Inhalable Gene Delivery System Using a Cationic RAGE-Antagonist Peptide for Gene Delivery to Inflammatory Lung Cells

Abstract

Acute lung injury (ALI) is a severe lung inflammatory disease. In ALI, the receptor for advanced glycation end-products (RAGE) is overexpressed in lung epithelial cells and involved in inflammatory reactions. A previous report showed that a RAGE-antagonist peptide (RAP), from high-mobility group box-1, bound to RAGE and reduced inflammatory reactions. RAP has high levels of positive amino acids, which suggests that RAP may form a complex with plasmid DNA (pDNA) by charge interactions. Because the charge density of RAP is lower than polyethylenimine (25 kDa, PEI25k), it may be able to avoid capture by the negatively charged mucus layer more easily and deliver pDNA into RAGE-positive lung cells of ALI animals by RAGE-mediated endocytosis. To prove this hypothesis, RAP was evaluated as a delivery carrier of adiponectin plasmid (pAPN) in lipopolysaccharide (LPS)-induced ALI animal models. In vitro transfection assays showed that RAP had lower transfection efficiency than PEI25k in L2 lung epithelial cells. However, in vivo administration to ALI animal models by inhalation showed that RAP had higher gene delivery efficiency than PEI2Sk. Particularly, due to a higher expression of RAGE in lung cells of ALI animals, the gene delivery efficiency of RAP was higher in ALI animals than that in normal animals. Delivery of the pAPN/RAP complex had anti-inflammatory effects, reducing pro-inflammatory cytokines. Hematoxylin and eosin staining confirmed that pAPN/RAP decreased inflammation in ALI models. Therefore, the results suggest that RAP may be useful as a carrier of pDNA into the lungs for ALI gene therapy.