Journal
CRISPR JOURNAL
Volume 4, Issue 1, Pages 69-81Publisher
MARY ANN LIEBERT, INC
DOI: 10.1089/crispr.2020.0095
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Funding
- DOD [W81XWH2010265]
- Bruce D. and Susan J. Meyer Charitable Fund
- state of North Carolina [330054]
- U.S. Department of Defense (DOD) [W81XWH2010265] Funding Source: U.S. Department of Defense (DOD)
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This study developed a delivery method to reduce RNA off-target activities of ABEs using electroporation or lentiviral capsids. These methods enabled efficient guided base editing with stable detectable RNA off-target activities. This approach makes ABE closer to possible therapeutic applications.
Adenine base editors (ABEs) can correct gene mutations without creating double-strand breaks. However, in recent reports, these editors showed guide-independent RNA off-target activities. This work describes our development of a delivery method to minimize ABEs' RNA off-target activity. After discovering a RNA off-target hot spot for sensitive detection of RNA off-target activities, we found that delivering ribonucleoproteins (RNPs) by electroporation generated undetectable non-specific RNA editing, but on-target base editing activity was also relatively low. We then explored a lentivirus capsid-based delivery strategy to deliver ABE. We used aptamer/aptamer-binding protein (ABP) interactions to package ABE RNPs into lentiviral capsids. Capsid RNPs were delivered to human cells for highly efficient guided base editing. Importantly, RNA off-target activities from the capsid RNPs were undetectable. Our new lentiviral capsid-based ABE RNP delivery method with minimal RNA off-target activities makes ABE one step closer to possible therapeutic applications.
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