[IBS 코로나19 리포트 시즌2] How mRNA was developed as a vaccine and what more can be done: Dong-A Science

From mRNA, COVID-19 vaccine to gene therapy

The first mRNA vaccine developed by mankind

An unfamiliar substance called the’mRNA (messenger RNA) vaccine’ first passed the finish line of the most fierce vaccine competition in human history. BNT162b2, jointly developed by Bioentech of Germany and Pfizer of the United States in November 2020, and mRNA-1273, jointly developed by Modena of the United States and the National Institute of Allergy and Infectious Diseases (NIAID), received FDA approval in December. As a result, the two types of mRNA vaccines outperformed traditional vaccines and advanced in the speed of development. The mRNA vaccine is also a technology that can respond by making a new vaccine the fastest even if another mutant virus appears.

Corona 19 was the first case that mRNA was used as a vaccine. But how did he achieve such a remarkable success? In this report, we will take a closer look at the definition of mRNA vaccine, the principle of inducing immunity, production method, advantages and disadvantages, and future prospects.

The process by which mRNA vaccines induce immune phenomena

mRNA is short for messenger ribonucleic acid (messenger RNA). It contains the genetic information that can synthesize proteins and acts as a messenger to deliver them. mRNA has a long chain structure in which hundreds to thousands of units are connected like beads. There are 4 types of units (A, G, C, U), and different genetic information is contained depending on how these 4 types of units are arranged.

The mRNA vaccine consists of an mRNA molecule and a lipid layer surrounding it (Figure 2). The mRNA is a material that contains information, and the lipid layer is a moving device that protects and puts the messenger (mRNA) into the cell. In the case of Sascoronavirus-2 mRNA vaccine, it contains the genetic information that makes the spike protein on the surface of the virus. In other words, when this mRNA enters a human cell, a spike protein is produced. Spike proteins produced by vaccines function as’antigens’ that induce antibody formation.

The human body’s immune cells make the mistake of being infected with a virus due to an artificially made protein and produce antibodies against the spike protein. Not only B immune cells that make antibodies but also T immune cells that help them are activated. This reaction is called’acquired immunity’. In other words, through’indirect experience’, the human body gains immunity against viruses. After that, even if the actual virus enters the body, the antibody can wrap around the virus to prevent infection. In addition, when a cell is infected with a virus and makes a spike protein, the immune cells recognize and attack the cell and kill it, preventing the growth of the virus.

Conditions for a successful mRNA vaccine

The mRNA used in vaccines is an artificial RNA made by mimicking natural mRNA. Two things are important for an mRNA vaccine to work successfully. First, it is similar to natural mRNA. Because it has to make proteins well, and be able to trick cells as if they were real viral mRNAs. Second, the innate immune response should not occur excessively. When cells recognize the vaccine’s mRNA itself as an invader and an excessive immune response occurs, the production of proteins that act as antigens is disrupted. This means that immunity’advance learning’ is impossible.

Let’s look at the structure of mRNA (Figure 3). The coding sequence containing protein information is the core of mRNA. In the case of an mRNA vaccine, the coding sequence contains information that creates an antigen that will trigger an immune response (a spike protein in the case of a Corona 19 vaccine). The untranslated region (UTR) surrounding the coding sequence helps protein production.

On the other hand, the end where RNA starts is called the 5’end, and there is an important structure called’cap’. The cap helps protein production and prevents mRNA from being destroyed. It also prevents mRNA from causing excessive innate immune responses. The other end is called the 3’end. There is an important structure here called’poly(A) tail’. It aids in protein production and plays a role in stably maintaining mRNA.

The structures described above are those originally present in natural mRNA. In the case of mRNA vaccines, excessive immune reactions are avoided and proteins are replaced by artificial derivatives such as pseudouridine, N1-methyl-pseudouridine, and methylcytosine (5mC). Design to make production work well.

Method for preparing mRNA vaccine

mRNA vaccines are produced through several enzymatic reactions (Figure 3). First of all, to make RNA, you need to make DNA that acts as a mold (template). RNA polymerase, RNA units and cap analogs are added to DNA and reacted to make mRNA. In this process, a polyA tail is created and a uridine derivative is inserted to increase the stability of the mRNA. In some cases, after RNA synthesis, a cap is attached and appropriate modifications are applied. After that, DNA and by-products are decomposed and impurities are removed. If the double-stranded RNA, which is generated as an impurity in the synthesis process, is left unattended, excessive innate immune reactions can occur, which can be a problem.

The mRNA must enter the cell before it can make a protein. However, RNA is charged and has a large molecular weight, so it cannot pass through the cell membrane and enter the cell. Protective devices are also needed to prevent degradation by RNA degrading enzymes in our body. For this reason, efforts have been made over a long period of time to develop technologies that protect RNA and deliver it to cells. Currently, liquid nanoparticle technology is the most widely used.

Lipid nanoparticles are made by mixing several types of substances. Phospholipids and ionizable lipids (ionizable lipids) are the main constituents of particles and are fused with cell membranes to deliver RNA. Cholesterol maintains the shape of the particles and aids the movement of RNA into the cytoplasm. Polyethylene glycol (PEG) increases the hydrophilicity of particles and helps lipid nanoparticles to stay in the body for a long time. The lipid nanoparticles made in this way are usually about 100 nm in diameter (a nanometer·1 nm is 1 billionth of a meter), and are similar in size to virus particles.

Advantages and limitations of RNA vaccines

As seen in the case of the COVID-19 vaccine, the advantage of an mRNA vaccine is, above all, its speed and flexibility. By knowing the genetic information of the pathogen, it can be designed and produced quickly. After the genetic information of Sascoronavirus-2 was released on January 10, 2020, it took only 25 days for Modena to produce the vaccine required for the phase 1 clinical trial.(Due to other issues, the actual clinical trial was conducted on March 16. Started).

Moreover, this period could be shortened further if the vaccine development platform is overhauled. No matter what new pathogens emerge, they can make a vaccine and enter clinical trials within a month with only the genetic information. Since the time and cost for initial development are low, it can be prepared for infectious diseases with relatively few patients. In addition, compared to existing drugs (proteins or low-molecular compounds), it can be produced at a small scale. Depending on the design of the production pipeline, the production volume can be flexibly adjusted from small to large. It is also possible to produce several types of mRNA, mix and introduce.

Stability is also a big strength. Since mRNA is a substance in our body, it is not toxic by itself. In the manufacturing process, instead of using cells, purified enzymes are used, so there is almost no risk of introducing dangerous impurities. This is why it is evaluated to be safer than any type of vaccine used previously. However, there is a possibility that polyethylene glycol, a component of lipid nanoparticles that transfers mRNA, causes side effects. However, with the widespread evaluation and validation of the Corona 19 vaccine, concerns about safety will soon come to a conclusion.

The disadvantage is thermal stability. Other vaccines can be stored at 4°C or room temperature. However, modders or vaccines must be stored in a freezer, and Pfizer vaccines must be distributed at cryogenic temperatures (-70°C). The purified mRNA itself is a very stable material, but it is presumed that the lipid nanoparticles are unstable and require low temperature storage. The key is to improve the lipid nanoparticles or to develop alternative technologies.

There is also a lot of room for improvement in mRNA design. By studying each of the components of RNA, we need to figure out how to make the RNA stay in the cell longer and produce more protein. It is also necessary to find the best solution that can induce an appropriate immune response while avoiding hyperimmunity. If the amount of RNA used can be reduced, it is expected that the vaccine will be delivered faster and more safely to more people at lower prices. Indeed, in the case of Curevac’s mRNA vaccine in Germany, which is currently undergoing phase 2 clinical trials, it is known that it is effective even at 12 micrograms and can be distributed in refrigeration, so the mRNA vaccine technology seems to have great room for further development.

The medical potential and impact of mRNA

mRNA has become a powerful tool to save mankind from the crisis of infectious diseases after over half a century of research since its discovery in 1961. As the safety and effectiveness of mRNA has been proven in the Corona 19 outbreak, it is clear that mRNA vaccines will be widely used in preventing infectious diseases in the future. Since Sascoronavirus-2 mutates and will continue to plague humans, mRNA vaccines can be of great help in dealing with mutant viruses. Of course, there is no guarantee that mRNA vaccines will work against all pathogens. However, it is highly likely to increase the ability to respond to new pathogens. It may provide a solution to the localized infection in underdeveloped countries that have been neglected without measures. Clinical trials of mRNA vaccines against Zika virus, influenza, and malaria are already in progress.

In addition to vaccines for preventing infectious diseases, it is also highly likely to be used as a cancer vaccine. Cancer cells make abnormal proteins unlike normal cells. When a cancer-specific protein is produced in the body by administration of an mRNA-based cancer vaccine, immune cells can then recognize the cancer protein and selectively destroy only cancer cells. Cancer cell proteins can vary from patient to patient, so it is desirable to develop personalized cancer vaccines. The mRNA is relatively easy to design, enabling the production of personalized cancer vaccines. In fact, the primary target of mRNA vaccines prior to the coronavirus outbreak was cancer, and a number of clinical trials are currently in progress.

Beyond vaccines, mRNA will have a broad and profound impact on medicine and biotechnology as a’gene carrier’. mRNA can theoretically pass any gene into our body. It is possible to’gene therapy’ by introducing a gene that is insufficient in our body using mRNA. mRNA is easy and fast to design and produce. Therefore, if the platform is well equipped, it can cope with various diseases within months. That is, if only the genetic cause of the disease is identified, it is relatively easy to develop vaccines and therapeutic agents to respond to it, and the initial development cost is low. Therefore, we can hope for the emergence of treatments for rare diseases, which were difficult to develop due to the small market size. It is a revolutionary change compared to the cost of more than a decade and hundreds of billions of won to develop existing drugs.

To make the possibilities of mRNA a reality

These rosy dreams do not come true on their own. More research is needed to make the potential of mRNA a reality, and there is a lot to do.

First of all, more research on RNA itself is needed. We need to make sure that RNA stays in our body for a long time and makes proteins well, while at the same time not triggering excessive immune responses. This requires a deeper understanding of RNA at a higher level than it is now. Second, RNA delivery technology should be further improved. To deliver enough mRNA anywhere in our body, efficient technology is needed to send RNA to specific organs and cells. Development of a safer carrier with high thermal stability is also key. Third, it is necessary to know exactly the genetic cause of the disease. Only then can you know which genes to pass. Therefore, molecular genetic studies on the cause of the disease must be preceded. Finally, human networks and material infrastructures that can organically connect these multidisciplinary studies are needed. There should be a system that fosters professional manpower and helps basic laboratories, hospitals, pharmaceutical industry, and government to cooperate closely.

Of course it is not easy. However, as the Corona 19 vaccine was developed quickly and successfully, it is unlikely that it will take too long for this dream to come true. In the near future, we will meet RNA technology that directly connects genetic information to therapeutic agents. Perhaps this is the greatest gift the 21st century science gives to mankind.

※references

Verbeke et al., Nano Today, 28:1 (2019), Three decades of messenger RNA vaccine development

Linares-Fernández et al., Trends in Molecular Medicine, 26:311 (2019), Tailoring mRNA vaccine to balance innate/adaptive immune response.

Topol, Cell, (2021), Messenger RNA vaccines against SARS-CoV-2.

Jackson et al., NPJ Vaccines 11:1 (2020), The promise of mRNA vaccines: a biotech and industrial perspective.

Polack et al., NEJM 383:2603 (2020), Safety and efficacy of the BNT162b2 mRNA Covid-19 vaccine.

Jackson et al., NEJM 383:1920 (2020), An mRNA vaccine against SARS-CoV-2-Preliminary report.

Bitnae-ri Kim, Director of the RNA Research Center, Institute for Basic Sciences (IBS), Chair Professor at Seoul National University, Department of Life Sciences (RNA Biology)

※ Source of original text: Institute for Basic Science

The Institute of Basic Science (IBS) publishes a series of’Corona 19 Science Report 2’to seek scientific understanding of SARS-CoV-2 and ways to overcome it, following last year. In this series, we will focus on research trends and issues related to the development of vaccines and treatments, as well as viral mutations that have recently aroused global interest. We hope that the front-line knowledge and information delivered by IBS scientists and domestic experts will help end coronavirus infection-19 (COVID-19, hereinafter referred to as COVID-19).

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