Engineering CAR-Mucosal-Associated Invariant T (MAIT) cells for cancer therapy

Abstract

The advent of chimeric antigen receptor (CAR) technology has revolutionized cancer immunotherapy by empowering T cells to recognize and eliminate tumour cells in a highly specific and efficient manner. In recent years, there has been a heightened interest in harnessing unconventional T cell subsets as potential candidates for CAR-based therapies, aiming to address the existing challenges associated with autologous CAR-T therapy. This thesis investigates the feasibility and functions of mucosal-associated invariant T (MAIT)- based cells as a novel therapeutic strategy for cancer immunotherapy. Primary human MAIT cells from peripheral blood mononuclear cells (PBMCs) were genetically engineered with a third-generation CAR targeting CD19, incorporating a truncated epidermal growth factor receptor (tEGFR) as a tracking marker. Upon CAR activation, CAR-MAIT cells demonstrated high levels of cytotoxicity toward target B cells in vitro, comparable to conventional CAR-T cells. The mixed lymphocyte reaction (MLR) assay revealed low alloreactivity of MAIT cells in response to allogeneic stimuli, suggesting their potential for allogeneic therapy. Besides, the study explored optimised manufacturing protocols for generating CAR-MAIT cells, including using retroviral and lentiviral platforms to ensure efficient gene delivery and sustained CAR expression. Additionally, MAIT cell-specific in vitro expansion strategies, employing the synthesized major histocompatibility complex (MHC) class-I related protein 1 (MR1) ligand 5- OP-RU and MR1 tetramer-loaded artificial antigen presenting cell (APC) beads, were explored. Investigations into the effects of different cytokine combinations and serum conditions on MAIT cell expansion provided valuable insights into optimising culture conditions for CAR-MAIT cell production. Together, these findings highlight the potential of CAR-MAIT cells as a promising off-the-shelf approach to cell-based cancer immunotherapy, paving the way for more effective and universally applicable CAR-MAIT cell-based cellular therapies.