(611b) Optimizing T Cell Expansion for Cell Therapy By Regulating IL-2, IL-7 and IL-15

CAR-T cell therapy is regarded as one of the most promising approaches for cancer treatment, but its high cost, partly due to slow T-cell expansion, presents a significant challenge. Cytokines like IL-2, IL-7, and IL-15 are known to influence T-cell growth; however, a comprehensive understanding of their systems-level effects on T cell growth remains unexplored. To address this, we investigated how these specific cytokines regulate the growth of CD4+ naïve and memory T cells, which are key components of adaptive immunity. To achieve this, we isolated CD4+ T cells, activated them, and cultured them in media supplemented with varying concentrations of IL-2, IL-7, and IL-15. Cell counts were performed on Day 3 through Day 7 to track growth rates, and RNA-Seq extraction was conducted on Days 4, 5 and 7 to identify cytokine-regulated genes. The complex interactions between these cytokines and their impact on T cell growth were further analyzed using a fuzzy logic model, a machine learning approach that mimics human reasoning and handles non-linear relationships. The results of our study indicated that cytokine combinations such as high IL-2, high IL-7, median IL-2/IL-7, and median IL-7/IL-15 enhanced T cell expansion compared to controls. The fuzzy logic model that was developed to predict T cell growth under various cytokine concentrations can generally capture synergistic effects—particularly between IL-2 and IL-7, and IL-7 and IL-15. RNA-Seq analysis further identified genes regulated by individual cytokines in CD4+ naïve T cells: IL-2 upregulated chemotactic and angiogenic genes such as CXCL8, CXCL9, and VEGFC; IL-7 influenced ion transport-related genes like TMEM213 and CLCN4; and IL-15 modulated mitochondrial and metal ion regulation genes including TRPM5, MRM1, and MT1H. Compared to CD4+ naïve T cells, the three cytokines had a weaker impact on the growth of CD4+ memory T cells, with minimal synergistic effects observed among them in this subset. These findings pave the way for optimizing cytokine combinations to accelerate T cell expansion, and future studies will extend this systems-level approach to other lymphocyte cells to reduce production costs in immunotherapy.