Multi-objective optimization reveals time- and dose-dependent inflammatory cytokine-mediated regulation of human stem cell derived T-cell development

Abstract

The generation of T-cells from stem cells in vitro could provide an alternative source of cells for immunotherapies. T-cell development from hematopoietic stem and progenitor cells (HSPCs) is tightly regulated through Notch pathway activation by Delta-like (DL) ligands 1 and 4. Other molecules, such as stem cell factor (SCF) and interleukin (IL)-7, play a supportive role in regulating the survival, differentiation, and proliferation of developing T-cells. Numerous other signaling molecules influence T-lineage development in vivo, but little work has been done to understand and optimize their use for T-cell production. Using a defined engineered thymic niche system, we undertook a multi-stage statistical learning-based optimization campaign and identified IL-3 and tumor necrosis factor α (TNFα) as a stage- and dose-specific enhancers of cell proliferation and T-lineage differentiation. We used this information to construct an efficient three-stage process for generating conventional TCRαβ+CD8+ T-cells expressing a diverse TCR repertoire from blood stem cells. Our work provides new insight into T-cell development and a robust system for generating T-cells to enable clinical therapies for treating cancer and immune disorders.

Sex biased human thymic architecture guides T cell development through spatially defined niches

“…These data show that thymic cells from female and male donors show significant differences in gene expression. Given that higher levels of cytokine signaling has been previously shown to influence thymus and T cell biology 44,45 , our data suggests male and female T cells develop in different signaling environments and may respond differently to cytokine stimuli. This suggests that biological sex should be included as a variable in studies on immune cell and thymic organoid screening models as a critical parameter affecting cell growth, differentiation, and function.…”

“…We first analysed the ability of the four Notch ligands present in the thymus to induce T-lineage commitment or alternative lineage development from cord-blood derived CD34 + hematopoietic stem and progenitor cells (HSPCs) in a defined, feeder-free culture system 44 (Figure 2H). We included titrated concentrations of granulocyte-macrophage colony-stimulating factor (GM-CSF), which is produced by mast cells at the CMJ, and has been shown previously to support development of DCs 51 .…”

A Serum- and Feeder-Free System to Generate CD4 and Regulatory T Cells from Human iPSCs

“…It is worth noting that while the process yielded a mixture of CD34 + and CD34cells, no enrichment or purification for CD34 + CD45 + target cells was required for further differentiation. Indeed, we found that by separation of the free-floating progenitor cells from the main EB we were able to generate lymphoid progenitor cells that co-expressed CD5 and CD7 marking their T cell commitment 49,50 . Continued differentiation of the lymphoid progenitor cells resulted in CD4 and CD8 DP T cells again, without enrichment of the starting population.…”

In Vitro Human Haematopoietic Stem Cell Expansion and Differentiation

“…Other factors: Other factors found to have an influence on T cell lymphopoiesis include IL-7, SCF, IL-3, IL-15, CXCL12 and TNFa. While IL-3 alone appears to stimulate myeloid-biased differentiation, when paired with TNFa there appears to be a strong proliferative effect towards lymphoid-biased differentiation [ 351 ]. CXCL12 (which binds CXCR4) has been found to be expressed on TECs [ 352 ], and promotes Notch-dependent differentiation [ 353 ].…”