P3-01-05: Wnt Signaling: A New Target for Treatment and Prevention of Endocrine Resistant Breast Cancer?
Authors: Micallef RA , Smith C , Barrow D , Nicholson RI , Gee JMW , Barratt-Lee PJ , Hiscox SE 
Published: June 2012

Background: Wnt signaling is altered in nearly half of all breast cancers. Both up regulation of Wnt pathway activators and down regulation of pathway inhibitors have been identified in breast cancer. Previously we have reported that activity of β-catenin is altered in acquired tamoxifen-resistant (TamR) breast cancer cells compared to endocrine-sensitive parental MCF7 cells, with an increase in β-catenin-mediated gene transcription. Data is lacking on the role of Wnt signaling in acquired endocrine resistance in breast cancer. Endocrine relapse predicts a poor clinical outcome and remains an important target for future drug development. In this study we investigated the role of Wnt signaling in acquired tamoxifen resistance and its contribution to the associated aggressive phenotype in vitro. Materials/Methods: We performed exploratory Affymetrix HGU-133A gene microarray analysis of key Wnt pathway components in log-phase TamR vs. MCF7 cell lines, as well as comparing MCF7 cells +/− 10 day tamoxifen exposure. This was paralleled by western blot (WB) analysis. Effects of Wnt signaling inhibition on growth and cell migration were explored using two commercially available inhibitors: IWP2 (inhibits Wnt production, pLRP6) and PNU74654 (interferes with β-catenin-Tcf/Lef binding). MTT cell proliferation assays (6 days) and cell migration assays (24hrs) were carried out.

Results: Microarray analysis of key Wnt pathway components revealed up regulation of wnts 3, 4 and 6; up regulation of Wnt pathway activators LRP5 and LRP6; down regulation of pathway inhibitors (DKK1, DKK4); and down regulation of Axinl (destruction complex component). Changes are &gt;1.5-fold. WB analysis confirmed up regulation of total β-catenin and LRP6 in TamR versus MCF7 cells as well as activity of LRP6. In accordance with this, inhibition of Wnt signaling using IWP2 10uM or PNU74654 10uM, whilst having no effect on MCF7 cell proliferation, significantly suppressed the growth of TamR cells (p&lt;0.001). Inhibition of Wnt signaling in TamR cells also suppressed cell migration (p&lt;0.001). Gene microarray data for MCF7 cells treated with tamoxifen for 10 days showed up regulation of LRP6 and wnt4 and down regulation of LRP5, wnt6, Axin1 and DKK4 compared to control.

Discussion: These observations suggest that deregulated Wnt signaling may play a role in acquired tamoxifen resistance in breast cancer where it may act to promote growth and the development of a more aggressive phenotype. Its interaction with other pathways is important in the prevention of resistance with co-treatment strategies. Wnt1 can rescue MCF7 cells from growth suppression caused by 4-hydroxytamoxifen. Given the early changes in the Wnt pathway in MCF7 cells treated with tamoxifen, further exploration in the context of earlier Wnt targeting alongside antihormone treatment is required. Monitoring of Wnt signaling within clinical samples from patients who have relapsed after hormone treatment is urgently warranted given that the Wnt inhibitors showed promise in controlling this state in vitro.

Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P3-01-05.