Characteristics of the AURKA_207-215–specific CTL clone AUR-2. (A) Representative flow cytometry plots showing staining of AUR-2 with the HLA-A*0201/AURKA_207-215 tetramer (left) and the irrelevant HLA-A*0201/Gag_77-85 tetramer (negative control; right). (B) The cyotoxic activity of AUR-2 was measured in ⁵¹Cr-release assays against C1R-A2 or C1R (negative control) cells loaded with a range of AURKA_207-215 peptide concentrations as indicated. E/T indicates effector:target ratio. (C) IFN-γ ELISPOT assays were conducted using C1R-A2 target cells loaded with 1μM AURKA_207-215 peptide and AUR-2 CTL at different input numbers as shown. (D) ⁵¹Cr-release assays were conducted using AUR-2 CTL with unpulsed or AURKA_207-215 peptide-pulsed (1μM) HLA-A*0201⁺ autologous or allogeneic B-LCLs, C1R-A2 cells or HLA-A*0201⁻ allogeneic B-LCLs as indicated. E/T indicates effector:target ratio. (E) The cytotoxic activity of AUR-2 CTL against the indicated leukemia cell lines was measured in ⁵¹Cr-release assays. GANMO-1, HLA-A*0201⁺; MEG01 and K562, HLA-A*0201⁻. Expression of AURKA mRNA and AURKA protein in these leukemia cell lines is shown in supplemental Figure 2. E/T indicates effector:target ratio. (F) Construction of a novel retroviral vector encoding full-length AURKA–specific TCR α and β genes derived from AUR-2. MoMLV indicates Moloney murine leukemia virus; LTR, long terminal repeat; EF1a, elongation factor 1a; PGK, phosphoglycerate kinase promoter; and MSCV, murine stem cell virus. Error bars represent SDs.