Background

Conventional messenger RNA (mRNA) vaccine technology delivers genetic material encoding specific antigens into cells, inducing the synthesis of proteins. In this process, the overall expression level of the antigen protein and its secretion efficiency are critical factors determining the vaccine's prophylactic efficacy and immunogenicity. However, existing vaccine designs have primarily focused on optimizing the amino acid sequence of the target antigen itself, leading to limitations in controlling the protein's movement along the secretory pathway after successful synthesis within the cell.

To address this, intracellular protein transport pathways are gaining attention. Signal peptides (SP), which guide proteins to the endoplasmic reticulum (ER), act as address labels for protein delivery. These peptides serve as signposts, guiding proteins produced through translation in the cytoplasm to pass through the ER and be efficiently released outside the cell. Recent research has focused on precisely manipulating these SPs to maximize the extracellular secretion efficiency of antigens. This study aims to present a novel molecular engineering strategy that can further enhance the performance of mRNA vaccines by improving the inefficient intracellular secretory pathways.

Key Findings

Chinese researchers used the receptor-binding domain (RBD) of SARS-CoV-2 as a model antigen and designed a panel of candidate SPs derived from highly secreted proteins in the human body for screening. The analysis included SPs from complement 3 (C3), which is involved in immune function in the blood, as well as interleukin-12 (IL-12) and interleukin-20 (IL-20), which are closely related to immune regulation in the body.

Experimental results with various SP combinations showed that RBD antigens containing SPs derived from C3, IL-12, and IL-20 exhibited significantly higher antigen expression and extracellular secretion levels compared to the control group. To understand how the modified SPs function specifically within cells, the researchers used fluorescence confocal microscopy to observe the changes at the subcellular level. The results showed that mRNA equipped with the engineered SPs exhibited a dramatically increased efficiency and targeting ratio of movement from the cytoplasm to the ER compared to the control group.

The practical efficacy of this mRNA vaccine candidate, incorporating the optimized intracellular transport design, was also demonstrated in animal model experiments. After administering the modified vaccine candidate to mice and analyzing the immune response, the results showed that it induced significantly stronger humoral and cellular immune responses compared to mice vaccinated with the conventional vaccine. This demonstrates that an approach that optimizes the intracellular transport of proteins, rather than simply modifying the antigen sequence, can be an effective key to improving the overall performance of vaccines.

Significance and Prospects

This research has significant academic value in that it overcomes the limitations of the mRNA vaccine platform and establishes a practical gene engineering methodology to improve expression and secretion efficiency. In the future, it is expected to facilitate the development of next-generation cancer mRNA vaccines that require high concentrations of antigen release, as well as the development of protein replacement therapies that require precise control of intracellular protein secretion for the treatment of specific protein deficiencies.

However, this study has the limitation that the screening was conducted using a specific viral RBD antigen and a limited number of cell lines. It is essential to verify in the future whether the same level of secretion enhancement effect can be reproduced when applied to various disease-specific antigens. Furthermore, subsequent verification procedures to confirm the long-term safety in vivo and the potential cytotoxicity caused by excessive protein expression and secretion are also necessary before it can smoothly enter the commercialization stage. This paper was published in the journal 'Acta Biochimica et Biophysica Sinica', and detailed information can be found on PubMed (https://pubmed.ncbi.nlm.nih.gov/42550669/).