Fig. 2: Induction of NK cell reactivity by Fc-optimized CD276 antibody against AML cell lines.

PBMC from healthy donors were co-cultured (E:T 2.5:1) in the presence or absence of 8H8_SDIE antibody or the MOPC_SDIE control (both 1 µg/mL). A NK cell activation was analyzed by CD69 expression after 24 h. The left panels show representative flow cytometric results of THP-1 cells, the right panels show separate and pooled data of the indicated leukemic cell lines incubated with PBMC (n = 4). B NK cell activation was determined by CD25 expression after 72 h. The left panels show representative flow cytometric results of THP-1 cells, the right panels show individual and pooled data of the indicated leukemic cell lines incubated with PBMC (n = 4). C NK cell degranulation was analyzed by CD107a expression after 4 h. The left panels show representative flow cytometric results of THP-1 cells, the right panels show individual and pooled data of the indicated leukemic cell lines with PBMC (n = 4). D Intracellular IFNγ expression of NK cells within PBMC was characterized by CD3^-CD56⁺CD16⁺ counterstaining and determined by flow cytometry after 4 h. The left panel shows exemplary data from THP-1 cells, the right panels show individual and pooled data of the indicated leukemic cell lines incubated with PBMC (n = 4). E, F Supernatants of the respective co-cultures were analyzed after 24 h for the release of the immunoregulatory molecules TNF, IL-2, IFNγ and for the effector molecules granzyme A (GrzA), granzyme B (GrzB), perforin (PFN) and granulysin (Grly) by Legendplex assay. The heatmap diagrams show individual results for the leukemic cell lines indicated and different PBMC (n = 4).