Combining computational modeling and experimental library screening to affinity-mature VEEV-neutralizing antibody F5

Area of Science:

• Biotechnology
• Immunology
• Virology

Background:

• Engineered monoclonal antibodies are effective therapeutics against viral outbreaks.
• Rapid adaptation and affinity enhancement of antibodies remain challenges.
• Venezuelan Equine Encephalitis Virus (VEEV) poses a significant public health threat.

Purpose of the Study:

• To develop a pipeline for increasing the binding affinity of the F5 monoclonal antibody to VEEV antigen.
• To utilize molecular modeling and experimental screening for antibody optimization.
• To assess the impact of enhanced affinity on antibody neutralization and cross-reactivity.

Main Methods:

• Molecular modeling (Rosetta-based and dTERMen) was used to predict mutations for improved F5 antibody binding.
• Phage display and error-prone PCR generated mutant antibody libraries.
• Screening identified favorable mutations, which were incorporated into human-IgG1 variants.

Main Results:

• The best antibody variant showed a 63-fold increase in binding affinity (KD from 0.63 nM to 0.01 nM).
• Enhanced affinity did not improve neutralization or therapeutic potency against the specific VEEV strain (IAB).
• Improved antibody variants demonstrated increased cross-reactivity to other VEEV strains.

Conclusions:

• The developed pipeline successfully enhanced antibody affinity and cross-reactivity to related VEEV strains.
• This method shows potential for rapidly adapting existing antibody therapeutics to emerging viral strains.
• Further research is needed to correlate enhanced affinity with improved therapeutic efficacy.