2013, XV, 371 p. 62 illus., 19 illus. in color.
- Includes cutting-edge methods and protocols
- Provides step-by-step detail essential for reproducible results
- Contains key notes and implementation advice from the experts
The major reason for the elevated costs of drug development in the pharmaceutical industry is the high attrition rate. In Target Identification and Validation in Drug Discovery: Methods and Protocols, expert researchers in the field detail many of the methods which are now commonly used to identify and validate new target. These include methods and approaches covering biochemical, cell based, in vivo models and translational methods. Chapters also include selected case reports that demonstrate the integration of these technologies to real life experiences and to demonstrate the multiple use of more than one technology to increase knowledge on a specific target. These Written in the highly successful Methods in Molecular Biology™ series format, the chapters include the kind of detailed description and implementation advice that is crucial for getting optimal results in the laboratory.
Thorough and intuitive, Target Identification and Validation in Drug Discovery: Methods and Protocols contains a comprehensive list of essential methods and clear protocols to follow.
Content Level » Professional/practitioner
Keywords » Omics - Oncology - biochemical - cell based - cell culture technologies - drug development - in vivo models - in-silico - translational methods
Contents
Preface. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . v
Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . vii
Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . xiii
PART I BACKGROUND
1 The Path to Oncology Drug Target Validation: An Industry Perspective . . . . . 3
Marta Cortés-Cros, Tobias Schmelzle, Volker M. Stucke,
and Francesco Hofmann
PART II DNA AS A TOOL TO MODULATE DRUG TARGETS
2 Identification of Aptamers as Specific Binders and Modulators
of Cell-Surface Receptor Activity. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
Henning Ulrich and Carsten Wrenger
3 The Design and Structure–Functional Properties
of DNA-Based Immunomodulatory Sequences . . . . . . . . . . . . . . . . . . . . . . . . 41
Nikolai V. Kuznetsov
PART III RNA INTERFERENCE: FROM DESIGN TO DATA ANALYSIS
4 siRNA Design Principles and Off-Target Effects . . . . . . . . . . . . . . . . . . . . . . . 59
Sebastian Petri and Gunter Meister
5 Western Blot Evaluation of siRNA Delivery by pH-Responsive Peptides . . . . . 73
Wanling Liang, A. James Mason, and Jenny K.W. Lam
6 High-Throughput RNAi Screening for the Identification
of Novel Targets . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89
Meredith C. Henderson and David O. Azorsa
7 Integration of RNAi and Small Molecule Screens to Identify Targets
for Drug Development . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97
Konstantinos Drosopoulos and Spiros Linardopoulos
8 CellProfiler and KNIME: Open Source Tools for High
Content Screening . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 105
Martin Stöter, Antje Niederlein, Rico Barsacchi, Felix Meyenhofer,
Holger Brandl, and Marc Bickle
9 PARP Inhibition as a Prototype for Synthetic Lethal Screens . . . . . . . . . . . . . . 123
Xuesong Liu x Contents
PART IV PROTEIN-FOCUSED TECHNOLOGIES
10 Structure-Based Target Druggability Assessment. . . . . . . . . . . . . . . . . . . . . . . 141
Jean-Yves Trosset and Nicolas Vodovar
11 Validating Pharmacological Disruption of Protein–Protein Interactions
by Acceptor Photobleaching FRET Imaging . . . . . . . . . . . . . . . . . . . . . . . . . . 165
Janos Roszik, Gábor Tóth, János Szöllősi, and György Vereb
12 Systematic Analysis of Complex Signal Transduction Pathways
Using Protein Fragment Complementation Assays . . . . . . . . . . . . . . . . . . . . . 179
Thomas I. Koblizek, Ann Siehoff, and Anthony Pitt
13 Reverse Phase Protein Microarrays and Their Utility in Drug
Development . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 187
Maria Isabella Sereni, Mariaelena Pierobon, Roberto Angioli,
Emanuel F. Petricoin III, and Mitchell J. Frederick
PART V INNOVATIVE CELL CULTURE TECHNIQUES TO MIMIC TISSUE
MICROENVIRONMENTS
14 A Cell Culture System That Mimics Chronic Lymphocytic Leukemia
Cells Microenvironment for Drug Screening and Characterization . . . . . . . . . 217
Alessandro Natoni, Michael O’Dwyer, and Corrado Santocanale
15 Two-Dimensional vs. Three-Dimensional In Vitro Tumor Migration
and Invasion Assays. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 227
Miriam Zimmermann, Carol Box, and Suzanne A. Eccles
16 Tumor Spheroid-Based Migration Assays for Evaluation
of Therapeutic Agents. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 253
Maria Vinci, Carol Box, Miriam Zimmermann,
and Suzanne A. Eccles
17 The Neurosphere Assay Applied to Neural Stem Cells
and Cancer Stem Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 267
Rossella Galli
PART VI GENETICALLY ENGINEERED ANIMAL MODELS TO STUDY GENE
FUNCTIONS
18 Genetically Engineered Animal Models for In Vivo Target
Identification and Validation in Oncology . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281
Gemma Texidó
19 Target Validation in Mice by Constitutive and Conditional RNAi . . . . . . . . . . 307
Aljoscha Kleinhammer, Wolfgang Wurst, and Ralf Kühn
20 In Vivo Target Validation by Inducible RNAi in Human
Xenograft Mouse Models . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 325
Marco Mazzoletti and Gemma TexidóContents xi
PART VII TRANSLATIONAL METHODS TO VALIDATE BIOMARKERS
21 Bright-Field In Situ Hybridization Methods to Discover Gene
Amplifications and Rearrangements in Clinical Samples . . . . . . . . . . . . . . . . . . 341
Hiroaki Nitta and Thomas M. Grogan
22 Combined MicroRNA In Situ Hybridization and Immunohistochemical
Detection of Protein Markers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 353
Boye Schnack Nielsen and Kim Holmstrøm
Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 367
