Pharmacokinetic-Pharmacodynamic and Pharmacogenetic Studies of Flavopiridol and its Glucuronide Metabolite DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Wenjun Ni Graduate Program in Pharmacy The Ohio State University 2011 Dissertation Committee: Dr. Mitch A. Phelps, Advisor Dr. M. Guillaume Wientjes Dr. Robert Lee Copyright by Wenjun Ni 2011 Abstract Chronic lymphocytic leukemia (CLL) is the most common adult leukemia in Western world. Flavopiridol (Alvocidib, NSC 649890), as a pan cyclin-dependent kinases inhibitor (CDKI), initiates cell cycle arrest and p53-independent apoptosis through down- regulation of Mcl-1 and X-linked inactivator of apoptosis (XIAP). A novel pharmacokinetically (PK)-based dosing schedule with a 30-minute intravenous bolus loading dose (IVB) followed by a 4-hour continuous intravenous infusion (CIVI) in patients with refractory CLL produced an approximately 50% overall response rate. A major flavopiridol metabolite, flavopiridol glucuronide (flavo-G), was as also evaluated in this study. In order to fully understand the between-subject variability (BSV) between patients, a phase 1 CLL patient data set (OSU0055) was evaluated and a two- compartment flavopiridol PK model followed by first-order elimination was developed by nonlinear mixed effects modeling. Bilirubin level was shown as a significant covariate, and OATP1B1 was first time discovered having a significant correlation with flavopiridol PK parameters. A functional analysis in vitro study was done to confirm that flavopiridol and flavo-G are substrates of OATP1B1. Since cellular redox status is important on cell survival and previous studies showed that flavopiridol can induce the decrease of intracellular GSH levels in transformed cells, glutathione (GSH) level was evaluated among leukemia cell models and patient’s CLL cells. The change in GSH level ii compared to baseline after flavopiridol treatment varied among cell models and individual patient. In order to evaluate flavo-G with flavopiridol treatment, a linked parent-metabolite population PK model was developed. This model was expanded to include a pharmacodynamic logistic-regression component linked to flavopiridol exposure level. This developing PK-PD-PG model of flavopiridol will help to characterize the factors associated with inter-individual variability in drug disposition and outcomes, and provide us better understanding of the mechanisms and factors governing the balance between safety and efficacy. iii Dedication This document is dedicated to my family. iv Acknowledgments This thesis would not have been possible without the support of many people. First and foremost, I would like to thank my adviser, Dr. Mitch A. Phelps, for his intellectual supervision, continuous support, extreme patience, motivation and encouragement throughout my graduate studies. I am very grateful to Ms. Kathy Brooks for her administrative help and support, and we will all miss her so much. I would also like to express my thanks to all of my friends and colleagues in College of Pharmacy for their encouragement and invaluable scientific discussions. Finally, my special thanks go to my family. I am deeply thankful to my parents for their everlasting love, support and encouragement. “It's not what I am underneath, but what I do, that defines me.” v Vita Sep 1997-Jun 2001………………… B.S. Pharm, the Second Military Medical University, Shanghai, P.R. China Sep 2003-Jun 2006……………………. Master of Science, Cincinnati Children’s Hospital Medical Center, Cincinnati OH Sep 2006 – Mar 2011………….…….. Doctor of Philosophy, The Ohio State University Publications MANUSCRIPTS • Ni W., Ji J., Dai Z., Papp A., Johnson AJ, Ahn S., Farley KL, Lin TS, Dalton JT, Li X., Jarjoura D., Byrd JC, Sadee W, Grever MR, Phelps MA. Flavopiridol Pharmacokinetics and Pharmacogenetics: Clinical and Functional Evidence for the Role of SLCO1B1/OATP1B1 in Flavopiridol Disposition. PLoS ONE. 2010:e13792. • Ni W., Riesen, S.C., Carnes C.A., Lindsey K.J., Phelps M.A., Schober K.E. Pharmacokinetics of Oral Ivabradine in Healthy Cats. The Journal of Veterinary Pharmacology and Therapeutic. 2010 Dec 1. doi: 10.1111/j.1365-2885.2010.01253.x. [Epub ahead of print] • Ramaswamy B., Phelps M.A., Baiocchi R., Bekaii-Saab T., Ni W., Lai J., vi Wolfson A., Lustberg M.E., Wei L., Wilkins D., Campbell A., Arbogast D., Doyle A., Byrd J.C., Grever M.R., and Shah M.H. A dose-finding, pharmacokinetic and pharmacodynamic study of a novel schedule of flavopiridol in patients with advanced solid tumors. Invest New Drugs. 2010 Oct 12. [Epub ahead of print] • Blum W., Phelps M.A., Klisovic R.B., Rozewski D.M., Ni W., Albanese K.A., Rovin B., Kefauver C., Devine S.M., Lucas D.M., Johnson A., Schaaf L.J., Byrd J.C., Marcucci G., Grever M.R. Phase I clinical and pharmacokinetic study of a novel schedule of flavopiridol in relapsed or refractory acute leukemias. Haematologica. 2010 Jul;95(7):1098-105. • Bosco EE, Ni W., Wang L., Guo F, Johnson JF, and Zheng Y. Rac1 targeting suppresses p53 deficiency–mediated lymphomagenesis. Blood. 2010; 115(16):3320-8. • Yan J, He C, Wang X, Bao X, Ni W., and Lu C, The effect of CNTF on glutamate-induced increase in intracellular free Ca2+ in hippocampal neurons. NeuroReport 2000; 11:3439-41. CONFERENCE POSTERS/PODIA • “Influence of Glutathione-S-Transferase Pharmacogenetics in a Population Pharmacokinetic-Pharmacodynamic Model of Flavopiridol and Its Metabolite” (Poster). vii Ni W., Ji J., Mahoney E, Smith LL, Dai Z., Papp A., Johnson AJ, Ahn S., Farley KL, Lin TS, Dalton JT, Li X., Jarjoura D., Byrd JC, Sadee W, Grever MR, Phelps MA. AAPS Annual Meeting 2010, New Orleans, LA. • “Population Pharmacokinetics and Pharmacogenetics of Flavopiridol in Chronic Lymphocytic Leukemia Patients and Functional Analysis of SLCO1B1” (Podium). Ni W., Ji J., Dai Z., Papp A., Johnson AJ, Ahn S., Farley KL, Lin TS, Dalton JT, Li X., Jarjoura D., Byrd JC, Sadee W, Grever MR, Phelps MA. The 42nd Annual Pharmaceutics Graduate Student Research Meeting (PGSRM) 2010, Columbus, OH. • “Clinical Pharmacokinetics and Pharmacogenetics of Flavopiridol” (Poster). Ni W., Dai Z., Papp A., Johnson AJ, Ahn S., Farley KL, Lin TS, Dalton JT, Li X., Jarjoura D., Byrd JC, Sadee W, Grever MR, Phelps MA. AAPS Annual Meeting 2009, Los Angeles, CA. Fields of Study Major Field: Pharmacy Studies on preclinical, clinical pharmacokinetics/pharmacodynamics, pharmacogenetics/pharmacogenomics, drug transporters and bioanalytical method development. viii Table of Contents Abstract ............................................................................................................................... ii Dedication .......................................................................................................................... iv Acknowledgments............................................................................................................... v Vita ..................................................................................................................................... vi Publications ........................................................................................................................ vi Fields of Study ................................................................................................................. viii Table of Contents ............................................................................................................... ix List of Tables .................................................................................................................... xv List of Figures .................................................................................................................. xvi Chapter 1: BACKGROUND AND INTRODUCTION ..................................................... 1 1.1. Background .............................................................................................................. 1 1.1.1. Flavopiridol ....................................................................................................... 1 1.1.2. Clinical Studies of Flavopiridol for Treatment of CLL ..................................... 2 1.1.3. Disposition of Flavopiridol and its Glucuronide Metabolits ............................. 3 1.1.4. Flavopiridol and Glutathione ............................................................................. 4 1.2. Significance .......................................................................................................... 5 ix Chapter 2: CLINICAL POPULATION PHARMACOKINETIC OF FLAVOPIRIDOL IN PATIENTS WITH CHRONIC LYMPHOCYTIC LEUKEMIA ...................................... 8 2.1. Introduction .................................................................................................................. 8 2.2. Methods........................................................................................................................ 9 2.2.1. Study Protocols and Clinical Design ................................................................. 9 2.2.2. Structural Model Development ......................................................................... 9 2.2.3. Final Model Development ............................................................................... 11 2.2.4. Internal Model validation ................................................................................ 13 2.2.5. External Model Validation .............................................................................