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Abstract Disclosure: C.A. Ufondu: None. M. Musick: None. M.J. Sikora: None. J.H. Ostrander: None. Estrogen receptor (ER) expression and E-cadherin (CDH1) loss are the two major molecular hallmarks of invasive lobular carcinoma (ILC) of the breast; however, it is unclear how ER signaling and E-cadherin loss crosstalk during ILC tumor formation. Furthermore, the risk of ILC is increased by estrogen-only hormone replacement treatment, but not that of the more frequent invasive ductal carcinoma (IDC), suggesting that ER may have a unique role in the initiation and progression of ILC. We hypothesize that E-cadherin loss accelerates estrogen-driven tumorigenesis by reprogramming ER DNA binding through the dysregulation of pathways and transcription factors linked to E-cadherin. To test the effect of E-cadherin loss during estrogen driven tumorigenesis, we established a CRISPR/Cas9 approach for knocking out CDH1 in primary human mammary epithelial cells (HMECs). Since primary HMECs did not show robust ER expression, we also re-expressed ER using lentiviral transduction of ESR1 into HMECs. We performed RNAseq from two independent CDH1 targeting gRNAs vs non-targeting gRNA (NTG) control (-/+ ER). Importantly, when comparing the two independent CDH1 gRNAs to NTG there were 2882 shared genes (1436 upregulated only, 1407 downregulated only) that were differentially regulated. Analysis of these shared genes showed the enrichment of estrogen response, PI3K signaling and cell cycle pathways in the CDH1 knockout cells vs control. We also observed the repression of epithelial mesenchymal transition, hypoxia-related genes and p53 in the CDH1 knockout samples vs control. Overall, analysis of the RNAseq data suggests that the loss of E-cadherin in HMECS impacts proliferation and EMT, which will be explored in future studies. In addition, we hope to identify mediators that drive estrogen-induced tumorigenesis using the mouse mammary intraductal (MIND) model, which better recapitulates the progression of ILC and response to estrogen in vivo. We injected control and CDH1 knockout cells into the mammary glands of adult female NSG mice, with estrogen supplemented in their drinking water, and assessed for engraftment using bioluminescence imaging (BLI). BLI suggests enhanced engraftment of CDH1 knockout cells compared to CDH1 wild-type HMECs and a survival advantage for HMECs expressing ESR1. Importantly, analysis of mammary glands two-month post-MIND injections showed engraftment of human cells positive for cytokeratin-5 and lamin a/c. Ongoing studies aim to assess the estrogen response in the CDH1-knockout versus control HMECs in vitro and in vivo to determine whether loss of E-cadherin contributes to ILC-specific ER activity. Understanding the unique interactions between E-cadherin loss and ER in ILC could lead to novel therapeutic approaches for the prevention and treatment of ILC. Presentation: 6/2/2024
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