Development of affinity-based delivery systems for angiogenic growth factors - PROJECT SUMMARY Angiogenesis is a coordinated regenerative process during which the temporally regulated presentation of many signaling proteins stimulates cellular recruitment, patterning, and morphogenesis. Disruptions in this healing cascade from chronic diseased tissues or severe injuries can easily impair the natural sequence of protein presentation, resulting in poor healing outcomes and further propagation of diseased states. The long-term objective of this research is to develop tunable multi-protein delivery systems capable of phased therapeutic angiogenic protein delivery to reconstitute and enhance angiogenesis. To achieve this objective, this work will integrate specific reversible affinity-based interactions between small protein domains called affibodies and therapeutic angiogenic proteins to temporally regulate multi-protein delivery from hydrogel. Building from the well characterized roles of vascular endothelial growth factor (VEGF), fibroblast growth factor-2 (FGF2), and platelet derived growth factor (PDGF) in stimulating the angiogenic cascade, we will sequentially deliver these three growth factors from this controlled delivery platform. We expect this approach will more accurately recapitulate the natural, staggered presentation of growth factors seen during natural angiogenic healing responses, providing the necessary stimuli to enhance the regeneration of healthy mature vasculature. This research will target two major knowledge gaps: 1) How do affinity-based delivery systems modulate the delivery and bioactivity of VEGF, FGF-2, and PDGF when released from hydrogels? 2) How is the sequential presentation of VEGF, FGF-2, and PDGF implicated in mediating angiogenesis? Facilitating this research, we have applied directed evolution and computational protein design approaches to develop affibodies with high specificity and a broad range of affinity strengths towards VEGF, FGF-2, and PDGF. Focusing on the implementation of these affibodies within a hydrogel network, my overarching hypothesis is that temporally regulated sequential delivery of VEGF, FGF-2, and PDGF will reconstitute the angiogenic cascade. I will test this hypothesis through two specific aims: 1) Develop hydrogels that sequentially release VEGF, FGF-2, and PDGF, 2) Probe how the sequence of VEGF, FGF-2, and PDGF presentation mediates angiogenesis. Ultimately, we expect to achieve tunable sequential controlled release of multiple therapeutically relevant angiogenic growth factors from the same delivery vehicle. Overall, this work will expand our understanding of applying affinity-based protein-protein interactions in hydrogel as temporally regulated controlled release systems. Moreover, this work will provide new therapeutic approaches for stimulating revascularization of wounded or diseased tissues.
