Journal: Scientific Reports

Article Title: Bromodomain and extra-terminal protein inhibitors modulate natural killer cell function and differentiation

doi: 10.1038/s41598-025-33437-1

Figure Lengend Snippet: BETi impact function of primary human NK cells. ( A ) Human primary NK cells isolated from healthy donors ( n = 4) unstimulated or stimulated with IL-12 + IL-15 for 24 h with prior (24 h) treatment with BETi (AZD5153 or CPI-203) compared to no drug treatment control. Bar graph of levels of IFNγ secreted into the cell culture supernatant measured by ELISA reported in pg/ml. ** P ≤ 0.01, * P ≤ 0.05, Ratio paired t-test. ( B ) Percentage of live CD14 − CD19 − cells and of total NK cells (live CD14 − CD19 − CD56 + CD16 + ) within the lymphocyte gate (Supplementary Fig. 2) in PBMCs from healthy donors ( n = 6) incubated in medium only (open circles) or with CPI-203 (black circles) or with AZD5153 (blue circles), in the presence or absence of IL-12 plus IL-18, for 48 h, and then co-cultured with CD4.221 cells for a further 5 h and analyzed. ( C ) Representative histogram overlays illustrating expression of granzyme B in NK cells in the indicated conditions, and bar graphs showing the median fluorescence intensity (MFI) of granzyme B staining in NK cells after culture of PBMCs from healthy donors ( n = 6) for 48 h in medium with or without IL-12 + IL-18 in the absence (open circles) or presence of BET inhibitors CPI-203 (black circles) or AZD5153 (blue circles), followed by co-culture for 5 h with CD4.221 cells in the presence of Brefeldin A. ( D – F ) Percentages of CD107a + ( D ), of IFN-γ + ( E ) and of TNF-α + cells ( F ) in total NK cells within PBMCs from healthy donors ( n = 6) incubated in medium only (open circles) or with CPI-203 (black circles) or with AZD5153 (blue circles), in the presence or absence of IL-12 plus IL-18, for 48 h, and then co-cultured with CD4.221 cells in the presence of Brefeldin A for a further 5 h and analyzed. In ( B – F ), each symbol represents data from a single donor; error bars indicate mean ± sem of n = 6 samples. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.00001; one way-ANOVA with Tukey’s post-test.

Article Snippet: Taqman probes for NCAM1 (Hs00941830_m1), KLRK1 (Hs00183683_m1) and 18s (Hs99999901_s1) RNA were purchased (Thermo Fisher Scientific).

Techniques: Isolation, Control, Cell Culture, Enzyme-linked Immunosorbent Assay, Incubation, Expressing, Fluorescence, Staining, Co-Culture Assay

Journal: Scientific Reports

Article Title: Bromodomain and extra-terminal protein inhibitors modulate natural killer cell function and differentiation

doi: 10.1038/s41598-025-33437-1

Figure Lengend Snippet: Epigenetic regulators control NK cell lineage fate commitment from bone marrow hematopoietic stem cells. ( A ) Schema of ex vivo culture to promote NK cell fate from human bone marrow CD34 + cells ( n = 4–5 donors per condition). ( B ) Bar graphs showing expression of NK lineage marker genes at day 0 and day 21 of culture by quantitative PCR. Error bars indicate mean ± sem. ** P ≤ 0.01, Wilcoxon-Mann-Whitney. ( C ) Bar graphs showing cell viability at day 21 of culture after harvest. ( D ) Bar graph showing overall cell number (cell count) of each culture condition at day 21. Error bars indicate mean ± sem. ** P ≤ 0.01,*** P ≤ 0.0001, Unpaired t-test. ( E , F ) Bar graphs of gene expression of ( E ) NCAM1 and ( F ) KLRK1 determined by qPCR at day 21 of culture. Error bars indicate mean ± sem. *** P ≤ 0.0001, ** P ≤ 0.01,* P ≤ 0.05, Unpaired t-test. ( G , H ) Volcano plot of Log2 fold-change and significance of differentially expressed genes from ( G ) Givinostat or ( H ) EZH2i compared to control determined by RNA-seq of differentiated NK cells from CD34 + cells at day 21( n = 4 per group). Blue dots represent significantly downregulated genes, and red dots represent significantly upregulated genes. Select individual genes are labeled. ( I ) Significantly enriched pathways in EZH2i-treated samples compared to control using GSEA. Number of genes (size), enrichment and normalized enrichment scores (ES, NES), and nominal and FDR-corrected p values shown (NOM p-val, FDR q-val).

Article Snippet: Taqman probes for NCAM1 (Hs00941830_m1), KLRK1 (Hs00183683_m1) and 18s (Hs99999901_s1) RNA were purchased (Thermo Fisher Scientific).

Techniques: Control, Ex Vivo, Expressing, Marker, Real-time Polymerase Chain Reaction, MANN-WHITNEY, Cell Characterization, Gene Expression, RNA Sequencing, Labeling

Journal: Scientific Reports

Article Title: Bromodomain and extra-terminal protein inhibitors modulate natural killer cell function and differentiation

doi: 10.1038/s41598-025-33437-1

Figure Lengend Snippet: Diversity of NK-lineage cells produced from bone marrow precursors. ( A ) UMAP plot showing expression level of the mature NK cell lineage marker CD56 ( NCAM1 ) that was used to sub-select cells for downstream analysis. ( B ) UMAP plots showing individual cells from each control (Control-1 or Control-2) or BETi (CPI203 or AZD5153) conditions of re-clustered NCAM1 -postive only cells. ( C ) Graph-based clustering used to identify unique clusters of cells based on transcriptome expression. 20 distinct clusters were identified. ( D ) Heatmap showing top genes upregulated in each cluster of cells and identification of four major groups of clusters (precursor/myeloid, IL-1R1 + high-expressing cells, CD16- NK cells and mature NK/T cells). ( E ) Bar graphs showing cell frequency in each cluster by sample group. Only clusters with at least 1% of cells shown (Cluster1-14). ( F ) Feature plots of UMAPs showing expression levels of NK cell marker genes.

Article Snippet: Taqman probes for NCAM1 (Hs00941830_m1), KLRK1 (Hs00183683_m1) and 18s (Hs99999901_s1) RNA were purchased (Thermo Fisher Scientific).

Techniques: Produced, Expressing, Marker, Control