characterization of car t cells manufactured using genetically engineered artificial antigen presenting cells

Objective: chimeric antigen receptor (car) t cell therapy has recently emerged as a promising approach for thetreatment of different types of cancer. improving car t cell manufacturing in terms of costs and product quality is animportant concern for expanding the accessibility of this therapy. one proposed strategy for improving t cell expansionis to use genetically engineered artificial antigen presenting cells (aapc) expressing a membrane-bound anti-cd3 fort cell activation. the aim of this study was to characterize car t cells generated using this aapc-mediated approachin terms of expansion efficiency, immunophenotype, and cytotoxicity.materials and methods: in this experimental study, we generated an aapc line by engineering k562 cells to expressa membrane-bound anti-cd3 (mokt3). t cell activation was performed by co-culturing pbmcs with either mitomycinc-treated aapcs or surface-immobilized anti-cd3 and anti-cd28 antibodies. untransduced and cd19-cartransducedt cells were characterized in terms of expansion, activation markers, interferon gamma (ifn-γ) secretion,cd4/cd8 ratio, memory phenotype, and exhaustion markers. cytotoxicity of cd19-car t cells generated by aapcsand antibodies were also investigated using a bioluminescence-based co-culture assay.results: our findings showed that the engineered aapc line has the potential to expand car t cells similar to thatusing the antibody-based method. although activation with aapcs leads to a higher ratio of cd8+ and effector memoryt cells in the final product, we did not observe a significant difference in ifn-γ secretion, cytotoxic activity or exhaustionbetween car t cells generated with aapc or antibodies.conclusion: our results show that despite the differences in the immunophenotypes of aapc and antibody-based car tcells, both methods can be used to manufacture potent car t cells. these findings are instrumental for the improvementof the car t cell manufacturing process and future applications of aapc-mediated expansion of car t cells.