Affimer reagents as tool molecules to modulate platelet GPVI-ligand interactions and specifically bind GPVI dimer

Area of Science:

• Biochemistry
• Molecular Biology
• Hematology

Background:

• Glycoprotein VI (GPVI) is crucial for collagen-mediated platelet aggregation.
• Engineered Affimers offer an alternative to antibodies for protein binding studies.

Purpose of the Study:

• To screen and characterize GPVI-binding Affimers.
• To evaluate their potential as tools for probing GPVI function and as therapeutic agents.

Main Methods:

• Affimer screening and characterization.
• Inhibition assays for GPVI-ligand interactions and platelet aggregation.
• In vitro thrombus formation studies under flow.
• Epitope mapping and binding site analysis.

Main Results:

• High-affinity GPVI-binding Affimers (M17, D22, D18) were identified.
• Affimers inhibited GPVI-ligand interactions, platelet aggregation, and D22 inhibited thrombus formation.
• Affimers bind distinct sites on GPVI D1 and D2 domains, with D18 targeting a dimer-specific epitope.
• D18 can detect GPVI dimerization, suggesting potential for antithrombotic drug development.

Conclusions:

• Affimers effectively modulate GPVI-ligand interactions and platelet function.
• Distinct binding sites on GPVI offer opportunities for targeted therapeutic interventions.
• D18's dimer-specific binding highlights its utility for studying GPVI dimerization and developing novel antithrombotic agents.