Gelatin- hydroxyapatite- calcium sulphate based biomaterial for long term sustained delivery of bone morphogenic protein-2 and zoledronic acid for increased bone formation: In-vitro and in-vivo carrier properties

In this study, a novel macroporous composite biomaterial consisting of gelatin-hydroxyapatite-calcium sulphate for delivery of bone morphogenic protein-2 (rhBMP-2) and zoledronic acid (ZA) has been developed. The biomaterial scaffold has a porous structure and functionalization of the scaffold with rhBMP-2 induces osteogenic differentiation of MC3T3-e1 cells seen by a significant increase in biochemical and genetic markers of osteoblastic differentiation. In-vivo muscle pouch experiments showed higher mineralization using scaffold + rhBMP-2 when compared to an approved absorbable collagen sponge (ACS) + rhBMP-2 as verified by micro-CT. Co-delivery of rhBMP-2 + ZA via the novel scaffold enabled a reduction in the effective rhBMP-2 doses. The presence of tartrate resistant acid phosphatase staining in the rhBMP-2 group indicates osteoclastic resorption, which could be stalled by adding ZA, which by speculation could explain the net increase in mineralization. The new scaffold allowed for slow release of rhBMP-2 in-vitro (3.3 ± 0.1%) after 4 weeks. Using single photon emission computed tomography (SPECT), the release kinetics of ¹²⁵I–rhBMP-2 in-vivo was followed for 4 weeks and a total of 65.3 ± 15.2% ¹²⁵I–rhBMP-2 was released from the scaffolds. In-vitro ¹⁴C–ZA release curve shows an initial burst release on day 1 (8.8 ± 0.7%) followed by a slow release during the following 4 weeks (13 ± 0.1%). In-vivo, an initial release of 43.2 ± 7.6% of ¹⁴C–ZA was detected after 1 day, after which the scaffold retained the remaining ZA during 4-weeks. Taken together, our results show that the developed biomaterial is an efficient carrier for spatio-temporal delivery of rhBMP-2 and ZA leading to increased bone formation compared to commercially available carrier for rhBMP-2.