Lipid nanoparticle mRNA delivery preserves CAR T cell cytotoxicity and limits exhaustion compared to electroporation

-transcribed (IVT) mRNA enables transient, non-integrating CAR expression with improved safety and scalability, making it particularly suited for non-malignant indications where prolonged persistence may not be required. Here, we systematically compare two IVT mRNA delivery platforms, electroporation and lipid nanoparticles (LNPs), for transient CAR T cell engineering in primary human T cells using single-cell transcriptomics and functional cell assays. We show that electroporation yields higher transfection efficiency and more sustained CAR surface expression, whereas LNP delivery reduces stress- and senescence-related transcriptional signatures as well as exhaustion marker expression, while enhancing antigen-driven activation, chemotactic responses, and cytotoxic function. Our comparative analysis highlights that the mode of mRNA delivery is associated with distinct transcriptional signatures and functional properties of CAR T cells, providing a framework to guide future development of mRNA-based approaches. These insights support LNP-mediated delivery as a functionally favorable strategy for transient CAR T cell engineering in autoimmune disease and beyond.