The future of vaccines? Japan’s breakthrough in producing 100% pure mRNA

Covered pure mRNA vaccine

A pure capped mRNA vaccine opens the door to more effective vaccines with lower chances of infection. Credit: Mizuki Tada

A Japanese research team has introduced the “Purecap” method, which enables the production of high-purity mRNA vaccines.

A research group from Japan has developed a method for producing highly active mRNA vaccines of high purity using a unique cap to easily separate the desired capped mRNA. This “Purecap” technology extracted up to 100% pure Cap2 mRNA, which has been shown to produce 3-4 times better production of the protein that stimulates the immune system. These findings open up the possibility of cleaner vaccines with a lower risk of infection from impurities. Their findings were recently published in the journal Nature Communications.

The potential of mRNA vaccines

mRNA vaccines have been used successfully as a treatment against coronavirus variants. This has given the researchers hope for their future use as a cancer vaccine. However, the purity of the vaccines hinders this goal because the impurities can stimulate the immune system. This may cause inflammation around the injection site, which is a common side effect of vaccination.

Understanding the Implications of Vaccines

Impurities are often introduced into mRNA vaccines at the capping stage. During this stage, the cap structure that improves mRNA translation, protection, and stabilization is added. Capsules can only be added to mRNA, so ideally the vaccine should contain 100% mRNA. However, there may be unwanted double strands of mRNA, which reduces its purity.

Since single and double-stranded mRNA have different properties, they can be separated using a technique called Reverse-phase high-performance liquid chromatography (RHB-HPLC). This technique separates mRNAs on the basis of their hydrophobicity or hydrophilicity, that is, their repulsion or attraction to water.

Research methodology and results

A research group led by Professor Hiroshi Abe, Project Associate Professor Masahito Inagaki, and Project Associate Professor Naoko Abe of the Graduate School of Science, Nagoya UniversityIn collaboration with the Tokyo University of Medicine and Dentistry, the unique PureCap method was used to introduce a hydrophobic tag into the dam stage. The tagged mRNAs were easily separated at the stage of RP-HPLC. The tag was then easily removed by photocuring, resulting in a 98%-100% pure vaccine.

“We were very excited about the result when we saw on the graph that the RP-HPLC process completely separated the capped and unfolded RNA,” said Hiroshi Abe. “For coronavirus mRNA, which is 4247 bases long, we successfully used the PureCap method to produce capped mRNA with a purity of more than 98%.”

The research group paid particular attention to a group of cap structures found in animal and plant cells, called Cap0, Cap1 and Cap2. Although Cap2 is present in animal and plant cells, evaluating its function has been difficult because there was no way to obtain pure, capped mRNA to ensure a fair test.

“The Cap structure used in mRNA vaccines has so far been limited to the Cap0 and Cap1 types. However, we used our technology to fabricate the Cap0, Cap1, and Cap2 types.” Highly purified mRNA from Cap0, Cap1, and Cap2, synthesized using the PureCap method, showed have lower immunostimulatory activity compared to mRNAs synthesized using conventional techniques illustrating their potential use in pharmaceuticals.”

Since viruses produce mostly Cap1 mRNA, the immune system is less stimulated by Cap2. This indicates that a vaccine using Cap2 will be less likely to cause unwanted side effects such as inflammation when injected. However, it will still be able to create viral proteins when copied that make the vaccine effective.

Benefits of the Cap2 structure

The group used Purecap to generate Cap2 mRNA and analyze its ability for protein synthesis. They found that Cap2 mRNA produced 3-5 times more protein than Cap1 mRNA, which would boost the immune response. They also showed that Cap2 mRNAs induced less stimulation of the inflammatory response than mRNAs synthesized using conventional techniques.

“Traditional mRNA vaccine production methods have not been able to prepare capped mRNA with high purity, which raises concerns about decreased protein synthesis and impurity-derived inflammatory reactions,” Abe said.

“The PureCap method solves these problems by selectively purifying only the capped mRNA. Furthermore, the Cap2-type construct generated with this technology is more efficient in protein synthesis and less irritating to the immune system. This technology has the potential to improve the safety and efficacy of vaccines.” mRNA It is a revolutionary advance towards the practical application of mRNA medicine, in addition to deepening our understanding of the fundamentals of mRNA science.

Reference: “Cap representations with a hydrophobic photocleavable tag enabling easy purification of fully capped mRNAs with different cap structures” by Masahito Inagaki, Naoko Abe, Zinmin Lee, Yoko Nakashima, Soset Acharya, Kazuya Ogawa, Daisuke Kawaguchi, Haruka Hiraoka, Ayaka Bano, Xiu Meng, Mizuki Tada, Tatsuma Ishida, Bengxu Liu, Kengo Kokubo, Hirotaka Murase, Fumitaka Hashiya, Yasuaki Kimura, Satoshi Uchida and Hiroshi Abe May 11, 2023 Nature Communications.
doi: 10.1038/s41467-023-38244-8

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