Pentoxifylline As an Adjuvant Treatment in Renal Cell Carcinoma
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Pentoxifylline As An Adjuvant Treatment In Renal Cell Carcinoma Item Type text; Electronic Dissertation Authors Mastrandrea, Nicholas Joseph Publisher The University of Arizona. Rights Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author. Download date 06/10/2021 16:37:11 Link to Item http://hdl.handle.net/10150/337293 PENTOXIFYLLINE AS AN ADJUVANT TREATMENT IN RENAL CELL CARCINOMA by Nicholas J. Mastrandrea A Dissertation Submitted to the Faculty of the DEPARTMENT OF PHARMACOLOGY AND TOXICOLOGY In Partial Fulfillment of the Requirements For the Degree of DOCTOR OF PHILOSOPHY In the Graduate College THE UNIVERSITY OF ARIZONA 2014 1 THE UNIVERSITY OF ARIZONA GRADUATE COLLEGE As members of the Dissertation Committee, we certify that we have read the dissertation prepared by Nicholas J Mastrandrea, titled Pentoxifylline as an Adjuvant Treatment in Renal Cell Carcinoma and recommend that it be accepted as fulfilling the dissertation requirement for the Degree of Doctor of Philosophy. ________________________________________________________ Date: 9/25/14 Serrine S. Lau, Ph. D. ________________________________________________________ Date: 9/25/14 Terrence J. Monks, Ph. D. ________________________________________________________ Date: 9/25/14 Donna Zhang, Ph. D. ________________________________________________________ Date: 9/25/14 Richard Vaillancourt, Ph. D. ________________________________________________________ Date: 9/25/14 George Tsaprailis, Ph. D. Final approval and acceptance of this dissertation is contingent upon the candidate’s submission of the final copies of the dissertation to the Graduate College. I hereby certify that I have read this dissertation prepared under my direction and recommend that it be accepted as fulfilling the dissertation requirement. ________________________________________________________ Date: 9/25/14 Dissertation Director: Serrine S. Lau 2 STATEMENT BY AUTHOR This dissertation has been submitted in partial fulfillment of the requirements for an advanced degree at the University of Arizona and is deposited in the University Library to be made available to borrowers under rules of the Library. Brief quotations from this dissertation are allowable without special permission, provided that an accurate acknowledgement of the source is made. Requests for permission for extended quotation from or reproduction of this manuscript in whole or in part may be granted by the copyright holder. SIGNED: Nicholas J. Mastrandrea 3 ACKNOWLEDGEMENTS First and foremost, I sincerely thank my mentor, Dr. Serrine Lau, for her guidance, patience and support throughout my graduate career. Dr. Lau provided me with enough freedom and funding to allow me to make this project my own. At times, she was the gas pedal in a stalled car. Other times, she was the brake pedal in a car out of control. Always, though, Dr. Lau put my training ahead of everything else. The independence and critical thinking I have gained here is a direct result of Dr. Lau's advising strategy; one that I am infinitely grateful for. I would also like to thank my dissertation committee members, Dr. Terrence Monks, Dr. Donna Zhang, Dr. Richard Vaillancourt, and Dr. George Tsaprailis for their continuous advice and guidance throughout the years. They helped me to rethink about my data and suggested key experiments to perform in order to effectively demonstrate my hypothesis. Thank you for the time and effort each one of you have put into me and this project. In addition, I am grateful for the numerous past and present members of the Lau and Monks labs for their training, friendship and support. Also, I would like to thank Dr. Nagle, Jaime Gard, David Chafin, and Ventana Medical Systems for our exciting, albeit challenging, collaboration. I would like to thank the staff at the shared cytometry, genomics, genetics, proteomics and animal cores here at the University of Arizona. Without your expertise and techniques, much of my data would not exist. A special thanks goes to Wesley Cai who worked closely with me during the four years of his undergraduate career. It didn't take long to train Wesley, nor did it take long for him to demonstrate excellent scientific ability. Thus, I expected more from Wesley than is typical of an undergraduate and Wesley continually exceeded those expectations. More importantly, though, Wesley has been a great friend. I am very proud of him and his accomplishments and I look forward to what discoveries he will make in his career. Finally, I would like to thank my family and friends for their support through these challenging six years. Thank you, Mom, Dad, and Gayle, for helping to form the man I am today. Thank you, Chris and Micah, Adrienne, Brad and Amanda, for your unconditional love and support. I couldn't have done this without the help of you all. To my friends, Patrick, Kuhlman, Pablo, Eric, Jon, Wnek and Wesley, thank you for all the shared experiences we had. I wish to never forget the memories we formed. 4 TABLE OF CONTENTS LIST OF FIGURES AND TABLES .............................................................................. 10 LIST OF ABBREVIATIONS ........................................................................................ 13 ABSTRACT ..................................................................................................................... 17 CHAPTER 1:INTRODUCTION ................................................................................... 19 1.1 General Comments................................................................................................. 19 1.2 Kidney Anatomy and Physiology .......................................................................... 20 1.3 Kidney Carcinogenesis .......................................................................................... 24 1.3.1 von Hippel-Lindau Disease ............................................................................ 25 1.3.2 Tuberous Sclerosis Complex .......................................................................... 26 1.4 Cell Cycle and Its Regulation ................................................................................ 27 1.4.1 Restriction Point Regulation ........................................................................... 30 1.5 Cyclin D1 Regulation and Role in Cancer ............................................................. 31 1.5.1 CCND1 Transcription via MAPK Signaling .................................................. 33 1.5.2 Cyclin D1 Translation via PI3K/AKT/mTOR/4EBP1 Signaling ................... 34 1.5.3 Cyclin D1 Nuclear Export and Proteasomal Degradation via GSK-3β Signaling ........................................................................................................................ 37 1.6 The Eker Rat and Production of the QTRRE Cell Model ..................................... 41 1.6.1 Hydroquinone and Resulting Nephrotoxic Metabolites ................................. 42 1.6.2 QTRRE Cell Model ........................................................................................ 44 1.7 Human RCC Cell Models ...................................................................................... 45 1.8 Anticancer Therapy ................................................................................................ 46 1.8.1 Current FDA-Approved Targeted Therapies for RCC ................................... 50 1.8.2 Sorafenib ......................................................................................................... 53 1.8.3 Current RCC Adjuvant Treatment Outlook.................................................... 54 1.9 Pentoxifylline ......................................................................................................... 55 1.10 Dissertation Aims................................................................................................... 61 CHAPTER 2: PENTOXIFYLLINE DECREASES CYCLIN D1, pRB LEVELS AND ARRESTS RCC CELL MODELS IN THE G1 PHASE .................................... 64 2.1 Introduction ............................................................................................................ 64 5 2.2 Materials and Methods ........................................................................................... 67 2.2.1 Cell Culture..................................................................................................... 67 2.2.2 Western Blot Analysis .................................................................................... 67 2.2.3 QTRRE Expression Microarray Analysis ...................................................... 68 2.2.4 RT-PCR .......................................................................................................... 69 2.2.5 Cell Cycle Analysis ........................................................................................ 70 2.2.6 MTS Cell Viability Assay .............................................................................. 70 2.2.7 Statistics .......................................................................................................... 71 2.3 Results .................................................................................................................... 72 2.3.1 PTX, but not db-cAMP or theophylline, decreases cyclin D1 in QTRRE cells 72 2.3.2 PTX induces gene