Loss of androgen receptor signaling in prostate cancer‐associated fibroblasts (CAFs) promotes CCL2‐ and CXCL8‐mediated cancer cell migration

Fibroblasts are abundantly present in the prostate tumor microenvironment (TME), including cancer‐associated fibroblasts (CAFs) which play a key role in cancer development. Androgen receptor (AR) signaling is the main driver of prostate cancer (PCa) progression, and stromal cells in the TME also express AR. High‐grade tumor and poor clinical outcome are associated with low AR expression in the TME, which suggests a protective role of AR signaling in the stroma against PCa development. However, the mechanism of this relation is not clear. In this study, we isolated AR‐expressing CAF‐like cells. Testosterone (R1881) exposure did not affect CAF‐like cell morphology, proliferation, or motility. PCa cell growth was not affected by culturing in medium from R1881‐exposed CAF‐like cells; however, migration of PCa cells was inhibited. AR chromatin immune precipitation sequencing (ChIP‐seq) was performed and motif search suggested that AR in CAF‐like cells bound the chromatin through AP‐1‐elements upon R1881 exposure, inducing enhancer‐mediated AR chromatin interactions. The vast majority of chromatin binding sites in CAF‐like cells were unique and not shared with AR sites observed in PCa cell lines or tumors. AR signaling in CAF‐like cells decreased expression of multiple cytokines; most notably CCL2 and CXCL8 and both cytokines increased migration of PCa cells. These results suggest direct paracrine regulation of PCa cell migration by CAFs through AR signaling. Biopsies were taken directly after prostatectomy from locations of prostate cancer identified by multiparametric MRI and palpation of the tumor and immediately transferred to the laboratory. Primary fibroblast cultures were established as previously described (Villegas and McPhaul, 2005). Prostate cancer-derived fibroblast cells were hormone‐deprived for 3 days and subsequently treated for 8 hours with R1881 (synthetic testosterone) or DMSO control. Then, cells were fixed for 10 minutes with 1% formaldehyde, quenched with glycine and lysed in LB1, LB2 and LB3 (Schmidt et al, 2009). Lysates were incubated with antibodies for AR or H3K27ac.