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Epoxyeicosanoids (Eets) Promote the Cancer Stem Cell State by Preventing Breast Cancer Cells Differentiation

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Epoxyeicosanoids (Eets) Promote the Cancer Stem Cell State by Preventing Breast Cancer Cells Differentiation University of Calgary PRISM: University of Calgary's Digital Repository Graduate Studies The Vault: Electronic Theses and Dissertations 2014-05-21 Epoxyeicosanoids (EETs) Promote The Cancer Stem Cell State by Preventing Breast Cancer Cells Differentiation El Kadiri, Zineb El Kadiri, Z. (2014). Epoxyeicosanoids (EETs) Promote The Cancer Stem Cell State by Preventing Breast Cancer Cells Differentiation (Unpublished master's thesis). University of Calgary, Calgary, AB. doi:10.11575/PRISM/25928 http://hdl.handle.net/11023/1536 master thesis University of Calgary graduate students retain copyright ownership and moral rights for their thesis. You may use this material in any way that is permitted by the Copyright Act or through licensing that has been assigned to the document. For uses that are not allowable under copyright legislation or licensing, you are required to seek permission. Downloaded from PRISM: https://prism.ucalgary.ca UNIVERSITY OF CALGARY Epoxyeicosanoids (EETs) Promote The Cancer Stem Cell State by Preventing Breast Cancer Cells Differentiation by Zineb El Kadiri A THESIS SUBMITTED TO THE FACULTY OF GRADUATE STUDIES IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE DEPARTMENT OF BIOLOGICAL SCIENCE CALGARY, ALBERTA MAY, 2014 © Zineb El Kadiri 2014 Abstract Our goal is to examine the role of epoxyeicosatrienoic acids (EETs) – omega-6 poly-unsaturated fatty acid metabolites - in modulating breast cancer cell plasticity manifest in the discrete attractor state transitions between cancerous and differentiated states. We used an in vitro breast cancer cell differentiation system, MDA-MB231 that exhibited two quasi-discrete subpopulations representing a stem-like and a differentiated state. We found that elevated EETs suppressed the vitaminD-induced shift of the MDA-MB231 cells towards the differentiated state. Moreover, subpopulation gene expression profiles of cells exposed to EETs robustly identified stem cell-like and differentiated subpopulations as two discrete clusters, regardless of EETs. Interestingly, EETs promoted “stemness” and metastasis genes, while suppressed differentiation related genes, suggesting that EETs not only blocked the switch between the two attractors but also affected cell-intrinsic attractor properties. Our findings suggest that anti-EETs agents may have therapeutic or preventive utility against breast cancer that could be further explored. ii Acknowledgements I wish to thank my thesis supervisor, Dr. Sui Huang for the opportunity to participate in this research project. I am grateful for his mentoring and patience as we explored new ideas together and defined the direction of my research. I would like to thank all those who have contributed to my work and supported me during my study including, Wei Wu, Mitra Mojtahedi, Ivan Gomez and Rebecca Leong. A special thank to Dr. Joseph Zhou for his help in analyzing the microarray data. The work in this thesis would not have been possible without the generous help and support I have received from my husband, my little ones Ilyas and Sulayman, my mom and sisters. Thank you all for your unwavering love and encouragement. iii Table of Contents Abstract............................................................................................................................... ii Acknowledgements............................................................................................................ iii Table of Contents............................................................................................................... iv List of Tables .................................................................................................................... vii List of Figures.................................................................................................................. viii List of Symbols, Abbreviations and Nomenclature........................................................... xi CHAPTER ONE: INTRODUCTION..................................................................................1 1.1 Systems view of the gene regulatory network: cancer cells as cancer attractors ......1 1.1.1 System biology approach ..................................................................................1 1.1.2 Theory of gene regulatory network and attractors ............................................2 1.1.3 Abnormal cell types as cancer attractors...........................................................4 1.1.4 The concept of cancer stem cell: refining of cancer cell hypotheses vs. clonal evolution ............................................................................................................6 1.2 Comparing the normal stem cells (NSC) with the cancer stem cells (CSC): the hallmark properties, the cell of origin and the surrounding microenvironment ......7 1.2.1 Normal stem cells: self-renewal and differentiation potential ..........................8 1.2.2 The cancer stem cell of origin ...........................................................................9 1.3 Breast cancer is a stem cell disease that could be influenced by environmental factors ..................................................................................................................................9 1.3.1 Environmental factors account to a certain degree for breast cancer disparities9 1.3.2 Mammary stem cells in normal development .................................................11 1.3.3 Mammary cancer stem cells in solid tumors ...................................................12 1.3.4 Breast cancer is a disease with functional inter-tumor and intra-tumor heterogeneity....................................................................................................13 1.3.4.1 Breast cancer cell population heterogeneity is reflected by their stem cell marker expression: ..................................................................................15 1.3.4.2 Functional heterogeneity in CD44 high human breast cancer stem cell like compartment: ..........................................................................................16 1.3.4.3 CD44+/CD24- pattern correlates with the basal breast cancer subtype rather than tumorigenisis:..................................................................................17 1.4 Molecular biology of breast cancer .........................................................................19 1.4.1 Cell signalling pathways regulating self-renewal and differentiation of Breast normal and cancer stem cells ...........................................................................19 1.4.1.1 The canonical Wnt signalling pathway:.................................................20 1.4.1.2 The Notch signalling pathway:..............................................................21 1.4.1.3 The Hedgehog signalling pathway: .......................................................22 1.4.1.4 The PTEN PI3K/Akt signalling pathway: .............................................22 1.4.2 Cancer stem cell resist therapeutic challenges and drive the relapse of the initial heterogeneous tumor........................................................................................23 1.4.3 Basal subclass of breast cancer is associated with high recurrence rate and metastatic spread..............................................................................................24 iv 1.5 Induced differentiation of human breast cancer cells reflects the dynamics of non- genetic heterogeneity .............................................................................................26 1.5.1 Disrupting the immature state: differentiation of breast cancer cells..............26 1.5.2 Vitamin D metabolism and mechanism of action ...........................................28 1.5.3 The role of vitamin D in repressing the progression and promoting differentiation of basal subtype of breast cancer cells.............................................................29 1.6 The cytochrome P450-derived eicosanoids (EETs) pathway in conjunction with cancer ................................................................................................................................30 1.6.1 Epoxyeicosatrienoic acids: formation, metabolism, and mechanism..............31 1.6.1.1 Production of EETs:...............................................................................32 1.6.1.2 Mechanism of action of EETs: ..............................................................34 1.6.1.3 Metabolism of EETs:.............................................................................35 1.6.2 EETs signalling in cancer................................................................................35 1.6.2.1 High levels of EET induce the activation of pathways involved in proliferation, angiogenesis and anti-apoptosis:.......................................35 1.6.2.2 Synthetic EETs trigger tumor initiation and metastasis in animal models: .................................................................................................................38 CHAPTER TWO: GOALS AND APPROACHES OF THE PROJECT ..........................40 2.1 Overall goal and biological rationale.......................................................................40 2.2 Experimental system................................................................................................44 2.2.1 MDA-MB-231 cell line as a surrogate for primary breast tumor cells ...........44 2.2.2 Vitamin D-induced differentiation highlights the micro-heterogeneity
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