1-7F9 increases NK cell–mediated lysis of primary AML blasts. (A) Lysis of primary AML cells by IL-2–activated NK cells of a healthy, KIR ligand-matched donor, in the presence of 1-7F9 at 10 μg/mL (▨) or without antibody (□), measured in standard 51Cr assay at the indicated E:T ratios. Differences were analyzed by Student t test (***P < .005). Experiment shown is representative of different experiments using different effector and target blast cells. (B) Patient NK cells expanded in IL-2 were mixed with freshly thawed, autologous AML blasts obtained at diagnosis. The 1-7F9, control hIgG4, or a mixture of mouse anti-KIR2DL1 and -2/3 F(ab′)2 fragments was added to each well (each at 30 μg/mL final concentration), and after 4 hours, percent specific lysis was measured by a flow cytometry–based assay. Results shown are representative of 4 independent experiments. (C) Patient NK cells (n = 4 for the E:T 30:1 level, plus n = 5 at E:T 15:1) were incubated with 1-7F9 or hIgG4 isotype control mAb and cocultured in the presence of autologous AML blasts. Cytotoxicity, measured as granzyme B release, is shown. Statistically significant differences in cytotoxicity (*P = .01; **P < .03), as a function of granzyme B release were observed at multiple E:T ratios as shown (■ = 1-7F9; ▩ = IgG4 isotype control). No appreciable granzyme B release was found in effectors and targets cultured alone (negative controls for the assay). As a positive control for the ELISPOT granzyme B assay readout, an equivalent number of NK cells from a healthy donor was cultured in 30:1 ratio against the NK cell–sensitive K562 cell line (■). Results are representative of 2 independent experiments in 2 patients.
