(436f) Systemic RNA Delivery By Phosphatidylserine Lipid Nanoparticles for Efficient Immunotherapy

The development of mRNA-lipid nanoparticles (mRNA-LNP) marks a revolutionary advance in medical science by enabling diverse therapeutic applications, including cancer vaccines and immune cell engineering, through targeted and efficient mRNA delivery to specific cells without adverse immune responses. Systemic RNA delivery to the secondary lymphoid organs (SLOs) for immunotherapy offers distinct advantages over traditional intramuscular methods, including an increased amount of antigen administration and direct access to most immune cells and lymphoid organs in the circulatory system. Current strategies of SLO-targeted delivery primarily rely on a passive mechanism that involves introducing an excessive amount of anionic charge to the formulation. However, this method comes at the cost of reduced transfection efficiency. Here, we report a new LNP system incorporating phosphatidylserine (PS) for the efficient systemic delivery of mRNA to SLOs. PS is known to bind endocytosis receptors on macrophages and is used by many viruses for enhanced cellular uptake. Our initial result demonstrated that the PS-LNP performed efficient mRNA expression in the spleen and lymph nodes. Mechanistic studies have verified its SLOs-targeting effect is mediated by monocytes/macrophages. A Design of Experiment (DOE) approach was further implemented to optimize the PS-LNP for improved mRNA transfection. The optimized formulation was validated in primary macrophages and in vivo, exhibiting more potent luciferase-mRNA expression than commercial formulations. Furthermore, it was validated to perform macrophage-specific expression by Cre-mediated editing as well as Chimeric-antigen engineering experiment, indicating promising in vivo macrophage engineering applications. Additionally, it induced potent antigen-specific T-cell response upon systemic vaccination, suggesting its viability as a promising platform for systemic immunotherapy.