IFN-γ secretion by CD8⁺cells. CD3⁺ T cells from patient 1 (Pt. 1) 1 and patient 14 (Pt. 14) were stimulated with WT1-A (native) peptide. IFN-γ–secreting T cells were measured by ELISPOT assay after challenge with indicated peptides. Controls were no peptide (only CD14⁺ antigen-presenting cells) or irrelevant Ewing sarcoma–derived peptide (EW). Data are mean ± SD from quadruplicate cultures from before GPS administration (time 0), after 6 GPS vaccinations (time 6), and after 12 GPS vaccinations (time 12). A positive response was defined as meeting all of the following criteria: (1) a twofold increase in the IFN-γ–secreting cells and in frequencies of CD8⁺ WT1-A tetramer–positive cells over the controls, (2) P < .05, and (3) >30 spots per 10⁵ cells. Results indicate that a WT1-A–specific response can be generated by challenge to both native and heteroclitic peptide, suggesting processing and presentation of the antigen. In patient 1, a stronger response was seen after 6 vaccinations and then faded. In patient 14, a stronger response was seen after 12 vaccinations.