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Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics
Open accessPreprint — published as 10.1038/s41467-022-28776-w · Paper page
Abstract
SUMMARY: Therapeutic mRNAs and vaccines are being developed for a broad range of human diseases, including COVID-19. However, their optimization is hindered by mRNA instability and inefficient protein expression. Here, we describe design principles that overcome these barriers. We develop a new RNA sequencing-based platform called PERSIST-seq to systematically delineate in-cell mRNA stability, ribosome load, as well as in-solution stability of a library of diverse mRNAs. We find that, surprisingly, in-cell stability is a greater driver of protein output than high ribosome load. We further introduce a method called In-line-seq, applied to thousands of diverse RNAs, that reveals sequence and structure-based rules for mitigating hydrolytic degradation. Our findings show that “superfolder” mRNAs can be designed to improve both stability and expression that are further enhanced through pseudouridine nucleoside modification. Together, our study demonstrates simultaneous improvement of mRNA stability and protein expression and provides a computational-experimental platform for the enhancement of mRNA medicines.
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Metadata
- Authors
- Leppek, K., Byeon, G. W., Kladwang, W., Wayment-Steele, H. K., Kerr, C. H., Xu, A. F., Kim, D. S., Topkar, V. V., Choe, C., Rothschild, D., Tiu, G. C., Wellington-Oguri, R., Fujii, K., Sharma, E., Watkins, A. M., Nicol, J. J., Romano, J., Tunguz, B., Participants, E., Barna, M., & Das, R.
- Affiliations
- Stanford University (US)
- Eterna Massive Open Laboratory (US)
- Nvidia (United States) (US)
- Corresponding author
- Barna, Maria
- Das, Rhiju
- ORCID
- Venue
- Nature Communications
- Publisher
- openRxiv
- Published
- 2021-03-30
- Volume
- 13
- Issue
- 1
- Pages
- 1536-1536
- ISSN
- 2041-1723 (ISSN-L)
- PMC
- PMC8940940
- OpenAlex
- W4220991830
- Cited by
- 399
- References
- 117
- Citations by year
- 2026: 88 · 2025: 127 · 2024: 99 · 2023: 53 · 2022: 23 · 2021: 8
- FWCI
- 24.66
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- Nature Communications (publishedVersion, cc-by)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- PubMed Central (submittedVersion, cc-by-nc)
- RePEc: Research Papers in Economics (submittedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
- Funders
- National Science Foundation
- Damon Runyon Cancer Research Foundation (DRG-2237-15)
- Pfizer
- Oregon State University
- Baidu
- National Institutes of Health
- University of California, San Francisco
- Stanford Bio-X
- Chemistry, Engineering and Medicine for Human Health, Stanford University
- Canadian Institutes of Health Research
- National Cancer Institute (R21CA219847)
- National Institute of General Medical Sciences (R35GM122579)
- Eunice Kennedy Shriver National Institute of Child Health and Human Development (F30HD100123, R01HD086634)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
- Keywords
- mRNA stability, mRNA structure, mRNA vaccines, RNA therapeutics, protein expression, hydrolytic degradation, pseudouridine modification, mRNA design, COVID-19 vaccines
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