Costimulation of IDO-specific T cells reduced Treg numbers while boosting IL-17, IL-6, and TNF-α production. PBMCs from patients with HLA-A2⁺ melanoma cancer stimulated in vitro with MART-1_26-35 (EAAGIGILTV) peptide either in coculture with IDO5 peptide or an irrelevant peptide (HIV-1 pol_476-484). The percentage of CD4⁺CD25^highCD127⁻Foxp3⁺ Tregs (A-B) and IL-17A–producing CD4⁺ T cells (C-D) in each culture was identified by flow cytometry with intracellular staining for Foxp3 and IL-17A, respectively. For comparison, cells were stained with isotype controls. Examples of Treg staining (A) and IL-17A staining (C) of PBMCs stimulated in vitro with MART-1_26-35 peptide either in coculture with an irrelevant peptide (HIV-1 pol_476-484; top) or IDO5 peptide (bottom). Examples shown are from 1 patient, representative of 4 different patients (A,C). Distribution of CD4⁺ and CD8⁺ T cells in the cultures (E). Secreted cytokines (IL-10, IL-17A, IL-6, and TNF-α) in cell culture supernatants quantified by ELISA (F). Data are mean ± SD (n = 4 patients). White bars indicate MART-1_26-35–stimulated PBMCs in coculture with an irrelevant peptide (HIV-1 pol_476-484); black bars, MART-1_26-35–stimulated PBMCs in coculture with IDO5 peptide.