Evolutionarily conserved sequence motif analysis guided development of chemically defined hydrogels for therapeutic angiogenesis

Biologically ligands (e.g., RGDS from fibronectin: Fn-RGD) play a critical role in the development of chemically defined biomaterials. However, there has been limited progress in recent decades towards discovering novel extracellular-matrix-protein-derived ligands for translational applications. Here, by combining motif analysis of evolutionarily conserved RGD-containing regions in laminin (Ln) with functional microarray screening, we identified a Ln-derived angiogenic peptide (LDAP) that showed enhanced proangiogenic activities over commonly used Fn-RGD. Mechanistic studies using RNA-sequencing of LDAP functionalized hydrogels showed an improved angiogenic transcriptome over Fn-RGD functionalized hydrogels and high similarity to Matrigel, attributed to the ability of LDAP to engage both Ln- and Fn-binding integrins. Injectable hydrogels functionalized with LDAP, along with MMP-QK (a VEGF-mimetic peptide), exhibited increased functional recovery over decellularized extracellular matrix (dECM) and alginates functionalized with Fn-RGD and MMP-QK in a mouse ischemic hindlimb model, illustrating the power of the strategy to rapidly develop potent chemically-defined biomaterials for therapeutic applications. HUVECs (p2-p3) were encapsulated into 0.125mmol/g peptide-modified alginate (at alginate concentration = 1% (w/v)) at the density of 2 million cells/mL with EGM for 24 hours. Groups include HUVECs cultured in LDAP(laminin derived angiogenic peptide)-peptide modified alginate compared to in RGDS-peptide modified alginate hydrogels.