Designed ankyrin repeat proteins are effective targeting elements for chimeric antigen receptors.

Area of Science:

• Immunology
• Oncology
• Biotechnology

Background:

• Adoptive cell transfer using tumor-specific T cells is a promising cancer treatment.
• Chimeric antigen receptors (CARs) engineered onto T cells target tumor antigens.
• Traditionally, CARs utilize single-chain antibodies (scFvs) for antigen targeting.

Purpose of the Study:

• To investigate the utility of designed ankyrin repeat proteins (DARPins) as an alternative targeting domain in CARs.
• To compare the functionality of DARPin-based CARs against traditional scFv-based CARs.

Main Methods:

• Second-generation anti-HER2 CARs were constructed using either DARPin or scFv targeting domains.
• CARs were engineered into human and murine T cells.
• CAR-mediated cytokine production, degranulation, and cytotoxicity were assessed.

Main Results:

• DARPin CARs showed reduced surface expression in murine T cells but equivalent expression in human T cells compared to scFv CARs.
• Both DARPin and scFv CARs demonstrated comparable functionality in triggering cytokine and cytotoxic responses in both human and murine systems.
• Potential processing differences in murine T cells were suggested.

Conclusions:

• Designed ankyrin repeat proteins (DARPins) are effective targeting agents for CAR development.
• DARPin-based CARs offer a viable alternative to scFv-based CARs.
• This approach enables the generation of CARs with novel antigen-binding capabilities for cancer immunotherapy.