Biparatopic affibody engineering enables high-affinity sortilin blockade and progranulin elevation.

Area of Science:

• Biochemistry
• Neuroscience
• Protein Engineering

Background:

• Loss-of-function mutations in progranulin (PGRN) are a primary cause of frontotemporal dementia.
• Therapeutic strategies aim to increase PGRN levels by inhibiting its clearance via sortilin.

Purpose of the Study:

• To design and optimize small biparatopic sortilin-binding proteins using an affibody scaffold.
• To evaluate the efficacy of these engineered proteins in increasing extracellular PGRN levels.

Main Methods:

• Genetically fused two anti-sortilin affibody molecules into heterodimeric constructs.
• Systematically varied domain orientations, linker lengths, and helix truncations.
• Assessed binding affinities and functional activity in PGRN clearance assays.

Main Results:

• Optimized dimers achieved subnanomolar sortilin affinities, a 45-fold improvement over monomers.
• The lead 18.6-kDa dimer increased extracellular PGRN levels with high potency (EC50 = 0.32 nM).
• Demonstrated effective blockade of sortilin-mediated clearance and modulation of sortilin levels.

Conclusions:

• Systematic combination and geometric optimization of affibodies yield potent biparatopic inhibitors.
• Compact affibody molecules are versatile for therapeutic protein design.
• Biparatopic affibody inhibitors offer potential for research and drug development in neurodegeneration, inflammation, and cancer.