Polyglutamate directed coupling of bioactive peptides for the delivery of osteoinductive signals on allograft bone

Abstract

Allograft bone is commonly used as an alternative to autograft, however allograft lacks many osteoinductive factors present in autologous bone due to processing. In this study, we investigated a method to reconstitute allograft with osteoregenerative factors. Specifically, an osteoinductive peptide from collagen I, DGEA, was engineered to express a heptaglutamate (E7) domain, which binds the hydroxyapatite within bone mineral. Addition of E7 to DGEA resulted in 9× greater peptide loading on allograft, and significantly greater retention after a 5-day interval with extensive washing. When factoring together greater initial loading and retention, the E7 domain directed a 45-fold enhancement of peptide density on the allograft surface. Peptide-coated allograft was also implanted subcutaneously into rats and it was found that E7DGEA was retained in vivo for at least 3 months. Interestingly, E7DGEA peptides injected intravenously accumulated within bone tissue, implicating a potential role for E7 domains in drug delivery to bone. Finally, we determined that, as with DGEA, the E7 modification enhanced coupling of a bioactive BMP2-derived peptide on allograft. These results suggest that E7 domains are useful for coupling many types of bone-regenerative molecules to the surface of allograft to reintroduce osteoinductive signals and potentially advance allograft treatments.

Enhancement of the Regenerative Potential of Anorganic Bovine Bone Graft Utilizing a Polyglutamate-Modified BMP2 Peptide with Improved Binding to Calcium-Containing Materials

“…15,[22][23][24] To determine whether the E7 domain similarly directs enhanced binding of the BMP2pep, equimolar solutions of E7BMP2pep-FITC and BMP2pep-FITC were incubated with ABB for 30 min and then samples were imaged. In addition to fluorescing, the FITC tag has a yellow tint that can be seen by the naked eye.…”

“…Large quantities of highly pure peptides can be produced in a costefficient manner. Moreover, polyglutamate-modified peptides exhibit near maximal binding to graft materials within 30-60 min, 15,22 suggesting that off-the-shelf graft materials could be conjugated with the peptides shortly before surgical placement. In addition, the length of the polyglutamate domain can be varied to tailor peptide release kinetics.…”

“…Quantitative measurements indicate that the binding/retention of peptides can be improved between 14-and 63-fold by adding an E7 domain to the peptide, depending upon the specific carrier substrate. 22,24 The current study investigated the regenerative potential of a commercial xenograft material, ABB, functionalized with E7-modified osteoinductive peptides. ABB is widely used for bone grafting, particularly in some European countries that prohibit the use of allograft materials.…”

“…We and others have focused on modifying bioactive peptides with a calcium-binding domain to anchor the peptides onto the surface of calcium-containing bone replacement materials. [15][16][17][18][19][20][21][22][23][24][25][26] Specifically, our group has engineered osteoinductive peptides with a heptaglutamate (E7) domain. The E7 domain, which is a natural motif found within bone sialoprotein, binds tightly and specifically to calcium phosphate crystals within bone (hydroxyapatite [HA]) or synthetic biomaterials designed for bone applications.…”

“…In prior studies we used a number of approaches to establish that E7 domains are highly effective in anchoring a wide variety of peptides to multiple types of substrates, including allograft, xenograft, and alloplast. 15,[22][23][24]26 For example, E7-modified peptides display tight coupling to synthetic HA disks, b-tricalcium phosphate, electrospun scaffolds with nanoHA particles, calcium sulfate bone cement, anorganic bovine bone (ABB; BioOss), and two types of allograft (OraGRAFT and MinerOss). Quantitative measurements indicate that the binding/retention of peptides can be improved between 14-and 63-fold by adding an E7 domain to the peptide, depending upon the specific carrier substrate.…”