IDX combined with cis enhances T cells expansion with cytolytic
potential in killing tumor cells
(A) To capture and visualize all mononuclear cell subpopulations in a
single two-dimensional (2D) map, representation of t-distributed
Stochastic Neighbor Embedding (t-SNE) analysis of concatenated
mononuclear cells (3,000 cells/sample) from PBMCs collected from
cocultures was applied. T-SNE represent phenotypic markers defining
specific cells and are assigned specific colors (CD19 B cells in purple;
CD56 natural killer cells in pink; CD14 monocyte in green; CD4 T helper
cells in orange; CD8 cytotoxic T cells in blue). (B) PBMCs cocultured
with tumor cells following treatments as followed: IDX at 1 µM and 2 µM
(IDX1 in orange and IDX2 in green) or cis at 1.2 µM (cis1.2 in pink),
combinatorial treatment with IDX1 µM and cis 1.2 µM (IDX1+cis1.2 in
purple) or IDX 2 µM and cis 1.2 µM (IDX2+cis1.2 in dark green), or left
untreated with DMSO (blue). Differential expression patterns are shown.
(C) Flow cytometry analysis of the frequencies of CD4+and CD8+ cells in the cocultures. (D) CD8/CD4 ratio
was calculated according to the cellular percentage changes between
different treatments. (E) Representative flow cytometry plots of NPC
cell line C17 (target cell; T) stained with CFSE before getting in
contact with PBMCs (effector cell; E) collected from cocultures at
indicated effector:target ratio (E:T) for 3 days. Cells were then
collected and stained with fixable viability dye (FVD) and the dead
cells (%) were measured by flow cytometry. CFSE+ cells are target cells
and FVD+ cells are dead cells, such that dead target cells are
represented by the CFSE + FVD +. (F) Quantitative data of % lysis (in
terms of CFSE+FVD+ cells) was obtained from three independent
experiments. Statistical significance was analyzed using 2-way ANOVA, *p
< 0.05; when compared with control (DMSO).