Fig. 6

Anti-leukemia effects against patient leukemia cells. a-f Primary AML cells of patients were incubated with PBMC of healthy donors (E:T ratio 2.5:1) in the presence or absence of NKG2D-CD16, NKG2D-CD3 or control as indicated (all 10 μg/mL). a, b The expression of CD69 and CD25 as markers for early and intermediate activation, respectively, and CD107a as marker for degranulation were determined after 24 h, 48 h and 4 h, respectively, on (a) NK cells after counterstaining for CD56⁺CD3⁻ or (b) T cells as identified by counterstaining for CD3, CD4 and CD8 by flow cytometry. Combined data for NK cells obtained with 7 different PBMC donors (CD69 and CD107a) and 13 different PBMC donors for CD25 and with 8 different donors (CD69 and CD25) and 4 different donors (CD107a) for T cells are shown (Mean ± SEM). c Intracellular granzyme B and perforin expression in T cells (identified by counterstaining for CD3, CD4 and CD8) was analyzed by flow cytometry (n = 4, Mean ± SEM). d Lysis of leukemia cells was determined by 2 h cytotoxicity assays. Exemplary results obtained at the indicated E:T ratios (left) and combined data obtained with PBMC of 12 different healthy donors at an E:T ratio of 80:1 (right) are shown (Mean ± SEM). e Killing of leukemia cells was determined by flow cytometry based lysis assays at an E:T ratio of 5:1 after 96 h using PBMC of 7 different donors (Mean ± SEM). f Supernatants of cultures were harvested after 4 h and IFN-γ levels were measured by ELISA (n = 10 different donors, Mean ± SEM). g PBMC of AML patients with moderate blast counts (26–75%) were left untreated or incubated with BFP (10 μg/mL each) for 96 h. Then AML cell lysis by autologous NK cells and T cells was determined by flow cytometry. Combined data obtained with samples of 8 different AML patients are shown (Mean ± SEM)