Engineering a Low-Immunogenic Mirror-Image VHH against Vascular Endothelial Growth Factor

Area of Science:

• Biotechnology
• Immunology
• Protein Engineering

Background:

• Protein therapeutics can trigger antidrug antibodies (ADAs), reducing efficacy and causing adverse events.
• Mirror-image proteins (d-amino acids) resist degradation and may reduce immunogenicity.
• Identifying mirror-image antibodies for natural ligands is challenging.

Purpose of the Study:

• To develop a novel screening platform for identifying d-VHH specific for vascular endothelial growth factor A (VEGF-A).
• To create a d-VHH candidate with high affinity for native VEGF-A and low immunogenicity.

Main Methods:

• Developed and utilized a mirror-image screening platform with T7 phage display.
• Screened two synthetic VHH libraries against a mirror-image VEGF-A target (d-VEGF-A).
• Synthesized and characterized a d-VHH candidate for binding to native VEGF-A (l-VEGF-A).
• Evaluated immunogenicity of the d-VHH candidate in mouse immunization studies.

Main Results:

• Identified VHH sequences binding to d-VEGF-A.
• Developed a d-VHH candidate with submicromolar affinity for l-VEGF-A.
• Demonstrated that the d-VHH candidate elicited no antidrug antibodies (ADAs) in mice, unlike its natural counterpart (l-VHH).

Conclusions:

• The novel d-VHH screening platform is effective for identifying mirror-image antibodies.
• This approach facilitates the development of protein therapeutics with reduced immunogenicity and enhanced efficacy.
• d-VHH therapeutics represent a promising strategy to overcome limitations of traditional protein drugs.