Enhancing CAR‐NK Cells Against Solid Tumors Through Chemical and Genetic Fortification with DOTAP‐Functionalized Lipid Nanoparticles

Abstract Natural killer (NK) cells are crucial in the innate immune response and show promise in cancer immunotherapy, but face challenges in activation and targeted gene delivery. In this study, bifunctional lipid nanoparticles (DLNPs) containing 1,2‐dioleoyl‐3‐trimethylammonium‐propane (DOTAP), designed to bolster the antitumor efficacy of chimeric antigen receptor‐modified NK (CAR‐NK) cells by facilitating activation and efficient CAR mRNA delivery are introduced. The DLNPs, created by functionalizing FDA‐approved LNP compositions, exhibit excellent mRNA encapsulation and colloidal stability. NK cells primed with DLNPs show increased cytotoxicity against cancer cells via extracellular signal‐regulated kinase/Mitogen‐Activated Protein Kinase pathway modulation and mitochondrial dynamics changes. The DLNPs enter NK cells predominantly through clathrin‐mediated endocytosis, boosting mRNA delivery and overcoming NK cells' resistance to genetic manipulation. CAR‐NK cells targeting glypican‐3, prevalent in hepatocellular carcinoma, show significant therapeutic effects in an orthotopic mouse model. These findings underscore the potential of DLNPs in enhancing CAR‐NK cell therapy for solid tumors, marking a significant stride in NK cell‐based cancer immunotherapy and broadening prospects for NK cell‐related disease interventions.