MASS SPECTROMETRY IN DRUG METABOLISM AND DISPOSITION ffirs 1 March 2011; 11:28:26 WILEY SERIES ON PHARMACEUTICAL SCIENCE AND BIOTECHNOLOGY: PRACTICES, APPLICATIONS AND METHODS Series Editor: Mike S. Lee Milestone Development Services Mike S. Lee  Integrated Strategies for Drug Discovery Using Mass Spectrometry Birendra Pramanik, Mike S. Lee, and Guodong Chen  Characterization of Impurities and Degradants Using Mass Spectrometry Mike S. Lee and Mingshe Zhu  Mass Spectrometry in Drug Metabolism and Disposition: Basic Principles and Applications ffirs 1 March 2011; 11:28:26 MASS SPECTROMETRY IN DRUG METABOLISM AND DISPOSITION Basic Principles and Applications Edited by Mike S. Lee Milestone Development Services Mingshe Zhu Bristol-Myers Squibb ffirs 1 March 2011; 11:28:26 Copyright r 2011 by John Wiley & Sons, Inc. All rights reserved. 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Pharmacokinetics. QV 38] RM301.55.M367 2011 615u.7—dc22 2010028341 Printed in Singapore 10987654321 ffirs 1 March 2011; 11:28:26 CONTENTS FOREWORD ix Tom Baillie PREFACE xi Mike Lee and Mingshe Zhu CONTRIBUTORS xv PART I BASIC CONCEPTS OF DRUG METABOLISM AND DISPOSITION 1 1 Progression of Drug Metabolism 3 Ronald E. White 2 Common Biotransformation Reactions 13 Bo Wen and Sidney D. Nelson 3 Metabolic Activation of Organic Functional Groups Utilized in Medicinal Chemistry 43 Amit S. Kalgutkar 4 Drug-Metabolizing Enzymes, Transporters, and Drug–Drug Interactions 83 Steven W. Louie and Magang Shou 5 Experimental Models of Drug Metabolism and Disposition 151 Gang Luo, Chuang Lu, Xinxin Ding, and Donglu Zhang 6 Principles of Pharmacokinetics: Predicting Human Pharmacokinetics in Drug Discovery 197 Takehito Yamamoto, Akihiro Hisaka, and Hiroshi Suzuki 7 Drug Metabolism Research as Integral Part of Drug Discovery and Development Processes 229 W. Griffith Humphreys v TOC 1 March 2011; 11:35:40 PART II MASS SPECTROMETRY IN DRUG METABOLISM: PRINCIPLES AND COMMON PRACTICE 255 8 Theory and Instrumentation of Mass Spectrometry 257 Ge ´ rard Hopfgartner 9 Common Liquid Chromatography–Mass Spectrometry (LC–MS) Methodology for Metabolite Identification 291 Lin Xu, Lewis J. Klunk, and Chandra Prakash 10 Mass Spectral Interpretation 321 Li-Kang Zhang and Birendra N. Pramanik 11 Techniques to Facilitate the Performance of Mass Spectrometry: Sample Preparation, Liquid Chromatography, and Non-Mass-Spectrometric Detection 353 Mark Hayward, Maria D. Bacolod, Qing Ping Han, Manuel Cajina, and Zack Zou PART III APPLICATIONS OF NEW LCÀMS TECHNIQUES IN DRUG METABOLISM AND DISPOSITION 383 12 Quantitative In Vitro ADME Assays Using LC–MS as a Part of Early Drug Metabolism Screening 385 Walter Korfmacher 13 High-Resolution Mass Spectromet ry and Drug Metabolite Identification 407 Russell J. Mortishire-Smith, Haiying Zhang, and Kevin P. Bateman 14 Distribution Studies of Drugs and Metabolites in Tissue by Mass Spectrometric Imaging 449 Richard F. Reich, Daniel P. Magparangalan, Timothy J. Garrett, and Richard A. Yost 15 Use of Triple Quadrupole–Linear Ion Trap Mass Spectrometry as a Single LC–MS Platform in Drug Metabolism and Pharmacokinetics Studies 483 Wenying Jian, Ming Yao, Bo Wen, Elliott B. Jones, and Mingshe Zhu 16 Quantitative Drug Metabolism with Accelerator Mass Spectrometry 525 John S. Vogel, Peter Lohstroh, Brad Keck, and Stephen R. Dueker 17 Standard-Free Estimation of Metabolite Levels Using Nanospray Mass Spectrometry: Current Statutes and Future Directions 567 Jing-Tao Wu vi CONTENTS TOC 1 March 2011; 11:35:40 18 Profiling and Characterization of Herbal Medicine and Its Metabolites Using LC–MS 579 Zeper Abliz, Ruiping Zhang, Ping Geng, Dongmei Dai, Jiuming He, and Jian Liu 19 Liquid Chromatography Mass Spectrometry Bioanalysis of Protein Therapeutics and Biomarkers in Biological Matrices 613 Fumin Li and Qin C. Ji 20 Mass Spectrometry in the Analysis of DNA, Protein, Peptide, and Lipid Biomarkers of Oxidative Stress 645 Stacy L. Gelhaus and Ian A. Blair 21 LC–MS in Endogenous Metabolite Profiling and Sm all-Molecule Biomarker Discovery 685 Michael D. Reily, Petia Shipkova, and Serhiy Hnatyshyn Appendix 723 Index 727 CONTENTS vii TOC 1 March 2011; 11:35:40 FOREWORD Studies in the areas of drug metabolism and pharmacokinetics have assum ed progressively greater importance in pharmaceutical research over the pa st two decades, reflecting an increased awareness of the critical impact on successful drug development of the absorption, metabolism, distribution, elimination, and toxicity (ADMET) properties of candidate therapeutic agents. Indeed, the role of drug metabolism studies in the pharmaceutical industry, formerly limited to later phases of the development process, now spans the continuum from early discovery efforts through lead optimization, preclinical development, clinical trials, and postmarketing surveillance. Information on the identities and exposure levels of drug metabolites, first in animals and subsequently in human subjects, represents an essential component of preclinical and clinical safety assessment programs and, in those cases where circulating metabolites are pharmacologically active, provides the basis for assessing their pharmacoki- netic/pharmacodynamic (PK/PD) relationships and contribution to the effects of the parent drug. Chemically reactive drug metabolites, which can be detected and characterized through specialized in vitro “trapping” techniques, generally are viewed as risk facto rs in drug development in light of their association with several forms of drug-induced toxicities, and early information on their identities is key to the design of optimized new chemical entities that lack this potential liability. The detection, structural characterization, and quantitative analysis of drug metabolites in complex biological matrices often is a challenging endeavor, given the low levels that derive from highly potent parent compounds that are administered at doses of a few milligrams per day or less. As a result, stringent demands are placed on the analytical methodology employed for drug metabolism studies conducted either in vitro or in vivo, in terms of sensitivity and specificity of detection, and of wide dynamic range. In this regard, mass spectrometry, which always has been an important technique in drug metabo- lism studies, rapidly became the dominant technology in the field following the introduction, in the early 1990s, of the first co mmercial LCÀMS/MS systems. Over the past decade, remarkable technical advances have been made in ion source design and ionization methods, rapid scanning and highly sensitive mass analyzers, efficient methods for indu cing fragmentation of parent ions, rapid- response detectors with wide dynamic range, and powerful data acquisition and processing systems with sophisticated software packages and expert systems designed specifically for investigations in drug metabolism. The evolution of ix fbetw 1 March 2011; 11:27:46 hybrid mass analyzers for MS/MS studies, and ancillary techniques such as ion mobility spectrometry, have added new dimensions to the mass spectrometry experiment, while the advent of high mass resolution capabilities on an LC time scale (even with “fast” chromatography) is having a truly revolutionary impact on the utility of LCÀMS/MS in this field. Mass Spectrometry in Drug Metabolism and Disposition: Basic Principles and Applications addresses each of these areas through a series of chapters authored by eminent scientists well versed in the application of contemporary mass spectrometry techniques to problems in drug metabolism and pharmacoki- netics, with an emphasis on issues in drug discovery and development. The reader cannot help but be impressed by the capabilities of the current generation of LCÀMS/MS instruments, which provide a combination of sensitivity, specificity, versatility, and speed of analysis that was difficult to envisage only a few years ago, and which have transformed the way drug metabolism studies are conducted. One can only wonder what lies in the years ahead! Thomas A. Baillie Seattle, WA August, 2010 x FOREWORD fbetw 1 March 2011; 11:27:46 PREFACE Two decades ago, drug metabolism research in the pharmaceutical industry was limited to radiolabeled in vivo drug disposition studies conducted in late stages of development. Drug metabolite identification was accomplished via a long and tedious process: metabolite separation and isolation followed by mass spectrometric and nuclear magnetic resonance analysis. Now, drug metabolism plays a critical role in the drug discovery and development process from lead optimization to clinical drug–drug interaction studies. Commercialized liquid chromatography/mass spectrometry (LC/MS) platforms have become the dominant analytical instrument employed in drug metabolism and pharmaco- kinetics (DMPK) studies and revolutionized the productivity of drug metabo- lism research. Certainly, the need for fast, sensitive and accurate measurements of drugs and metabolites in complex biological matrices has driven the con- tinued development of novel LC/MS technology. Drug metabolism science and mass spectrometry technology have been integrated into an inseparable arena in drug discovery and development as well in related academic research activities. This book provides a unique thesis on mass spectrometry in drug metabolism with specific emphasis on both principles and applications in drug design and development. We believe that this integration will provide a unique contribution to the field that details both drug metabolism and analytical perspectives. Therefore, this book can be used as a teaching and/or reference tool to delineate and understand the “why” and “how” with regard to the many creative uses of mass spectrometry in drug metabolism and disposi- tion studies. This work, authored by internationally renowned researchers, represents a combination of complementary backgrounds to bring technical and cultural awareness to this very important endeavor while serving to address needs within academia and industry. The book is organized into three parts. Part I provides the reader with the basic concepts of drug metabolism and disposition. These concepts are intended to build a unique foundation of knowledge for drug metabolism and the subsequent studies performed during drug discovery and drug devel- opment endeavors. The book begins with an elegant perspective on drug metabolism. This review or perhaps “state of the union” provides unique insight into where we are, how we got there, and where we appear to be headed. Next we delve into the details of drug metabolism with a chapter on common biotransformation reactions. Further detail is provided in Chapter 3 from a medicinal chemistry perspective as the metabolic activation of organic xi fpref 1 March 2011; 11:35:10 [...]... the mass spectrometer Part III of the book focuses on LC/MS techniques in drug metabolism and disposition The application of quantitative LC/MS in drug metabolism and disposition is highlighted in Chapter 12 Critical studies that are routinely performed in drug discovery that involve metabolic stability, enzyme kinetics, metabolizing enzyme inhibition and induction, permeability and absorption, and in. .. will obtain insight into the strategies used to design experiments for characterizing drug metabolism properties and addressing drug metabolism related issues from drug discovery to regulatory registrations Part II of the book highlights the principles and common practices of mass spectrometry in drug metabolism The basic concepts and theory of mass spectrometry are presented in Chapter 8 In this chapter... in drug design Chapter 4 provides an overview on metabolizing enzymes, transporters, and their involvement with drug drug interactions In vitro experimental approaches to assess and predict drug drug interactions in humans are elaborated Chapter 5 starts with DMPK strategies in a drug discovery setting followed by a comprehensive overview of various experimental models applied in drug metabolism and. .. background and advantages of modern high-resolution mass spectrometry along with the use of newly developed data-mining tools for in vitro and in vivo drug metabolite identification The understanding of the tissue distribution of a drug and the corresponding metabolites is illustrated in Chapter 14 The recent applications of imaging mass spectrometry for these studies are described Novel instrumentation and mass. .. Scatina J Obtaining exposures of metabolites in preclinical species through plasma pooling and quantitative NMR: Addressing metabolites in safety testing (MIST) guidance without using radiolabeled compounds and chemically synthesized metabolite standards Chem Res Toxicol 2009;22:311À322 White RE Short and long-term projections about the involvement of drug metabolism in drug discovery and development Drug. .. its core, and the backbone technology remains mass spectrometry (MS) However, if we consider the more complete picture, DM has expanded enormously in scope and level of understanding in those 50 years While the chemistry-based core remains intact and actively growing, several other kinds of DM studies have layered over the core, all coexisting and relevant in contemporary DM science Thus, in addition... continually increases with time but abruptly increases about 2010 due to enhanced regulatory surveillance of metabolites Starting around 1975, biochemistry (B, dark gray) begins to be included in the DM characterization and slowly increases with time Genetics information (G, light gray) continues to increase, mainly in clinical trials The apparent jump in B and G work around 2010 is due to the abrupt increase... instantly recognized and understood by DM scientists from 50 years ago This is not the case with most other disciplines in the life sciences Mass Spectrometry in Drug Metabolism and Disposition: Basic Principles and Applications, First Edition Edited by Mike S Lee and Mingshe Zhu r 2011 John Wiley & Sons, Inc Published 2011 by John Wiley & Sons, Inc 3 CH001 1 March 2011; 10:32:13 4 PROGRESSION OF DRUG. .. relate to safety and efficacy When designing a practical clinical medicine, we need to establish a balance between too rapid metabolism, leading to reduced efficacy, and too sluggish metabolism, leading to accumulation and possible toxicity In the clinic, we need to determine whether metabolism decreases or increases the desired pharmacological effect (i.e., active metabolites) And, finally, in this holistic... for microdosing and DMPK studies in early clinical studies is described Chapter 17 provides a provocative description of novel approaches, typically used in the field of protein identification, to obtain standard-free estimation of metabolite levels using nanospray mass spectrometry A unique perspective on the profiling and characterization of Chinese herbal medicine and their metabolites using LC/MS is . Spectrometry Mike S. Lee and Mingshe Zhu  Mass Spectrometry in Drug Metabolism and Disposition: Basic Principles and Applications ffirs 1 March 2011; 11:28:26 MASS SPECTROMETRY IN DRUG METABOLISM AND DISPOSITION Basic. Quadrupole–Linear Ion Trap Mass Spectrometry as a Single LC–MS Platform in Drug Metabolism and Pharmacokinetics Studies 483 Wenying Jian, Ming Yao, Bo Wen, Elliott B. Jones, and Mingshe Zhu 16. Utilized in Medicinal Chemistry 43 Amit S. Kalgutkar 4 Drug- Metabolizing Enzymes, Transporters, and Drug Drug Interactions 83 Steven W. Louie and Magang Shou 5 Experimental Models of Drug Metabolism and