Tyrosine kinase inhibitor dasatinib blocked the activation, cytokine production, proliferation, and cytolytic activity of resting and, to a lesser extent, activated CAR T cells in vitro by blocking the phosphorylation of LCK, CD3ζ, and ZAP70, and limiting the induction of NFAT. Dasatinib inhibited CAR T cell function even in the presence of endogenous TCR stimulation. The induced “off” state persisted for up to 7 days, was rapidly and completely reversible upon dasatinib discontinuation, and did not reduce CAR T cell viability. In mice, dasatinib reversibly paused CAR T cell antitumor activity and prevented early-onset cytokine release syndrome.
Immunotherapy with chimeric antigen receptor (CAR)-engineered T cells can be effective against advanced malignancies. CAR T cells are "living drugs" that require technologies to enable physicians (and patients) to maintain control over the infused cell product. Here, we demonstrate that the tyrosine kinase inhibitor dasatinib interferes with the lymphocyte-specific protein tyrosine kinase (LCK) and thereby inhibits phosphorylation of CD3zeta and zeta-chain of T cell receptor-associated protein kinase 70 kDa (ZAP70), ablating signaling in CAR constructs containing either CD28_CD3zeta or 4-1BB_CD3zeta activation modules. As a consequence, dasatinib induces a function-off state in CD8(+) and CD4(+) CAR T cells that is of immediate onset and can be sustained for several days without affecting T cell viability. We show that treatment with dasatinib halts cytolytic activity, cytokine production, and proliferation of CAR T cells in vitro and in vivo. The dose of dasatinib can be titrated to achieve partial or complete inhibition of CAR T cell function. Upon discontinuation of dasatinib, the inhibitory effect is rapidly and completely reversed, and CAR T cells resume their antitumor function. The favorable pharmacodynamic attributes of dasatinib can be exploited to steer the activity of CAR T cells in "function-on-off-on" sequences in real time. In a mouse model of cytokine release syndrome (CRS), we demonstrated that a short treatment course of dasatinib, administered early after CAR T cell infusion, protects a proportion of mice from otherwise fatal CRS. Our data introduce dasatinib as a broadly applicable pharmacologic on/off switch for CAR T cells.
Author Info: (1) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (2) Center for Cell Engineering and Immunology Program, Sloan Kettering Institute
Author Info: (1) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (2) Center for Cell Engineering and Immunology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (3) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (4) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (5) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (6) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (7) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (8) Center for Cell Engineering and Immunology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA. (9) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. (10) Medizinische Klinik und Poliklinik II, Universitatsklinikum Wurzburg, 97080 Wurzburg, Germany. hudecek_m@ukw.de.
