Integrating binding affinity and tonic signaling enables a rational CAR design for augmented T cell function

Area of Science:

• Immunology
• Cancer Biology
• Biochemistry

Background:

• Chimeric antigen receptor (CAR) T cell therapy shows promise for hematological malignancies but faces challenges in solid tumors.
• Optimizing CAR T cell functionality is essential for improving efficacy against solid tumors.

Purpose of the Study:

• To investigate the relationship between CAR binding affinity, tonic signaling, and T cell functionality in solid tumor treatment.
• To explore the role of hydrophobic interactions in CAR T cell avidity and signaling.

Main Methods:

• Utilized a translational pipeline involving biophysical characterization and structural prediction of CAR binding moieties.
• Evaluated cellular avidity, synapse formation, T cell motility, and functional capacities under repetitive target challenge.
• Assessed sustained tumor control in preclinical models.

Main Results:

• Derived anti-Her2 CARs with a 4-log affinity range, demonstrating that scFv mutations impact affinity, avidity, and tonic signaling.
• Observed a non-linear relationship between increased affinity and functional avidity above a minimum threshold.
• Found that hydrophobic interactions within the scFv augment affinity, non-specific binding, and tonic signaling, influencing T cell functionality.

Conclusions:

• Tonic signaling can be driven by hydrophobic interactions in the scFv, not solely by positive charge.
• CAR binding affinity above a threshold and tonic signaling are critical for sustained T cell functionality during antigen rechallenge and long-term tumor control.