Zinc Finger E‐Box Binding Homeobox 1 Promotes Cholangiocarcinoma Progression Through Tumor Dedifferentiation and Tumor–Stroma Paracrine Signaling

Abstract

BaCKgRoUND aND aIMS:Zinc finger E-box binding homeobox 1 (ZEB1) is a transcription factor that promotes metastatic and stem cell features, which has been associated with poor prognosis in cholangiocarcinoma (CCA), a desmoplastic cancer enriched in cancer-associated fibroblasts (CAFs). We aimed to define ZEB1 regulatory functions in malignant and stromal compartments of CCA.appRoaCH aND ReSUltS: Bioinformatic and immunohistochemical analyses were performed to determine correlations between ZEB1 and markers of progressiveness in human intrahepatic CCA (iCCA). Gain-of-function and loss-of-function models were generated in CCA cells and liver myofibroblasts as a model of CAFs. Conditioned media (CM) was used to unravel tumor-stroma interplay. In vivo experiments were performed using a xenograft CCA model. ZEB1 expression in tumor cells of human iCCA was associated with undifferentiated tumor and vascular invasion. In vitro, ZEB1 promoted epithelial-mesenchymal transition and stemness in tumor cells, leading to cell migration and spheroid formation. In vivo, ZEB1-overexpressing CCA cells formed larger tumors with more abundant stroma. Expression of cellular communication network factor 2 (CCN2, encoding connective tissue growth factor [CTGF]) was increased in tumor cells from ZEB1overexpressing xenografts and correlated with ZEB1 expression in human tumors. In vitro, CM from ZEB1-overexpressing tumor cells or recombinant CTGF induced myofibroblast proliferation. ZEB1 was also expressed by CAFs in human CCA, and its expression correlated with CCN2 in myofibroblasts and CCA stroma. In mice, cotransplantation of CCA cells with ZEB1-depleted myofibroblasts reduced CCA progressiveness compared to CCA cells/ZEB1-expressing myofibroblasts. Furthermore, ZEB1 controls the expression of paracrine signals (i.e., HGF and IL6) in tumor cells and myofibroblasts.CoNClUSIoNS: ZEB1 plays a key role in CCA progression by regulating tumor cell-CAF crosstalk, leading to tumor dedifferentiation and CAF activation. (Hepatology 2021;74:3194-3212). C holangiocarcinoma (CCA) includes a heterogeneous group of malignancies that develop from the epithelial layer of the biliary tract. (1,2) CCA is the second most common primary liver tumor after hepatocellular carcinoma,

CHST4 might promote the malignancy of cholangiocarcinoma

“…At the same time, the incidence rate of CCA has increased significantly, especially in western countries. CCA is a nonpainful but lethal tumor, so at an advanced stage it is usually diagnosed if surgery isn’t available [ 21 ]. Surgical resection is the only effective treatment option, which is suitable for only 20% of cases [ 22 ].…”

Cancer-Associated Fibroblasts and Tumor Cells in Pancreatic Cancer Microenvironment and Metastasis: Paracrine Regulators, Reciprocation and Exosomes

“…In cholangiocarcinoma, overexpression of the transcription factor zinc finger E-box-binding homeobox 1 (ZEB1) in tumor cells leads to increased CTGF expression. Culture medium from ZEB1-overexpressing tumor cells induces proliferation of myofibroblasts (Figure 1) [13]. In human cholangiocarcinoma, ZEB1 is expressed in CAFs, correlating with cellular communication network factor 2 (CCN2) gene (encoded for CTGF) expression (Figure 1) [13].…”

“…Culture medium from ZEB1-overexpressing tumor cells induces proliferation of myofibroblasts (Figure 1) [13]. In human cholangiocarcinoma, ZEB1 is expressed in CAFs, correlating with cellular communication network factor 2 (CCN2) gene (encoded for CTGF) expression (Figure 1) [13]. ZEB1 regulates the expression of paracrine signals such as hepatocyte growth factor (HGF) and interleukin 6 (IL-6) in tumor cells and CAFs, highlighting that the ZEB1-CTGF axis plays a key role in regulating the network between tumor cells and CAFs [13].…”

“…In human cholangiocarcinoma, ZEB1 is expressed in CAFs, correlating with cellular communication network factor 2 (CCN2) gene (encoded for CTGF) expression (Figure 1) [13]. ZEB1 regulates the expression of paracrine signals such as hepatocyte growth factor (HGF) and interleukin 6 (IL-6) in tumor cells and CAFs, highlighting that the ZEB1-CTGF axis plays a key role in regulating the network between tumor cells and CAFs [13]. Dysregulation of phosphatidylinositol-4,5-bisphosphatase 3-kinase (PI3K) and Hippo signaling pathways synergistically induce Hippo effector YES-associated protein (YAP), which in turn upregulates CTGF for PSC activation (Figure 1) [12].…”

Cancer Stem Cells in Intrahepatic Cholangiocarcinoma; Their Molecular Basis, and Therapeutic Implications

“…ZEB1 induces pluripotent factors such as SOX2, NANOG, and OCT4 through miR-200 in prostate cancer cells ( Kong et al, 2010 ). In iCCA, ZEB1 is expressed both in tumor and stroma, and plays a key role in the induction of EMT in tumor cells ( Lobe et al, 2021 ). Expression of ZEB1 in tumor cells leads to induction of EMT and stemness, and functions at both sides of the tumor-stroma interface by regulating the production of connective tissue growth factor (CTGF), hepatocyte growth factor (HGF), and interleukin 6 (IL6), to boost tumor progression.…”