Improved translation efficiency of therapeutic mRNA
Abstract
Recent developments in the field of the messenger RNA and its advantages versus DNA have led to a renewed interest in mRNA-based technologies. Despite its advantages, mRNA therapy has a number of drawbacks including low amount of mRNA production, short-term existence of mRNA and mRNA-mediated protein within the cell, severe mRNA cytotoxicity, and immune response activation following mRNA transfection. Here, we applied untranslated regions of human beta-globin to increase the stability and translation efficiency of a destabilized GFP mRNA. In order to suppress the innate immune response, which is the main barrier of mRNA therapy, we used the vaccinia virus derived capping enzyme and substituted standard nucleotides with modified nucleotides. At the end, the Kozak sequence of human beta-globin was replaced with the strongest sequence for the further improvement of mRNA translation. Overall, these modifications with native Kozak (K1) sequence of human beta-globin enhanced the stability of destabilized GFP mRNA up to 48 h and no increase in the level of interferon-α and -β was found. The GFP expression of mRNA with modified Kozak (K2) sequence initiated earlier than mRNA and plasmid DNA with K1 sequence. In contrast to mRNA with K1 sequence, the cells containing mRNA with K2 sequence remained positive for GFP expression up to 72 h post-transfection. Interestingly, transfection efficiency and mean fluorescence intensity (MFI) of mRNA with K2 sequence were higher than mRNA and plasmid DNA with K1 sequence. Taken together, these results provide valuable information for the optimization of mRNA stability and translation. Therefore, the methods used in the current study can successfully be applied for reprogramming, gene editing, trans-differentiation, tumour therapy, and gene therapy.
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mRNA contains 5′ and 3′ UTRs, whose functions are related, respectively, to regulating translation and maintaining mRNA stability.
“mRNA contains 5′ and 3′ UTRs, whose functions are related, respectively, to regulating translation and maintaining mRNA stability.131”
From Advances in COVID-19 mRNA vaccine development (2022) · cites this paper as [131]
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An Overview of the Progress Made on the Coronavirus Vaccine
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- Authors
- Zarghampoor, F., Azarpira, N., Khatami, S. R., Behzad-Behbahani, A., & Foroughmand, A. M.
- Affiliations
- Shahid Chamran University of Ahvaz (IR)
- Shiraz University (IR)
- Corresponding author
- Azarpira, Negar
- Venue
- Gene
- Publisher
- Elsevier BV
- Published
- 2019-07
- Volume
- 707
- Pages
- 231-238
- ISSN
- 0378-1119 (ISSN-L)
- OpenAlex
- W2944311326
- Cited by
- 72
- References
- 36
- Citations by year
- 2026: 9 · 2025: 14 · 2024: 11 · 2023: 14 · 2022: 10 · 2021: 3 · 2020: 11
- FWCI
- 1.99
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Gene (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institute for Medical Research Development (958923)
- Iranian National Science Foundation (94808805)
- Shiraz University of Medical Sciences
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
- Keywords
- mRNA translation efficiency, mRNA stability, Kozak sequence, mRNA capping, modified nucleotides, innate immune response, interferon-alpha, interferon-beta, mRNA transfection
- MeSH terms
- Animals, Gene Expression Regulation, Humans, RNA, Messenger, Transfection, 3' Untranslated Regions, RNA Stability, Green Fluorescent Proteins, beta-Globins, Translational Research, Biomedical, HEK293 Cells
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