Cancer immunotherapy has become a standard pillar of cancer treatment throughout the last decade. Immune checkpoint inhibitors (ICIs) and adoptive cell therapies (ACTs) such as tumor-infiltrating lymphocytes, tumor-specific T-cell receptor-modified T-cells, and chimeric antigen receptor (CAR)-engineered T-cells have proven successful in initiating an immunologic response against various cancers. Unfortunately, not all patients respond to ICIs and ACTs requires cumbersome and expensive processes.For currently approved CAR-T therapies, T-cells are first extracted from the patient by leukapheresis followed by being transduced with lentivirus that encodes the CAR. CAR-expressing T-cells are then expanded over a two-week period before finally being re-administrated to the patient. Besides the substantial manufacturing costs associated with this therapy, CAR-T cells often cause toxicity-related side effects upon infusion.This thesis aims to develop a strategy that allows for the generation of transient CAR-transfected T-cells in situ to treat hematological malignancies and thus overcome the lim-itations associated with currently approved CAR-T therapies. The technology uses in vitro transcribed synthetic mRNA that is formulated in lipid nanoparticles (LNPs) as a systemically injectable drug. The LNPs are directed to circulating T-cells via an antibody-based targeting moiety to transiently express disease-specific receptors, thereby bypassing the need to extract, culture, and reinfuse lymphocytes into patients. The thesis then aims to demonstrate the broad applicability of the technology beyond hematological malignancies by targeting other immune cell subsets to overcome solid tumors.First, we demonstrate that the technology can be used to deliver reporter mRNA specifi-cally to human T-cells. We then show that the platform enables specific delivery of functional CAR-encoding mRNA to circulating T-cells directly in vivo. Finally, we extend the application of the technology to reprogram T-cells to become bi-specific T-cell engager-secreting factories, thereby reducing solid tumor burden in mice.Overall, this thesis demonstrates the development and potential applications of a novel groundbreaking immunotherapeutic technology. This technology holds strong promise to be employed in the clinic to treat various cancers and thus improve the lives of patients worldwide
