Abstract
Two key challenges in translating messenger RNA-based lipid nanoparticle (mRNA-LNP) cancer vaccines to clinical use are limited mRNA expression and unavoidable inflammatory responses. Here we develop a membrane-destabilizing zwitterionic ionizable lipid that enhances mRNA expression by promoting endosomal escape while reducing inflammatory reactogenicity. This lipid features a pyridine-based carboxybetaine (PyCB) zwitterionic headgroup, biodegradable multitailed alkyl chains and a tertiary amine linker. The PyCB headgroup forms a zwitterionic PyCB–water complex that protonates to a positively charged state below pH 6.8. This allows for good biocompatibility at physiological pH and strong protonation in endosomes, enabling earlier and more efficient mRNA release when synergistic with the tertiary amine and the tail moiety. Incorporating membrane-destabilizing zwitterionic lipids into the LNP formulation used in a commercially available mRNA vaccine significantly boosts mRNA expression in antigen-presenting cells within lymph nodes, enhancing cytotoxic T cell activation. In addition, these membrane-destabilizing zwitterionic lipid-containing nanoparticles show reduced inflammation and neutrophil infiltration at the injection site due to their zwitterionic property. These lipids are also compatible with existing targeted nanoparticle formulations, further improving mRNA delivery.
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All data supporting the findings of this study are available within the Article and its supplementary files. Any additional requests for information can be directed to and will be fulfilled by the corresponding author. Source data are provided with this paper.
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Acknowledgements
We acknowledge financial support from the National Institute of Allergy and Infectious Disease (1R01AI178125-01A1 and 1R01AI193334-01) and start-up support from Cornell University, including Robert Langer ’70 Family and Friends Professorship, Cornell NEXT Nano Initiative and Cornell Engineering’s inaugural Sprout Award. Animal IVIS images were acquired through the Cornell Institute of Biotechnology’s Imaging Facility with NIH S10OD025049. We thank the Weiss lab at Cornell for the kind gift of EO771 cells; T. Totman, E. Feldman and F. Burgus at Cornell CARE for service and advice on caring for animals; the Cornell IACUC for help with composing and managing animal protocols; and the NIH Tetramer Core Facility (NIH Contract 75N93020D00005 and RRID:SCR_026557) for providing anti-H2Kb OVA tetramer-SIINFEKL antibody. Elements in Supplementary Fig. 7 were created with BioRender.com.
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Y.Z. and S.J. devised the project and wrote the paper. Y.Z., R.L., P.L., J.W., Y.C., Y.M., Z.C., M.C. and S.L. carried out the experimental work and analysed the data. E.W. and A.L. provided the supplies for cell experiments. Y.D., Y.H., Z.T., C.T., S.C., H.Y., D.L., W.G. and S.B. contributed to the discussion of the experimental results.
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S.J. and Y.Z. are authors of a US Provisional Application (No. 63/723,698) related to this work filed on 22 November 2024 and S.J., Y.Z., Y.H. and P.L. an International (PCT) Application (No. PCT/US2025/056506) filed on 21 November 2025 by Cornell University. The other authors declare no competing interests.
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Zhao, Y., Li, R., Liu, P. et al. Low reactogenicity and high tumour antigen expression from mRNA-LNPs with membrane-destabilizing zwitterionic lipids. Nat. Biomed. Eng (2025). https://doi.org/10.1038/s41551-025-01577-4
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DOI: https://doi.org/10.1038/s41551-025-01577-4
