Lipid nanoparticles outperform electroporation in mRNA-based CAR T cell engineering

Engineered T cells expressing chimeric antigen receptors (CARs) have been proven as efficacious therapies against selected hema-tological malignancies. However, the approved CAR T cell ther-apeutics strictly rely on viral transduction, a time-and cost-inten-sive procedure with possible safety issues. Therefore, the direct transfer of in vitro transcribed CAR-mRNA into T cells is pur-sued as a promising strategy for CAR T cell engineering. Electro-poration (EP) is currently used as mRNA delivery method for the generation of CAR T cells in clinical trials but achieving only poor anti-tumor responses. Here, lipid nanoparticles (LNPs) were examined for ex vivo CAR-mRNA delivery and compared with EP. LNP-CAR T cells showed a significantly prolonged effi- cacy in vitro in comparison with EP-CAR T cells as a result of extended CAR-mRNA persistence and CAR expression, attrib-uted to a different delivery mechanism with less cytotoxicity and slower CAR T cell proliferation. Moreover, CAR expression and in vitro functionality of mRNA-LNP-derived CAR T cells were comparable to stably transduced CAR T cells but were less exhausted. These results show that LNPs outperform EP and un-derline the great potential of mRNA-LNP delivery for ex vivo CAR T cell modification as next-generation transient approach for clinical studies.