The delivery of proteins has been widely explored to promote vascularization and tissue repair because of their ability to initiate pro-angiogenic and pro-repair signaling cascades. The Segura Laboratory has explored the delivery of growth factors in both a nanoencapsulated and a tethered approach. The goal of the encapsulated approach is to deliver growth factors in a dormant, protected form that could become activated by the protease wound microenvironment in a controlled manner. Although the delivery of factors in clinical trials has required supra-physiological levels of the proteins, we believe that we can reduce the large dose and unwanted side effect by delivering growth factors in dormant conformations that are subsequently activated when needed.
We have demonstrated that functional proteins, including growth factors, can be encapsulated within degradable nanocapsules using in situ radical polymerization. The approach is to first surface absorb a monomer using charge interactions (negative surface protein and positive monomer), mix in neutral monomer, mix in a degradable crosslinker and start the polymerization through the addition of radical initiators. The effect of this process is the polymerization of a polymeric network around the protein. By controlling the concentration of the protein, monomers and crosslinker, nanoparticles (nanocapsules) can be generated that are stable at physiological conditions. The choice of the crosslinker is critical for inducing release of the growth factor cargo in the extracellular space such that the growth factor can interact with surface receptors. We chose a plasmin protease degradable crosslinker that can release the cargo at wound sites. Last, to modulate growth factor release rate from the nano capsules, we mixed L-chirality and D-chirality peptides, which degrade at different rates to modulate the capsule degradation rate. Using this approach, we can achieve either sustained or sequential delivery of proteins.
VEGF is the master regulator of angiogenesis. Ineffective VEGF signaling is directly responsible for lack of blood supply in cardiovascular disease that leads to organ and tissue damage. Thus, much effort has been placed on delivering VEGF to restore blood flow. However, delivery of VEGF has been plagued with limited therapeutic benefit in clinical trials and serious side effects.
The Segura Laboratory is interested in delivering VEGF from our materials in a safe and effective manner. When we started our work in this field, it was known that the implantation of hydrogel scaffolds with bound VEGF (electrostatic or covalently bound) results in vessels that are more highly branched, perfused and mature than when soluble VEGF is encapsulated in the hydrogel. Moreover, although it was (and still is) common practice in bioengineered materials to covalently bind bioactive signals to the matrix, little is done to know the biological ramifications of such immobilization. We found that covalently bound VEGF can induce the phosphorylation of VEGFR-2 and that release of the VEGF ligand was not necessary. In fact, the ligand stays behind on the surface and can induce the phosphorylation of VEGFR-2 on a new set of cells. Further, we found covalently bound VEGF led to differential activation of endothelial cells compared to soluble VEGF. These experiments also demonstrated that bound VEGF clusters its receptor and recruits’ integrin receptors during endothelial cell activation. Using this molecular understanding the Segura laboratory has design VEGF loaded hydrogel biomaterials that lead to improved tissue repair and regeneration.