Discovery and Characterization of a Potent and Specific Peptide Ligand Targeting Endothelial Progenitor Cells and Endothelial Cells for Tissue Regeneration

Endothelial progenitor cells (EPcs) and endothelial cells (ECs) play a vital role in endothelialization and vascularization for tissue regeneration. Various EPC/EC targeting biomolecules have been investigated to improve tissue regeneration with limited success often due to their limited functional specificity and structural stability. one-bead one-compound (OBOC) combinatorial technology is an ultrahigh throughput chemical library synthesis and screening method suitable for ligand discovery Against a wide range of biological targets, such as integrins. In this study, using primary human EPCs/ECs as living probes, we identified an alpha v beta 3 integrin ligand LXW7 discovered by OBOC combinatorial technology as a potent and specific EPC/EC targeting ligand. LXW7 overcomes the major barriers of other functional biomolecules that have previously been used to improve vascularization for tissue regeneration and possesses optimal stability, EPC/EC specificity, and functionality. LXW7 is a disulfide cyclic octa-peptide (CORGDdvc) containing unnatural amino acids flanking both sides of the main functional motif; therefore it will be more resistant to proteolysis and more stable in vivo compared to linear peptides and peptides consisting of only natural amino acids. Compared with the conventional alpha v beta 3 integrin ligand GRGD peptide, LXW7 showed stronger binding affinity to primary EPCs/ECs but weaker binding to platelets and no binding to THP-1 monocytes. In addition, ECs bound to the LXW7 treated culture surface exhibited enhanced biological functions such as proliferation, likely due to increased phosphorylation of VEGF receptor 2 (VEGF-R2) and activation of mitogen-activated protein kinase (MAPK) ERK1/2. Surface modification of electrospun microfibrous PLLA/PCL biomatetial scaffolds with LXW7 via Click chemistry resulted in significantly improved endothelial coverage. LXW7 and its derivatives hold great premise for EPC/EC recruitment and delivery and can be widely applied to functionalize various biological and medical materials to improve endothelialization and vascularization for tissue regeneration applications.