Targeting DNA-LNPs to Endothelial Cells Improves Expression Magnitude, Duration, and Specificity

Marzolini, N.; Brysgel, T. V.; Essien, E.-O.; Rahman, R. J.; Wu, J.; Majumder, A.; Patel, M. N.; Tiwari, S.; Espy, C. L.; Shuvaev, V. V.; Hood, E. D.; Chase, L. S.; Katzen, J. B.; Frank, D. B.; Bennett, M. L.; Marcos-Contreras, O. A.; Myerson, J. W.; Muzykantov, V. R.; Reyes-Esteves, S.; Brenner, J. S.

doi:10.1101/2025.07.09.663747

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bioRxiv · 10.1101/2025.07.09.663747

Targeting DNA-LNPs to Endothelial Cells Improves Expression Magnitude, Duration, and Specificity

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Abstract

DNA-lipid nanoparticles (DNA-LNPs) loaded with inhibitors of the cGAS-STING pathway enable safe and effective delivery of DNA in vivo. Herein, we report the first instances of extrahepatic DNA-LNP targeting. DNA-LNPs conjugated to antibodies against PECAM-1 or VCAM-1 target the endothelium of the lungs and brain/spleen, respectively. These LNPs drive robust transgene expression in their target organs, with greater magnitude and duration than untargeted LNPs. Lung specificity of PECAM-targeted transgene expression increases over two weeks, resulting in markedly higher lung-to-liver expression ratios than our previous PECAM-targeted mRNA-LNPs. Off-target liver DNA expression declines to undetectable levels but persists in the lungs, while mRNA expression uniformly decreases due to its short half-life. We further improve this expression specificity by replacing full-length antibodies with Fab fragments. Single-cell analysis reveals a key mechanism underlying the improvements in organ-specificity: target organ expression is dominated by long-lived endothelial cells, while off-target liver delivery and expression are in non-endothelial cells with shorter half-lives. Collectively, these studies demonstrate that targeted DNA-LNPs achieve high levels of organ- and cell-type-specific transgene expression and thus provide a therapeutic platform for dozens of endothelial-centric diseases.

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Marzolini, N., Brysgel, T. V., Essien, E.-O., Rahman, R. J., Wu, J., Majumder, A., Patel, M. N., Tiwari, S., Espy, C. L., Shuvaev, V. V., Hood, E. D., Chase, L. S., Katzen, J. B., Frank, D. B., Bennett, M. L., Marcos-Contreras, O. A., Myerson, J. W., Muzykantov, V. R., Reyes-Esteves, S., Brenner, J. S.. 2025-07-12. Targeting DNA-LNPs to Endothelial Cells Improves Expression Magnitude, Duration, and Specificity. https://doi.org/10.1101/2025.07.09.663747

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