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Nanoparticle stereochemistry-dependent endocytic processing improves in vivo mRNA delivery

Nature Chemistry volume 15, pages 508–515 (2023)Cite this article

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Abstract

Stereochemistry can alter small-molecule pharmacokinetics, safety and efficacy. However, it is unclear whether the stereochemistry of a single compound within a multicomponent colloid such as a lipid nanoparticle (LNP) can influence its activity in vivo. Here we report that LNPs containing stereopure 20α-hydroxycholesterol (20α) delivered mRNA to liver cells up to 3-fold more potently than LNPs containing a mixture of both 20α- and 20β-hydroxycholesterols (20mix). This effect was not driven by LNP physiochemical traits. Instead, in vivo single-cell RNA sequencing and imaging revealed that 20mix LNPs were sorted into phagocytic pathways more than 20α LNPs, resulting in key differences between LNP biodistribution and subsequent LNP functional delivery. These data are consistent with the fact that nanoparticle biodistribution is necessary, but not sufficient, for mRNA delivery, and that stereochemistry-dependent interactions between LNPs and target cells can improve mRNA delivery.

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Fig. 1: In vivo LNP-mediated mRNA delivery varies batch-to-batch.
Fig. 2: Quantifying in vivo LNP-mediated mRNA delivery at single-cell resolution.
Fig. 3: The cellular response to 20α and 20mix LNPs.
Fig. 4: Stereochemistry-dependent mRNA delivery is not LNP specific.

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Data availability

All RNA sequencing data are available at GEO (GSE181333). All other data are represented in the main figures or supplementary figures.

Code availability

All code used to analyse the data is available at https://github.com/Jack-Feldman/barcode_count.

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Acknowledgements

The authors thank S. Durham, R. Hughley, D. Gulick, N. Djeddar at Georgia Tech, and D. Alexis at the Emory Winship Cancer Institute. The authors thank K. Tiegren. This work was funded by the National Institutes of Health (R01GM132985, awarded to J.E.D., and UG3-TR002855, awarded to J.E.D. and P.J.S.).

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Author notes

  1. These authors contributed equally: Marine Z. C. Hatit, Curtis N. Dobrowolski.

Authors and Affiliations

  1. Wallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory University School of Medicine, Atlanta, GA, USA

    Marine Z. C. Hatit, Curtis N. Dobrowolski, Melissa P. Lokugamage, David Loughrey, Huanzhen Ni, Alejandro J. Da Silva Sanchez, Afsane Radmand, Sebastian G. Huayamares, Ryan Zenhausern, Kalina Paunovska, Jinwhan Kim, Manaka Sato, Jacob I. Feldman, Michael-Alexander Rivera, Ana Cristian, YongTae Kim, Philip J. Santangelo & James E. Dahlman

  2. Parker H. Petit Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, GA, USA

    Chiara Zurla, Hannah E. Peck & YongTae Kim

  3. School of Electrical & Computer Engineering, Georgia Institute of Technology, Atlanta, GA, USA

    Jinwhan Kim

  4. George W. Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, GA, USA

    YongTae Kim

  5. Institute for Electronics and Nanotechnology, Georgia Institute of Technology, Atlanta, GA, USA

    YongTae Kim

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Contributions

M.Z.C.H., C.Z., P.J.S. and J.E.D. designed the experiments. All authors helped to perform the experiments. M.Z.C.H. and J.E.D. wrote the initial draft, which was sent to the other authors.

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Correspondence to James E. Dahlman.

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J.E.D. is an advisor to GV. All other authors declare no competing interests.

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Nature Chemistry thanks Hideyoshi Harashima and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Statistical data for Figs. 1 and 4 and Supplementary Figs. 3, 4 and 11.

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Hatit, M.Z.C., Dobrowolski, C.N., Lokugamage, M.P. et al. Nanoparticle stereochemistry-dependent endocytic processing improves in vivo mRNA delivery. Nat. Chem. 15, 508–515 (2023). https://doi.org/10.1038/s41557-023-01138-9

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