Disruption of LAMP1 expression inhibits cytotoxic activity of NK cells. (A) YTS (top) or ex vivo isolated NK cells (bottom) were untransduced (UT; YTS cells), mock-transduced (mock; NK cells), or transduced with control (CTRL) or LAMP1 RNAi. Levels of LAMP1 transcripts in the indicated cells were analyzed by real-time PCR; relative expression of LAMP1, normalized to actin, is shown. Data are represented as mean + standard deviation (SD) from 6 (YTS) or 3 (NK) analyses. **P < .01; ***P < .001; one-way analysis of variance (ANOVA). (B) Protein level of LAMP1 in the indicated cells was analyzed by western blot (left; actin was used as a loading control) and by flow cytometry (right). (C) Cytotoxic activity of YTS and NK cells, untransduced, mock-transduced, or transduced with CTRL or LAMP1 RNAi. The percentages of 721.221 target-cell lysis for different effector-to-target (E:T) ratios are shown as mean values + SD determined from at least 5 experiments. *P < .05; ***P < .001; 2-way ANOVA. (D) Ab-dependent cell-mediated cytotoxicity. Ex vivo isolated NK cells, either mock-transduced or transduced with CTRL or LAMP1 RNAi, were incubated with SK-OV3 target cells, in the presence of either human IgG (negative control) or anti-HER2 Ab. The percentage of target-cell lysis was determined from 3 experiments with different donors and is shown as means + SD. ***P < .001; 2-way ANOVA.