A Broad-specificity Neutralizing Nanobody against Hepatitis E Virus Capsid Protein

Area of Science:

• Virology
• Immunology
• Biotechnology

Background:

• Hepatitis E virus (HEV) poses a significant global public health challenge.
• Understanding HEV biology is crucial for developing effective vaccines and antiviral drugs.
• Nanobodies are emerging as valuable tools for studying viral pathogens.

Purpose of the Study:

• To develop and characterize novel nanobodies against Hepatitis E virus (HEV).
• To identify and analyze the epitope targeted by the developed nanobodies.
• To enhance nanobody properties for improved HEV neutralization and therapeutic potential.

Main Methods:

• Immunization of a Bactrian camel with HEV capsid proteins from multiple genotypes.
• Construction of a phage display library and screening for HEV-specific nanobodies.
• Characterization of nanobody binding, epitope mapping, and in vitro/in vivo neutralization assays.
• Engineering nanobodies into ferritin-based nanocages (fenobodies) to enhance affinity and half-life.

Main Results:

• Screened 12 nanobodies against HEV genotype 3 truncated capsid protein (g3-p239).
• Identified g3-p239-Nb55, which cross-reacts with multiple HEV genotypes and blocks viral infection.
• Discovered a novel, conserved conformational epitope on the HEV surface targeted by g3-p239-Nb55.
• Engineered fenobody-55 demonstrated significantly increased affinity (∼20x) and half-life (9x) compared to the original nanobody.
• Fenobody-55 completely neutralized HEV infection in a rabbit model upon preincubation with viral particles.

Conclusions:

• Developed fenobody-55, a potent nanobody-based therapeutic candidate against HEV.
• Identified a novel, conserved conformational epitope crucial for HEV infectivity.
• Demonstrated the potential of nanobodies and fenobodies for HEV research and therapeutic intervention.
• Provided a new tool for studying HEV biology and developing strategies against HEV infection.