Codon-optimized Npc1 mRNA corrects Niemann-Pick type C1 disease phenotypes in vitro and in vivo.

Niemann-Pick type C1 disease is a lysosomal storage disorder caused by mutations in the NPC1 gene, resulting in the accumulation of unesterified cholesterol in multiple tissues. Despite its severity, therapeutic options remain limited. Using codon-optimized Npc1 mRNA delivered by lipid nanoparticles (Co-Npc1:LNPs), we achieved enhanced NPC1 protein expression and prolonged therapeutic activity in vitro, correcting both primary and secondary disease defects. In an Npc1 -/- mouse model, a single intravenous injection of Co-Npc1:LNP restored hepatic NPC1 protein levels, normalized autophagic flux, improved lipid abnormalities, and altered markers of liver injury. To interpret transcriptional changes post-Co-Npc1:LNP administration, we performed bulk RNA sequencing and applied MENTOR, a network-based clustering algorithm, and MENTOR-IA, an unbiased functional annotation tool. We identified transcriptionally restored pathways including cholesterol metabolism, lysosomal and mitochondrial function, and liver homeostasis. Overall, the Npc1 -/- mouse liver showed a transcriptional shift toward a more Npc1 +/+ state post-Co-Npc1:LNP treatment. Integration of single-nucleus RNA sequencing with bulk transcriptomics further revealed cell-type-specific correction of disease-associated gene expression across disease-relevant hepatic cell types. Together, we report the development and in vivo validation of the first mRNA-based therapeutic for Niemann-Pick type C1 disease.