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Lipid Nanoparticle-Enabled Intracellular Delivery of Prime Editors


Prime editing is an advanced gene editing platform with potential to correct almost any disease-causing mutation. As genome editors have evolved, their size and complexity have increased, hindering delivery technologies with low-carrying capacity and endosomal escape. We formulated an array of lipid nanoparticles (LNPs) containing prime editors (PEs). We were able to encapsulate PEs in LNPs and confirmed the presence of PE mRNA and two different guide RNAs using HPLC. In addition, we developed a novel reporter cell line for rapid identification of LNPs suited for prime editing. A 54% prime editing rate was observed with enhanced LNPs (eLNPs) containing the cholesterol analog β-sitosterol at optimal ratios of RNA cargoes. eLNPs displayed a polyhedral morphology and a more fluid membrane state that led to improved endosomal escape, eventually causing onset of editing within 9 h and reaching maximum efficiency after 24 h. Hence, PEs delivered using LNPs can propel a new wave of therapies for many additional targets potentially enabling a range of new applications.

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prime editor


prime editor and pegRNA


prime editor with pegRNA and nsgRNA


prime editing guideRNA


nicking single guideRNA


emerald green fluorescent protein


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We thank the Multiscale Microscopy Core (MMC) with technical support from the Oregon Health and Science University (OHSU) and the OHSU Center for Spatial Systems Biomedicine (OCSSB) for assistance with cryoTEM. We thank Michael Henderson for assistance with cell culture for cell screens and Antony Jozic with technical assistance. We would like to thank Anindit Mukherjee for initial help in designing HAP1 reporter with G.S and M.H.B. We acknowledge expert technical assistance by the staff in the Advanced Multiscale Microscopy Shared Resource, supported by the OHSU Knight Cancer Institute (NIH P30 CA069533) and the Office of the Senior Vice President for Research. Equipment purchases included support by the OHSU OCSSB, the MJ Murdock Charitable Trust, and the Collins Foundation.


This project was supported through funding from the National Heart Lung and Blood Institute (NHLBI) R01HL146736-01 (G.S), SAHAY 19XX0 (G.S) and National Eye Institute (NEI) 1R01EY033423-01A1 (G.S.). This study was also supported by Independent Research Fund Denmark (grant number DFF-9041-00198B to C.F.). We are grateful to the Graduate School of Health and Medical Sciences, University of Copenhagen, for funding the stay of A.L. in G.S.´s lab.

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G.S. conceived of the idea and directed research. G.S. and C.F. acquired funding. G.S. and M.H-B. designed the experiments. M.H-B., M.G., A.L., K. V performed the research. M.H-B, M.G., A.L. and K.V. contributed to data analysis. M.H-B. and G.S. wrote the manuscript with input from all authors.

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Correspondence to Gaurav Sahay.

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G.S. is an inventor in patent application US20200129445A1 that details LNP-Sito.

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Herrera-Barrera, M., Gautam, M., Lokras, A. et al. Lipid Nanoparticle-Enabled Intracellular Delivery of Prime Editors. AAPS J 25, 65 (2023).

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