Introduction
The mRNA vaccines utilize a messenger RNA (mRNA) version to initiate an immune reaction. This vaccine introduces antigen-carrying mRNA molecules into our immune cells. These cells then read the mRNA instructions to construct a foreign protein, which is usually made by harmful pathogens like viruses or cancer cells. This newly created protein activates an adaptive immune response, training the body to recognize and eliminate the related pathogen or cancer cells in the future. Lipid nanoparticles are used to envelop and protect the mRNA, ensuring its safe delivery into the cells.
Regarding side effects, mRNA vaccines are comparable to traditional non-RNA vaccines. However, individuals prone to autoimmune reactions might experience adverse reactions to mRNA vaccines. When comparing mRNA vaccines to their conventional counterparts, they stand out due to their rapid design process, cost-effectiveness, and capability to induce cellular and antibody-mediated immunity. Moreover, they don’t interfere with our DNA. One drawback, for example, with the Pfizer–BioNTech COVID-19 mRNA vaccine is its need for freezing storage. On the other hand, Moderna, CureVac, and Walvax mRNA vaccines for COVID-19 don’t have such stringent storage conditions.
The potential of mRNA vaccines became especially evident during the COVID-19 pandemic. In December 2020, Pfizer–BioNTech and Moderna received approvals for their mRNA COVID-19 vaccines. The UK’s MHRA was the first to approve the Pfizer–BioNTech vaccine on 2 December. Subsequently, on 11 December, the US FDA provided emergency use authorization for the same vaccine, and soon after, the Moderna vaccine received a similar nod. Recognizing the groundbreaking work in mRNA vaccine development, the 2023 Nobel Prize in Physiology or Medicine was presented to Katalin Karikó and Drew Weissman for their pivotal role in creating effective mRNA vaccines against COVID-19.
History and Evolution of mRNA Vaccines
Origins and Preliminary Findings
The journey of mRNA vaccines began in 1989 when a significant scientific achievement was made. Researchers successfully introduced mRNA designed and encapsulated within a lipid nanoparticle into a cell. A subsequent experiment in 1990 saw the injection of “naked” or non-encapsulated, laboratory-created mRNA into mouse muscles. These initial experiments confirmed that synthesized mRNA carrying a selected gene could transfer genetic information to produce a specific protein in living tissue. This groundbreaking realization formed the foundation for the idea of messenger RNA vaccines.
In 1993, another pivotal moment they occurred when scientists demonstrated that liposome-wrapped mRNA encoding a viral antigen could activate T cells in mice. Progress continued quickly: by 1994, self-amplifying mRNA was created. This form of mRNA incorporated both a viral antigen and a gene encoding replicase. This innovative approach provoked mice’s antibody-mediated (humoral) and cell-mediated immune response against viral infections. In 1995, the scope expanded when mRNA encoding a tumour-specific antigen was employed to trigger an immune response targeting mouse cancer cells.
Advances and Clinical Applications
The dawn of the 21st century marked the commencement of the first-ever human clinical trial that used ex vivo dendritic cells loaded with mRNA that encoded tumour antigens. This was the inception of therapeutic mRNA vaccines aimed at treating cancer. By 2005, scientists made another advancement by utilizing modified nucleosides to facilitate mRNA entry into cells without tripping the body’s alarm system. Results from clinical trials in 2008 revealed that directly injecting mRNA vaccines into the human body could target cancer cells effectively.
During this dynamic exploration and progress period, two major biotech companies emerged: BioNTech, founded in 2008, and Moderna, established in 2010. Both were committed to harnessing and advancing mRNA technologies. Parallelly, the US defence research agency DARPA initiated the ADEPT program, focusing on groundbreaking biotechnologies for defence applications. Recognizing the transformative potential of nucleic acid technologies to combat pandemics, DARPA began significant investments in the area. This bolstered confidence in the technology, prompting other governmental bodies and private investors to fund mRNA research. Notably, Moderna received a generous $25 million grant from DARPA.
The fight against infectious diseases using mRNA saw significant progress in 2013, with the commencement of the first human clinical trials against rabies using an mRNA vaccine. In subsequent years, clinical trials began for various viruses, including influenza, Zika, cytomegalovirus, and Chikungunya. By March 2022, Moderna announced it was venturing into developing mRNA vaccines for 15 diseases, ranging from COVID-19 and Ebola to Malaria and Tuberculosis.
Global Impact and Rapid Advancements
The world’s confrontation with the COVID-19 pandemic in 2020 brought mRNA vaccines to the forefront of global attention. With the sequencing of the SARS-CoV-2 virus in early 2020, there was an unprecedented acceleration in mRNA vaccine development. BioNTech and Moderna received approval for their respective COVID-19 mRNA vaccines in December 2020. The UK’s MHRA set a historical precedent on 2 December by becoming the first international medicinal regulatory body to approve an mRNA vaccine—specifically Pfizer–BioNTech’s BNT162b2 COVID-19 vaccine, only a week after its final trial. Following closely, the US FDA granted emergency authorizations for the Pfizer–BioNTech and Moderna COVID-19 vaccines. This swift progression in mRNA vaccines continues, with many more in various stages of research and development.
Mechanism of Action for Vaccines: Traditional vs. mRNA
The Aim of Vaccination
Vaccines are designed to prime the adaptive immune system, enabling it to produce antibodies tailored to recognize and combat specific pathogens. The particular components of the pathogen identified by the antibodies are called antigens.
Traditional Vaccination Methods
The classical approach to vaccination involves directly injecting the body with antigens. This can be attenuated (weakened), inactivated (dead) pathogens, or a viral vector engineered to carry and express the desired antigen. Antigens or pathogenic entities are cultured and grown externally before being administered to individuals to prepare these vaccines.
mRNA Vaccines: A Revolutionary Approach
mRNA vaccines employ a significantly different strategy. Instead of injecting ready-made antigens, they introduce a synthesized RNA fragment from the targeted virus into the person’s body. This mRNA is soon absorbed by immune cells, particularly dendritic cells, through a process known as phagocytosis. Within these cells, ribosomes – the cellular protein factories – interpret the mRNA sequence to synthesize the associated viral antigen. The body naturally breaks down these mRNA fragments within days of their introduction.
Notably, while a variety of non-immune cells can also internalize and use the mRNA from the vaccine to produce and showcase antigens on their surfaces, dendritic cells have a greater propensity to uptake these mRNA particles. Notably, mRNA translation into proteins occurs in the cell’s cytoplasm, ensuring that the body’s genetic material in the cell nucleus remains untouched and unaffected by the vaccine.
Post the production of viral antigens by cells, the typical processes of the adaptive immune response come into play. The newly formed antigens undergo degradation by structures known as proteasomes. Both Class I and Class II MHC (Major Histocompatibility Complex) molecules then bind to these antigen fragments, effectively “activating” the dendritic cells. Once primed, these dendritic cells travel to lymph nodes, displaying the antigen to other immune cells, specifically T and B cells. This interaction produces antibodies that identify and neutralize the pathogen, bestowing individual immunity.
mRNA
The core element of an mRNA vaccine is its constructed mRNA sequence. This mRNA, synthesized in the lab, originates from a specially designed plasmid DNA equipped with an RNA polymerase promoter and a line matching the desired mRNA construct. Mixing T7 phage RNA polymerase with the plasmid DNA produces mRNA. The vaccine’s effectiveness hinges on the stability and architecture of the created mRNA.
This lab-produced mRNA mirrors the structural attributes of natural mRNA found in eukaryotic cells. It consists of a 5′ cap, a 5′-untranslated region (UTR), a 3′-UTR, an open reading frame (ORF) encoding the target antigen, and a 3′-poly(A) tail. By tweaking various sections of the artificial mRNA, its stability and translation efficiency can be boosted, enhancing the vaccine’s overall effectiveness.
The mRNA’s potency can be augmented by employing synthetic 5′-cap analogues, which bolster its stability and amplify protein translation. Likewise, adjusting regulatory elements in both the 5′- and 3′- untranslated regions and fine-tuning the poly(A) tail length can fortify the mRNA and escalate protein synthesis. Modifications to the mRNA nucleotides can temper innate immune responses and lengthen the mRNA’s lifespan within the recipient cell. The choice of nucleic acid sequences and codons has implications for protein translation. Increasing the guanine-cytosine content in the series enhances the mRNA’s stability, lifespan, and resultant protein synthesis. Swapping less common codons with synonymous, host cell-preferred codons further amplifies protein synthesis.
Delivery
For a vaccine to be successful, sufficient mRNA must enter the host cell cytoplasm to stimulate the production of the specific antigens. Entry of mRNA molecules, however, faces several difficulties. Not only are mRNA molecules too large to cross the cell membrane by simple diffusion, but they are also negatively charged like the cell membrane, which causes a mutual electrostatic repulsion. Additionally, mRNA is easily degraded by RNAases in skin and blood.
Various methods have been developed to overcome these delivery hurdles. The vaccine delivery method can be broadly classified by whether mRNA transfer into cells occurs within (in vivo) or outside (ex vivo) the organism.
Ex Vivo Methods
Dendritic cells act as antigen-presenters on their surfaces, setting off interactions with T cells that kick-start the immune response. These cells can be extracted from patients, manipulated with specific mRNA, and reintroduced into the patients to evoke an immune reaction.
In the lab, dendritic cells can uptake mRNA via endocytosis, although this method is not highly efficient. This efficiency can be enhanced through a technique known as electroporation.
In Vivo Approaches
Since it was understood that the direct introduction of lab-synthesized mRNA could result in antigen expression within a body, in vivo methods have been under the spotlight. These strategies present several benefits over ex vivo techniques, mainly because they bypass the expense and complexity of extracting and adjusting dendritic cells. Additionally, they mimic natural infections more closely.
The method of administering the mRNA, whether via skin, bloodstream, or muscles, can result in varied mRNA uptake levels. Specific research has shown, for example, that direct injection into lymph nodes can yield the most potent T-cell response.
Delivery by Naked mRNA Injection
Introducing “naked” mRNA means administering the vaccine in a basic buffer solution. This concept dates back to the 1990s. Pioneering global clinical trials involved intradermal injections of this form of mRNA. While this technique does trigger an immune reaction, the response is generally subdued, and the mRNA tends to break down swiftly post-injection.
Polymer and Peptide Carriers
By merging mRNA with cationic polymers, protective shells known as polyplexes are formed. These shields guard the artificial mRNA against destruction by ribonucleases and facilitate its entry into cells. An example of this approach is using protamine, a naturally occurring cationic peptide, to encapsulate mRNA.
Lipid Nanoparticle Delivery
The FDA’s first-ever approval of lipid nanoparticles for drug delivery came in 2018 with the green light for the siRNA drug Onpattro. The advent of housing mRNA within lipid nanoparticles was a game-changer for the potential of mRNA vaccines, breaking down several pivotal technical barriers. The research in lipid delivery for siRNA laid the groundwork for its application with mRNA. Still, novel lipids were essential to envelop the longer mRNA sequences.
At its core, the lipid protects against decay, ensuring a more consistent translational result. Also, modifying the exterior of the lipid enables targeting specific cell types via ligand interactions. However, research indicates a disparity between in vivo and in vitro nanoparticle applications, particularly concerning cellular absorption. Depending on the requirement, nanoparticles can be administered intravenously or channelled through the lymphatic system.
Scaling up lipid nanoparticles poses challenges, particularly when it comes to microfluidics. These microscale systems are complex to upscale due to their inherent nature. For instance, Pfizer tackled this issue by running numerous microfluidic chambers simultaneously, demanding specialized equipment.
Moreover, sourcing the unique lipids in these nanoparticles, especially ionizable cationic lipids, is challenging. The surge in demand post-2020 forced companies to upscale their lipid production capabilities significantly.
Viral Vector Approaches
Beyond non-viral delivery mechanisms, engineered RNA viruses have been exploited to elicit similar immunological outcomes. Common RNA viruses utilized as vectors encompass retroviruses, lentiviruses, alphaviruses, and rhabdoviruses. Their structural and functional attributes can vary, but they have been employed in clinical trials against various diseases using model animals like mice and primates.
Advantages Of Different Vaccine Types
Compared to Traditional Vaccines:
- Safety and Infectious Risk: mRNA vaccines present a clear advantage in terms of security. Unlike traditional vaccines that often use live, attenuated (weakened), or inactivated (killed) pathogens, mRNA vaccines use a synthetic fragment of the virus’s genetic code. This means they are non-infectious. Producing large quantities of live pathogens, as required for traditional vaccines, risks potential localized outbreaks at manufacturing sites.
- Dual Immunity Stimulation: mRNA vaccines introduce a unique mechanism whereby the antigens are created within the recipient’s cells. This stimulates cellular and humoral immunity, providing a more comprehensive immune response than some traditional vaccines.
- Rapid Development and Production: mRNA technology allows for the swift design of vaccines. For instance, Moderna developed its mRNA-1273 vaccine against COVID-19 in just two days. Additionally, mRNA vaccines can be produced more quickly, affordably, and consistently than their traditional counterparts. This was evident with the Pfizer–BioNTech vaccine, which initially took 110 days for large-scale production but was later optimized to a 60-day process. Notably, the show only spans approximately 22 days, with rigorous quality control stages throughout the manufacturing process.
Compared to DNA Vaccines:
- No Risk of Integration: Since mRNA operates in the cell’s cytoplasm, it doesn’t need to enter the cell nucleus. This eliminates the risk of integration into the host’s genome, a concern associated with DNA-based approaches.
- Enhanced Stability and Efficiency: mRNA molecules can incorporate modified nucleosides, like pseudouridines or 2′-O-methylated nucleosides. This improves stability by reducing the likelihood of an immediate immune response. The result is a longer-lasting effect with superior translation efficiency.
- Customizability: mRNA sequences offer flexibility. The open reading frame (ORF) and untranslated regions (UTR) can be tailored for specific purposes through sequence engineering. This might involve increasing the guanine-cytosine content or selecting UTRs that boost translation. Moreover, adding an ORF for replication can amplify the antigen production, magnifying the immune response and potentially reducing the required dose.
Limitations and Concerns
Storage Challenges
The fragile nature of mRNA demands stringent storage conditions for certain vaccines to prevent degradation and ensure adequate immunization. The BNT162b2 mRNA vaccine by Pfizer–BioNTech necessitates temperatures ranging from −80 to −60 °C (−112 to −76 °F). In contrast, Moderna’s mRNA-1273 vaccine can be stored at temperatures typical of household freezers, between −25 and −15 °C (−13 and 5 °F), and maintains stability at 2 to 8 °C (36 to 46 °F) for up to a month. As reported by Nature in November 2020, despite potential differences in LNP formulations or mRNA structures, it’s believed that these vaccines might eventually exhibit similar storage needs and shelf lives across various temperatures. Research is ongoing for formulations that can withstand warmer storage conditions.
Novelty
Before 2020, no mRNA-based drugs or vaccines had received human-use authorization. This novelty introduced potential uncertainties about unforeseen effects. Amid the 2020 COVID-19 pandemic, the accelerated production potential of mRNA vaccines appealed to health authorities, igniting discussions about the appropriate initial approvals, such as emergency use or expanded access, following the two-month observation post-trial phase.
Potential Side Effects
The reactogenicity of mRNA vaccines is on par with many traditional vaccines. Nevertheless, those prone to autoimmune reactions might exhibit adverse responses. The mRNA sequences in the vaccine might trigger unintended immune reactions, causing individuals to experience flu-like symptoms. To curtail these reactions, mRNA vaccine sequences are tailored to resemble those our cells generate.
In clinical trials for the COVID-19 mRNA vaccines, some participants experienced robust yet temporary reactogenic responses, like fever and fatigue. Though most won’t encounter severe side effects, they’re characterized as symptoms that disrupt daily activities.
Efficacy Uncertainties
The mRNA vaccines for COVID-19, developed by Moderna and Pfizer–BioNTech, demonstrated impressive efficacy rates between 90 to 95 per cent. However, earlier trials on mRNA drugs for pathogens other than COVID-19 weren’t successful and were halted during the initial trial phases. The reasons behind the new mRNA vaccines’ effectiveness remain a topic of discussion.
Margaret Liu, a physician-scientist, suggested that the success of the new mRNA vaccines might stem from the vast resources poured into their development. Alternatively, the vaccines might amplify the immune response due to a lingering inflammatory reaction to the mRNA. While the modified nucleoside approach has reduced inflammation, it hasn’t been entirely eradicated. This could also account for the pronounced responses, such as body aches and fevers, reported by some vaccine recipients. While these reactions can be intense, they are temporary and might also be attributed to the lipid delivery mechanisms.
Misinformation about Vaccine
There is misinformation implying that mRNA vaccines could alter DNA in the nucleus. mRNA in the cytosol is rapidly degraded before it would have time to enter the cell nucleus. MRNA vaccines must be stored at low temperatures and free from RNA to prevent mRNA degradation. Retrovirus can be single-stranded RNA (just as many SARS-CoV-2 vaccines are single-stranded RNA), which enters the cell nucleus and uses reverse transcriptase to make DNA from the RNA in the cell nucleus. A retrovirus has mechanisms to be imported into the heart, but other mRNA (such as the vaccine) lacks these mechanisms. Once inside the middle, the creation of DNA from RNA cannot occur without a reverse transcriptase and appropriate primers, which both accompany a retrovirus but would not be present for another exogenous mRNA (such as a vaccine) even if it could enter the nucleus.
mRNA Vaccine Types: An Overview
Types of mRNA vaccines
There are two primary categories of mRNA vaccines – non-amplifying (conventional) and self-amplifying mRNA. The Pfizer–BioNTech and Moderna vaccines belong to the non-amplifying category. Both types are scrutinised for their potential use against various pathogens and even in cancer treatments.
Non-amplifying mRNA vaccines: These vaccines utilize a straightforward mRNA structure. In this configuration, the mRNA has just one open reading frame dedicated to encoding the target antigen. The quantity of mRNA accessible to the cell directly corresponds to the amount introduced by the vaccine. There’s a limitation on dosage strength based on the maximum mRNA quantity the vaccine can deliver. To mitigate potential toxicity, these vaccines substitute uridine with N1-Methylpseudouridine.
Self-amplifying mRNA vaccines: These vaccines can reproduce their mRNA post-cell entry. They contain two open reading frames. While the first is similar to its non-amplifying counterpart, coding for the desired antigen, the second frame encodes for an RNA-dependent RNA polymerase (and associated proteins). This ensures the replication of the mRNA structure within the cell, permitting lower vaccine doses. Due to its distinct mechanisms and larger molecular size, the assessment criteria for self-amplifying mRNA may differ from conventional mRNA.
Examples of ongoing research in saRNA vaccines include efforts towards a malaria vaccine. In 2021, Gritstone Bio initiated a phase 1 trial for a self-amplifying mRNA COVID-19 vaccine, intended as a booster shot. This vaccine aims not just at the spike protein of the SARS‑CoV‑2 virus but also at other viral proteins less susceptible to genetic changes, enhancing protection against various SARS‑CoV‑2 strains. For replication to take place in saRNA vaccines, the use of uridine is essential.
