DNA methylation variations are required for epithelial-to-mesenchymal transition induced by cancer-associated fibroblasts in prostate cancer cells

Abstract

Widespread genome hypo-methylation and promoter hyper-methylation of epithelium-specific genes are hallmarks of stable epithelial-to-mesenchymal transition (EMT), which in prostate cancer (PCa) correlates with castration resistance, cancer stem cells generation, chemoresistance and worst prognosis. Exploiting our consolidated 'ex-vivo' system, we show that cancer-associated fibroblasts (CAFs) released factors have pivotal roles in inducing genome methylation changes required for EMT and stemness in EMT-prone PCa cells. By global DNA methylation analysis and RNA-Seq, we provide compelling evidence that conditioned media from CAFs explanted from two unrelated patients with advanced PCa, stimulates concurrent DNA hypo- and hyper-methylation required for EMT and stemness in PC3 and DU145, but not in LN-CaP and its derivative C4-2B, PCa cells. CpG island (CGI) hyper-methylation associates with repression of genes required for epithelial maintenance and invasion antagonism, whereas activation of EMT markers and stemness genes correlate with CGI hypo-methylation. Remarkably, methylation variations and EMT-regulated transcripts almost completely reverse qualitatively and quantitatively during MET. Unsupervised clustering analysis of the PRAD TCGA data set with the differentially expressed (DE) and methylated EMT signature, identified a gene cluster of DE genes defined by a CAF+ and AR- phenotype and worst diagnosis. This gene cluster includes the relevant factors for EMT and stemness, which display DNA methylation variations in regulatory regions inversely correlated to their expression changes, thus strongly sustaining the ex-vivo data. DNMT3A-dependent methylation is essential for silencing epithelial maintenance and EMT counteracting genes, such as CDH1 and GRHL2, that is, the direct repressor of ZEB1, the key transcriptional factor for EMT and stemness. Accordingly, DNMT3A knock-down prevents EMT entry. These results shed light on the mechanisms of establishment and maintenance of coexisting DNA hypo- and hyper-methylation patterns during cancer progression, the generation of EMT and cell stemness in advanced PCa, and may pave the way to new therapeutic implications.

CD66b+ neutrophils and α‐SMA+ fibroblasts predict clinical outcomes and benefits from postoperative chemotherapy in gastric adenocarcinoma

“…Cancer‐associated fibroblasts (CAFs) are spindle‐shaped fibroblast‐like interstitial cells expressing α‐smooth muscle actin (α‐SMA), constitute a primary fraction of the carcinoma stroma, and are frequently exposed to different inflammatory cells and mediators in the TME . Thus, they may obtain new features that are not present in conventional fibroblasts, and these features tend to mediate reshaping of the TME and ultimately impact cancer evolution . CAFs have an active role in mutual bidirectional interactions with tumor cells and other cell types in the TME, thereby promoting the niche that allows the tumor and promoting tumor development.…”

Treatment with Cannabinoids as a Promising Approach for Impairing Fibroblast Activation and Prostate Cancer Progression

“…We previously demonstrated that CAFs reactivity is crucial to induce an epithelial to mesenchymal transition (EMT) of PC-3 cells gaining them with pro-invasive capabilities [14,29,30]. Thus, to evaluate if cannabinoids could affect also the pro-invasive activity of CAFs, PC-3 cells were exposed for 48 h to the conditioned media obtained from CAFs treated or not with cannabinoids.…”

Concise Review: Prostate Cancer Stem Cells: Current Understanding

“…This includes a shift of CAFs metabolism toward glycolysis with accelerated expression of glucose transporter GLUT1 and increased lactic acid production and secretion. In turn, CAF-generated lactate stimulates mitochondrial biogenesis and aerobic metabolism in PCa cells (reverse Warburg effect) associated with a decrease in expression of GLUT1 transporter and activation of lactate upload [117][118][119][120]. Notably, lactate-mediated tumor acidification has been associated with suppression of anticancer immune response mediated by the loss of cytotoxic T-cell and NK cell function, reduction of dendritic cell maturation and increased helper cell activities as reviewed by Choi and coauthors [121].…”