The STING agonist cGAMP enhances AMG 330–mediated cytotoxicity against AML cells. (A-B) Flow cytometric analysis of AMG 330–mediated (5 ng/mL) cytotoxicity after 72 hours against HL-60 cells in cocultures with either CD8⁺ (A) or CD4⁺ (B) human T cells (n = 4). Specific lysis was calculated relative to the c-BiTE condition. The concentration of added cGAMP in cocultures was 40 μg/mL. (C-D) Effector-to-target cell ratio–dependent cytotoxicity after 72 hours against HL-60 cells in cocultures with either CD8⁺ (C) or CD4⁺ (D) human T cells treated as indicated (n = 3). (E) Percentage of caspase-3–positive HL-60 cells measured by intracellular staining and flow cytometry after 72 hours of coculture with human Pan–T cells treated as indicated (n = 3). (F) The percentage of granzyme B⁺ T cells, determined by intracellular staining and flow cytometry after 72 hours in cocultures with HL-60 cells treated as indicated (n = 3). (G) The percentage of TRAIL⁺ T cells, determined by flow cytometry after 72 hours in cocultures with HL-60 cells (n = 6). (H) Flow cytometric analysis of T-cell degranulation measured by staining surface CD107a after 72 hours in cocultures with HL-60 cells treated as indicated (n = 3). (I-J) Secretion of IFN-γ and TNF, determined after 72 hours by cytometric bead array (CBA) analysis, from cocultures of human T cells and HL-60 cells treated as indicated (n = 3). (K) Human T-cell proliferation expressed as fold change in CD2⁺ cells on day 6 of coculture with HL-60 cells in the presence of AMG 330 ± 10 μg/mL cGAMP (E:T of 1:20) normalized to c-BiTE conditions (n = 3). After 3 days, half of the medium was exchanged with fresh medium containing AMG 330 ± cGAMP. All graphs present the mean ± standard error of the mean (SEM). Statistical analysis was performed using ordinary 1-way analysis of variance (ANOVA) with the Tukey comparison or the 2-way ANOVA with Šidák correction (panels C-D). ns, P > .05; ∗P < .05; ∗∗P < .01; ∗∗∗P < .001. E:T, effector-to-target cell ratio; ns, not significant.