NanoengineeredSelf-AssemblingPeptides with IncreasedProteolytic Stability Promote Wound Healing

The copper complex of the tripeptide glycine–histidine–lysine (GHK) has proven benefits in wound healing and tissue remodeling by promoting blood vessel growth and increasing skin oxygen levels, but its activity is reduced in body fluids due to fast proteolytic cleavage. Herein, we designed several peptides that bear the GHK sequence and can self-assemble into supramolecular nanostructures aiming for enhanced bioactivity. The design involves a phenylalanine (F) backbone known for its ability to form supramolecular assemblies. We tested either coassembly between the structural peptide F₄D and the functional sequence GHK or assembly of covalently bound peptides, in which the GHK is bound via the glycine (F₄D-GHK) or lysine (F₄D-KHG, i.e., inverted GHK sequence). All tested peptides assembled into nanotapes, but their resistance to proteolytic degradation was different: covalently bound peptides generated more stable assemblies. Wound healing assays demonstrated that the supramolecular structures have enhanced bioactivity when compared to GHK alone. Multiplex immunoassay analyses demonstrated the secretion of key regulators of the healing process, such as cytokines, matrix metalloproteinases, and growth factors. Altogether our data show that incorporation of GHK/KHG into supramolecular structures improves its stability, bioactivity, and efficacy in promoting wound healing.