Fig. 3.
Induction of cancer/testes antigen-specific CD4+ T cells by depletion of CCR4- or CD25-expressing T cells in healthy donors. (A) CD4+ T-cell responses to NY-ESO-1 peptides after depletion of CCR4+ or CD25+ T cells. CD4+ T cells prepared from PBMCs of healthy donors were presensitized with APCs pulsed with NY-ESO-1 peptide covering the entire sequence of NY-ESO-1. Results of 2 (HD#1 and HD#7) among 16 healthy donors are shown. The numbers of IFN-γ–secreting CD4+ T cells were assessed by ELISpot assay. (B) Intracellular cytokine secretion of CD4+ T cells shown in A. The numbers in figures indicate the percentage of gated CD4+ T cells. (C) Peptide dose-dependent recognition of NY-ESO-1–specific IFN-γ–secreting CD4+ T cells. NY-ESO-1–specific CD4+ T cells derived from CCR4+ or CD25+ T-cell–depleted cells (CCR4 dep and CD25 dep, respectively) were cultured with autologous activated T-cell APCs pulsed with graded amounts of NY-ESO-1 peptides and assessed for the number of IFN-γ–secreting cells as in A. Triangles indicate responses to control peptide at 10 μM. (D) Recognition of naturally processed NY-ESO-1 protein antigen by NY-ESO-1–specific CD4+ T cells derived from whole CD4+, CCR4+ cell-depleted, or CD25+ cell-depleted cells. NY-ESO-1–specific CD4+ T cells from two healthy donors were cultured with autologous dendritic cells pulsed with NY-ESO-1 or control protein, or with NY-ESO-1 or control peptide. The experiments were independently performed twice with similar results.