Understanding the molecular reach of therapeutic CD3-targeting bi-specific antibodies to optimise efficacy

Antibodies are soluble proteins that contribute to immune responses by directly targeting pathogens. This ability critically depends on bivalent binding whereby a single antibody binds two antigens on the pathogen. Beyond their native roles, antibodies are also developed as clinical tools for diagnostics and as biological drugs to treat transplant rejections, cancers, and infections.

Their clinical success has inspired the development of synthetic bivalent bispecifc molecules that can target two different antigens with >60 in clinical trials and several EMA/FDA approvals. Again, their activity depends on their ability to simultaneously bind (and hence reach) their two antigens to maintain them in proximity.

Although the activity of antibodies and other synthetic bivalent molecules critically depends on their ability to simultaneously bind their target antigens, current methods to develop and optimise them largely rely on monovalent binding assays. We have recently developed a new mathematical analysis of bivalent antibody binding generated using surface plasmon resonance (SPR).

This new analysis provides the standard monovalent kinetics (kon, koff, KD) and new bivalent binding parameters, including the bivalent on-rate and the molecular reach. The molecular reach is the maximum distance that two antigens can be placed that would still allow a single antibody to bind them both simultaneously. We recently analysed 80 RBD-specific antibodies finding that the molecular reach was the best predictor of SARS-CoV-2 neutralisation.

In this project, we will partner with GSK who has expertise in generating and functionally analysing bi-specific antibodies that re-direct T cells to kill target cells (e.g. cancer cells). These molecules bind the CD3 molecule on the TCR complex and a second antigen that is displayed on the target cell surface. The key aims of the project will be to 1) analyse the bivalent binding of therapeutic bi-specific antibodies using SPR, 2) determine the relationship between molecular reach and efficacy, and 3) rationally design improved therapeutics.

Together, these aims will provide the community with design principles for producing and optimising bi-specific therapeutics.