A Protease-Mediated Regulatory Chimeric Antigen Receptor (Car): "Double Arm" Car System Improves Tumor-Cell-Specificity Of Car-T Cell Therapy

【Introduction】 CAR-T cell therapy has shown excellent therapeutic effects against some malignancies including refractory B cell lymphoma or leukemia. This adoptive T cell transfer therapy is an attractive methodology; however, there are several disadvantages to be overcome. Current CAR-T cell therapy targets single cell-surface molecule, which leads to damage of normal cells expressing the target protein. Such “On-target / off-tumor” effect is one of the major adverse effects. Improvement of target-cell-specificity is needed to avoid this serious adverse effect and to expand target diseases of this therapy. Here, we show the protease-mediated “Double-Arm” CAR-T cell system, which improved the specificity of CAR-T cell therapy by recognizing two distinct cell-surface proteins. We designed two types of CARs: “Effector CAR” and “Scissors CAR”. The “Effector CAR” is constituted of a single chain Fv fragment (scFv) targeting a cell-surface protein (protein X) on tumor cells, Human Immunodeficiency Virus protease (HIVPR) recognition polypeptide sequence, and a functional domain of CD3-zeta. The “scissors CAR” is constituted of a recognition portion targeting another protein (protein Y) and HIVPR. The HIVPR induces cleavage of the recognition polypeptide sequence in the effector CAR leading to inactivation of the effector CAR when the CAR-T cells contact with cells expressing both proteins X and Y.