Abstract
The genetic modification of T cells has advanced cellular immunotherapies, yet the delivery of biologics specifically to T cells remains challenging. Here we report a suite of methods for the genetic engineering of cells to produce extracellular vesicles (EVs)—which naturally encapsulate and transfer proteins and nucleic acids between cells—for the targeted delivery of biologics to T cells without the need for chemical modifications. Specifically, the engineered cells secreted EVs that actively loaded protein cargo via a protein tag and that displayed high-affinity T-cell-targeting domains and fusogenic glycoproteins. We validated the methods by engineering EVs that delivered Cas9–single-guide-RNA complexes to ablate the gene encoding the C-X-C chemokine co-receptor type 4 in primary human CD4+ T cells. The strategy is amenable to the targeted delivery of biologics to other cell types.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 397-414 |
| Number of pages | 18 |
| Journal | Nature Biomedical Engineering |
| Volume | 8 |
| Issue number | 4 |
| DOIs | |
| State | Published - Apr 2024 |
Funding
We thank R. D\u2019Aquila for his support and guidance in starting this project. We thank I. Clerc for her assistance with the measles virus glycoproteins. J.N.L. discloses support for the research described in this study from Third Coast Center for AIDS Research, an NIH-funded centre (P30 AI117943), NIH grants R01AI165236 and R01AI150998 (J.F.H.), National Science Foundation (NSF) award 1844219 (J.N.L. and N. P. Kamat), Kairos Ventures (gift), and Syenex. This work was also supported by NSF Graduate Research Fellowship awards DGE-1324585 (to D.M.S.) and DGE-1842165 (to B.N.D.). Sanger sequencing was performed through the Northwestern University Sequencing Core (NUSeq) Core Facility of Northwestern\u2019s Center for Genetic Medicine and a partnership with ACGT. NanoSight analysis was performed in the Analytical bioNanoTechnology Core Facility (ANTEC) of the Simpson Querrey Institute at Northwestern University. ANTEC is currently supported by the Soft and Hybrid Nanotechnology Experimental Resource (NSFECCS-1542205). We thank C. Wilke for her assistance with TEM. TEM was performed at the BioCryo facility of Northwestern University\u2019s Atomic and Nanoscale Characterization Experimental (NUANCE) Center, which has received support from the Soft and Hybrid Nanotechnology Experimental Resource (NSF ECCS-1542205); the Materials Research Science and Engineering Centers (MRSEC) program (NSF DMR-1720139) at the Materials Research Center; the International Institute for Nanotechnology; and the State of Illinois, through the International Institute for Nanotechnology. It also made use of the CryoCluster equipment, which has received support from the Major Research Instrumentation (MRI) program (NSF DMR-1229693). We thank H. Edelstein for her assistance with confocal microscopy. Microscopy was performed at the Biological Imaging Facility at Northwestern University (RRID:SCR_017767), graciously supported by the Chemistry for Life Processes Institute, the Northwestern University Office for Research, and the Department of Molecular Biosciences. We thank P. Mehl for his assistance with FACS. Flow cytometry was performed at the Northwestern University Robert H. Lurie Comprehensive Cancer Center (RHLCCC) Flow Cytometry Facility, which is supported by a Cancer Center Support Grant (NCI CA060553). We thank J. Brink and S. Hockema at 496code for their assistance with HTS data analysis.
ASJC Scopus subject areas
- Biotechnology
- Bioengineering
- Medicine (miscellaneous)
- Biomedical Engineering
- Computer Science Applications