Enhancement of periosteal bone formation by basic fibroblast-derived growth factor containing polycystic kidney disease and collagen-binding domains fromClostridium histolyticumcollagenase

Abstract

Recombinant basic fibroblast growth factor (bFGF) is a potent mitogen for mesenchymal cells that accelerates bone union and repair when applied locally at defect sites. However, because bFGF diffuses rapidly from bone defect sites, repeated dosing is required for sustained therapeutic effect. We previously fused the collagen-binding domain (CBD) and polycystic kidney disease (PKD) domain of Clostridium histolyticum class II collagenase (ColH) to bFGF and demonstrated that the fusion protein markedly enhances bone formation when loaded onto collagen materials used for grafting. However, systemic injection of a fusion protein consisting of parathyroid hormone (PTH) and a CBD was shown to accelerate bone formation in an osteoporosis model more rapidly than treatment with a PTH-PKD-CBD fusion protein. Here, we compared the biological properties of two collagen-binding forms of bFGF, bFGF-CBD and bFGF-PKD-CBD. Both fusion proteins promoted the in vitro proliferation of periosteal mesenchymal cells, indicating that they had biological activity similar to that of native bFGF. In vivo periosteal bone formation assays in rat femurs showed that both bFGF-CBD and bFGF-PKD-CBD induced periosteal bone formation at higher rates than collagen sheet alone and bFGF. However, bFGF-PKD-CBD markedly enhanced bone formation and had higher collagen-binding ability than bFGF-CBD in in vitro protein release assays. Taken together, these results suggest that the PKD domain increases the retention of bFGF at graft sites by enhancing collagen-binding affinity. Therefore, bFGF-PKD-CBD-collagen composite appears to be a promising material for bone repair in the clinical setting. Copyright © 2015 John Wiley & Sons, Ltd.

Acceleration of bone formation during fracture healing by poly(pro–hyp–gly)10 and basic fibroblast growth factor containing polycystic kidney disease and collagen‐binding domains from Clostridium histolyticum collagenase

“…We previously established a non‐invasive method for accelerating periosteal bone formation using collagen materials anchored with collagen‐binding bFGF protein fused with a PKD and CBD derived from C. histolyticum ColH . When this method was applied to bone fracture models, the bFGF‐PKD‐CBD/collagen composite was shown to have superior bone growth‐promoting ability compared to a bFGF/collagen composite .…”

“…We previously established a non-invasive method for accelerating periosteal bone formation using collagen materials anchored with collagen-binding bFGF protein fused with a PKD and CBD derived from C. histolyticum ColH. 31,32 When this method was applied to bone fracture models, the bFGF-PKD-CBD/collagen composite was shown to have superior bone growth-promoting ability compared to a bFGF/collagen composite. 33 In this study, bFGF-PKD-CBD/poly(Pro-Hyp-Gly) 10 had higher chondrogenesis-and bone formationpromoting effects compared to bFGF/poly(Pro-Hyp-Gly) 10 , a finding that is similar to a previous study comparing bFGF/collagen and bFGF-PKD-CBD/collagen composites.…”

“…Clostridium histolyticum class II collagenase (ColH) contains two polycystic kidney disease (PKD) domains and one collagen‐binding domain (CBD), which are necessary for the catalytic activity and collagen‐binding affinity of this enzyme . We previously fused the CBD and PKD domain of ColH to bFGF and demonstrated that the resulting bFGF‐PKD‐CBD fusion protein had higher in‐vitro collagen‐binding ability compared to bFGF‐CBD and enhanced bone formation compared to native bFGF and bFGF‐CBD when grafted together with collagen in a rat femur . In addition, the combination of injectable collagen powder and bFGF‐PKD‐CBD markedly accelerated bone formation in a mouse femur fracture model compared to treatment with bFGF alone .…”

“…29,30 We previously fused the CBD and PKD domain of ColH to bFGF and demonstrated that the resulting bFGF-PKD-CBD fusion protein had higher in-vitro collagen-binding ability compared to bFGF-CBD and enhanced bone formation compared to native bFGF and bFGF-CBD when grafted together with collagen in a rat femur. 31,32 In addition, the combination of injectable collagen powder and bFGF-PKD-CBD markedly accelerated bone formation in a mouse femur fracture model compared to treatment with bFGF alone. 33 As we previously reported that the CBD of ColG also binds to ((Pro-Hyp-Gly) 10 ) 3, 34 the combination of bFGF-PKD-CBD and poly(Pro-Hyp-Gly) 10 has the potential to promote the retention of bFGF at bone injury sites and accelerate bone repair with a similar efficacy as animal-derived collagen.…”

Design and Use of Chimeric Proteins Containing a Collagen-Binding Domain for Wound Healing and Bone Regeneration

“…For example, TKKTLRT-bFGF was shown to promote a stronger neovascularization than WREPSFMALS-bFGF (132), in good agreement with TKKTLRT displaying a higher affinity for collagen than WREPSFMALS (Table 1). Similarly, bFGF fused to the S2b+S3 domains of ColH markedly enhanced bone formation in comparison with bFGF fused to the S3 domain alone (133), which has a lower affinity for collagen (Table 1). Notwithstanding, opposite results were obtained with CBD-PTH chimeras: the PTH-S2b+S3 fusion protein was not as effective as the PTH-S3 fusion protein in improving the bone mineral density of mice.…”