Lung epithelial binding peptide-linked high mobility group box-1 A box for lung epithelial cell-specific delivery of DNA.

High mobility group box-1 A box (HMGB1A) is an anti-inflammatory peptide originating from HMGB1. A previous report demonstrated that recombinant HMGB1A could deliver DNA into cells. Lung epithelial-specific gene delivery is required for the gene therapy of various lung diseases such as acute lung injury. In this study, a lung epithelial-specific DNA carrier was produced by linking the lung epithelial binding peptide (LEBP) to HMGB1A. An LEBP-linked HMGB1A (LEBP-HMGB1A) expression vector, pET21a-LEBP-HMGB1A, was constructed. LEBP-HMGB1A was expressed in BL21 strain and purified by consecutive applications of nickel affinity chromatography and cationic exchange chromatography. In a gel retardation assay, LEBP-HMGB1A completely retarded DNA at a 5:1 weight ratio (peptide:DNA). LEBP-HMGB1A/DNA complexes were prepared at various weight ratios, to which a fixed amount of polyethylenimine (2 kDa, PEI2k) was added to increase the proton buffering effect of the complex. LEBP-HMGB1A had the highest transfection efficiency to L2 lung epithelial cells at a 20:1 weight ratio (peptide:DNA). At this ratio, LEBP-HMGB1A had a higher transfection efficiency than poly-L-lysine (PLL) as well as HMGB1A without LEBP. A cytotoxicity assay showed that LEBP-HMGB1A was not toxic to L2 cells. Therefore, LEBP-HMGB1A may be useful in developing gene therapies for lung diseases.