Improved transfection using epithelial cell line-selected ligands and fusogenic peptides

Synthetic gene transfer vectors can be optimised by combining DNA-binding peptides, cell surface receptor ligands, and fusogenic and nuclear localisation peptides. We have used the phage display technique to identify ligands of the tracheal epithelial cell line CFT-2. The peptides harboured by two phages were selected for transfection studies: peptide 7 (GRGDGDV) that contained the integrin-binding motif RGD, and peptide 9 (RFDSLKV) that was found in six out of 24 phages analysed. Both peptides, fused with the DNA-binding peptide P2 (SPKRSPKRSPKR), enhanced transfection efficiency in cell lines CFT-2, NT-1, NIH-3T3 and ECV-304. In particular, peptide P2-7 increased transfection efficiency from 36. 5% to 44.8% in NIH-3T3 cells and from 10.9% to 14.4% in CFT-2 cells, when compared to transfections performed with peptide P2. Two fusogenic peptides, HA (GLFEAIAEFIEGGWEGLIEGC) and JTS-1 (GLFEALLELLESLWELLLEA), were then added to the complexes and shown to improve transfection efficiency to the same extent. For instance, when combined to peptide P2-7, transfection levels of 54.1% and 55. 2% were attained in NIH-3T3 cells with HA and JTS-1, respectively. The addition of the ligands and fusogenic peptides thus allowed us to construct greatly improved transfection reagents.