Inhibition of natural cytotoxicity by anti-NKp44L mAb. Primary NK cells from healthy donors were purified, activated by IL-2 for 7 to 10 days, and then analyzed for cytotoxic activity against various tumors cell lines. (A) NK cytotoxicity of the 721.221 and K562 target cell lines, at different effector/target cell ratios. Open squares: IgM-isotype control-treated target cells. Closed squares: anti–NKp44L-treated target cells. Open circles: IgG1-isotype control-treated effector cells. Closed circles: anti-NKp44-treated effector cells. (B) Killing pattern of different target cells: WM1361, 1106mel, LB33mel-B1, HeLa, H441, and HEK-293 cell lines were tested for their sensitivity to NK lysis after treatment with anti-NKp44L mAb (closed squares) or IgM-isotype control (open squares), at different effector/target cell ratios. (C) NK lysis sensitivity of other hematologic (MT2, H9, Mono Mac, and U937), and nonhematologic (HM11, Mel1080, SkMel5, and JAR) tumor cells, after treatment with anti-NKp44L mAb (closed bars) or IgM-isotype control (open bars). Data are shown for an effector/target (E/T) ratio of 5:1. (D) Cytotoxicity of NK-resistant EL4 cells stably transfected with NKp44L. EL4 cells were either transfected either with the full length (triangles or diamonds), or the C-terminal-deleted (Δ21spe) (circles) sequence of NKp44L, or the control vector (squares). Transfected cells were tested for their NK sensitivity in the presence of IL2-activated NK cells without (plain lines and open symbols) or after pretreatment with specific mAbs (dotted lines and closed symbols). EL4 cells were treated (dotted lines) or not (plain lines) with anti-NKp44L mAbs (+αNKp44L) (left panel), whereas in the right panel, NK cells were treated (dotted lines) or not (plain lines) with anti-NKp44 mAbs (+αNKp44). More than 30% of CD3^-CD56⁺ cells expressed NKp44. Data are representative of 2 independent experiments.