Fig. 3: CS of biological ligands on PDAC organoids reveals major axes of transcriptional response that are recapitulated in single-ligand validation screens with clinical relevance.

a, Overview of a CS designed to explore the effects of macrophage-derived ligands on patient-derived PDAC organoid transcriptional states. b, Overview of experimental setup for the 68 biological ligand CS (with select single-ligand landmark perturbations) on PDAC organoids with a scRNA-seq readout. c, Overview of computational analyses on scRNA-seq results from the CS used to identify GEPs and the ligands that induce them. d, Scatter plot of significant ligand effects on cNMF modules (deconvolution regression coefficients) from two CSs with distinct random pooling. e, Heat map of the mean significant ligand effects on cNMF modules over both CS runs. f, Top, overview of a single-ligand validation experiment. The top 11 hits (adjusted P value < 0.05) that emerged in both CSs were validated by performing single-ligand treatments in duplicate. Hits are grouped by the GEPs they induce, and landmark perturbations are noted with an asterisk. DMSO-treated wells served as negative controls. Bottom, Heat map visualizing the Pearson correlation between the effects of each ligand on the CS cNMF GEPs in the CS (rows) and the effects of the same ligands on the CS cNMF GEPs in the single-ligand experiment (columns). g, Top, Venn diagrams depicting the number of shared and unique genes between the cNMF PDAC IL-4 and IL-13 response GEP and corresponding signatures in MsigDB. Bottom, Kaplan–Meier survival plots of TCGA PAAD cohort (n = 182, bulk RNA-seq expression data) based on (left to right) our PDAC IL-4 and IL-13 response module or three gene signatures (Reactome, Biocarta and Lu et al. IL-4) from MsigDB.