CP-690,550 inhibited the proliferation of the cytokine-dependent NK92 cell line mediated by cytokines that signal through Jak3/STAT5 but not by cytokines that use other pathways. (A) After cytokine starvation for 24 hours, NK92 cells were stimulated by the addition of human IL-2 for 48 hours with and without the addition of serially increasing concentrations of CP-690,550. ³H-thymidine was added during the last 6 hours of the cultures. Cells were then harvested and analyzed for ³H-thymidine incorporation. (B) Cytokine-starved NK92 cells were stimulated with human IL-2, combined IL-6/IL-6R, or IL-12 for 48 hours with and without serially increasing concentrations of CP-690,550. (C) The NK92 cells were treated as those in panel B with and without a single 50nM dose of CP-690,550 (CP). Data are presented as means ± SD (A-C) and are representative of 3 independent experiments. (D) After cytokine starvation for 24 hours, NK92 cells were stimulated with 30 ng/mL of IL-2, 100 ng/mL of combined IL-6/IL-6R, or 100 ng/mL of IL-12 for 1 hour with and without the addition of CP-690,550 (CP). The cell lysates were immunoblotted with an anti–phospho-STAT5 monoclonal antibody and an anti-STAT5 antibody. β-Actin was used as an input control. Data are representative of 3 independent experiments.