TY - JOUR
T1 - A synthetic peptide vector system for optimal gene delivery to corneal endothelium
AU - Collins, L
AU - Fabre, J W
PY - 2004/2
Y1 - 2004/2
N2 - Background Efficient and non-toxic gene delivery, preferably with non-viral DNA vectors readily transferable to clinical practice, is generally regarded as a major limitation for gene therapy. Methods A 31 amino acid, integrin-targeted bifunctional synthetic peptide (polylysine-molossin), and two (Lys)(16)-containing control peptides, were assessed for ex vivo gene delivery to the rabbit cornea. Critical physical properties of polylysine-molossin/DNA complexes were evaluated and both chloroquine and a 20 amino acid fusogenic peptide were used to promote endocytic exit. Results Polylysine-molossin/DNA complexes and (Lys)16/DNA complexes at 10 mug/ml of DNA were much smaller and much more positively charged in non-ionic isotonic medium (5% dextrose or 5% dextrose buffered to pH 7.4 in 10 mM Tris) when compared with culture medium or phosphate-buffered saline (PBS). Addition of the fusogenic peptide (net charge -5) reversed the positive charge of complexes in PBS, and reduced the strong positive charge of polylysine-molossin/DNA complexes in dextrose. Polylysine-molossin/DNA complexes in 5% dextrose were much more effective for gene delivery to the cornea when compared with complexes in culture medium, and the fusogenic peptide was much more effective than chloroquine for promoting gene delivery. The optimal DNA/polylysine-molossin/fusogenic peptide w/w ratio was 1: 3: 2 at 10 mug/ml of DNA. Essentially 100% of corneal endothelial cells were transfected under these optimal conditions, without any evidence of toxicity. Integrin-targeting did not contribute significantly to gene delivery in this system. Conclusions This DNA vector system, consisting entirely of synthetic peptides, is ideally suited for clinical applications of gene therapy of the corneal endothelium. Copyright (C) 2004 John Wiley Sons, Ltd.
AB - Background Efficient and non-toxic gene delivery, preferably with non-viral DNA vectors readily transferable to clinical practice, is generally regarded as a major limitation for gene therapy. Methods A 31 amino acid, integrin-targeted bifunctional synthetic peptide (polylysine-molossin), and two (Lys)(16)-containing control peptides, were assessed for ex vivo gene delivery to the rabbit cornea. Critical physical properties of polylysine-molossin/DNA complexes were evaluated and both chloroquine and a 20 amino acid fusogenic peptide were used to promote endocytic exit. Results Polylysine-molossin/DNA complexes and (Lys)16/DNA complexes at 10 mug/ml of DNA were much smaller and much more positively charged in non-ionic isotonic medium (5% dextrose or 5% dextrose buffered to pH 7.4 in 10 mM Tris) when compared with culture medium or phosphate-buffered saline (PBS). Addition of the fusogenic peptide (net charge -5) reversed the positive charge of complexes in PBS, and reduced the strong positive charge of polylysine-molossin/DNA complexes in dextrose. Polylysine-molossin/DNA complexes in 5% dextrose were much more effective for gene delivery to the cornea when compared with complexes in culture medium, and the fusogenic peptide was much more effective than chloroquine for promoting gene delivery. The optimal DNA/polylysine-molossin/fusogenic peptide w/w ratio was 1: 3: 2 at 10 mug/ml of DNA. Essentially 100% of corneal endothelial cells were transfected under these optimal conditions, without any evidence of toxicity. Integrin-targeting did not contribute significantly to gene delivery in this system. Conclusions This DNA vector system, consisting entirely of synthetic peptides, is ideally suited for clinical applications of gene therapy of the corneal endothelium. Copyright (C) 2004 John Wiley Sons, Ltd.
UR - http://www.scopus.com/inward/record.url?scp=3142773814&partnerID=8YFLogxK
U2 - 10.1002/jgm.482
DO - 10.1002/jgm.482
M3 - Article
VL - 6
SP - 185
EP - 194
JO - JOURNAL OF GENE MEDICINE
JF - JOURNAL OF GENE MEDICINE
IS - 2
ER -