Abstract 2030: Mechanisms of E2F-mediated centrosome amplification in HER2 positive breast cancer

Abstract

Abstract Centrosome amplification associates with most human cancers and has been suggested to trigger tumor initiation and progression. Centrosomes are the major microtubule organizing centers in animal cells. They are duplicated once in each cell cycle and duplicated centrosomes establish mitotic spindle bipolarity. Centrosome duplication is tightly regulated because deregulated centrosome duplication leads to centrosome amplification, multipolar and pseudobipolar mitotic spindles that potentiate chromosome instability and may trigger tumorigenesis. One of the tumor types in which the relationship between centrosome amplification and tumorigenesis is well known are breast cancers, as most breast tumors exhibit centrosome amplification. Centrosome amplification arises through various mechanisms, such as deregulation of cell cycle regulators, oncogenes and tumor suppressor genes. Our laboratory discovered that HER2 positive breast cancer cells and K-RasG12D pre-malignant mouse mammary lesions displayed overexpressed E2F1-3 transcriptional activators. Although E2Fs are involved in various cellular processes, including the regulation of the cell and centrosome duplication cycles and differentiation, the role of E2Fs in centrosome amplification in breast tumors is unknown. To study the role of the E2F activators in centrosome amplification, we first assayed centrosome amplification in the E2F-overexpressing HER2 positive breast cancer cells, HCC1954, JIMT1 and SKBR3. There was increased centrosome amplification in the HER2 positive cell lines HCC1954, JIMT1 or SKBR3 relative to MCF10A control mammary epithelial cells. To find if centrosome amplification shown in HER2 positive cells overexpressing E2F1-3 was E2F-specific, the siRNA-mediated silencing technique was applied to transiently knock-down the E2F activators (E2F1-3a) and cells were assayed for centrosome amplification. In addition, to permanently knockdown E2Fs in HER2 positive cells, shRNA-mediated silencing technique will be applied. Western blots confirmed that E2Fs knockdown was efficient, followed by decreased centrosome amplification in HER2 positive cells downregulated for E2F1 and E2F3a; results for E2F2 are still awaiting. These data suggested that centrosome amplification detected in HER2 positive cells is specific to the E2F activators. Future experiments will involve the identification of E2F-dependent transcriptional targets mediating centrosome amplification in HER2 positive cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2030. doi:1538-7445.AM2012-2030