Molecular Pathology in Drug Discovery and Development

MOLECULAR PATHOLOGY IN DRUG DISCOVERY AND DEVELOPMENT......Page 5
CONTENTS......Page 9
Preface......Page 17
Contributors......Page 19
1.1. General Pathology......Page 23
1.3. Molecular Pathology, the Molecular Way......Page 24
1.3.1. Loss of Gene Expression......Page 25
1.3.3. Detection of Pathogens......Page 27
1.3.4. Forensic Identification......Page 28
1.3.5. Protein Changes......Page 29
1.4. Application of Molecular Pathology......Page 30
1.5. Molecular Pathology in Drug Development......Page 32
1.5.1. Most Important Molecular Pathologic Consideration......Page 33
1.6.2. Drug Discovery and Development......Page 34
References......Page 40
2.1. Introduction......Page 43
2.2. History of Chemotherapy and Cancer Drug Discovery......Page 44
2.3. Target-Based Drug Discovery......Page 48
2.4. Utilization of Molecular Pathology in the Discovery of Novel Cancer Targets......Page 51
2.5. Hit Identification and In Vitro Lead Optimization......Page 56
2.6. Implications for Molecular Pathology in Cancer Drug Development and Use......Page 61
2.7. Summary and Future Considerations......Page 63
References......Page 64
3.2. Biomarkers in Clinical Setting and in Early Drug Development......Page 69
3.3. Advantages of Biomarker Implementation......Page 73
3.4. Changing Paradigm in Clinical Drug and Biomarker Development......Page 75
3.5. Promises of Transcriptional Profiling......Page 77
3.6. Biomarker Development and Validation Using Microarray Analysis......Page 81
3.7. Neoadjuvant Chemotherapy as an Intriguing Model for Biomarker Development......Page 84
3.8. Transcriptional Profiling for Identification of Individual Genes as Biomarkers......Page 86
3.9. Transcriptional Profiling for the Definition of Multigene Predictors Using Transcriptional Profiling......Page 88
3.9.1. Multigene Predictors of Chemotherapy Response......Page 91
3.9.2. Multigene Predictor of Response to a Targeted Therapy Agent......Page 93
3.9.3. Novel Approaches to Phase II Clinical Trial Design......Page 94
3.10. Novel Tools for Pathway Analysis......Page 96
3.11. Implementation of Biomarkers into the Clinical Setting......Page 97
References......Page 99
4.1. Introduction......Page 107
4.2. Drug Development......Page 109
4.2.1. Immunohistochemistry in Drug Development......Page 111
4.3.1. Target Validation and Target-Related Safety Concerns......Page 114
4.3.2. Off-Target Effects......Page 118
4.4. Biopharmaceuticals......Page 128
References......Page 131
5.1. Introduction......Page 133
5.2.1. RNA Quality......Page 135
5.2.2. Hybridization Platforms......Page 136
5.3.1. Normalization and Filtering......Page 139
5.3.2. Analyzing for Differences between Experimental Samples......Page 140
5.3.4. Resampling......Page 141
5.3.6. Pathway Analysis......Page 142
5.3.8. Analyzing Relatedness of Gene Expression Profiles......Page 143
5.4.1. Discovery......Page 149
5.4.3. Idiosyncratic Toxicity......Page 150
5.5.1. In Vitro Studies......Page 151
5.5.2. In Vivo Studies......Page 153
5.6.1. Data Submission......Page 154
5.6.2. Voluntary Genomic Data Submission......Page 155
5.7. Summary......Page 156
References......Page 157
6.1. Introduction......Page 165
6.2. Anti-EGFR-Targeted Therapies: The Pharmacodynamic Experience......Page 167
6.3. Molecular Pathology with Small Molecules Gefitinib and Erlotinib......Page 170
6.4. Molecular Pathology with Cetuximab and Other Monoclonal Antibodies to EGFR......Page 174
6.5. Proteasome Inhibitors: Pharmacodynamics on Blood Samples......Page 177
6.6. Pharmacodynamics with Rapamycin Analogs......Page 178
6.7. Second Generation of Targeted Therapies: Multitarget Agents......Page 181
6.8. Conclusions and Perspectives: Phase 0 Clinical Trials......Page 185
References......Page 186
7.2. Molecular Pathology Techniques......Page 191
7.3.1. Selection of Methylating Chemotherapeutic Regimen......Page 194
7.3.2. Epidermal Growth Factor Receptor Expression Is Important Biomarker in Many Tumors......Page 197
7.3.3. Gastrointestinal Stromal Tumor and KIT Evaluation......Page 202
7.3.4. Chronic Myeloid Leukemia and BCR-ABL Protooncogene......Page 204
7.4. Conclusion......Page 206
References......Page 207
8.1. Introduction......Page 217
8.2.1. Mutation Compensation......Page 218
8.2.2. Gene Therapy in Drug Development......Page 224
8.2.3. Immunogene Therapy......Page 227
8.2.4. Oncolytic Viruses......Page 228
8.2.5. Antiangiogenic Therapy......Page 229
8.2.6. Engineered RNA Gene Therapy......Page 230
8.3. Molecular Therapy Clinical Trials......Page 231
References......Page 233
9 MOLECULAR PATHOLOGY: IMMUNOHISTOCHEMISTRY ASSAYS IN DRUG DEVELOPMENT PERFORMED BY A CONTRACT RESEARCH LABORATORY......Page 243
9.1. Immunohistochemistry Is the Technique of Microscopic Visualization of Target Proteins in Tissue Sections Using Specific Antibodies......Page 244
9.2. Basics of the IHC Assay......Page 247
9.2.1. Formalin Fixation and Paraffin Embedding of Tissues......Page 248
9.2.3. Blocking Step......Page 249
9.2.4. Primary Antibody......Page 250
9.2.7. Avidin–Biotin Complex Peroxidase......Page 251
9.2.10. Coverslip......Page 252
9.3. Immunohistochemistry Assay Development......Page 253
9.4. Sending a Study to a Contract Laboratory vs. Running In-house......Page 254
9.4.2. In-house Expertise......Page 255
9.4.3. Throughput Capacity and Range of Services......Page 257
9.4.4. Laboratory Infrastructure......Page 258
9.4.5. Costs......Page 259
9.4.6. Lab Certifications......Page 260
9.4.8. Future Considerations for Partnering......Page 261
9.5.1. Finding the Right Laboratory, Asking the Right Questions......Page 262
9.5.2. What to Look for When Choosing/Evaluating a Laboratory to Outsource IHC Studies......Page 263
9.5.3. Relationship between Sponsor and Contract Laboratory......Page 265
9.5.4. Roles for a Successful Relationship......Page 266
9.6.1. Confidentiality Agreement......Page 269
9.6.3. Project Initiation and Study Protocol......Page 270
9.6.7. Scoring of Study Samples......Page 271
9.6.8. Reporting and Documentation of Outsourced IHC Studies......Page 272
9.6.9. Quality Assurance Review of GLP Studies......Page 273
9.6.11. Data Integration and Follow-up......Page 274
9.7.2. Preclinical Testing......Page 275
9.7.4. Companion Diagnostics......Page 277
References......Page 279
10 QUANTIFICATION OF MOLECULAR PATHOLOGY: COLORIMETRIC IMMUNOHISTOCHEMISTRY......Page 281
10.1. Introduction......Page 282
10.2. Imaging Devices and Systems......Page 287
10.3. Quantification: Introduction to Colorimetric Image Analysis......Page 290
10.3.2. Resolution and Magnification......Page 291
10.3.3. Sensor Size......Page 292
10.3.4. Two Major Camera Technologies Available for Color Image Acquisition......Page 293
10.4.2. Koehler Illumination......Page 294
10.4.3. Additive Property of the Lambert–Beer Law......Page 296
10.4.4. Correction of Chromatic Aberration......Page 297
10.5.1. Looking at Each Pixel as a Set of Linear Equations......Page 298
10.5.2. Shading Correction......Page 300
10.5.3. Solution of Linear Algebraic Equations—Matrices......Page 302
10.5.5. Example......Page 304
10.5.6. Generating Artificial Marker Images......Page 305
10.6. Measuring Information......Page 306
10.6.2. Cell Scoring......Page 307
10.6.3. Multiplexing and Tissue Microarrays......Page 309
10.7. Conclusion......Page 315
References......Page 316
11.1. Introduction......Page 317
11.2.1. Staining Methodology......Page 320
11.2.3. Automated Fluorescence Microscopy and Image Acquisition......Page 323
11.2.4. Automated Image Validation......Page 324
11.2.5. AQUA (Image) Analysis—Producing an AQUA Score......Page 325
11.3. Standardization......Page 330
11.4. Quantification......Page 336
11.5. Localization......Page 343
11.6. Multiparametric Analysis......Page 344
11.7. Application of AQUA Technology to Drug Discovery and Companion Diagnostics......Page 347
11.8. Summary and Conclusions......Page 349
References......Page 350
Index......Page 357