid-based vaccines and recent research in developing self-amplifying mRNA vaccines. These vaccines promise the flexibility of plasmid DNA vaccines with enhanced immunogenicity and safety. The key to realizing the full potential of these vaccines is efficient delivery of nucleic acid to the cytoplasm of a cell, where it can amplify and express the encoded antigenic protein. The hydrophilicity and strong net negative charge of RNA impedes cellular uptake. To overcome this limitation, electrostatic complexation with cationic lipids or polymers and physical delivery using electroporation or ballistic particles to improve cellular uptake has been evaluated. This chapter highlights the rapid progress made in using nonviral delivery systems for RNA-based vaccines. Initial preclinical testing of self-amplifying mRNA vaccines has shown nonviral delivery to be capable of producing potent and robust innate and adaptive immune responses in small animals and nonhuman primates. Historically, the prospect of developing mRNA vaccines was uncertain due to concerns of mRNA instability and the feasibility of large-scale manufacturing. Today, these issues are no longer perceived as barriers in the widespread implementation of the technology. Currently, nonamplifying mRNA vaccines are under investigation in human clinical trials and can be produced at a sufficient quantity and quality to meet regulatory requirements. If the encouraging preclinical data with self-amplifying mRNA vaccines are matched by equivalently positive immunogenicity, potency, and tolerability in human trials, this platform could establish nucleic acid vaccines as a versatile new tool for human immunization.
Subject(s)
RNA, Messenger/administration & dosage , Vaccines/administration & dosage , Animals , Antigens/genetics , Electroporation , Humans , Nanoparticles/administration & dosage , Nanoparticles/chemistry , RNA, Messenger/adverse effects , RNA, Messenger/genetics , Vaccines/adverse effects , Viral VaccinesABSTRACT
INTRODUCTION: Self-amplifying mRNA vaccines are being developed as a platform technology with potential to be used for a broad range of targets. The synthetic production methods for their manufacture, combined with the modern tools of bioinformatics and synthetic biology, enable these vaccines to be produced rapidly from an electronic gene sequence. Preclinical proof of concept has so far been achieved for influenza, respiratory syncytial virus, rabies, Ebola, cytomegalovirus, human immunodeficiency virus and malaria. AREAS COVERED: This editorial highlights the key milestones in the discovery and development of self-amplifying mRNA vaccines, and reviews how they might be used as a rapid response platform. The paper points out how future improvements in RNA vector design and non-viral delivery may lead to decreases in effective dose and increases in production capacity. EXPERT OPINION: The prospects for non-viral delivery of self-amplifying mRNA vaccines are very promising. Like other types of nucleic acid vaccines, these vaccines have the potential to draw on the positive attributes of live-attenuated vaccines while obviating many potential safety limitations. Hence, this approach could enable the concept of vaccines on demand as a rapid response to a real threat rather than the deployment of strategic stockpiles based on epidemiological predictions for possible threats.
Subject(s)
Drug Delivery Systems , RNA, Messenger/administration & dosage , Technology, Pharmaceutical , Vaccines, Synthetic/administration & dosage , Humans , RNA, Messenger/immunology , Technology, Pharmaceutical/methods , Technology, Pharmaceutical/trendsABSTRACT
Self-amplifying messenger RNA (mRNA) of positive-strand RNA viruses are effective vectors for in situ expression of vaccine antigens and have potential as a new vaccine technology platform well suited for global health applications. The SAM vaccine platform is based on a synthetic, self-amplifying mRNA delivered by a nonviral delivery system. The safety and immunogenicity of an HIV SAM vaccine encoding a clade C envelope glycoprotein formulated with a cationic nanoemulsion (CNE) delivery system was evaluated in rhesus macaques. The HIV SAM vaccine induced potent cellular immune responses that were greater in magnitude than those induced by self-amplifying mRNA packaged in a viral replicon particle (VRP) or by a recombinant HIV envelope protein formulated with MF59 adjuvant, anti-envelope binding (including anti-V1V2), and neutralizing antibody responses that exceeded those induced by the VRP vaccine. These studies provide the first evidence in nonhuman primates that HIV vaccination with a relatively low dose (50 µg) of formulated self-amplifying mRNA is safe and immunogenic.