Abstract P5-09-03: Endocrine response in invasive lobular carcinoma is characterized by unique estrogen-mediated gene expression and de novo tamoxifen resistance
Authors: Sikora MJ , Cooper KL , Bahreini A , Luthra S , Chandran UR , Wang G , Dabbs DJ , Welm AL , Oesterreich S 
Published: August 2014

Abstract
Invasive lobular carcinoma (ILC) represents ∼10% of newly diagnosed breast tumors, or ∼30,000 cases annually in the US. However, ILC-specific signaling and endocrine responsiveness are not well characterized. Retrospective analyses suggest that ILC patients treated with endocrine therapy have poorer outcomes than invasive ductal carcinoma (IDC) patients with similar biomarkers, and that ILC patients may not benefit from adjuvant tamoxifen. We hypothesize that estrogen receptor-alpha (ER) regulated gene expression is unique in ILC cells and drives endocrine resistance.
The ER-positive ILC cell lines MDA MB 134VI and SUM44PE were used as in vitro models of cell growth and ER-regulated gene expression in response to estradiol (E2). To examine the ER-regulated transcriptome, we performed gene expression microarray analyses and ER ChIP-Seq following E2 treatment. In parallel, E2 response was assessed in vivo in the primary ILC xenograft HCI-013. Response to endocrine therapies, tamoxifen (Tam), 4-hydroxytamoxifen (4OHT), endoxifen (Bx), and fulvestrant (ICI), were also examined in ILC cell lines.
We observed that E2 induced growth and ER target gene expression in MDA MB 134VI and SUM44PE. We compared our ILC microarray data to published data from ER-positive IDC cell lines (MCF-7, T47D, BT474), and identified 254 genes that were E2-regulated in all 5 cell lines (e.g. GREB1, MYC). 915 genes were E2-regulated only in both ILC cell lines. Consistent with this, roughly half of ER binding sites identified in MDA MB 134VI ChIP-Seq were unique versus published MCF-7 data. We chose a subset of ILC-specific and common E2-regulated genes (n = 107) to assess in vivo using Nanostring gene expression analyses of HCI-013. E2-regulation was observed for 32/107 target genes (30%), suggesting that these genes may be E2-regulated in vivo in ILC patient tumors.
Consistent with clinical data, both ILC cell lines presented de novo tamoxifen resistance. SUM44PE were growth-inhibited by ICI, but unaffected by Tam, 4OHT, or Bx. Similarly, ICI blocked E2-induced growth in MDA MB 134VI, but Tam, 4OHT, and Bx acted as partial agonists, inducing ∼25% growth. Partial agonism was not limited to tamoxifen, as other SERMs (e.g. raloxifene) also induced growth. We then measured ER-regulated gene expression in MDA MB 134VI following tamoxifen treatment. Tam, 4OHT, and Bx acted as agonists for 38/107 genes, whereas ICI acted as an antagonist. All 38 genes were E2-repressed targets (e.g. CCNG2), suggesting that ER-mediated gene repression may be critical to tamoxifen-resistance in ILC. Finally, we observed that FGFR1, frequently amplified in ILC, may be critical for ILC cell survival in the presence of tamoxifen.
These data support the hypothesis that unique ER-mediated gene expression in ILC cells drives endocrine resistance. The de novo tamoxifen resistance observed in ILC cells may correlate with the worse outcomes in ILC patients recently reported. We hypothesize that genes regulated by tamoxifen as an agonist may play a role in tamoxifen-induced growth or serve as biomarkers of resistance. Targeting growth factor signaling using FGFR1 inhibitors may block survival pathways required by ILC cells and reverse tamoxifen resistance.
Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P5-09-03.