Influence of the Nuclear Hormone Receptor Axis in the Progression and Treatment of Hormone Dependent Cancers
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Influence of the nuclear hormone receptor axis in the progression and treatment of hormone dependent cancers A dissertation submitted to the Division of Research and Advanced Studies at the University of Cincinnati in partial fulfillment of the requirements for the degree of Doctorate of Philosophy (Ph.D.) In the Department of Cell and Cancer Biology of the College of Medicine 2007 by Janet K. Hess-Wilson B.A., Wittenberg University, 2000 Committee Chair: Karen E. Knudsen, Ph.D. Abstract Due to its pivotal role in prostatic growth and survival, the androgen receptor (AR) is the primary target of disseminated prostate cancer (CaP), as achieved via androgen deprivation therapy (ADT). Unfortunately, ADT is circumvented by restoration of AR activity, resulting in ADT resistant tumors for which there is no alternate treatment option. Through multiple mechanisms, reactivation of the AR specifically underlies the progression to therapy resistant tumors. The environmentally prevalent endocrine disrupting compound (EDC), bisphenol A (BPA) is able to activate specific somatically mutated ARs commonly found in CaP, resulting in androgen-independent proliferation of CaP cells. To directly assess the effect of BPA on ADT, we used an in vivo xenograft model of CaP that expresses a BPA-sensitive mutant AR, and mimics standard human cytotoxic response to ADT, followed by subsequent tumor re-growth. When tumor- bearing animals were exposed to environmentally relevant levels of BPA during ADT, the tumors failed therapy more rapidly (compared to placebo controls), with AR re- activation and concomitant increased tumor cell proliferation. These data suggest that environmentally relevant exposure to EDCs may reduce the efficacy of mainline ADT for CaP. We next determined that the environmentally persistent pesticide, DDE, was able to activate select AR mutants, and in in vitro models of CaP, DDE induces cellular proliferation in the absence of androgen, demonstrating that this observed response was not unique to BPA. Strikingly, the mechanism of DDE impact on CaP cells was distinct from BPA, in that at low doses this agent also activated the MAPK pathway, and requires this activation for mitogenesis. Given the context specific impact of AR activation by EDCs, and the deleterious effect of exposure to BPA on ADT in vivo, we next addressed the impact of AR action on taxane-based therapy. These cytotoxic agents have recently been shown to improve survival outcome for patients with advanced CaP, and are potentially new second line therapies, however the impact of AR action on this cytotoxic modality had yet to be assessed. Contrary to the stigma of AR as a survival factor, we found that AR activation by both endogenous and exogenous agonists (i.e. DHT and BPA), synergized with taxanes to decrease cell survival, through p53-mediated, caspase dependent apoptosis. This synergistic action was attributed directly to the AR-dependent mitogenic capacity of these agents. Importantly, these data further support the conclusion that the environmental impact on AR action and CaP therapeutic outcome is context specific. We further evaluated the complexity of EDC affects by assessing the impact of these agents under clinically relevant conditions in another hormone-dependent tissue, breast cancer. BPA and the phytoestrogen, coumestrol (COU), were able to activate the estrogen receptor (ERα), but this activation was restricted to estrogen-depleted conditions, and could be blocked by the standard ER antagonist, tamoxifen. Additionally, BPA and COU have disparate affects in multiple analyses including mutant ERα activation, and specific coactivator deregulation. In conclusion, the environmental impact on nuclear receptor signaling and hormone dependent cancer progression and treatment is highly molecular context specific. Therefore, the impact of EDCs on hormone dependent cancer treatment needs to be identified under specific and well defined, clinically relevant, molecular contexts. Acknowledgements I would like to thank my family, friends, and co-workers for their support and encouragement. I would like to especially thank my wife, Carrie, and daughter Kaia, for their patience, understanding, and unwavering support. They are my daily examples of the truly important things in life that no publication or degree could replace. My family, parents and brother have shown me unconditional love and support, and without that, I would not have had the confidence or stamina to push through. In addition, I could not have succeeded without the help from my fellow lab mates, past and present – thanks to them for their constructive criticism, technical support, and friendships. I also want to thank my committee members for their ideas, comments and guidance. Finally, I would like to thank my advisor, Karen Knudsen, for everything that she has taught me over the last four and a half years, and for all of the opportunities that she has provided me. I truly admire her knowledge, strength, commitment to science and her lab, and who she is as a person. Thank you! Table of Contents List of Figures and Tables ………………………………………………………….. 2 Chapter I: Introduction A: Prostate cancer: the role of the androgen receptor (AR) in disease progression and treatment…………………………………………… 4 B: Endocrine disrupting compounds and prostate cancer……………….. 11 Chapter II: Bisphenol A facilitates bypass of androgen ablation therapy in prostate cancer A: Abstract…………………………………………………………………….. 24 B: Introduction………………………………………………………………… 24 C: Materials and Methods…………………………………………………… 25 D: Results……………………………………………………………………… 27 E: Discussion………………………………………………………………….. 29 F: Acknowledgements……………………………………………………….. 32 G: References………………………………………………………………… 32 Chapter III: DDE modulates proliferation of prostate cancer cells through MAPK and mutant AR pathways A: Abstract…………………………………………………………………….. 35 B: Introduction………………………………………………………………… 38 C: Materials and Methods…………………………………………………… 40 D: Results……………………………………………………………………… 45 E: Discussion………………………………………………………………….. 57 F: Acknowledgements……………………………………………………….. 61 G: References………………………………………………………………… 61 Chapter IV: Mitogenic action of the androgen receptor sensitizes prostate cancer cells to taxane-based cytotoxic insult A: Abstract…………………………………………………………………….. 75 B: Introduction………………………………………………………………… 75 C: Materials and Methods…………………………………………………… 75 D: Results……………………………………………………………………… 76 E: Discussion…………………………………………………………………. 82 F: Acknowledgements…………………………………………………….…. 84 G: References………………………………………………………………… 84 Chapter V: Xenoestrogen action in breast cancer: impact on ER-dependent transcription and mitogenesis A: Abstract…………………………………………………………………….. 87 B: Introduction………………………………………………………………… 87 C: Materials and Methods…………………………………………………… 88 D: Results……………………………………………………………………… 90 E: Discussion………………………………………………………………….. 95 F: Acknowledgements……………………………………………………….. 98 G: References………………………………………………………………… 98 Chapter VI: Summary and Future Directions…………………………………….. 102 List of Tables and Figures Chapter I-A: Figure 1 ……………………………………………………………………… 5 Figure 2 ……………………………………………………………………… 7 Chapter I-B: Figure 1 ……………………………………………………………………… 12 Table 1.……………………………………………………………………… 14 Figure 2 ……………………………………………………………………… 16 Chapter II: Figure 1 ……………………………………………………………………… 27 Figure 2 ……………………………………………………………………… 28 Figure 3 ……………………………………………………………………… 28 Figure 4 ……………………………………………………………………… 30 Figure 5 ……………………………………………………………………… 31 Chapter III: Figure 1 ……………………………………………………………………… 67 Figure 2 ……………………………………………………………………… 68 Figure 3 ……………………………………………………………………… 69 Figure 4 ……………………………………………………………………… 70 Figure 5 ……………………………………………………………………… 71 Figure 6………………………………………………………………………. 72 Figure 7……………………………………………………………………….. 73 Chapter IV: Figure 1 ……………………………………………………………………… 77 Figure 2 ……………………………………………………………………… 78 Figure 3 ……………………………………………………………………… 79 Figure 4 ……………………………………………………………………… 80 Figure 5 ……………………………………………………………………… 82 Figure 6………………………………………………………………………. 83 Chapter V: Figure 1 ……………………………………………………………………… 89 Figure 2 ……………………………………………………………………… 91 Figure 3 ……………………………………………………………………… 92 Table 1 ……………………………………………………………………….. 93 Figure 4 ……………………………………………………………………… 93 Figure 5………………………………………………………………………. 94 Figure 6………………………………………………………………………. 95 Table 2 ………………………………………………………………………. 95 Chapter VI: Figure 1.................................................................................................. 106 Chapter I: Introduction A. Prostate cancer: the role of the androgen receptor (AR) in disease progression and treatment B. Endocrine disrupting compounds and prostate cancer I-A: Prostate cancer: the role of the androgen receptor (AR) in disease progression and treatment Prostate Cancer Prevalence and Risk: In the United States, prostate cancer (CaP) is the most commonly diagnosed malignancy, and the second leading cause of cancer related deaths in men (1). The causes of CaP remain undefined; however, the most significant risk factor is age (2). CaP is regulated to older men, as incidence of prostate cancer in men under the age of 40 is rare and the risk for development of this disease in men over the age of 65 is a striking 1 in 9 (2). Other known risk factors include race, diet, family history and exposure to environmental contaminants. African American men have the highest incidence of CaP; however in general, all Western populations have the highest prevalence, most likely due to diet, life span, and overall higher health screening practices