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New Mechanism Based Approaches for Treating Prostate Cancer Rayna Rosati Wayne State University

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New Mechanism Based Approaches for Treating Prostate Cancer Rayna Rosati Wayne State University Wayne State University Wayne State University Dissertations 1-1-2017 New Mechanism Based Approaches For Treating Prostate Cancer Rayna Rosati Wayne State University, Follow this and additional works at: https://digitalcommons.wayne.edu/oa_dissertations Part of the Oncology Commons Recommended Citation Rosati, Rayna, "New Mechanism Based Approaches For Treating Prostate Cancer" (2017). Wayne State University Dissertations. 1865. https://digitalcommons.wayne.edu/oa_dissertations/1865 This Open Access Dissertation is brought to you for free and open access by DigitalCommons@WayneState. It has been accepted for inclusion in Wayne State University Dissertations by an authorized administrator of DigitalCommons@WayneState. NEW MECHANISM BASED APPROACHES FOR TREATING PROSTATE CANCER by RAYNA C. ROSATI DISSERTATION Submitted to the Graduate School of Wayne State University, Detroit, Michigan in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY 2017 MAJOR: CANCER BIOLOGY Approved By: Advisor Date DEDICATION This dissertation is dedicated to my family, who made me who I am today with all of their love and support. To my grandmother, Lucy, who just recently lost her battle with lung cancer and who would send me newspaper clippings in the mail about new topics of prostate cancer research. To my siblings, who have always been my best friends. To my father, Daniel, who has been my greatest mentor in life. To my mother, Nanci, who has been there for me through everything and who gave me my creative bone. ii ACKNOWLEDGEMENTS First off, I would like to thank my mentor, Dr. Manohar Ratnam, whose enthusiasm for science is undeniable. Thank you for always being so optimistic and believing I could accomplish this very challenging project. Thank you for teaching me how to ask the proper questions and how to be a good scientist. Thank you for your dedication to my project and your dedication in preparing me for my future. I would like to thank my committee members, Dr. Larry Matherly, Dr. Izabela Podgorski, and Dr. Elisabeth Heath for all of their continued support and advice over the past five years and for being so accommodating for meetings. I would like to thank all of my lab members, both past and present, Dr. Mugdha Patki, Dr. Venkatesh Chari, Yanfang Huang, and especially Thomas McFall, for not only providing me with exceptional laboratory support but also for his friendship. I would like to thank the cancer biology graduate program, Drs. Matherly and Brush as well as Nadia Daniel. Thank you for taking a chance on me and accepting me into the program and for all of your guidance over the past few years. Thank you to all of our collaborators, Dr. Zhihui Qin from Wayne State University, Dr. Peter Shaw from Nottingham University, and Dr. Hiroki Kakuta from Okayama University. Thank you to Dr. Andrew Fribley and Martha Larsen for everything involving my small molecule screening optimization and help with performing the screen. Thank you to Dr. Lisa Polin for all of your contribution to the animal studies involved with my project, it truly would have not been possible without you. Thank you to Dr. Selvakumar Dakshnamurthy for performing the SPR studies over the past year that has contributed to this project. Thank you to Dr. Jing Li for all of the PK/PD support on this project. iii Thank you to my funding, the 5T32CA009531 fellowship and the Thomas C. Rumble University Graduate Fellowship. Lastly, thank you to everyone at Wayne State University School of Medicine that has contributed to my career and time here at the University. iv TABLE OF CONTENTS DEDICATION ................................................................................................................... ii ACKNOWLEDGEMENTS ............................................................................................... iii LIST OF TABLES ............................................................................................................ xi LIST OF FIGURES ......................................................................................................... xii LIST OF ABBREVIATIONS ........................................................................................... xiv CHAPTER 1 – INTRODUCTION………………………………………………………………1 1.1 Prostate Cancer –a 2017 overview ................................................................. 1 1.2 The normal prostate: structure and function……………………………………..1 1.2.1 Prostate zones ................................................................................. 2 1.2.2 Aspects of endocrine action, the androgen receptor and the prostate gland .......................................................................................................... 4 1.3 Prostate Disease ........................................................................................... 9 1.3.1 Prostatitis .......................................................................................... 9 1.3.2 Benign Prostatic Hyperplasia ............................................................ 9 1.3.3 Prostate Cancer .............................................................................. 10 1.4 Prostate Cancer-diagnosis ........................................................................... 12 1.4.1 Prostate Cancer Grading .............................................................. 12 1.4.2 Prostate Cancer Staging ............................................................... 12 1.5 Prostate cancer treatment ............................................................................ 13 1.5.1 Androgen deprivation therapy and anti-androgen therapy ............. 13 1.5.2 Available Treatments for CRPC ..................................................... 14 1.6 Other Major Limitations of Testosterone Suppression in Prostate Cancer – Effects on Normal Tissues ................................................................................. 15 1.6.1 Hot Flashes ..................................................................................... 16 v 1.6.2 Hyperlipidemia ................................................................................ 16 1.6.3 Sexual Dysfunction ......................................................................... 17 1.6.4 Anemia ............................................................................................ 17 1.6.5 Skeletal Complications .................................................................... 18 1.6.6 Effects on the cardiovascular system .............................................. 19 1.6.7 Cognitive and Psychological Impairment ........................................ 20 1.7 Mechanisms of restoration of AR signaling in CRPC .................................... 21 1.7.1 Intratumoral Androgen Synthesis .................................................... 21 1.7.2 Overexpression of AR ..................................................................... 22 1.7.3 Imbalance of AR co-regulator expression ....................................... 22 1.7.4 Dysregulation of growth factors and crosstalk with other signaling pathways .................................................................................................. 23 1.7.5 Mutations in AR and Splice Variants of AR ..................................... 23 1.8 Targeting the amino-terminal domain of AR ................................................. 26 1.8.1 Bromodomain inhibitors .................................................................. 27 1.8.2 EPI family small molecules ............................................................. 28 1.8.3 Sintokamides .................................................................................. 30 1.8.4 Bispecific antibodies to inhibit N-terminal AR activity ...................... 30 1.9 Targeting AR tethering proteins ................................................................... 31 1.9.1 HoxB13 ........................................................................................... 32 1.9.2 C/EBPα ........................................................................................... 33 1.9.3 ELK1 and other TCF subfamily members ....................................... 34 1.9.4 ELK1 as an AR tethering protein essential for PC/CRPC growth ... 35 1.10 The ELK1-AR complex as a critical and selective therapeutic target in prostate cancer ................................................................................................... 38 vi CHAPTER 2-THESIS OUTLINE ................................................................................... 40 CHAPTER 3- THE AMINO-TERMINAL DOMAIN OF THE ANDROGEN RECEPTOR CO-OPTS ERK DOCKING SITES IN ELK1 TO INDUCE SUSTAINED GENE ACTIVATION THAT SUPPORTS PROSTATE CANCER CELL GROWTH .................. 41 3.1 Introduction ................................................................................................... 41 3.2 Experimental Procedures ............................................................................. 43 3.2.1 Cell culture and reagents ................................................................ 43 3.2.2 Purified proteins .............................................................................. 45 3.2.3 Surface plasmon resonance ........................................................... 45 3.2.4 Plasmids ......................................................................................... 46 3.2.5 siRNA mediated gene knockdown .................................................. 48 3.2.6 Co-Immunoprecipitation .................................................................
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