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Published: 12 January 2018
mRNA vaccines — a new era in vaccinology
Norbert Pardi, Michael J. Hogan, Frederick W. Porter & Drew Weissman
Nature Reviews Drug Discovery volume 17, pages261–279 (2018)Cite this article

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Key Points
Recent improvements in mRNA vaccines act to increase protein translation, modulate innate and adaptive immunogenicity and improve delivery.

mRNA vaccines have elicited potent immunity against infectious disease targets in animal models of influenza virus, Zika virus, rabies virus and others, especially in recent years, using lipid-encapsulated or naked forms of sequence-optimized mRNA.

Diverse approaches to mRNA cancer vaccines, including dendritic cell vaccines and various types of directly injectable mRNA, have been employed in numerous cancer clinical trials, with some promising results showing antigen-specific T cell responses and prolonged disease-free survival in some cases.

Therapeutic considerations and challenges include scaling up good manufacturing practice (GMP) production, establishing regulations, further documenting safety and increasing efficacy.

Important future directions of research will be to compare and elucidate the immune pathways activated by various mRNA vaccine platforms, to improve current approaches based on these mechanisms and to initiate new clinical trials against additional disease targets.

Abstract
mRNA vaccines represent a promising alternative to conventional vaccine approaches because of their high potency, capacity for rapid development and potential for low-cost manufacture and safe administration. However, their application has until recently been restricted by the instability and inefficient in vivo delivery of mRNA. Recent technological advances have now largely overcome these issues, and multiple mRNA vaccine platforms against infectious diseases and several types of cancer have demonstrated encouraging results in both animal models and humans. This Review provides a detailed overview of mRNA vaccines and considers future directions and challenges in advancing this promising vaccine platform to widespread therapeutic use.

Main
Vaccines prevent many millions of illnesses and save numerous lives every year1. As a result of widespread vaccine use, the smallpox virus has been completely eradicated and the incidence of polio, measles and other childhood diseases has been drastically reduced around the world2. Conventional vaccine approaches, such as live attenuated and inactivated pathogens and subunit vaccines, provide durable protection against a variety of dangerous diseases3. Despite this success, there remain major hurdles to vaccine development against a variety of infectious pathogens, especially those better able to evade the adaptive immune response4. Moreover, for most emerging virus vaccines, the main obstacle is not the effectiveness of conventional approaches but the need for more rapid development and large-scale deployment. Finally, conventional vaccine approaches may not be applicable to non-infectious diseases, such as cancer. The development of more potent and versatile vaccine platforms is therefore urgently needed.

Nucleic acid therapeutics have emerged as promising alternatives to conventional vaccine approaches. The first report of the successful use of in vitro transcribed (IVT) mRNA in animals was published in 1990, when reporter gene mRNAs were injected into mice and protein production was detected5. A subsequent study in 1992 demonstrated that administration of vasopressin-encoding mRNA in the hypothalamus could elicit a physiological response in rats6. However, these early promising results did not lead to substantial investment in developing mRNA therapeutics, largely owing to concerns associated with mRNA instability, high innate immunogenicity and inefficient in vivo delivery. Instead, the field pursued DNA-based and protein-based therapeutic approaches7,8.

Over the past decade, major technological innovation and research investment have enabled mRNA to become a promising therapeutic tool in the fields of vaccine development and protein replacement therapy. The use of mRNA has several beneficial features over subunit, killed and live attenuated virus, as well as DNA-based vaccines. First, safety: as mRNA is a non-infectious, non-integrating platform, there is no potential risk of infection or insertional mutagenesis. Additionally, mRNA is degraded by normal cellular processes, and its in vivo half-life can be regulated through the use of various modifications and delivery methods9,10,11,12. The inherent immunogenicity of the mRNA can be down-modulated to further increase the safety profile9,12,13. Second, efficacy: various modifications make mRNA more stable and highly translatable9,12,13. Efficient in vivo delivery can be achieved by formulating mRNA into carrier molecules, allowing rapid uptake and expression in the cytoplasm (reviewed in Refs 10,11). mRNA is the minimal genetic vector; therefore, anti-vector immunity is avoided, and mRNA vaccines can be administered repeatedly. Third, production: mRNA vaccines have the potential for rapid, inexpensive and scalable manufacturing, mainly owing to the high yields of in vitro transcription reactions.

The mRNA vaccine field is developing extremely rapidly; a large body of preclinical data has accumulated over the past several years, and multiple human clinical trials have been initiated. In this Review, we discuss current mRNA vaccine approaches, summarize the latest findings, highlight challenges and recent successes, and offer perspectives on the future of mRNA vaccines. The data suggest that mRNA vaccines have the potential to solve many of the challenges in vaccine development for both infectious diseases and cancer.

Basic mRNA vaccine pharmacology
mRNA is the intermediate step between the translation of protein-encoding DNA and the production of proteins by ribosomes in the cytoplasm. Two major types of RNA are currently studied as vaccines: non-replicating mRNA and virally derived, self-amplifying RNA. Conventional mRNA-based vaccines encode the antigen of interest and contain 5′ and 3′ untranslated regions (UTRs), whereas self-amplifying RNAs encode not only the antigen but also the viral replication machinery that enables intracellular RNA amplification and abundant protein expression.

The construction of optimally translated IVT mRNA suitable for therapeutic use has been reviewed previously14,15. Briefly, IVT mRNA is produced from a linear DNA template using a T7, a T3 or an Sp6 phage RNA polymerase16. The resulting product should optimally contain an open reading frame that encodes the protein of interest, flanking UTRs, a 5′ cap and a poly(A) tail. The mRNA is thus engineered to resemble fully processed mature mRNA molecules as they occur naturally in the cytoplasm of eukaryotic cells.

Complexing of mRNA for in vivo delivery has also been recently detailed10,11. Naked mRNA is quickly degraded by extracellular RNases17 and is not internalized efficiently. Thus, a great variety of in vitro and in vivo transfection reagents have been developed that facilitate cellular uptake of mRNA and protect it from degradation. Once the mRNA transits to the cytosol, the cellular translation machinery produces protein that undergoes post-translational modifications, resulting in a properly folded, fully functional protein. This feature of mRNA pharmacology is particularly advantageous for vaccines and protein replacement therapies that require cytosolic or transmembrane proteins to be delivered to the correct cellular compartments for proper presentation or function. IVT mRNA is finally degraded by normal physiological processes, thus reducing the risk of metabolite toxicity.

Recent advances in mRNA vaccine technology
Various mRNA vaccine platforms have been developed in recent years and validated in studies of immunogenicity and efficacy18,19,20. Engineering of the RNA sequence has rendered synthetic mRNA more translatable than ever before. Highly efficient and non-toxic RNA carriers have been developed that in some cases21,22 allow prolonged antigen expression in vivo (Table 1). Some vaccine formulations contain novel adjuvants, while others elicit potent responses in the absence of known adjuvants. The following section summarizes the key advances in these areas of mRNA engineering and their impact on vaccine efficacy.

Table 1 mRNA vaccine complexing strategies for in vivo use
Full size table
Optimization of mRNA translation and stability
This topic has been extensively discussed in previous reviews14,15; thus, we briefly summarize the key findings (Box 1). The 5′ and 3′ UTR elements flanking the coding sequence profoundly influence the stability and translation of mRNA, both of which are critical concerns for vaccines. These regulatory sequences can be derived from viral or eukaryotic genes and greatly increase the half-life and expression of therapeutic mRNAs23,24. A 5′ cap structure is required for efficient protein production from mRNA25. Various versions of 5′ caps can be added during or after the transcription reaction using a vaccinia virus capping enzyme26 or by incorporating synthetic cap or anti-reverse cap analogues27,28. The poly(A) tail also plays an important regulatory role in mRNA translation and stability25; thus, an optimal length of poly(A)24 must be added to mRNA either directly from the encoding DNA template or by using poly(A) polymerase. The codon usage additionally has an impact on protein translation. Replacing rare codons with frequently used synonymous codons that have abundant cognate tRNA in the cytosol is a common practice to increase protein production from mRNA29, although the accuracy of this model has been questioned30. Enrichment of G:C content constitutes another form of sequence optimization that has been shown to increase steady-state mRNA levels in vitro31 and protein expression in vivo12.

Although protein expression may be positively modulated by altering the codon composition or by introducing modified nucleosides (discussed below), it is also possible that these forms of sequence engineering could affect mRNA secondary structure32, the kinetics and accuracy of translation and simultaneous protein folding33,34, and the expression of cryptic T cell epitopes present in alternative reading frames30. All these factors could potentially influence the magnitude or specificity of the immune response.