Abstract
Chimeric antigen receptor (CAR) T cell therapy has revolutionised treatment for B cell malignancies. Despite this, it remains constrained by relapse, antigen escape, T cell exhaustion, and safety concerns such as cytokine release syndrome (CRS). NK cells provide innate cytotoxicity, minimise CRS- associated cytokines, and secrete cytokines to support CAR T cell persistence. To improve upon the shortcomings of CAR T cell therapy, we developed a cooperative CAR T and CAR NK cell treatment with dual antigen targeting and complementary effector functions.
To guide the selection of CD19 CAR T and CD20 CAR NK cells, surface profiling of the Raji non- Hodgkin lymphoma cell line was performed, confirming high expression of CD19 and CD20. T and NK cells were isolated from peripheral blood mononuclear cells, with NK cells being expanded in vitro using artificial antigen-presenting cells expressing membrane-bound interleukin-15, interleukin-21, CD86, and 4-1BB ligand. Cells were transduced using a third-generation lentiviral process, resulting in high CAR expression and cell viability.
Functional assays in vitro revealed that CD19 CAR T cells and CD20 CAR NK cells were cytotoxic against Raji cells. To evaluate combined efficacy in vivo, NSG mice were intravenously engrafted with Raji to model disseminated lymphoma prior to treatment with the combination therapy. This revealed early tumour clearance and increases in survival when receiving the combination compared with single- agent treatment.
These findings provide a rationale for a cooperative CAR platform utilising both NK and T cells. This approach addresses recognised limitations of CAR therapy and provides a framework for the next generation of cellular immunotherapies against B cell malignancies.