Screening for Inhibitors of ZEB1, a Key Regulator of Epithelial-to-Mesenchymal Transition (EMT) in Breast Cancer Cells

Abstract

Breast cancer is a global healthcare burden. This type of malignancy accounts for approximately 670,000 deaths annually. The primary cause of mortality is metastasis. Metastasis is dissemination of cancer cells from primary tumor to other parts of the body. The molecular mechanism that promotes dissemination of cancer cells is mediated by Epithelial-to-Mesenchymal Transition (EMT) enabling cellular motility. Several transcription factors, including Zeb1, regulate the initiation of EMT. This project aimed to screen for small-molecule inhibitors targeting Zeb1 activity. We utilized a Zeb1-negative epithelial MCF-7 breast cancer cell line with Tetracycline-inducible Zeb1 expression. Western Blot analysis confirmed the validity of the model. The protein profile demonstrated that Zeb1 induction inversely correlates with E-cadherin expression. Downregulation of E-cadherin is a fundamental hallmark of the transition into a motile state. We evaluated candidate small-molecule compounds for their impact on Zeb1-driven apoptosis and cell cycle arrest using Flow Cytometry. However, preliminary results indicated limited efficacy of the selected compounds. We attempted to build a rescue model by stably transfecting Zeb1-knockout MDA-MB-231 cells. While control GFP transfection was successful, stable integration of Zeb1 plasmids via lipofection proved inefficient under antibiotic selection. We plan to employ viral transduction to generate more robust, stable lines.