Moderate-Affinity Affibodies Modulate the Delivery and Bioactivity of Bone Morphogenetic Protein-2

Area of Science:

• Biomaterials Science
• Protein Engineering
• Regenerative Medicine

Background:

• Uncontrolled release of bone morphogenetic protein-2 (BMP-2) can cause adverse effects, necessitating controlled delivery systems.
• Affibodies, engineered protein binders, offer a potential solution for targeted BMP-2 modulation.
• Understanding BMP-2 binding kinetics and mechanisms is crucial for therapeutic applications.

Purpose of the Study:

• To identify and characterize BMP-2-specific affibodies with varying affinities.
• To investigate the binding interactions between affibodies and BMP-2 using biophysical methods.
• To evaluate the efficacy of affibody-conjugated hydrogels in controlling BMP-2 release and biological activity.

Main Methods:

• Yeast surface display for affibody library screening.
• Biolayer interferometry to determine binding kinetics (KD, koff).
• Computational modeling to predict binding sites.
• In vitro studies using C2C12 myoblasts to assess alkaline phosphatase (ALP) activity.
• Fabrication and evaluation of affibody-conjugated hydrogels for BMP-2 release.

Main Results:

• High- and low-affinity affibodies targeting distinct BMP-2 sites were identified.
• Affibody binding reduced BMP-2-induced alkaline phosphatase (ALP) expression in myoblasts.
• Affibody-conjugated hydrogels enhanced BMP-2 uptake and controlled its release over four weeks.
• High-affinity hydrogels demonstrated significantly lower BMP-2 release compared to low-affinity and affibody-free hydrogels.
• BMP-2 loaded into hydrogels prolonged myoblast ALP activity.

Conclusions:

• Affibodies can be engineered to modulate BMP-2 binding affinity and delivery kinetics.
• Affibody-conjugated hydrogels provide a promising platform for controlled BMP-2 release.
• This approach offers potential for improved therapeutic outcomes in bone regeneration and other clinical applications.
• The differential binding and release characteristics highlight the tunable nature of this system.