AML cells convert CD4⁺CD25⁻ into CD4⁺CD25⁺ T cells through an IDO-dependent mechanism. (A) Flow cytometry analysis of CD4⁺CD25⁺ and CD4⁺CD25⁻ T cells before and after culture with allogeneic IDO-expressing AML cells in the presence and absence of 1-MT (1000 μM). Results are representative of 7 independent experiments. *P = .03, experimental versus control sample. (B) Annexin-V staining and flow-cytometry analysis of CD4⁺CD25⁺ and CD4⁺CD25⁻ cells after culture with IDO⁺ AML cells. Cells were analyzed at different time points. Results are representative of 3 independent experiments. (C) Purity of CD4⁺CD25⁻ cells. Results are representative of 4 independent experiments. (D) Purified CD4⁺CD25⁻ T cells were incubated for 7 days with normal MNCs (CTR) or AML cells in the presence and absence of 1-MT. At the end of culture, CD4⁺ cells were gated and analyzed for the presence of converted CD4⁺CD25⁺ cells. Results are the mean ± SD of 4 independent experiments. (E) Purified CD4⁺CD25⁻ T cells were resuspended in the conditioned medium collected from IDO⁻ and IDO⁺ AML cells and stimulated with IDO⁻ AML cells for 7 days. At the end of culture, CD4⁺ cells were gated and analyzed for the presence of converted CD4⁺CD25⁺ cells. Results are representative of 3 independent experiments.