Delivery of Itgb1-siRNA by triptolide-modified and anti-Flt1 peptide-guided ionizable cationic LNPs for targeted therapy of corneal neovascularization.

Corneal neovascularization (CoNV) is a leading cause of visual impairment worldwide. However, CoNV remains challenging to cure clinically because of the limitations of current drugs. New and more effective therapeutic formulations for CoNV treatment are therefore urgently needed. Antisense oligonucleotide drugs hold great promise for the treatment of neovascular diseases, and ionizable cationic lipid nanoparticles (icLNPs) have shown excellent performance for nucleic acid delivery, with high encapsulation, good cellular uptake, and effective endosomal escape. In the present study, we identified integrin β1 (Itgb1) as a key gene involved in angiogenesis and revealed the significant upregulation of Flt1 in vascular endothelial cells and pericytes in CoNV using single-cell sequencing. Itgb1 knockdown significantly inhibited the proliferation and migration of vascular endothelial cells and CoNV in mice. Based on these findings, we developed Itgb1-small interfering RNA (siRNA)-loaded icLNPs, and conjugated anti-Flt1 peptide to their surface to improve CoNV targeting. Furthermore, because lipid nanoparticles reportedly trigger immune responses by upregulating pro-inflammatory cytokine expression, which may promote neovascularization, we modified triptolide (a compound with anti-inflammatory properties) into the icLNPs. The triptolide-modified, anti-Flt1 peptide-conjugated, and Itgb1-siRNA-loaded icLNPs (Itgb1-siRNA@TPL) effectively inhibited the proliferation and migration of vascular endothelial cells in vitro and CoNV in mice after eye drop administration. These effects occurred via downregulation of the PI3K/AKT and NF-κB signaling pathways. Finally, the biosafety of Itgb1-siRNA@TPL was tested, and the results revealed that it was not toxic to the cornea or major organs and had no impact on corneal epithelial healing. In conclusion, Itgb1-siRNA@TPL represent a novel, noninvasive, and effective approach for the treatment of CoNV.