Bone marrow or stem cell transplantation
Ex vivo Generated Lymphoid Progenitors for Immune Reconstitution in the Context of Allogeneic Transplantation and Gene Therapy
The thymic microenvironment supports T-cell commitment. An in vitro feeder-free cell culture based on Notch ligand DL-4 and cytokines has been developed in our laboratory leading to the generation in 7 days of T-cells progenitors from both human cord blood and adult CD34+ hematopoietic stem and progenitor cells (HSPCs).
Since TNFα is constitutively synthesized in the thymus, we explored its role during human early T-cell development. TNFα accelerated early T-cell differentiation and greatly increased the number of CD34-CD7+CD5- T-cell precursors generated in the in vitro DL-4 culture both for cord blood and adult CD34+ HSPCs. TNFα improved the production of CD34-CD7+CD5-/lo cells at the expense of myeloid and NK progenitors leading to a highly purified population of CD7+ T cell precursors. These T-cell precursors expressed early T-cell commitment markers as shown by extensive RNAseq analysis and had a high T-cell differentiation potential in vitro upon differentiation on OP9/DL1 cells and in vivo when transplanted to NOD/SCID/γc-/- mice. TNFα increased specifically the rate of proliferation in CD7+ T cell precursors between day 4 and 7 by promoting T-cell progenitors entry into the cell cycle through the activation of NFkB pathway.
We further demonstrated that this improved culture system can be combined with lentiviral gene transfer to produce large numbers of gene-corrected T-cell progenitors, opening new cell and gene therapy approaches for immune deficiencies.