In VitroExpansion of Antigen-Specific CD8+ T Cells from Human PBMCs Using Artificial T-cell Stimulating Microparticles

Abstract

Abstract While current T cell-based immunotherapies have shown moderate success, clinical translation of these methods has been hindered by biological variability, limited efficacy, and high expense. Here, we create a novel T cell stimulating platform for antigen-specific CD8+ T cell activation and expansion, eliminating the extensive ex vivoexpansion steps and thus making it accessible for many more cancer patients. The scaffold is formed using thiol-modified hyaluronic acid cross-linked with PEGDA, magnetic dextran nanoparticles, as well as DTT-reduced MHC-Ig dimer HLA-A*02 (A2) and anti-CD28. The gel is passed through a mesh device to form microparticles (MPs), which are then passively loaded with peptides (here, CMV and MART1) for ease of multiple expansions. They were plated with human PBMCs thawed overnight in culture media supplemented with human AB serum and a cocktail of cytokines (IL-1b, IL-2, IL-4, IL-6, and IFNγ). On days 7 and 14, cells were harvested and analyzed for proliferation of antigen-specific cells. After fourteen days of culturing PBMCs with A2 MPs, we could detect a significant expansion of antigen-specific T cells via flow cytometry. When stimulated with peptide-pulsed target cells for an intracellular cytokine assay, they expressed one or more of IFNγ, TNFα, and IL-2, demonstrating their polyfunctionality. We also observed a greater population of memory T cells, which indicates that these cells may persist longer in the body. We have developed a biomaterial-based T cell activation platform that provides critical signals, mimicking the lymph nodes. These MPs are effective in expanding antigen-specific T cells from human PBMCs, and these cells have robust killing of target splenocytes.