mRNA-Laden Lipid-Nanoparticle-Enabled in Situ CAR-Macrophage Engineering for the Eradication of Multidrug-Resistant Bacteria in a Sepsis Mouse Model

Sepsis, which is the most severe clinical manifestation of acute infection and has a mortality rate higher than that of cancer, represents a significant global public health burden. Persistent methicillin-resistant Staphylococcus aureus (MRSA) infection and further host immune paralysis are the leading causes of sepsis-associated death, but limited clinical interventions that target sepsis have failed to effectively restore immune homeostasis to enable complete eradication of MRSA. To restimulate anti-MRSA innate immunity, we developed CRV peptide-modified lipid nanoparticles (CRV/LNP-RNAs) for transient in situ programming of macrophages (M phi s). The CRV/LNP-RNAs enabled the delivery of MRSA-targeted chimeric antigen receptor (CAR) mRNA (SasA-CAR mRNA) and CASP11 (a key MRSA intracellular evasion target) siRNA to M phi s in situ, yielding CAR-M phi s with boosted bactericidal potency. Specifically, our results demonstrated that the engineered M phi s could efficiently phagocytose and digest MRSA intracellularly, preventing immune evasion by the "superbug" MRSA. Our findings highlight the potential of nanoparticle-enabled in vivo generation of CAR-M phi s as a therapeutic platform for multidrug-resistant (MDR) bacterial infections and should be confirmed in clinical trials.