(9a) Rational Engineering of an or-Gate CAR for Adoptive T-Cell Therapy Against B-Cell Malignancies

Chimeric antigen receptor (CAR)-based immunotherapy for treatment of cancer has shown great promise in recent years for the treatment of cancers resistant to surgery, chemotherapy and radiation. The most clinically successful CAR, targeting the B-cell antigen CD19, has been shown to be effective against both acute and chronic B-cell malignancies. However, the CD19 CAR can be susceptible to mutational escape as demonstrated by clinical trials in which multiple patients treated with the single-input CD19-targeting CAR relapsed due to the emergence of CD19-negative leukemic cells. As a solution to this challenge, receptors that trigger robust T-cell activation in response to multiple antigens can significantly reduce the probability of antigen escape and increase the robustness of CAR–T-cell therapy. Here, we present the development of OR-gate CARs targeting CD19 and CD20, two clinically relevant antigens associated with B-cell malignancies. Bispecific CARs were rationally designed and optimized based on the structural requirements of CD19 and CD20 CARs. Primary human T cells expressing CD19-OR-CD20 CARs demonstrate robust cytokine production and efficient lysis of Burkitt’s lymphoma cells, including tumor cells that have lost CD19 expression due to CRISPR/Cas9-guided genome engineering. To our knowledge, this is the first demonstration of a bi-specific CAR that effectively targets a clinically relevant pair of tumor antigens, and ongoing work aims to optimize the technology for clinical translation.