The generation of antigen-specific CD8+ T cells from stem cells for adoptive transfer immunotherapy
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Adoptive T cell transfer is a form of immunotherapy that has shown promise in treating several cancers and post-transplant lymphoproliferative diseases. This therapy relies on the unique ability of cytotoxic T lymphocytes to specifically recognize and eliminate pathogen-infected or malignant cells. Adoptive transfer involves the isolation of patient-derived T cells, followed by ex vivo expansion, and in some cases genetic manipulation, before infusion into the recipient. The procedure is often limited by the availability of donor cells, problems with primary cell expansion, the time required to generate adequate numbers of T cells, and the complications associated with using genetically modified cells in vivo. As a result, there is need for a high-throughput system from which large quantities of antigen-specific cytotoxic T cells can be generated. The multipotency of stem cells makes them attractive, scalable cell sources for adoptive transfer T lymphocytes. In this work, functional, antigen-specific CD8+ T cells were differentiated from human CD34+ cord blood-derived hematopoietic stem cells, in vitro, using exogenous Notch ligands and peptide-loaded human leukocyte antigen tetramers. Tetramer-differentiated, progenitor-derived, antigen-specific CD8+ T cells were then enriched and expanded using media supplemented with co-stimulatory molecules and proliferative cytokines. The enriched T cells remained functional, but did not undergo robust expansion, suggesting that they entered a state of dysfunction. Lastly, the effects of peptide major histocompatibility complex (pMHC) density and surface presentation on thymocyte TCR signaling and antigen-specific differentiation were studied. Microplates and microbeads, fabricated with varying densities of pMHC molecules, were used to stimulate and differentiate thymocytes. Plate- and bead-immobilized pMHCs were more efficient at stimulating thymocytes compared to soluble pMHC tetramers, and were capable of inducing antigen-specific T cell differentiation in a density-dependent manner. In conclusion, the findings of this research indicate that antigen-specific CD8+ T cells can be generated from progenitor cells in vitro, with the potential of high-throughput and large-scale production.