Identification of peptides that target the endothelial cell-specific LOX-1 receptor.
Current gene delivery vectors demonstrate inefficient and nonselective gene transfer to vascular endothelial cells, limiting their use in cardiovascular gene transfer and therapy. The lectinlike oxidized LDL receptor (LOX-1) is expressed selectively at low levels on endothelial cells but is strongly upregulated in dysfunctional endothelial cells associated with hypertension and atherogenesis. Using LOX-1 as a target receptor, we have sought to isolate peptide ligands that mediate binding to the extracellular domain of LOX-1 as a definitive step in the development of targeted gene transfer aimed at dysfunctional endothelium. To achieve this, we ectopically overexpressed LOX-1 in cells lacking endogenous LOX-1 by using an episomally maintained expression system and designed a novel subtractive phage display strategy to identify peptides selective for LOX-1. After extensive biopanning, we sequenced individual phage and identified 60 novel peptides. This population of peptides contained a number of potential consensus motifs. To define the selectivity of individual peptides for LOX-1 with the use of an independent gene transfer system, we developed a novel adenoviral vector to overexpress LOX-1 transiently in primary cells and cell lines. We then quantified recovery of each peptide from LOX-1-positive and LOX-1-negative cells after adenovirus-mediated gene transfer. This strategy confirmed selectivity to LOX-1 for many peptides and highlighted the peptides LSIPPKA, FQTPPQL, and LTPATAI as principal candidates. These peptides will be useful for the selective targeting of viral and nonviral gene transfer vectors to endothelial cells expressing the LOX-1 receptor in vitro and in vivo and in particular dysfunctional endothelial cells associated with hypertension and atherosclerosis.
Bristol Heart Institute, Bristol Royal Infirmary, University of Bristol, UK.
S J
SJ
White
S A
SA
Nicklin
T
T
Sawamura
A H
AH
Baker
D000256:Adenoviridae / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001161:Arteriosclerosis / Q000503:physiopathology; D015153:Blotting, Western; D002460:Cell Line; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism* / Q000503:physiopathology; D005455:Fluorescent Antibody Technique; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006973:Hypertension / Q000503:physiopathology; D008024:Ligands; D010455:Peptides / Q000737:chemistry / Q000235:genetics*; D011973:Receptors, LDL / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D051131:Receptors, Oxidized LDL; D020133:Reverse Transcriptase Polymerase Chain Reaction
D008024:Ligands; D010455:Peptides; D011973:Receptors, LDL; D051131:Receptors, Oxidized LDL