Reactivity of T790M-5- or T790M-7-stimulated T cells against HLA-A2⁺ NSCLC cells harboring EGFR T790M mutation.

<p>(A) CD8⁺ T cells (2×10⁴ cells/well) isolated from the T790M-5-stimulated PBMCs by magnetic beads were examined for their reactivity against different NSCLC cell lines (1×10⁴ cells/well), NCI-H1975 (HLA-A2⁻ T790M⁺), NCI-H1975-A2 (HLA-A2⁺ T790M⁺), or HCC827 (HLA-A2⁻ T790M⁻), by IFN-γ ELISPOT assay (left panel). The T790M-7-stimulated PBMCs (2×10⁴ cells/well) were similarly examined for their reactivity against the same tumor cell lines by IFN-γ ELISPOT assay (right panel). The assays were carried out in duplicate wells, and representative wells are shown. The numbers of spots are shown for each well. The experiments were repeated with blood from three different donors, and representative results are shown. (B) The T790M-5 and T790M-7-stimulated PBMCs (2×10⁴ cells/well) were examined for their reactivity against NCI-H1975-A2 cells (1×10⁴ cells/well) by IFN-γ ELISPOT assay in the absence or presence of 10 µg/ml of anti-HLA class I (W6/32) or anti-HLA-DR (L243) mAb. The assays were carried out in duplicate wells, and the means of spot numbers from duplicate wells are shown. The experiments were repeated with blood from three different donors, and a representative result is shown.