Figure 4.

Effect of ITF2357 on CD126 expression and function in MM cells. (A) MM cell lines were treated with increasing concentrations of ITF2357 for 24 h and stained with CD126-PE or control antibody. The percentage CD126 stained cells relative to untreated control are shown. All cell lines were analyzed in two independent experiments. (B) The H929 MM cell line was treated with medium (gray profile), 0.1 μM (thin lines) or 0.5 μM ITF2357 (thick lines) for 24 h. Cells were then stained with CD126-PE, CD130-PE or control antibodies, as indicated, and analyzed on the FACS. (C–F) The indicated MM cell lines (C–D) or purified CD138⁺ cells from a MM patient (Pt MM3, E–F) were treated for 24 h in the absence (continuous line) or presence (broken line) of 0.5 μM ITF2357 and then recombinant human IL-6 was added during the last 30 min of culture. Cells were then stained with CD126-PE antibody and analyzed on the FACS (C,E). Total cell extracts were analyzed in western blots for the indicated proteins (D,F). The data are representative of six different cell lines and freshly isolated cells from three patients. Most cell lines were analyzed in two independent experiments.