Structure-Guided Combinatorial Engineering Facilitates Affinity and Specificity Optimization of Anti-CD81 Antibodies

Area of Science:

• Immunology
• Virology
• Structural Biology

Background:

• Hepatitis C virus (HCV) infection affects over 71 million people globally, causing chronic hepatitis.
• Targeting the virus directly is challenging due to rapid mutation and serotype diversity.
• Host factors, such as the CD81 receptor, offer alternative therapeutic targets.

Purpose of the Study:

• To develop novel therapeutic antibodies targeting the host CD81 receptor to block HCV entry.
• To utilize structural information of the CD81 large extracellular loop (LEL) to guide antibody engineering.
• To enhance antibody affinity, efficacy, and cross-reactivity for improved therapeutic potential and preclinical testing.

Main Methods:

• Generation of four independent antibodies targeting human CD81.
• Crystallization of antibody single-chain variable fragments (scFvs) with the CD81 LEL.
• Affinity maturation of selected antibodies using phage display technology.
• Engineering antibodies for increased affinity to human CD81 and cross-reactivity with cynomolgus CD81.

Main Results:

• Structural analysis guided the affinity maturation process for two distinct antibodies.
• One antibody demonstrated increased affinity for human CD81 LEL, leading to enhanced efficacy.
• A second antibody was successfully engineered for cross-reactivity with cynomolgus CD81, enabling animal model studies.
• The study validated the combined utility of structural biology and phage display for antibody development.

Conclusions:

• Antibodies targeting CD81 represent a promising strategy to inhibit HCV entry.
• Structure-guided affinity maturation is a powerful approach for optimizing therapeutic antibodies.
• Engineered antibodies with enhanced affinity and cross-reactivity facilitate preclinical evaluation and therapeutic advancement.