In Vivo Endothelial Cell Gene Silencing by siRNA‐LNPs Tuned with Lipoamino Bundle Chemical and Ligand Targeting

Abstract

Although small‐interfering RNAs (siRNAs) are specific silencers for numerous disease‐related genes, their clinical applications still require safe and effective means of delivery into target cells. Highly efficient lipid nanoparticles (LNPs) are developed for siRNA delivery, showcasing the advantages of novel pH‐responsive lipoamino xenopeptide (XP) carriers. These sequence‐defined XPs are assembled by branched lysine linkages between cationizable polar succinoyl tetraethylene pentamine (Stp) units and apolar lipoamino fatty acids (LAFs) at various ratios into bundle or U‐shape topologies. Formulation of siRNA‐LNPs using LAF4‐Stp1 XPs as ionizable compounds led to robust cellular uptake, high endosomal escape, and successful in vitro gene silencing activity at an extremely low (150 picogram) siRNA dose. Of significance is the functional in vivo endothelium tropism of siRNA‐LNPs with bundle LAF4‐Stp1 XP after intravenous injection into mice, demonstrated by superior knockdown of liver sinusoidal endothelial cell (LSEC)‐derived factor VIII (FVIII) and moderate silencing of hepatocyte‐derived FVII compared to DLin‐MC3‐DMA‐based LNPs. Optimizing lipid composition following click‐modification of siRNA‐LNPs with ligand c(RGDfK) efficiently silenced vascular endothelial growth factor receptor‐2 (VEGFR‐2) in tumor endothelial cells (TECs). The findings shed light on the role of ionizable XPs in the LNP in vivo cell‐type functional targeting, laying the groundwork for future therapeutic applications.