Fig. 6: High throughput screening identifies signaling pathways involved in paracrine signaling by CAFs to luminal breast cancer cells.

Examples of positive hits from the drug screen showing how the drug dose response affects MCF7 ERE-luciferase activity in mono- or co-cultures with CAF2 cells, as well as viability of both cell types in co-cultures. 4-OH Tamoxifen is shown as an example of a drug that reduce ERE activity (A). Drugs that abrogate the reduced luciferase activity induced by CAFs include the topoisomerase II inhibitor etoposide (B), the HDAC inhibitor valproic acid (C), the CDK4/6 inhibitors palbociclib (D) and abemaciclib (E), the TGF-β inhibitor TEW-7197 (F) and the JAK inhibitor pacritinib (G). The left y-axis displays the ER-α activity (as measured by luciferase) of MCF7 cells alone (blue line) or with CAF2 co-culture (red line). The bar graphs represent the viability of MCF7 cells (blue column), or the CAF2 (red column) on the right y-axis. The effect of TGF-β type I receptor inhibition and JAK inhibition was validated through the use of other inhibitors of the TGF-β type I receptor (SB-431542; TGF-βi, 5 µM) (H) or pan-JAK inhibitor Pyridone 6 (JAKi, 75 nM) (I) on CAF2-mediated reduction of ER-α-activity in MCF7 cells. Pathway activity scores derived from PROGENy demonstrate a significant negative correlation between TGF-β (J) and JAK (K) signaling pathway activity with the expression of ESR1 in luminal breast cancers included in the TCGA dataset. Error bars: SEM with unpaired ordinary one-away ANOVA, Fisher’s LSD multiple comparisons test: *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.