Degron-Based bioPROTACs for Controlling Signaling in CAR T Cells.

Area of Science:

• Immunology
• Molecular Biology
• Biotechnology

Background:

• Chimeric antigen receptor (CAR) T cells offer potent cancer therapy but face challenges with signaling control, impacting safety and efficacy.
• Current methods for regulating CAR stability often rely on systemic small molecules, limiting precision.
• Genetic circuits present an alternative for autonomous, cell-by-cell control of CAR signaling.

Purpose of the Study:

• To develop a programmable protein degradation tool for precise control of CAR T cell signaling.
• To engineer novel bioPROTACs capable of targeting CARs and other proteins for degradation.
• To demonstrate the utility of bioPROTACs in a genetic circuit for antigen-specific CAR T cell inhibition.

Main Methods:

• Designed and synthesized novel bioPROTACs utilizing a compact four-residue degron.
• Employed nanobodies or synthetic leucine zippers as protein binders for target recognition.
• Tested bioPROTACs for degradation of cytosolic and membrane proteins, including CARs, in primary human T cells.
• Constructed a genetic circuit for antigen-specific degradation of ZAP70 kinase.

Main Results:

• Successfully demonstrated potent degradation of CARs and inhibition of CAR T cell signaling.
• Validated bioPROTACs for targeting both cytosolic and membrane proteins.
• Engineered a genetic circuit that specifically degrades ZAP70 upon antigen recognition, disrupting CAR T cell activity only in target cell populations.
• Showcased bioPROTACs as a versatile tool for CAR T cell engineering.

Conclusions:

• BioPROTACs offer a precise and programmable method for controlling CAR T cell signaling, overcoming limitations of current strategies.
• This technology enhances the safety and efficacy of CAR T cell therapies by enabling autonomous, cell-specific regulation.
• BioPROTACs significantly expand the engineering toolbox for developing advanced CAR T cell therapeutics.