Fig 2: Gemcitabine increases NK cell activating ligand and death receptor expression on OC cells. Phenotype of OC cell lines SKOV-3 (white), IGROV-1 (light gray) and OVCAR-3 (gray) treated with gemcitabine for 48 h (all n = 4). Delta Geometric Mean Fluorescence Intensity (ΔMFI) was determined based on the same sample stained with a live/dead dye only. Protein expression (mean + SD) of NKG2D ligands (a) MICA/B, (b) ULBP1, (c) ULBP2/5/6, (d) ULBP3, DNAM-1 ligands (e) CD112 (Nectin-2), (f) CD155 (PVR), and death receptors (g) CD261 (TRAIL-R1), (h) CD262 (TRAIL-R2), and (i) CD95 (Fas) is shown. Statistical significances were evaluated per cell line by One-way ANOVA with Bonferroni correction

Fig 3: Pre-treatment of SKOV-3 cells with gemcitabine does not improve HPC-NK cell functionality. A-B. Phenotype of SKOV-3 pre-treated with or without 8.8 nM gemcitabine for 48 h in a representative experiment. Overlay of (a) ULBP2/5/6 and (b) CD112 (nectin-2) expression is shown. The control sample contained a live/dead dye only. The numbers on the left show the median fluorescence intensity (MFI), the numbers on the right the delta MFI (ΔMFI), which was determined based on the control sample. C-F. Functionality of HPC-NK cells after 4 h stimulation with SKOV-3 cells pre-treated 48 h with or without 8.8 nM gemcitabine (GEM, n = 2). C-D. Percentage positivity of (c) CD107a (degranulation) and (d) IFNγ expression on living CD56+ perforin+ HPC-NK cells is shown in the absence or presence of an isotype control antibody, DNAM-1 and/or NKG2D blocking antibody. E-F. Percentage inhibition of (e) CD107a (degranulation) and (f) IFNγ positivity by blocking antibodies relative to isotype after 4 h stimulation with pre-treated (8.8 nM GEM) or untreated (0 nM GEM) SKOV-3 cells. Graphs show NK cell donors (NK8-9) with mean in panel C-F. Statistical significance was tested for C-F by repeated-measures Two-way ANOVA with Bonferroni correction