Fig. 4: CS of an MOA drug library identifies modulators of immune responses across human PBMCs.

a, Overview of CS for examining the immunomodulatory effects of a MOA drug library on the responses of human PBMCs to stimulation with either LPS or IFNβ. b, Experimental overview. c, Left, overview of data processing and analysis workflow. Right, overview of the computational workflow used to analyze the CS data. d, Summary statistics of the number of affected stimulant–response GEPs and number of affected cell types for each molecule in each stimulation condition (LPS or IFNβ). Permutation test P-value cutoff = 0.01. e, Classification of the impact of a compound on cell-type-specific immune modulation. f, Example of a compound (δ-tocotrienol) with diverse effects within the same cell type and across different cell types. The usage scores for each GEP were normalized against the median of the DMSO + IFNβ condition. A Mann–Whitney U-test (two-sided, no correction) was performed to test score differences (Methods). The bottom, center and top lines of the box plots indicate the 25th percentile, median and 75th percentile, respectively. Whiskers are drawn to the farthest datapoints within 1.5× the interquartile range from the nearest hinge. Sample size by stimulation condition: DMSO, n = 396 (monocytes), n = 1,730 (CD4) and n = 404 (NK cells); IFNβ, n = 197 (monocytes), n = 1,276 (CD4) and n = 265 (NK cells); IFNβ + δ-tocotrienol, n = 41(monocytes), n = 631 (CD4) and n = 53 (NK cells). g, Ruxolitinib acts as an IFNβ interferer for CD4 T cell GEP3. A Mann–Whitney U-test (two-sided, no correction) was performed to test score differences (Methods). The bottom, center and top lines of the box plots indicate the 25th percentile, median and 75th percentile, respectively. Whiskers are drawn to the farthest datapoints within 1.5× the interquartile range from the nearest hinge. Sample sizes: DMSO, n = 1,730; IFNβ, n = 1,276; IFNβ + ruxolitinib, n = 1,356. h, Heat map showing the effects of all 90 compounds on PBMC responses to IFNβ (top) and LPS (bottom). Color bars represent the strength of the effect. Negative values denote a negative potentiator or interferer while positive values denote a positive potentiator or interferer. Permutation test P-value cutoff = 0.01. i, Low-dimensional embedding (UMAP) of single-compound validation experiment of ruxolitinib’s effect on LPS responses. j, Evaluation of CS LPS response module (CD8 T cell GEP3) in ruxolitinib-only perturbation. The bottom, center and top lines of the box plots indicate the 25th percentile, median and 75th percentile, respectively. Whiskers are drawn to the farthest datapoints within 1.5× the interquartile range from the nearest hinge. Sample sizes by condition: DMSO, n = 112 (B cells), n = 648 (CD4), n = 376 (CD8), n = 27 (dendritic cells (DCs)), n = 215 (monocytes), n = 188 (NK cells) and n = 27 (γδ T cells); LPS, n = 130 (B cells), n = 1,004 (CD4), n = 478 (CD8), n = 10 (DCs), n = 48 (monocytes), n = 264 (NK cells) and n = 41 (γδ T cells); LPS + ruxolitinib, n = 62 (B cells), n = 475 (CD4), n = 299 (CD8), n = 5 (DCs), n = 19 (monocytes), n = 152 (NK cells) and n = 28 (γδ T cells).