Reprogramming the antigen specificity of B cells using genome-editing technologies.

Area of Science:

• Immunology
• Molecular Biology
• Genetics

Background:

• The human antibody repertoire is crucial for adaptive immunity.
• Developing novel antibodies, particularly for challenging targets like HIV, remains a significant scientific pursuit.
• Existing methods for antibody generation can be limited in scope and efficiency.

Purpose of the Study:

• To develop a method for introducing novel antibody specificities into the human B cell repertoire.
• To engineer B cells to express HIV-specific B cell receptors (BCRs) using genome editing.
• To assess the functionality and diversity of engineered BCRs for potential therapeutic applications.

Main Methods:

• Utilized CRISPR-Cas9 genome editing with homology-directed repair to replace immunoglobulin heavy chain variable regions in B cell lines.
• Introduced the heavy chain variable region from an HIV broadly neutralizing antibody (bnAb), PG9.
• Employed endogenous activation-induced cytidine deaminase (AID) for immunoglobulin class switching and affinity maturation.
• Applied the strategy to peripheral blood-derived primary B cells from multiple donors.

Main Results:

• Successfully engineered B cells expressed PG9 heavy chains paired with native light chains, leading to cell surface expression of HIV-specific BCRs.
• Engineered BCRs demonstrated binding to the PG9 epitope.
• Activation-induced cytidine deaminase facilitated Ig class switching, generating BCR variants with enhanced HIV neutralizing activity.
• Primary B cells from different donors were successfully edited, expressing PG9 heavy chains as multiple isotypes after stimulation.

Conclusions:

• Genome editing of mature B cells offers a viable strategy for introducing novel antibody specificities, such as those targeting HIV.
• This approach retains the natural genetic flexibility of B cells for affinity maturation, leading to improved antibody function.
• The engineered B cells and their expressed BCRs hold potential for developing new immunotherapies against HIV infection.