Fig. 3: Artificial tumor immune microenvironments.
From: Self-assembly of hybrid 3D cultures by integrating living and synthetic cells
A Schematic representation of the ART-TIME screening approach and the panel of immune ligands tested. B Differential gene expression analysis of Panc-1 tumoroids with integrated ART-TIMEs presenting varying combinations of immune ligands. DEGs compared to ART-TIMEs with only PD-1 are shown to derive transcriptional changes based on receptor co-signaling. C Pathway enrichment analysis from PD-1/CD2 DEGs in C. Pathways with the ten highest enrichment scores are shown. D Gene network analysis from PD-1/CD2 DEGs. 5 nodes with the highest degrees are highlighted. E Heat map of protein phosphorylation analysis from Panc-1 tumoroids with ART-TIMEs of PD-1, CD2 and PD-1/CD2. Phosphorylation levels are shown as compared to hybrid tumoroids with plain DSLBs. F Four exemplary protein phosphorylation levels from E, showing synergistic, paradox or de novo effect with PD-1/CD2 co-presentation. Results shown as mean from two antibody spots pooled from forty technical replicates. G Flow cytometry quantification of the fraction of CD25 and CD69 expressing primary human CD8 T cells incubated with pre-conditioned medium of Panc-1 tumoroids with varying ART-TIME compositions. Results are shown from n = 3 donors as mean ± SD from two replicates each. H Enzyme-linked immunosorbent assay-based quantification of soluble PD-L1 and CCL2 secreted from Panc-1 tumoroids with ART-TIMEs of varying composition after 48 hours of incubation. Results shown as mean ± SD from 2 biological replicates measured in technical triplicates. I Ven diagram of PD-L1 and CCL2 transcription factors identified as DEGs in PD-1/CD2 ART-TIME tumoroids. J Flow cytometry quantification of the fraction of CD25-expressing primary human CD8+ T cells incubated with ART-TIME containing Panc-1 3D cultures for 12 h in the presence of increasing bispecific T cell engager concentrations. Results are shown from n = 2 donors as mean ± SD from two replicates each. K Enzyme-linked immunosorbent assay-based quantification of three effector cytokines secreted from primary human CD8+ T cells incubated with Panc-1 tumoroids with ART-TIMEs of varying composition after 12 h of incubation with 5 pM bispecific T cell engager concentration. Results shown as mean ± SD from 2 biological replicates measured in duplicates. p-values are indicated from one-way ANOVA statistical testing.
