Reconstitution of EBV-directed T cell immunity by adoptive transfer of peptide-stimulated T cells in a patient after allogeneic stem cell transplantation for AITL
Fig 1
Manufacture of EBV-specific T cells.
(A) Composition of the apheresis product that served as starting material (day 0) and the resulting T cell culture after stimulation with EBV peptides (day 9). Left panel: Proportions of different cell types in CD45+ cells (monocytes: CD14+SSClow, NK cells: CD56+, B cells: CD19+, T cells: CD3+, rest of leukocytes). Right panel: Composition in absolute cell numbers. (B) Percentage of peptide-specific T cells assessed by flow cytometry using peptide-MHC multimers (RLR, HLA-A*03:01; QAK, RAK, HLA-B*08:01; YPL, HPV, EPL, HLA-B*35:01) on day 0 and day 9. (C) IFN-γ secretion of peptide-specific T cells after restimulation with single peptides on day 9, assessed by intracellular cytokine staining (neg.: unstimulated CD8+ T cells, pos.: stimulation of T cells with ionomycin). (D) Flow cytometric analysis of T cell memory/differentiation markers on day 0 and day 9. Plots on the right side are pre-gated on CD3+ cells. temra = terminally differentiated effector memory T cells (CCR7- CD45RA+), eff/em = effector/effector memory T cells (CCR7-CD45RA-), cm = central memory T cells (CCR7+ CD45RA-), naïve = naïve T cells (CCR7+CD45RA+). (E) Percentage of T cell subsets within CD4+ and CD8+ T cells. (F) Flow cytometric analysis of T cell activation markers CD25, HLA-DR and CD38 within the CD4+ and CD8+ T cell compartment.