The effects of IMD-0354 on NF-κB activity and cell survival in Epstein-Barr virus positive T- or NK-cells.

<p>(A) Dual luciferase assay for NF-κB in SNT8, SNT15, SNT16, and SNK6 cells treated with IMD-0354. The cells were transfected with 10 μg of pNF-κB-luc and 1 μg of pRLSV40. Twelve hours after transfection, cells were treated with IMD-0354 for 24 hours and harvested for the dual luciferase assay. Luciferase activity was normalized to <i>Renilla luciferase</i> activity and expressed as an increase relative to the control. (B) SNT8, SNT15, SNT16, and SNK6 cells were treated with IMD-0354 for 24 hours and the viable cell numbers were estimated using the XTT assay and expressed in arbitrary units. Data are shown as the means ± standard deviations of 3 independent experiments. (C) SNT8, SNT15, SNT16, and SNK6 cells were treated with IMD-0354 as indicated for 24 h and subjected to immunofluorescent staining. NF-κB protein expression and localization were examined by immunofluorescent staining using anti-p50, p52, RelA, and RelB antibodies as indicated. DAPI was used for nuclear staining. Cells were analyzed via confocal microscopy. (D) SNT8, SNT15, SNT16, and SNK6 cells were treated with IMD-0354 as indicated for 24h and used in the assay. Cells were stained with Annexin V and PI and subsequently analyzed by flow cytometry.