Abstract
Lipid nanoparticle (LNP)–mRNA complexes are transforming medicine. However, the medical applications of LNPs are limited by their low endosomal disruption rates, high toxicity and long tissue persistence times. LNPs that rapidly hydrolyse in endosomes (RD-LNPs) could solve the problems limiting LNP-based therapeutics and dramatically expand their applications but have been challenging to synthesize. Here we present an acid-degradable linker termed ‘azido-acetal’ that hydrolyses in endosomes within minutes and enables the production of RD-LNPs. Acid-degradable lipids composed of polyethylene glycol lipids, anionic lipids and cationic lipids were synthesized with the azido-acetal linker and used to generate RD-LNPs, which significantly improved the performance of LNP–mRNA complexes in vitro and in vivo. Collectively, RD-LNPs delivered mRNA more efficiently to the liver, lung, spleen and brains of mice and to haematopoietic stem and progenitor cells in vitro than conventional LNPs. These experiments demonstrate that engineering LNP hydrolysis rates in vivo has great potential for expanding the medical applications of LNPs.
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Data availability
The authors declare that the data supporting the findings of this study are available within the Article and its Supplementary Information files. Should any raw data files be needed in another format, they are available from the corresponding authors upon reasonable request. Source data are provided with this paper.
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Acknowledgements
N.M. thanks the California Institute for Regenerative Medicine (CIRM) DISC2-14045, and the NIH for NIAID award number UM1AI164559, co-funded by NHLBI, NIMH, NIDA, NIDDK and NINDS. N.M. also thanks NIH grants UG3NS115599, R33 and R61DA048444-01, R01MH125979-01, funding from the BAKAR Spark award, the Cystic Fibrosis Foundation, the Innovative Genomics Institute, the CRISPR Cures for Cancer Initiative and the Heritage Medical Research Institute. Cryo-TEM data were collected at the Cal-Cryo facility at the University of California, Berkeley Institute for Quantitative Biosciences (QB3). A.E.B. thanks the NIH for funding this work with a Pioneer Award, grant number 1DP1OD029517-01 and J. J. Truchard and the Truchard Foundation. J.E.N. was funded by grant NNF21OC0068675 from the Novo Nordisk Foundation and the Stanford Bio-X Program. A.W. thanks the NIH grant with number 1R21NS133881-01, California Institute for Regenerative Medicine (CIRM) DISC2-14097, Shriners Children’s basic research award 85400-NCA-24. We thank SLAC for SAXS beamtime, and T. Weiss for support during the SAXS experiment. Use of the Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, is supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences under contract no. DE-AC02-76SF00515. The SSRL Structural Molecular Biology Program is supported by the DOE Office of Biological and Environmental Research, and by the NIH, National Institute of General Medical Sciences (P30GM133894). We also thank the Doudna laboratory for help with the HSPCs transfection experiments.
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N.M., A.W. and H.H. conceived the study and supervised the project. S.Z. and M.S. synthesized compounds. S.Z. performed the characterizations of LNPs. H.H. performed cell culture experiments. H.H., K.G., B.Y., H.S., B.W.B. and A.W. designed and conducted most of the in vivo experiments and data analysis. A.P., J.T., M. G. Collins and Y.-J.C. performed brain editing experiments. H.L. supervised brain editing experiments. B.S.P. performed HSPC transfection experiments. O.H.A. performed the quantum dots/LNPs imaging experiments, and A.M.S. supervised these experiments. A.L., J.E.N. and S.P. performed the SAXS and cryo-TEM experiments, and A.E.B. supervised these experiments. N.M. wrote the paper with contributions from all authors, and N.M., S.Z., H.H. and R.S. finalized and corrected the paper with input from all authors.
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The authors declare the following competing interests: H.H., K.G. and N.M. own equity in Opus Biosciences. All the other authors declare no competing interests.
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Detailed methods of experiments in this paper; Supplementary Schemes 1–6, Figs. 3-1–3-4, 4-1, 5-1, 6-1–6-5, 7-1–7-5, 8-1, 9-1–9-10 and 10-1–10-8, Tables 1–11; NMR spectra of compounds 1–7; and high-resolution mass spectra of compounds 1–3.
Supplementary Video 1
The movement of quantum dots (QDs) delivered with ADA-LNPs in cell.
Supplementary Video 2
The movement of quantum dots (QDs) delivered with NDA-LNPs in cell.
Supplementary Video 3
The movement of quantum dots (QDs) delivered with Std-LNPs in cell.
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Zhao, S., Gao, K., Han, H. et al. Acid-degradable lipid nanoparticles enhance the delivery of mRNA. Nat. Nanotechnol. (2024). https://doi.org/10.1038/s41565-024-01765-4
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DOI: https://doi.org/10.1038/s41565-024-01765-4
