Autocrine WNT signaling contributes to breast cancer cell proliferation via the canonical WNT pathway and EGFR transactivation

Abstract

BackgroundDe-regulation of the wingless and integration site growth factor (WNT) signaling pathway via mutations in APC and Axin, proteins that target β-catenin for destruction, have been linked to various types of human cancer. These genetic alterations rarely, if ever, are observed in breast tumors. However, various lines of evidence suggest that WNT signaling may also be de-regulated in breast cancer. Most breast tumors show hypermethylation of the promoter region of secreted Frizzled-related protein 1 (sFRP1), a negative WNT pathway regulator, leading to downregulation of its expression. As a consequence, WNT signaling is enhanced and may contribute to proliferation of human breast tumor cells. We previously demonstrated that, in addition to the canonical WNT/β-catenin pathway, WNT signaling activates the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway in mouse mammary epithelial cells via epidermal growth factor receptor (EGFR) transactivation.MethodsUsing the WNT modulator sFRP1 and short interfering RNA-mediated Dishevelled (DVL) knockdown, we interfered with autocrine WNT signaling at the ligand-receptor level. The impact on proliferation was measured by cell counting, YOPRO, and the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) assay; β-catenin, EGFR, ERK1/2 activation, and PARP (poly [ADP-ribose]polymerase) cleavages were assessed by Western blotting after treatment of human breast cancer cell lines with conditioned media, purified proteins, small-molecule inhibitors, or blocking antibodies.ResultsPhospho-DVL and stabilized β-catenin are present in many breast tumor cell lines, indicating autocrine WNT signaling activity. Interfering with this loop decreases active β-catenin levels, lowers ERK1/2 activity, blocks proliferation, and induces apoptosis in MDA-MB-231, BT474, SkBr3, JIMT-1, and MCF-7 cells. The effects of WNT signaling are mediated partly by EGFR transactivation in human breast cancer cells in a metalloprotease- and Src-dependent manner. Furthermore, Wnt1 rescues estrogen receptor-positive (ER+) breast cancer cells from the anti-proliferative effects of 4-hydroxytamoxifen (4-HT) and this activity can be blocked by an EGFR tyrosine kinase inhibitor.ConclusionOur data show that interference with autocrine WNT signaling in human breast cancer reduces proliferation and survival of human breast cancer cells and rescues ER+ tumor cells from 4-HT by activation of the canonical WNT pathway and EGFR transactivation. These findings suggest that interference with WNT signaling at the ligand-receptor level in combination with other targeted therapies may improve the efficiency of breast cancer treatments.

Predominant role of PDGF receptor transactivation in Wnt3a-induced osteoblastic cell proliferation

“…With this report and previous studies describing transactivation of receptor tyrosine kinases by Wnt protein, (50)(51)(52) it seems that the network of signaling pathways activated by Wnt proteins is becoming more complex, especially when a Wnt protein activates several tyrosine kinase receptors in a particular cell type.…”

“…Clearly, PDGF-BB was more potent than PDGF-AA, confirming that signaling by PDGFRb plays a predominant role in controlling osteoblastic cell growth. (50,51) Downregulation of PDGF-Rb using an siRNA approach indicated that this receptor is a major contributor in Wnt3a-induced MC3T3-E1 cell proliferation. Whether activation of PDGF-Rs by Wnt3a in osteoblastic cells involves an autocrine production of PDGFs was assessed using either recombinant PDGF-Rb or PDGF-Ra Fc chimera that can sequester extracellular PDGFs.…”

“…To our knowledge, this is the first report on transactivation of PDGF-Rs by a Wnt protein, whereas transactivation of EGF-Rs has already been reported in mammary epithelial cells, (52) NIH3T3 cells, (50) and breast cancer cells. (51) The molecular mechanism by which Wnt proteins transactivate some receptor tyrosine kinases remains poorly understood. A study has shown that Wnt3a can stimulate Src docking to dishevelled-2 (Dvl2) and activates its tyrosine kinase activity.…”

Prodigiosin inhibits Wnt/β-catenin signaling and exerts anticancer activity in breast cancer cells

“…Although Wnt/β-catenin pathway mutations are rarely detected in breast tumors, elevated levels of nuclear and/or cytoplasmic β-catenin are frequently found and are associated with poor prognosis (40). Furthermore, DVL1 is up-regulated in 50% of human breast cancers, and phosphorylated DVL proteins have been detected in many breast tumor cell lines (41,42).…”

Downregulation of Dickkopf-1 is responsible for high proliferation of breast cancer cells via losing control of Wnt/β-catenin signaling

“…Next, we try to identify the downstream effectors of DKK-1 in promotion of proliferation of breast cancer cells. A hallmark of canonical Wnt pathway activation which can enhance proliferation ability of several tumor cell lines shows an increase in β-catenin-dependent transcription, primarily as a result of β-catenin protein stabilization [9,10,31,32] . Thus, we assessed the effects of DKK-1 on the phosphorylation of β-catenin which led to β-catenin degradation.…”