RNA squencing of human NK cells activated with IL-15 or IL-12, IL-15, and IL-18

NK cells are an emerging cancer cellular therapy and potent mediators of anti-tumor immunity. Cytokine-induced memory-like (ML) NK cellular therapy is safe and induces remissions in acute myeloid leukemia (AML) patients. However, the dynamic molecular changes that occur after memory-like differentiation in vitro are unclear. Here, control or ML NK cells purified from normal donor PBMC were generated in vitro. Briefly, RosetteSep-purified NK cells were incubated in IL-12, IL-15, and IL-18, or low-dose IL-15 as a control for 16-18 hours. Control or cytokine-activated NK cells were washed three times and cultured for 6 days in low-dose IL-15, which is required for NK cell survival. After 6 days, RNA was isolated from control and memory-like (ML) NK cells (IL12/15/18 activation) and RNA-sequencing performed. Because the transcription factor GATA-3 was increased specifically in ML NK cells, we hypothesized ML NK cells would exhibit a GATA-3 gene signature compared to control NK cells. Indeed, using GSEA, a significant gene signature was associated with ML NK cell differentiation. These data support the role for GATA-3 in regulating the ML NK cell molecular program. Overall design: NK cells were isolated from 3 distinct human donors (CD56+ CD3-), incubated with low-dose IL-15 (1 ng/mL) or IL-12, IL-15, and IL-18 (IL-12 - 10ng/mL, IL-15 - 50 ng/mL, IL-18 50ng/mL) for 16 hours, washed, and cultured in 1ng/mL IL-15 for 6 days before isolation for sequencing analysis