Engineering Chimeric Antigen T Cells Using Protein Delivery Mediated by Nanolipoprotein Particles 2197

Abstract Description Personalized cancer therapies using Chimeric Antigen Receptor T cells (CAR-T) have improved the survival rates of some blood cancers, but current technologies to manufacture CAR-T cells are expensive and time-consuming. Our group has developed a unique transgene-free manufacturing strategy, where a CAR protein enveloped in a nanolipoprotein particle (NLP) is delivered to human T cells in vitro. To test the viability of this engineering strategy, we treated human T cells with CAR-NLP particles in the presence of immune activating reagents. This resulted in detectable CAR uptake in > 50% of T cells at 24 hours as measured by flow cytometry. Cell viability remained high during CAR-NLP dosing (>80%), indicating low toxicity associated with delivery of the CAR protein. Treated T cells were stained with a fluorescent CD19 protein to test if integration of the CAR protein was successful. CD19 staining on CAR-NLP treated cells was comparable to positive antibody controls while cells treated with the vehicle (empty NLPs) showed very low CD19 binding, indicating that CAR-NLP treated T cells can bind the target ligand specific to the engineered CAR construct. These data indicate that delivery of a CAR protein encased in an NLP may be a viable cell engineering strategy. Overall, this approach does not require genetic modifications of cells and can be performed in < 24 hours. Thus, we believe it could enable faster, cheaper, and safer engineering of CAR-T cells to better treat cancers. Funding Sources Funding provided by LLNL LDRD 22-DR-001. Patent pending. This work performed under the auspices of the U.S. Department of Energy by Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344. Topic Categories Vaccines and Immunotherapy (VAC)