Abstract
Naturally generated lipid nanoparticles termed extracellular vesicles (EVs) hold significant promise as engineerable therapeutic delivery vehicles. However, active loading of protein cargo into EVs in a manner that is useful for delivery remains a challenge. Here, we demonstrate that by rationally designing proteins to traffic to the plasma membrane and associate with lipid rafts, we can enhance loading of protein cargo into EVs for a set of structurally diverse transmembrane and peripheral membrane proteins. We then demonstrate the capacity of select lipid tags to mediate increased EV loading and functional delivery of an engineered transcription factor to modulate gene expression in target cells. We envision that this technology could be leveraged to develop new EV-based therapeutics that deliver a wide array of macromolecular cargo.
| Original language | English (US) |
|---|---|
| Article number | 5618 |
| Journal | Nature communications |
| Volume | 15 |
| Issue number | 1 |
| DOIs | |
| State | Published - Dec 2024 |
Funding
We thank the Kamat and Leonard labs for providing feedback and proofreading the manuscript. We thank Jan Steink\u00FChler for helpful discussions on how to characterize lipid-protein interactions. We thank Patrick Donahue for generating the landing pad destination backbone used in this study. We thank the Tullman-Ercek Lab for the use of their chemiluminescent imager (Azure c600). This work was supported in part by the McCormick Research Catalyst Program at Northwestern University (J.N.L., N.P.K.); National Science Foundation under Grant No. 1844219 (N.P.K., J.N.L.) and Grant No. 2145050 (N.P.K., T.F.G.); J.A.P., T.F.G., and H. I. E. were supported by NSF Graduate Research Fellowships (DGE-1842165). Any opinions, findings, and conclusions or recommendations expressed in this material are those of the authors(s) and do not necessarily reflect the views of the National Science Foundation. J.A.P. gratefully acknowledges support from the Ryan Fellowship, and the International Institute for Nanotechnology at Northwestern University. T.F.G. was supported in part by the Northwestern University Graduate School Cluster in Biotechnology, Systems, and Synthetic Biology, which is affiliated with the Biotechnology Training Program. This work was supported by the Northwestern University\u2014Flow Cytometry Core Facility supported by Cancer Center Support Grant (NCI CA060553). This work made use of the BioCryo facility of Northwestern University\u2019s NUANCE Center, which has received support from the SHyNE Resource (NSF ECCS-2025633), the IIN, and Northwestern\u2019s MRSEC program (NSF DMR-1720139). This work was supported by the Northwestern University Sanger Sequencing Facility. This work made use of the Keck Biophysics Facility, a shared resource of the Robert H. Lurie Comprehensive Cancer Center of Northwestern University supported in part by the NCI Cancer Center Support Grant #P30 CA060553. This work was also supported by the Keck Biophysics Facility\u2019s Azure Sapphire Imager which is funded by NIH grant 1S10OD026963-01 NIH grant. Biological and chemical analysis was performed in the Analytical bioNanoTechnology Core Facility of the Simpson Querrey Institute at Northwestern University. The U.S. Army Research Office, the U.S. Army Medical Research and Materiel Command, and Northwestern University provided funding to develop this facility and ongoing support is being received from the Soft and Hybrid Nanotechnology Experimental (SHyNE) Resource (NSF ECCS-1542205). We thank the Kamat and Leonard labs for providing feedback and proofreading the manuscript. We thank Jan Steink\u00FChler for helpful discussions on how to characterize lipid-protein interactions. We thank Patrick Donahue for generating the landing pad destination backbone used in this study. We thank the Tullman-Ercek Lab for the use of their chemiluminescent imager (Azure c600). This work was supported in part by the McCormick Research Catalyst Program at Northwestern University (J.N.L., N.P.K.); National Science Foundation under Grant No. 1844219 (N.P.K., J.N.L.)\u00A0and Grant No.\u00A02145050 (N.P.K., T.F.G.); J.A.P., T.F.G., and H. I. E. were supported by NSF Graduate Research Fellowships (DGE-1842165). Any opinions, findings, and conclusions or recommendations expressed in this material are those of the authors(s) and do not necessarily reflect the views of the National Science Foundation. J.A.P. gratefully acknowledges support from the Ryan Fellowship, and the International Institute for Nanotechnology at Northwestern University. T.F.G. was supported in part by the Northwestern University Graduate School Cluster in Biotechnology, Systems, and Synthetic Biology, which is affiliated with the Biotechnology Training Program. This work was supported by the Northwestern University\u2014Flow Cytometry Core Facility supported by Cancer Center Support Grant (NCI CA060553). This work made use of the BioCryo facility of Northwestern University\u2019s NUANCE Center, which has received support from the SHyNE Resource (NSF ECCS-2025633), the IIN, and Northwestern\u2019s MRSEC program (NSF DMR-1720139). This work was supported by the Northwestern University Sanger Sequencing Facility. This work made use of the Keck Biophysics Facility, a shared resource of the Robert H. Lurie Comprehensive Cancer Center of Northwestern University supported in part by the NCI Cancer Center Support Grant #P30 CA060553. This work was also supported by the Keck Biophysics Facility\u2019s Azure Sapphire Imager which is funded by NIH grant 1S10OD026963-01 NIH grant. Biological and chemical analysis was performed in the Analytical bioNanoTechnology Core Facility of the Simpson Querrey Institute at Northwestern University. The U.S. Army Research Office, the U.S. Army Medical Research and Materiel Command, and Northwestern University provided funding to develop this facility and ongoing support is being received from the Soft and Hybrid Nanotechnology Experimental (SHyNE) Resource (NSF ECCS-1542205).
ASJC Scopus subject areas
- General Chemistry
- General Biochemistry, Genetics and Molecular Biology
- General Physics and Astronomy