(226g) cAMP-Incorporated Poly(lactide-co-glycolide) Sintered Microsphere Scaffolds for Bone Regenerative Engineering

The use of recombinant human bone morphogenetic proteins (rhBMPs) has become a common strategy to enhance bone formation within regenerative engineering scaffolds. While engineered constructs employing these osteoinductive growth factors have shown promise, disadvantages such instability, limited solubility, immunogenicity and high production cost have hindered their clinical viability. Previous research has identified the capability of a short-term treatment scheme (1 day) of the small molecule cAMP analogue, 8-bromoadenosine-3’-5’-cyclic monophosphate (8-Br-cAMP), to promote in vitro osteoblastic differentiation and adhesion of osteoprogenitor MC3T3-E1 cells, and well as in vitro angiogenesis through the cell-based production of VEGF [1][2]. However, the integration of 8-Br-cAMP within an osteoconductive bone graft and its efficacy to promote bone formation within a preclinical animal model has yet to be evaluated. The objective of the proposed research is to develop an osteoinductive, engineered bone graft using the small molecule, 8-Br-cAMP. By loading the small molecule onto the engineered construct through surface adsorption, we hypothesize that the rate of diffusion of the cAMP small molecule will be modulated to induce the desired localized, short-term treatment and therapeutic effect. PLGA sintered microsphere scaffolds were developed through a single emulsion, solvent evaporation, and heat-sintering technique previously optimized in our laboratory [3]. 100 μM concentration of small molecule 8-Br-cAMP was physically adsorbed onto the surface of the scaffold in order to provide a burst release into the surrounding medium. In vitro dissolution testing showed approximately 70% of 8-Br-cAMP was released from the scaffold within 30 minutes, and 80% of 8-Br-cAMP was released after 1 hour. When implanted into a mouse calvarial defect model for 6 weeks, x-ray and histological images demonstrated evidence of bone formation throughout the scaffold due to the localized interaction of 8-Br cAMP with the surrounding host cells. These results provide promising verification for the use of the small molecule, 8-Br-cAMP in future bone regenerative engineering studies.

References

[1] K.W. Lo, K.M. Ashe, H.M. Kan, D.A. Lee, C.T. Laurencin, Activation of cyclic amp/protein kinase: a signaling pathway enhances osteoblast cell adhesion on biomaterials for regenerative engineering, J Orthop Res 29(4) (2011) 602-8.

[2] K.W. Lo, H.M. Kan, K.A. Gagnon, C.T. Laurencin, One-day treatment of small molecule 8-bromo-cyclic AMP analogue induces cell-based VEGF production for in vitro angiogenesis and osteoblastic differentiation, J Tissue Eng Regen Med 10(10) (2016) 867-875.

[3] M. Borden, S.F. El-Amin, M. Attawia, C.T. Laurencin, Structural and human cellular assessment of a novel microsphere-based tissue engineered scaffold for bone repair, Biomaterials 24(4) (2003) 597 - 609.