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Nanoparticle single-cell multiomic readouts reveal that cell heterogeneity influences lipid nanoparticle-mediated messenger RNA delivery

Abstract

Cells that were previously described as homogeneous are composed of subsets with distinct transcriptional states. However, it remains unclear whether this cell heterogeneity influences the efficiency with which lipid nanoparticles (LNPs) deliver messenger RNA therapies in vivo. To test the hypothesis that cell heterogeneity influences LNP-mediated mRNA delivery, we report here a new multiomic nanoparticle delivery system called single-cell nanoparticle targeting-sequencing (SENT-seq). SENT-seq quantifies how dozens of LNPs deliver DNA barcodes and mRNA into cells, the subsequent protein production and the transcriptome, with single-cell resolution. Using SENT-seq, we have identified cell subtypes that exhibit particularly high or low LNP uptake as well as genes associated with those subtypes. The data suggest that cell subsets have distinct responses to LNPs that may affect mRNA therapies.

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Fig. 1: SENT-seq uses orthogonal capture sequences to generate tunable multiomic readouts.
Fig. 2: In vivo multiomic single-cell readouts of the transcriptome, functional LNP-mediated mRNA delivery and LNP-mediated DNA barcode delivery.
Fig. 3: Cell subsets differentially uptake LNPs.
Fig. 4: Endothelial cell subtypes show transcriptional differences that may dictate LNP-mediated mRNA delivery.
Fig. 5: Chemically distinct LNPs exhibit different tropism within the liver microenvironment.

Data availability

All raw sequencing data (GEO: GSE186395) are available online. All other data are provided in the main text or the Supplementary Information.

Code availability

The custom code used is available at https://github.com/Jack-Feldman/barcode_count.

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Acknowledgements

We thank K. Tiegren for copyediting the manuscript. Funding was provided by the National Institutes of Health (grant nos. UG3-TR002855 (J.E.D. and P.J.S.) and R01DE026941 (J.E.D.)).

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Contributions

Conceptualization: C.D., K.P. and J.E.D. Methodology: C.D., K.P. and J.E.D. Investigation: C.D., K.P., E.S.E., D.L., A.J.D.S.S., H.N., M.Z.C.H., M.P.L., Y.K., H.E.P., P.J.S. and J.E.D. Visualization: C.D., K.P. and J.E.D. Funding acquisition: J.E.D. and P.J.S. Project administration: J.E.D. Supervision: P.J.S. and J.E.D. Writing—original draft: C.D., K.P. and J.E.D. Writing—review and editing: C.D., K.P., E.S.E., D.L., A.J.D.S.S., H.N., M.Z.C.H., M.P.L., Y.K., H.E.P., P.J.S. and J.E.D.

Corresponding author

Correspondence to James E. Dahlman.

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C.D., K.P. and J.E.D. have filed intellectual property related to SENT-seq. J.E.D. is an advisor for GV. All other authors declare no competing interests.

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Dobrowolski, C., Paunovska, K., Schrader Echeverri, E. et al. Nanoparticle single-cell multiomic readouts reveal that cell heterogeneity influences lipid nanoparticle-mediated messenger RNA delivery. Nat. Nanotechnol. 17, 871–879 (2022). https://doi.org/10.1038/s41565-022-01146-9

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