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

Abstract

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 αvβ3 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 (cGRGDdvc) 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 αvβ3 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 biomaterial scaffolds with LXW7 via Click chemistry resulted in significantly improved endothelial coverage. LXW7 and its derivatives hold great promise 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.

Engineering mesenchymal stem cells to improve their exosome efficacy and yield for cell‐free therapy

“…MSC-derived exosomes (MSC-exosomes) have been used as a treatment for diseases and disorders of many systems such as the cardiovascular, neurological, musculoskeletal and immune systems [5,6,8,31]. Rats with traumatic brain injury exhibited significant functional recovery, increased neurite remodelling, angiogenesis and neurogenesis after treatment with exosomes [32].…”

“…A variety of advanced engineering approaches have been used to produce native ECM mimicking structures. For example, an ECM mimicking nanofibrous scaffold could be created by electrospinning from polymer blends of polylactic acid (PLA) and polycaprolactone (PCL) [31]. At the same time, such a scaffold could be potentially functionalised by presenting specific functional ligands to improve production of MSC-exosomes [31].…”

“…For example, an ECM mimicking nanofibrous scaffold could be created by electrospinning from polymer blends of polylactic acid (PLA) and polycaprolactone (PCL) [31]. At the same time, such a scaffold could be potentially functionalised by presenting specific functional ligands to improve production of MSC-exosomes [31]. Another advantage of the synthetic scaffold is the control over their structural pattern and composition.…”

Strategies in cell‐free tissue‐engineered vascular grafts

“…Click chemistry technology was also an effective method for peptide modification (Hao et al, 2017). Click chemistry technology was also an effective method for peptide modification (Hao et al, 2017).…”

“…The cell-free grafts were employed in phase-2 single-arm clinical trials as AVF grafts in patients of end-stage renal disease (Lawson et al, 2016). Antibodies, bioactive peptides, and cytokines were widely used to accelerate in situ regeneration (Antonova et al, 2016;Hao et al, 2017;Kanie et al, 2012;Lahtinen et al, 2007;Lu et al, 2013;Song et al, 2016;Sun et al, 2010;Yu et al, 2012). Antibodies, bioactive peptides, and cytokines were widely used to accelerate in situ regeneration (Antonova et al, 2016;Hao et al, 2017;Kanie et al, 2012;Lahtinen et al, 2007;Lu et al, 2013;Song et al, 2016;Sun et al, 2010;Yu et al, 2012).…”