Molecular understanding of tamoxifen resistance in breast cancer
Leeuw, R. de
Citation
Leeuw, R. de. (2012, February 9). Molecular understanding of tamoxifen resistance in breast cancer. Retrieved from
https://hdl.handle.net/1887/18458
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Molecular understanding of
tamoxifen resistance in breast cancer
Renée de Leeuw
Omslag ontworpen door: Theophile Suijkerbuijk (www.theophile.nl)
Molecular understanding of
tamoxifen resistance in breast cancer
proefschrift
ter verkrijging van de graad van Doctor aan de Universiteit Leiden,
op gezag van Rector Magnificus prof. mr. P.F. van der Heijden, volgens besluit van het College voor Promoties
te verdedigen op donderdag 9 februari 2012 klokke 11:15 uur
door
Renée de Leeuw
geboren te Breda in 1983
Promotiecommissie
Promotor Prof. Dr. JJ Neefjes Copromotor Dr. RJAM Michalides
Het Nederlands Kanker Instituut Overige leden Dr. WT Zwart
Het Nederlands Kanker Instituut
Prof. Dr. P ten Dijke Prof. Dr. B van de Water Prof. Dr. C de Vries
Universiteit van Amsterdam Prof. Dr. GA Meijer
Vrije Universiteit (VUMC) Dr. J Carroll
Cancer Research UK
(c) 2012 Renée de Leeuw Drukkerij: Ipskamp Drukkers
The work described in this thesis was performed in the division of Tumor and Cell Biology of the Netherlands Cancer Institute (NKI-AVL), Amsterdam, The Netherlands. This work was supported by a grant from Top Institute Pharma (TI Pharma). Financial support for the publication of this thesis was provided by the Netherlands Cancer Institute (NKI-AVL).
voor oma
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Table of contents
Scope of this thesis p9
Chapter 1. Introduction - review p11
A Role for Estrogen Receptor Phosphorylation in the Resistance to Tamoxifen
International Journal of Breast Cancer 2011, Article ID 232435
Chapter 2. Research article p27
The hinge region of the human estrogen receptor determines functional synergy between AF-1 and AF-2 in the quantitative response to estradiol and tamoxifen
Journal of Cell Science 2010, Vol.123, pp1253-1261
Chapter 3. Research article p49
A high-throughput coregulator peptide array detects estrogen receptor alpha Serine-305 phosphorylation that is associated with response to tamoxifen treatment
Molecular Cancer Therapeutics (in revision)
Chapter 4. Research article p67
PKA Phosphorylation redirects oestrogen receptor to promoters of a unique gene set to induce tamoxifen resistance
Submitted
Chapter 5. Summary & discussion p93
Nederlandse samenvatting & discussie p103
List of publications p113
Curriculum vitae p115
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Scope of this thesis
Estrogen receptor signalling,…
The estrogen receptor is a nuclear hormone receptor that is activated by estrogens.
The activated receptor dimerizes and binds to responsive elements on the DNA. Cofactors are recruited to regulate the transcription of genes. Thereby, estrogens can induce cell proliferation.
…breast cancer…
The clinic makes clever use of this characteristic in breast cancer treatment. In about 70% of all breast cancer cases, the tumor expresses the estrogen receptor. Patients diagnosed with this type of breast cancer commonly receive endocrine treatment after surgical removal of the tumor. Endocrine treatment can either target the receptor with competitive drugs (antiestrogens), or prevent formation of natural estrogens by inhibiting the aromatase enzyme (aromatase inhibitors). Because such tumors are dependent on estrogens for their growth, both treatment modalities should result in growth arrest and prevention of disease recurrence.
…and endocrine resistance
Endocrine treatment in breast cancer is often successful, yet a subset of patients develop therapeutic resistance. This thesis focuses on one type of endocrine resistance: the widely used antiestrogen tamoxifen. A number of factors have been attributed to tamoxifen resistance, among which phosphorylation of the estrogen receptor.
A general introduction into the role of phosphorylation of ERα in tamoxifen resistance is given in Chapter 1. In this review, biological and clinical data are discussed concerning known and putative phosphorylation sites of ERα. Phosphorylation is the addition of a phosphate group onto a protein, which is a bulky and negatively charged molecule. Phosphorylation can affect the structural conformation of the receptor. This can in turn influence ER function, e.g. by interfering with dimerization or by differential recruitment of coregulators. Kinases and molecular pathways involved in targeting these sites are briefly discussed. Some phosphorylation events are beneficial to tamoxifen response, whereas others may induce resistance.
Chapter 2 addresses a more fundamental question of how different structural domains of the estrogen receptor define its function. This was done by a comparison between the two receptor subtypes: ERα and ERβ. Both proteins contain two activation function domains, AF-1 and AF-2, separated by a hinge region. Although both are members of one family and are partially conserved, their differences result in different function. By swapping domains between ERα and ERβ, we discovered that the synergy between the AF-1 and AF-2 domains is dependent on the hinge region. Together, the domains determine the quantitative response to estradiol or tamoxifen.
The estrogen receptor requires coregulators to perform its transactivation function on DNA. By differential coregulator recruitment it determines which genes are transcribed and which are blocked. In Chapter 3 we applied a high-throughput chip containing coregulator motif peptides, to investigate coregulator binding by full-length ERα. This technique enabled us to study both overexpressed and endogenous proteins in lysates from different cell lines.
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Moreover, peptide binding was measured in crude lysates from patient tumor tissue.
Phosphorylation of the estrogen receptor can affect its function by altering coregulator recruitment. Therefore, we investigated if protein kinase A (PKA) induced phosphorylation of the ERα Serine 305 residue, which has been associated with tamoxifen resistance, influences coregulator binding. Phosphorylation resulted in enhanced coregulator interactions.
How does phosphorylation of ERα result in a conformational change and differential recruitment of coregulators ultimately lead to cell proliferation, tumor growth and tamoxifen resistance? Chapter 4 addresses these questions by genome wide analyses of the chromatin binding landscape and gene expression profile of cells expressing Serine 305 phosphorylated ERα. We discovered that one post-translational modification, i.e. ERα Ser305 phosphorylation, can dramatically influence the chromatin binding preferences of the receptor, here showing a strong enrichment at promotor start sites, 3’ and 5’ UTRs. We defined a 100-gene classifier, and determined distinct targets for the phosphorylated receptor, including MYC, a known oncogene. A differential regulation of the MYC pathway could very well explain resistance to tamoxifen. This suggestion is supported by the significant correlation of the classifier with poor disease outcome in two independent datasets of ERα positive, tamoxifen treated patients.
Finally, Chapter 5 puts the work presented in this thesis in perspective and provides an outlook for future studies.
