A Rapamycin-Activated Caspase 9-Based Suicide Gene

Area of Science:

• Immunology and Cancer Therapy
• Molecular and Cellular Biology

Background:

• Engineered T-cell therapies offer promise for refractory malignancies but carry risks of unpredictable toxicities.
• A safety mechanism for selective T-cell depletion post-administration is crucial for managing potential adverse events.
• Rapamycin, an immunosuppressant, facilitates protein-protein interactions via FKBP12-FRB binding.

Purpose of the Study:

• To develop a controllable suicide gene system for engineered T-cells.
• To create a rapamycin-inducible caspase 9 suicide gene for selective T-cell ablation.
• To validate the functionality of the rapamycin-induced caspase 9 system in preclinical models.

Main Methods:

• Engineered a two-protein system where caspase 9 activation was mediated by rapamycin-induced FKBP12 and FRB binding.
• Developed a single-protein construct, rapamycin caspase 9 (rapaCasp9), by fusing FKBP12 and FRB to the caspase 9 catalytic domain.
• Co-expressed rapaCasp9 with a CD19 chimeric antigen receptor (CAR) for functional assessment.

Main Results:

• Demonstrated rapamycin-inducible activation of caspase 9 using the two-protein system.
• Successfully generated and validated the compact single-protein rapaCasp9 construct.
• Confirmed the functionality of rapaCasp9 in controlling engineered T-cells both in vitro and in vivo when co-expressed with a CD19 CAR.

Conclusions:

• The developed rapamycin-induced caspase 9 suicide gene (rapaCasp9) provides a controllable safety switch for engineered T-cell therapies.
• This system enables selective depletion of T-cells, mitigating risks associated with prolonged persistence and potential toxicities.
• rapaCasp9 represents a promising tool for enhancing the safety profile of adoptive T-cell immunotherapy.