Intranasal CRISPR lipid nanoparticles targeting MAPK9 attenuate neuroinflammation after traumatic brain injury

Traumatic brain injury (TBI) induces a sustained neuroinflammatory response involving activated microglia and infiltrating myeloid cells, contributing to secondary brain damage and long-term neurological dysfunction. Modulating these inflammatory responses toward a more reparative phenotype represents a promising therapeutic strategy, but achieving targeted delivery within the injured brain remains a major challenge. Here, we developed a targeted, non-viral gene-editing platform using lipid nanoparticles (LNPs) encapsulating CRISPR-Cas12a components directed against MAPK9, a key mediator of inflammatory signaling. LNPs were functionalized with an Iba-1 antibody to enhance targeting of Iba-1 + myeloid cells following intranasal administration. In primary bone marrow-derived macrophages and primary microglia, CRISPR-mediated MAPK9 targeting reduced MAPK9 expression and suppressed pro-inflammatory activation, decreasing iNOS, NLRP3, CD80, and CCL2 while increasing the anti-inflammatory/reparative markers CD206 and Arg1. In a mouse model of TBI, intranasally delivered Iba-1-targeted CRISPR-LNPs showed preferential association with Iba-1 + cells compared with NeuN+ neurons in the injured cortex and reduced MAPK9 expression within Iba-1 + cells. CRISPR-LNP treatment attenuated microglial/macrophage activation, reduced pro-inflammatory cytokine expression, and decreased iNOS+/Iba-1 + cells while increasing CD206+/Iba-1 + cells in the peri-contusional cortex, supporting a shift toward a less inflammatory phenotype. Treatment also exhibited a favorable safety profile, with no detectable toxicity in the major organs examined. Together, these findings demonstrate that intranasal delivery of Iba-1-targeted CRISPR-LNPs enables effective MAPK9 modulation in Iba-1 + myeloid cells within the injured brain and attenuates acute neuroinflammation following TBI. This non-invasive therapeutic platform provides a promising approach for targeted modulation of neuroinflammatory responses after brain injury.