Fig. 3
From: CCR7 as a novel therapeutic target in t-cell PROLYMPHOCYTIC leukemia
Anti-CCR7 mAb mediates a strong CDC on T-PLL cells. a CDC on alemtuzumab-naïve patients. Leukemic cells from T-PLL patients (n = 8) were incubated with anti-CCR7, anti-CD52 (alemtuzumab), or an IC at the indicated concentrations and then exposed to rabbit complement for 1.5 h. The percentage of specific cell lysis was determined through quantification of 7-AAD incorporation by flow cytometry. b CDC on alemtuzumab-refractory patients. Specific lysis on T-PLL cells from 4 refractory patients was assayed as described in A. c Relative median fluorescence intensity (RMFI) of CCR7, CD52 and CCR4 in T-PLL cells from alemtuzumab-naïve or alemtuzumab-refractory patients (R/R). Cells obtained from PB samples of patients not treated with alemtuzumab (n = 8; black bars) or refractory to this mAb (n = 4; grey bars) were analyzed by flow cytometry to determine CCR7, CD52, and CCR4 surface density measured as MFI relative to a corresponding IC. d CCR7, CD52, and CCR4 surface expression in T-PLL cells from one representative alemtuzumab-naïve or -refractory patient. E) Proportion of CCR7+, CD52+ and CCR4+ in T-PLL cells from alemtuzumab-naïve or alemtuzumab–refractory patients. The percentage of positive cells was determined in naïve (n = 8; black bars) and refractory patients (n = 4; grey bars). f and g Anti-CCR7 antibody does not mediate antibody-dependent cell-mediated cytotoxicity (ADCC). T-PLL cells were incubated with media alone or in the presence of an IC, anti-CD52 (alemtuzumab) or anti-CCR7 antibodies. f Isolated and IL-2-stimulated human NK cells from a healthy donor were used as effector cells at an effector to target (E:T) ratio of 6:1 (n = 1). g Isolated and IL-2-stimulated wild-type mouse NK cells from spleen (n = 2) were used as effector cells at an effector to target (E:T) ratio of 6:1. After 4 h, the percentage of T-PLL cells killed by ADCC was determined through 7-AAD staining by flow cytometry. In A, B, C, E, F, and G, bars (or symbols) represent mean ± SEM. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ns, not significant