title: WNT4 mediates estrogen receptor signaling and endocrine resistance in invasive lobular carcinoma cell lines.
abstract: Invasive lobular carcinoma (ILC) of the breast typically presents with clinical biomarkers consistent with a favorable response to endocrine therapies, and over 90 % of ILC cases express the estrogen receptor (ER). However, a subset of ILC cases may be resistant to endocrine therapies, suggesting that ER biology is unique in ILC. Using ILC cell lines, we previously demonstrated that ER regulates a distinct gene expression program in ILC cells, and we hypothesized that these ER-driven pathways modulate the endocrine response in ILC. One potential novel pathway is via the Wnt ligand WNT4, a critical signaling molecule in mammary gland development regulated by the progesterone receptor.
  
  
  
  The ILC cell lines MDA-MB-134-VI, SUM44PE, and BCK4 were used to assess WNT4 gene expression and regulation, as well as the role of WNT4 in estrogen-regulated proliferation. To assess these mechanisms in the context of endocrine resistance, we developed novel ILC endocrine-resistant long-term estrogen-deprived (ILC-LTED) models. ILC and ILC-LTED cell lines were used to identify upstream regulators and downstream signaling effectors of WNT4 signaling.
  
  
  
  ILC cells co-opted WNT4 signaling by placing it under direct ER control. We observed that ER regulation of WNT4 correlated with use of an ER binding site at the WNT4 locus, specifically in ILC cells. Further, WNT4 was required for endocrine response in ILC cells, as WNT4 knockdown blocked estrogen-induced proliferation. ILC-LTED cells remained dependent on WNT4 for proliferation, by either maintaining ER function and WNT4 regulation or uncoupling WNT4 from ER and upregulating WNT4 expression. In the latter case, WNT4 expression was driven by activated nuclear factor kappa-B signaling in ILC-LTED cells. In ILC and ILC-LTED cells, WNT4 led to suppression of CDKN1A/p21, which is critical for ILC cell proliferation. CDKN1A knockdown partially reversed the effects of WNT4 knockdown.
  
  
  
  WNT4 drives a novel signaling pathway in ILC cells, with a critical role in estrogen-induced growth that may also mediate endocrine resistance. WNT4 signaling may represent a novel target to modulate endocrine response specifically for patients with ILC.
authors.affiliation: Women's Cancer Research Center, University of Pittsburgh, Pittsburgh, PA, USA. matthew.sikora@ucdenver.edu.|Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA. matthew.sikora@ucdenver.edu.|Present address: Department of Pathology, University of Colorado - Anschutz Medical Campus, Mail Stop 8104, Research Complex 1 South, Room 5117, 12801 East 17th Avenue, Aurora, CO, 80045, USA. matthew.sikora@ucdenver.edu.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Sikora
authors.affiliation: Department of Pathology, University of Colorado - Anschutz Medical Campus, Aurora, CO, USA.
authors.forename: Britta M
authors.initials: BM
authors.lastname: Jacobsen
authors.affiliation: Women's Cancer Research Center, University of Pittsburgh, Pittsburgh, PA, USA.|Department of Pathology, University of Pittsburgh, Pittsburgh, PA, USA.
authors.forename: Kevin
authors.initials: K
authors.lastname: Levine
authors.affiliation: Women's Cancer Research Center, University of Pittsburgh, Pittsburgh, PA, USA.
authors.forename: Jian
authors.initials: J
authors.lastname: Chen
authors.affiliation: Women's Cancer Research Center, University of Pittsburgh, Pittsburgh, PA, USA.|Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA.
authors.forename: Nancy E
authors.initials: NE
authors.lastname: Davidson
authors.affiliation: Women's Cancer Research Center, University of Pittsburgh, Pittsburgh, PA, USA.|Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA.
authors.forename: Adrian V
authors.initials: AV
authors.lastname: Lee
authors.affiliation: McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, WI, USA.
authors.forename: Caroline M
authors.initials: CM
authors.lastname: Alexander
authors.affiliation: Women's Cancer Research Center, University of Pittsburgh, Pittsburgh, PA, USA.|Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA.
authors.forename: Steffi
authors.initials: S
authors.lastname: Oesterreich
mesh_terms: D018931:Antineoplastic Agents, Hormonal / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D018275:Carcinoma, Lobular / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism; D009361:Neoplasm Invasiveness; D009367:Neoplasm Staging; D011485:Protein Binding; D011960:Receptors, Estrogen / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D060528:Wnt4 Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: D018931:Antineoplastic Agents, Hormonal; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016328:NF-kappa B; D011960:Receptors, Estrogen; D060528:Wnt4 Protein
keywords: Breast cancer; Endocrine resistance; Endocrine therapy; Estradiol; Estrogen receptor; Lobular carcinoma; Wnt signaling