"Building blocks" to make vaccines, monkeypox mRNA vaccine can "kill three birds with one stone"!

Produced by: Science Popularization China

Author: Du Pei (Institute of Microbiology, Chinese Academy of Sciences)

Producer: China Science Expo

In 2022, a new variant of the monkeypox virus broke through the restrictions of the long-term local epidemic in Africa and spread widely around the world, leading to a new global epidemic. On July 23, 2022, the World Health Organization (WHO) issued the highest level of alert for the monkeypox epidemic - a Public Health Emergency of International Concern (PHEIC). As the global monkeypox epidemic gradually subsided, the WHO announced on May 11, 2023 that monkeypox no longer constituted a PHEIC.

However, starting in 2024, new monkeypox variants appeared in Africa, causing higher mortality and new transmission characteristics. On August 14, 2024, the WHO had to sound the alarm again and declared the monkeypox outbreak a PHEIC for the second time.

Recently, new mutant strains have been introduced into many countries around my country, including Thailand and India. my country is facing severe monkeypox epidemic challenges and urgently needs monkeypox vaccine for prevention.

How about the monkeypox vaccine that "crossed over" from the smallpox vaccine?

Vaccinia virus, smallpox virus and monkeypox virus are close relatives. They all belong to the genus Orthopoxvirus in virus taxonomy and have relatively similar gene sequences. The gene sequence of an organism determines the characteristics of the protein, and protein, as an antigen, is a key molecule that stimulates the human body to produce antibodies and T cells. The antigenic characteristics of vaccinia virus are similar to those of smallpox virus and monkeypox virus, but it is less harmful to humans. Taking advantage of this similarity, smallpox virus has been studied at home and abroad through vaccinia virus for a long time, and vaccinia virus has also been used for attenuation to develop attenuated live vaccines for the prevention of smallpox and monkeypox. The Tiantan strain attenuated live vaccine, which was widely used in my country before the 1980s to prevent smallpox, belongs to this category.

At present, three live attenuated smallpox vaccines abroad have "crossed the border" to become monkeypox vaccines. Although they can solve the urgent problem of the epidemic, there are still some limitations in terms of safety and accessibility.

(1) Safety: The attenuated smallpox vaccine has been developed to the third generation, which can prevent the attenuated vaccinia virus from replicating in the body of the vaccine recipient. It is safer than the early replicable attenuated live vaccine, but the vaccinia virus can still produce many proteins in the body that inhibit the human immune response, which may lead to potential side effects. According to the WHO's recommendations for monkeypox vaccination, the third-generation attenuated live vaccine is still not safe enough for HIV-infected people whose AIDS has progressed to the point where their immune function is no longer sound (less than 200 CD4+ helper T cells per milliliter of blood). However, there is a large overlap between the population infected with monkeypox virus and HIV carriers. According to WHO statistics, about 52.1% of monkeypox virus-infected people carry HIV. HIV-infected people with impaired immune function are at high risk of monkeypox and need more protection.

(2) Accessibility: Vaccines can only play a role in controlling infectious diseases if they are actually administered to people. The production of live attenuated vaccines involves virus cultivation, which has complex production processes and high safety risks. It is difficult to quickly increase production capacity to vaccinate more people when an infectious disease breaks out. According to a report in Nature magazine in October 2024, the Africa Centers for Disease Control and Prevention estimates that the number of people in Africa who need to be vaccinated with monkeypox vaccine may be as high as 15 million, but only about 275,000 doses of monkeypox vaccine are available. In fact, since the outbreak of monkeypox in 2022, the world has continued to face the problem of insufficient monkeypox vaccine production capacity. The major pharmaceutical companies that produce third-generation live attenuated vaccines only supplied about 2 million doses of monkeypox vaccine to the world in the first eight months of 2024.

Therefore, the new generation of monkeypox vaccine must "eliminate the dross and retain the essence", remove the "bad" proteins, and only retain the "good" proteins as vaccine antigens to make the vaccine safer; it must also make full use of new vaccine technologies in recent years to make the vaccine more efficient and easier to produce.

How should a new generation of monkeypox vaccine be designed?

The new generation of monkeypox vaccine faces many challenges in eliminating the dross and retaining the essence of traditional live attenuated vaccines.

Which monkeypox virus proteins should be selected as antigens?

Poxvirus is a complex virus that contains more than 200 proteins, but only some of the key proteins that can stimulate the human immune response are suitable for use as antigens in vaccines. In recent years, virologists and immunologists at home and abroad have conducted in-depth research on the antigens of vaccinia virus and monkeypox virus, and have basically determined that several proteins are the key antigens of orthopoxvirus, providing a reference for the design of a new generation of monkeypox vaccine.

Monkeypox vaccines require a combination of multiple antigens to achieve optimal protection. How can we include multiple monkeypox antigens in one vaccine?

The simplest way is the "cocktail" solution. For mRNA vaccines, one mRNA is used to encode an antigen, and then multiple mRNAs are mixed into a vaccine like a cocktail. For recombinant protein vaccines, multiple antigen proteins are directly mixed and matched with other necessary ingredients to make a vaccine. However, the "cocktail" vaccine is not the best route, because it requires the production of multiple different mRNAs or proteins, which increases the complexity of the production process, makes quality control more challenging, and ultimately increases the cost of the vaccine. At the same time, the multiple different antigens in the "cocktail" may also produce uneven immune effects.

Which technology route of vaccine should we choose?

Different vaccine technology routes have different advantages. Among them, mRNA vaccines use mRNA molecules as carriers. After entering the body, they produce antigen proteins in cells and stimulate immune responses. Therefore, just like writing a computer program, a variety of vaccines can be produced by simply changing the sequence of mRNA, which has the advantages of fast development speed and high standardization of production lines. At the same time, mRNA vaccines can also stimulate antibodies and T cells at the same time, and have a good immune effect. During the COVID-19 pandemic, after large-scale vaccination among the population, the safety of mRNA vaccines has been recognized, which further highlights the advantages of mRNA vaccines. Recombinant protein vaccines use proteins to stimulate the human immune response, and also have good immune effects and safety. Since 2022, the new generation of monkeypox vaccines have included mRNA vaccines and recombinant protein vaccines, most of which are "cocktail" mRNA vaccines. Abroad, two monkeypox mRNA vaccines have entered clinical trials.

Use the "building blocks" approach to make vaccines!

Proteins have complex three-dimensional structures, but the three-dimensional structure of the same protein is relatively fixed, predictable, and can also be analyzed by experimental means. Through a reasonable scheme, it is possible to design a protein that contains multiple antigens and exists stably, which can not only stimulate antibodies to various antigens in a balanced manner, but also be encoded with one mRNA, reducing production costs and improving the accessibility of vaccines. This scheme is like "building blocks". According to the three-dimensional structure of the protein, multiple protein molecules are combined and spliced ​​into a larger protein. They are embedded and locked into a stable structure like Kongming lock blocks, and are reinforced on the binding surfaces of different proteins using various chemical bonds like Lego blocks. This vaccine design strategy based on protein structure is called structural vaccinology.

Diagram of building a vaccine with building blocks

(Image source: The left image is from 360 Encyclopedia, the right image is from Reference 1)

So, are there other vaccines designed using structural vaccinology? Yes.

In 2020, in the face of the COVID-19 pandemic, the team of Academician Gao Fu from the Institute of Microbiology, Chinese Academy of Sciences, used structural vaccinology to design a recombinant protein vaccine for the COVID-19 pandemic, and jointly developed it with Zhifei Biotechnology into the ZF2001® vaccine, which has been administered 350 million doses worldwide. Among them, the ZF2001® vaccine has achieved technology exports in Uzbekistan and has become the country's main vaccine to fight the COVID-19 pandemic. In his signed article "Working Together to Create a Better Future for China-Uzbekistan Relations" published during his visit to Uzbekistan on September 13, 2022, General Secretary Xi Jinping mentioned: "Chinese vaccines have become the main force in Uzbekistan's fight against the pandemic and have achieved joint production. Anti-COVID-19 drugs have been approved for clinical use in Uzbekistan, effectively protecting the lives of the people of the two countries."

After the outbreak of monkeypox, the team led by Academician Gao Fu designed a recombinant protein vaccine DAM that combines two monkeypox virus antigens. It can stimulate 28 times more monkeypox virus neutralizing antibodies than the Tiantan strain live attenuated vaccine, achieve complete protection against lethal doses of vaccinia virus infection, and quickly clear the virus from organs. As a single protein, the DAM vaccine has shown good safety and industrial development potential.

Monkeypox multi-antigen chimeric mRNA vaccine: from "three arrows, three kills" to "one arrow, three kills".

Using the structural vaccinology platform, the team of Academician Gao Fu and Researcher Wang Qihui from the Institute of Microbiology developed three monkeypox multi-antigen chimeric mRNA vaccines, using one mRNA to encode a chimeric protein containing three monkeypox virus antigens. After inoculation into mice, these vaccines can stimulate efficient humoral immunity (antibodies) and cellular immunity (T cells), and maintain antibody levels that basically do not decrease after nearly 6 months of immunization. **All mice vaccinated with these vaccines survived infection with a lethal dose of vaccinia virus. **After infection with the same dose of vaccinia virus, all mice vaccinated with the Tiantan strain attenuated live vaccine died. In the face of monkeypox virus, these vaccines also showed good protective effects. The viral load in multiple organs of mice infected with monkeypox virus was significantly lower than that of mice in the control group.

Three-dimensional structures of proteins encoded by three monkeypox multi-antigen chimeric mRNA vaccines

(Image source: Reference 2)

If the "cocktail" vaccine with three mRNAs encoding three antigens is "killing three birds with one stone", then these monkeypox multi-antigen chimeric mRNA vaccines achieve the effect of "killing three birds with one stone". Compared with the "cocktail" mRNA vaccine, the chimeric vaccine encoded by single-stranded mRNA has lower production difficulty and is expected to achieve greater cost advantages, simpler production processes and wider vaccine accessibility.

At present, the Institute of Microbiology of the Chinese Academy of Sciences has signed a technology licensing agreement with a pharmaceutical company, authorizing it to further develop, apply for clinical trials and market the vaccine. We look forward to the new monkeypox multi-antigen chimeric mRNA vaccine making a contribution to the prevention of monkeypox!

References:

1.Rational design of a 'two-in-one' immunogen DAM drives potent immune response against mpox virus

2.Single-chain A35R-M1R-B6R trivalent mRNA vaccines protect mice against both mpox virus and vaccinia virus