PU.1 is a mediator of the reactive stroma response associated with castration-resistant prostate cancer

Emerging evidence demonstrates the pivotal role played by the tumor microenvironment, particularly cancer-associated fibroblasts, during prostate cancer tumorigenesis and the development of castration-resistant prostate cancer. In this study, we aimed to characterize the stromal gene expression profile of advanced stages of prostate cancer to identify targetable factors associated with castration-resistant prostate cancer progression. Transcriptomic and histological analysis of the mouse stromal component of androgen-sensitive (PNPCa, BM18) and castration-resistant (LAPC9) patient-derived xenograft models representative of advanced prostate cancer was performed. Master regulator analysis of the patient-derived xenograft stromal gene expression profile was performed to identify targetable stromal-associated transcription factors. Patient-derived xenograft collagen-based organoid tumor-fibroblast 3D co-cultures were established to study tumor-fibroblast interactions and to test the effect of an identified fibroblast-targeting compound. Transcriptomic and histological analysis determined the presence of a “reactive” pro-fibrotic stroma in the castration-resistant (LAPC9) versus the castration-sensitive (PNPCa, BM18) xenograft models, characterized by alpha-smooth muscle actin-positive “myofibroblast-like” cancer-associated fibroblasts, high levels of collagen and tenascin C deposition, and upregulation of myofibroblast and inflammatory markers. Intra-tumoral collagen- and tenascin-positive stromal areas specifically correlate with higher tumor invasiveness. Master regulator analysis identified the transcription factor PU.1 as a mediator of the LAPC9 pro-fibrotic phenotype, whose transcriptional activity can be specifically inhibited by a small molecule (DB1976) that prevents its DNA binding. In LAPC9 collagen-based tumor-fibroblast 3D co-cultures, inhibition of stromal PU.1 activity reverted the fibroblast-associated myofibroblast phenotype and, in turn, reduced tumor organoid growth. In this study, we identified the transcription factor PU.1 as a novel targetable molecular player of the pro-fibrotic, cancer-associated fibroblast phenotype in PCa, and we highlighted the applicability of 3D tumor organoid-fibroblast co-cultures as in vitro tools to test the effect of stromal-targeting compounds.