Figure 4. CD64 can capture soluble therapeutic mAb to induce cell killing.

A. NK-92MI^CD64 cells preincubated with or without anti-TROP2 or B12 mAb (5 μg/mL) in SFM were stained with an APC-conjugated anti-human IgG secondary fab and analyzed by flow cytometry. Over 95% of NK-92MI^CD64 cells incubated with therapeutic mAb were positively stained compared with unstained NK-92MI^CD64 cells. Histogram is a representative of data from 2 independent experiments. B. mAb-directed killing effect was measured using the Delfia EuTDA cell cytotoxicity assay. Anti-TROP2-NK-92MI^CD64 cell killing of DU145 cells was significantly higher than NK-92MI^CD64 cells alone at the indicated E:T ratios (1 = 8x10³ cells/well) after 2 hours incubation. The experiment was repeated with B12-NK-92MI^CD64 cells cultured with hPrCSC-44 target cell and yielded similar results. Data are represented as mean ± SEM of n=3 technical replicates; n = 3 independent experiments; **, P ≤ 0.01; ****, P ≤ 0.0001 by ANOVA and Tukey’s post hoc for multiple comparisons. C. The experiment was repeated but NK-92MI^CD64 cells were preincubated with or without an isotype control mAb. Effector cells were combined with DU145 or hPrCSC-44 target cells at the indicated E:T ratios (1 = 8x10³ cells/well) and incubated for 2 hours. There was no difference in cell killing between the groups. Data are represented as mean ± SEM of n=3 technical replicates; n = 3 independent experiments.