A high throughput bispecific antibody discovery pipeline

Area of Science:

• Immunotherapy
• Antibody Engineering
• Single-Cell Analysis

Background:

• Bispecific antibodies (BsAbs) are promising immunotherapeutics but their discovery is hampered by inefficient methods.
• Limited clinical availability of BsAbs restricts their therapeutic potential.

Purpose of the Study:

• To develop a high-throughput, single-cell-based functional screening pipeline for efficient bispecific antibody discovery.
• To enable the identification and characterization of rare functional BsAb clones.

Main Methods:

• A novel pipeline combining molecular and cell engineering for generating BsAb library cells.
• Single-cell level functional interrogation, sorting, and downstream sequence identification.
• Utilized a CD19xCD3 bispecific T cell engager (BiTE) model for validation.

Main Results:

• Achieved high-throughput screening efficiency of up to 1.5 million variant library cells per run.
• Successfully isolated rare functional clones at abundances as low as 0.008% and 0.001%.
• Identified 98 unique CD19xCD3 BiTE clones from a library of ~22,300 variants, including novel functional properties.

Conclusions:

• The developed single-cell platform significantly enhances the discovery efficiency of novel immunotherapeutics.
• The platform facilitates the identification of generalizable design principles for BsAb development.
• Enables a deeper understanding of sequence-structure-function relationships in BsAbs.