Neutralizing Antibodies: Key Immune Barriers

Clinical promise aside, AAV capsids face challenges due to pre-existing NAb from individuals with prior exposure to wild-type AAV. These antibodies bind to viral capsids, impacting AAV-mediated gene transfer and diminishing gene therapy efficacy. 
Addressing the presence of NAb during the development of AAV-based therapy, it is crucial to develop strategies to overcome capsid immunity.  Strategies like epitope masking, directed evolution, plasmapheresis, IgG-degrading enzymes, and structural modifications have been developed to address capsid immunity.

        Strategies to overcome capsid immunity: 

• Epitope Masking: Coating the AAV surface with lipids or extracellular vesicles to mask NAb recognition sites.
• Directed Evolution: Identifying AAV variants less sensitive to NAbs through mutational screening.
• Plasmapheresis: Removing antibodies from the blood to minimize vector-NAb contact.
• IgG-degrading Enzymes: Temporarily eliminating IgG for a NAb-free window.
• Structural Modifications: Modifying NAb recognition sites without affecting production efficiency

Challenges of Accurate NAb Measurement

Accurate NAb measurement with the current AAV-luciferase gold standard method faces challenges due to various factors, including high variability in AAV production from one batch to another, complex incubation procedures that can lead to assay variations, the presence of empty capsids, and matrix effects that can falsely lower the NAb assay results and create false positives. 

Therefore, it is essential to have reproducible and specific in vitro assays for each AAV variant or serotype.

Furthermore, cell-based assays are crucial since they provide a direct and accurate evaluation of neutralization activity for each AAV serotype or variant.