TCR-transduced cells from responding patients persisted and showed antitumor activity ex vivo. Blood samples were taken from patients' cells before and after TCR-transduced cell infusion. PBMCs were evaluated for persistence of infused cells in peripheral blood after treatment by specific tetramer staining and were also used directly in coculture assays with mel624 tumors (MART1⁺, gp100⁺, HLA-A2⁺). Antitumor activity was evaluated by IFN-γ and IL-2 ELISPOT, and also by intracellular staining for IFN-γ production. (A) Persistence and activity of DMF5 patient treatment cells before, 2 weeks after, and 1 month after infusion. Responding (PR) and nonresponding (NR) patients are represented by solid and broken lines, respectively. (B) Comparison of PBMCs from patients treated with either DMF5 or gp100(154) TCR-transduced cells. Tetramer staining and ELISPOT analysis of IFN-γ and IL-2 production from nonresponding (NR) and responding (PR) patients at 1 month after treatment. All samples had less than 10 ELISPOTs (per 100 000 PBMCs) and less than 1% IFN-γ–positive cells, respectively, against the HLA-A*02⁻ mel888 tumor (data not shown). Patients with objective clinical responses (PR) had higher numbers of antitumor IFN-γ (P = .02) and IL-2 (P = .02) secreting cells than nonresponders (NR).