Abstract P3-07-04: WNT4 mediates endocrine response and resistance in invasive lobular carcinoma cells
Authors: Sikora MJ , Oesterreich S 
Published: March 2017

Abstract
Invasive lobular carcinoma (ILC) is a breast cancer subtype affecting ~30,000 U.S. women annually. Over 90% of ILC are estrogen receptor (ER)-positive; however, endocrine therapy may have poorer efficacy in a subset of ILC patients versus invasive ductal carcinoma (IDC) patients. This prompted us to assess global ER activity in ILC cell lines MDA MB 134VI (MM134) and SUM44PE (44PE) to identify novel mediators of ER signaling. These analyses identified the Wnt ligand WNT4 as an ILC-specific ER target gene, with an ILC-specific ER binding site (ERBS) at the WNT4 locus. Considering the critical role of WNT4 in normal mammary gland expansion, we hypothesize that ILC cells utilize WNT4 signaling to drive endocrine response and resistance.
We assessed whether WNT4 is necessary for ILC cell growth using siRNA. WNT4 knockdown completely blocked estrogen-induced growth in ILC cells but not IDC cells. In parallel, the WNT4 ERBS was only occupied in ILC cells in response to estrogen, but progesterone-induced WNT4 in IDC was not associated with this ERBS. This suggests that, via the ILC-specific WNT4 ERBS, ILC cells drive estrogen-regulated proliferation by hijacking a developmental Wnt pathway. Wnt pathways typically activate β-catenin; however, we observed β-catenin dysfunction in ILC cells and that WNT4 cannot activate β-catenin. Thus, WNT4 signals in ILC cells via a novel non-canonical pathway.
Using long-term estrogen-deprived (LTED) variants of MM134 and 44PE (4 and 2 lines, respectively), we assessed WNT4 in ILC endocrine resistance. WNT4 is over-expressed, but uncoupled from ER, in all MM134-LTED. Conversely, WNT4 is reduced in 44PE-LTED but remains ER-regulated; ER occupies the WNT4 ERBS only in 44PE-LTED cells and not MM134-LTED. Using siRNA, MM134-LTED (high WNT4) are growth-inhibited by WNT4 knockdown, while 44PE-LTED (low WNT4) are insensitive. However, WNT4 knockdown sensitizes 44PE-LTED to endocrine therapy. Taken together, uncoupling and upregulating WNT4 or WNT4/ER cross-talk may represent convergent endocrine resistance mechanisms in ILC. Further characterization of ILC-LTED cells demonstrated WNT4 expression is driven by activated NFκB signaling in MM134-LTED, and implicated the pluripotency factor Oct4 in regulating WNT4 in 44PE-LTED cells. In both parental ILC cells and ILC-LTED cells, WNT4 leads to suppression of CDKN1A/p21, which is critical for ILC cell proliferation; CDKN1A knockdown partially reverses the effects of WNT4 knockdown.
Clinical observations suggest that ER regulates unique pathways in ILC. We identified WNT4 as a downstream effector of endocrine signaling in ILC, with critical roles in both estrogen-induced growth and endocrine resistance. WNT4 signaling may represent a novel target to modulate endocrine response specifically for ILC patients.
Citation Format: Sikora MJ, Oesterreich S. WNT4 mediates endocrine response and resistance in invasive lobular carcinoma cells [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P3-07-04.