Targeted silencing of the oncogenic transcription factor SOX2 in breast cancer

Epigenetic Editing, Department of Pathology and Medical Biology, University Medical Center Groningen, University of Groningen, Hanzeplein 1, 9713 GZ Groningen, The Netherlands.
Nucleic Acids Research (Impact Factor: 9.11). 05/2012; 40(14):6725-40. DOI: 10.1093/nar/gks360
Source: PubMed

ABSTRACT The transcription factor (TF) SOX2 is essential for the maintenance of pluripotency and self-renewal in embryonic stem cells. In addition to its normal stem cell function, SOX2 over-expression is associated with cancer development. The ability to selectively target this and other oncogenic TFs in cells, however, remains a significant challenge due to the 'undruggable' characteristics of these molecules. Here, we employ a zinc finger (ZF)-based artificial TF (ATF) approach to selectively suppress SOX2 gene expression in cancer cells. We engineered four different proteins each composed of 6ZF arrays designed to bind 18 bp sites in the SOX2 promoter and enhancer region, which controls SOX2 methylation. The 6ZF domains were linked to the Kruppel Associated Box (SKD) repressor domain. Three engineered proteins were able to bind their endogenous target sites and effectively suppress SOX2 expression (up to 95% repression efficiencies) in breast cancer cells. Targeted down-regulation of SOX2 expression resulted in decreased tumor cell proliferation and colony formation in these cells. Furthermore, induced expression of an ATF in a mouse model inhibited breast cancer cell growth. Collectively, these findings demonstrate the effectiveness and therapeutic potential of engineered ATFs to mediate potent and long-lasting down-regulation of oncogenic TF expression in cancer cells.

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    • "Effective regulation has been achieved by targeting both core promoters and enhancer sequences [67]. "
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    ABSTRACT: Epigenetic modifications such as histone post-transcriptional modifications and DNA methylation organize the DNA in the nucleus of eukaryotic cells and are critical for the spatio-temporal regulation of gene expression. These epigenetic modifications are reversible and precisely regulated by epigenetic enzymes. In addition to genetic mutations, epigenetic modifications are highly disrupted in cancer relative to normal tissues. Many epigenetic alterations (epi-mutations) are associated with aberrations in the expression and/or activity of epigenetic enzymes. Thus, epigenetic regulators have emerged as prime targets for cancer therapy. Currently, several inhibitors of epigenetic enzymes (epi-drugs) have been approved for use in the clinic to treat cancer patients with hematological malignancies. However, one potential disadvantage of epi-drugs is their lack of locus-selective specificity, which may result in the over-expression of undesirable parts of the genome. The emerging and rapidly growing field of epigenome engineering has opened new grounds for improving epigenetic therapy in view of reducing the genome-wide “off-target” effects of the treatment. In the current review, we will first describe the language of epigenetic modifications and their involvement in cancer. Next, we will overview the current strategies for engineering of artificial DNA binding domains in order to manipulate and ultimately normalize the aberrant landscape of the cancer epigenome (epigenome engineering). Lastly, the potential clinical applications of these emerging genome-engineering approaches will be discussed.
    Frontiers in Oncology 01/2015; 5(22). DOI:10.3389/fonc.2015.00022
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    • "As described by Hanahan and Weinberg, cancer is a disease characterized by determined hallmarks some of which are: sustained proliferative signaling, activation of invasion and metastasis, and evasion of cell death [73]. Studies have strongly associated SOX2 to these respective cancer hallmarks and thus far SOX2 has been shown to promote cellular proliferation (breast, prostate, pancreatic and cervical cancers) [21,28,57,58], evade apoptotic signals (prostate, gastric cancer and NSCLC) [37,58,63] and promote invasion, migration and metastasis (melanoma, colorectal, glioma, gastric, ovarian cancer and hepatocellular carcinoma) [15,29,47,49,55]. We summarized SOX2 amplification and resulting alterations in cellular functions in all cancer types in Table 1 and showed examples of SOX2’s role in oncogenic signaling in Figure 2. "
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    ABSTRACT: SOX2 is a gene that encodes for a transcription factor belonging to the SOX gene family and contains a high-mobility group (HMG) domain, which permits highly specific DNA binding. Consequently, SOX2 functions as an activator or suppressor of gene transcription. SOX2 has been described as an essential embryonic stem cell gene and moreover, a necessary factor for induced cellular reprogramming. SOX2 research has only recently switched focus from embryogenesis and development to SOX2's function in disease. Particularly, the role of SOX2 in cancer pathogenesis has become of interest in the field. To date, studies have shown SOX2 to be amplified in various cancer types and affect cancer cell physiology via involvement in complicated cell signaling and protein-protein interactions. Recent reviews in this field have highlighted SOX2 in mammalian physiology, development and pathology. In this review, we comprehensively compile what is known to date about SOX2's involvement in cancer biology, focusing on the most recent findings in the fields of cellular signaling and cancer stem cells. Lastly, we underscore the role of SOX2 in the clinic and highlight new findings, which may provide novel clinical applications for SOX2 as a prognostic marker, indicator of metastasis, biomarker or potential therapeutic target in some cancer types.
    Clinical and Translational Medicine 07/2014; 3(1):19. DOI:10.1186/2001-1326-3-19
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    • "The OCT4 proximal and distal enhancers have been previously defined as key regions important in regulating OCT4 expression in different cell types (24,25,40). ZF-TFs and TALE-TFs that target the enhancer regions of SOX2 and OCT4, respectively, have been shown to activate gene expression (20,22). However, none of the previous studies comprehensively probed the intervening sequences for effective target sites. "
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    ABSTRACT: Artificial transcription factors are powerful tools for regulating gene expression. Here we report results with engineered zinc-finger transcription factors (ZF-TFs) targeting four protein-coding genes, OCT4, SOX2, KLF4 and c-MYC, and one noncoding ribonucleic acid (RNA) gene, the microRNA (miRNA) miR302/367 cluster. We designed over 300 ZF-TFs whose targets lie within 1 kb of the transcriptional start sites (TSSs), screened them for increased messenger RNA or miRNA levels in transfected cells, and identified potent ZF-TF activators for each gene. Furthermore, we demonstrate that selected ZF-TFs function with alternative activation domains and in multiple cell lines. For OCT4, we expanded the target range to −2.5 kb and +500 bp relative to the TSS and identified additional active ZF-TFs, including three highly active ZF-TFs targeting distal enhancer, proximal enhancer and downstream from the proximal promoter. Chromatin immunoprecipitation (FLAG-ChIP) results indicate that several inactive ZF-TFs targeting within the same regulatory region bind as well as the most active ZF-TFs, suggesting that efficient binding within one of these regulatory regions may be necessary but not sufficient for activation. These results further our understanding of ZF-TF design principles and corroborate the use of ZF-TFs targeting enhancers and downstream from the TSS for transcriptional activation.
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