Mass cytometry reveals alterations in PBMCs and the NK cell phenotype after ALT-803 therapy. (A) viSNE analysis of PBMCs with CD4, CD8, and Treg and B lymphocytes, NK cells, and monocytes annotated. Numbers represent the percentage of cells within the indicated viSNE cell-type island. At day 7 after ALT-803, there is a significant expansion of NK cells, with a significant decrease in CD4 T-cell frequencies and a trend toward lower Treg frequencies (repeated-measures ANOVA). (B) NK cells demonstrate marked phenotypic changes following ALT-803 therapy. viSNE density map analyses of the NK cells pretherapy and at days 5 and 7 posttherapy clustered on 20 markers. Composite of all time points with island gates shown (right). (C) Changes in the NK receptors NKp30 (C) and NKG2D (D) and TIM-3 (E) on NK cell subsets after ALT-803 SQ administration (paired Student t test). (F) Percentage of NK cells positive for GzmB contrasting pretherapy to posttherapy 3 weekly doses of ALT-803. Experiments were performed on PBMC samples from 5 patients after the first dose of 10 μg/kg SQ ALT-803.