Fig. 6.

Combination treatment with anti-PD-L1 Ab and Ro-61–8048 enhances T cell-and NK cell-mediated MM-specific cytotoxic activity (a) MM patient BM CD8⁺ T cells (n = 7) were cocultured with autologous pDCs (pDC:T; 1:10 ratio) in the presence of anti-PD-L1 Ab (5 µg/ml), Ro-61–8048 (100 nM), or Ro-61–8048 plus anti-PD-L1 Ab for 3 days. After washing to remove drugs, cells were cultured with autologous MM cells prestained with CellTracker Violet (T/MM; 10:1 ratio) for 24 h, followed by 7-AAD staining and quantification of CTLs-mediated MM cell lysis by FACS. Left panel: representative FACS scatter plot shows a decrease in number of viable CellTracker Violet-positive MM cells. Right panel: bar graph shows quantification of CD8⁺ CTLs-mediated MM cell lysis, reflected in % viable MM cells, using data obtained in left panel. Percentage of viable MM cells for each treatment versus control (isotype Ab) is presented (data obtained from seven MM patient BM samples; mean ± SD; p < 0.05). b NK cells from MM patient BM (n = 10) were cocultured with autologous pDCs (1:10 pDC: NK ratio) in the presence anti-PD-L1 Ab (5 µg/ml), Ro-61–8048 (100 nM), or Ro-61–8048 plus anti-PD-L1 Ab for 3 days. After washing to remove drugs, cells were cultured with autologous MM cells pre-stained with CellTracker Violet (10:1 NK: MM ratio) for 24 h, followed by 7-AAD staining and quantification of MM cell lysis by FACS. Left panel: representative FACS scatter plot shows the decrease in number of viable CellTracker Violet-positive MM cells. Right panel: bar graph shows quantification of NK-mediated MM cell lysis using data obtained in left panel. The fold change was obtained after normalization with control, and normalized MM cell lysis in each treated sample versus control (isotype Ab) is presented (data obtained from ten0 MM patient BM samples; mean ± SD; p < 0.05)