Skip to main content
bioRxiv
  • Home
  • About
  • Submit
  • ALERTS / RSS
Advanced Search
New Results

Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics

View ORCID ProfileKathrin Leppek, View ORCID ProfileGun Woo Byeon, Wipapat Kladwang, View ORCID ProfileHannah K. Wayment-Steele, View ORCID ProfileCraig H. Kerr, View ORCID ProfileAdele F. Xu, View ORCID ProfileDo Soon Kim, View ORCID ProfileVed V. Topkar, Christian Choe, View ORCID ProfileDaphna Rothschild, View ORCID ProfileGerald C. Tiu, View ORCID ProfileRoger Wellington-Oguri, View ORCID ProfileKotaro Fujii, View ORCID ProfileEesha Sharma, View ORCID ProfileAndrew M. Watkins, John J. Nicol, Jonathan Romano, Bojan Tunguz, Eterna Participants, View ORCID ProfileMaria Barna, View ORCID ProfileRhiju Das
doi: https://doi.org/10.1101/2021.03.29.437587
Kathrin Leppek
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Kathrin Leppek
Gun Woo Byeon
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Gun Woo Byeon
Wipapat Kladwang
2Department of Biochemistry, Stanford University, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Hannah K. Wayment-Steele
3Department of Chemistry, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Hannah K. Wayment-Steele
Craig H. Kerr
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Craig H. Kerr
Adele F. Xu
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Adele F. Xu
Do Soon Kim
2Department of Biochemistry, Stanford University, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Do Soon Kim
Ved V. Topkar
4Program in Biophysics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Ved V. Topkar
Christian Choe
5Department of Bioengineering, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Daphna Rothschild
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Daphna Rothschild
Gerald C. Tiu
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Gerald C. Tiu
Roger Wellington-Oguri
6Eterna Massive Open Laboratory
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Roger Wellington-Oguri
Kotaro Fujii
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Kotaro Fujii
Eesha Sharma
2Department of Biochemistry, Stanford University, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Eesha Sharma
Andrew M. Watkins
2Department of Biochemistry, Stanford University, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Andrew M. Watkins
John J. Nicol
6Eterna Massive Open Laboratory
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jonathan Romano
6Eterna Massive Open Laboratory
7Department of Computer Science and Engineering, State University of New York at Buffalo, Buffalo, New York, 14260, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Bojan Tunguz
2Department of Biochemistry, Stanford University, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Eterna Participants
6Eterna Massive Open Laboratory
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Maria Barna
1Department of Genetics, Stanford University, Stanford, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Maria Barna
  • For correspondence: mbarna@stanford.edu rhiju@stanford.edu
Rhiju Das
2Department of Biochemistry, Stanford University, California 94305, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Rhiju Das
  • For correspondence: mbarna@stanford.edu rhiju@stanford.edu
  • Abstract
  • Full Text
  • Info/History
  • Metrics
  • Supplementary material
  • Preview PDF
Loading

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.

Competing Interest Statement

Stanford University has submitted provisional patent applications related to use of the Hoxa9 P4 stem-loop and the CoV2 5 UTR, computational design of mRNAs, chemically modified nucleotides to stabilize RNA therapeutics, and PERSIST-seq.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC 4.0 International license.
Back to top
PreviousNext
Posted March 30, 2021.
Download PDF

Supplementary Material

Email

Thank you for your interest in spreading the word about bioRxiv.

NOTE: Your email address is requested solely to identify you as the sender of this article.

Enter multiple addresses on separate lines or separate them with commas.
Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics
(Your Name) has forwarded a page to you from bioRxiv
(Your Name) thought you would like to see this page from the bioRxiv website.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics
Kathrin Leppek, Gun Woo Byeon, Wipapat Kladwang, Hannah K. Wayment-Steele, Craig H. Kerr, Adele F. Xu, Do Soon Kim, Ved V. Topkar, Christian Choe, Daphna Rothschild, Gerald C. Tiu, Roger Wellington-Oguri, Kotaro Fujii, Eesha Sharma, Andrew M. Watkins, John J. Nicol, Jonathan Romano, Bojan Tunguz, Eterna Participants, Maria Barna, Rhiju Das
bioRxiv 2021.03.29.437587; doi: https://doi.org/10.1101/2021.03.29.437587
Reddit logo Twitter logo Facebook logo LinkedIn logo Mendeley logo
Citation Tools
Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics
Kathrin Leppek, Gun Woo Byeon, Wipapat Kladwang, Hannah K. Wayment-Steele, Craig H. Kerr, Adele F. Xu, Do Soon Kim, Ved V. Topkar, Christian Choe, Daphna Rothschild, Gerald C. Tiu, Roger Wellington-Oguri, Kotaro Fujii, Eesha Sharma, Andrew M. Watkins, John J. Nicol, Jonathan Romano, Bojan Tunguz, Eterna Participants, Maria Barna, Rhiju Das
bioRxiv 2021.03.29.437587; doi: https://doi.org/10.1101/2021.03.29.437587

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Subject Area

  • Molecular Biology
Subject Areas
All Articles
  • Animal Behavior and Cognition (4380)
  • Biochemistry (9571)
  • Bioengineering (7084)
  • Bioinformatics (24832)
  • Biophysics (12595)
  • Cancer Biology (9949)
  • Cell Biology (14344)
  • Clinical Trials (138)
  • Developmental Biology (7943)
  • Ecology (12095)
  • Epidemiology (2067)
  • Evolutionary Biology (15980)
  • Genetics (10915)
  • Genomics (14730)
  • Immunology (9862)
  • Microbiology (23636)
  • Molecular Biology (9472)
  • Neuroscience (50824)
  • Paleontology (369)
  • Pathology (1538)
  • Pharmacology and Toxicology (2678)
  • Physiology (4009)
  • Plant Biology (8653)
  • Scientific Communication and Education (1508)
  • Synthetic Biology (2389)
  • Systems Biology (6422)
  • Zoology (1345)