Fragment Based Drug Design

Tools, Practical Approaches, and Examples
1. Auflage 2011
ISBN: 978-0-12-381275-9
Verlag: Elsevier Science & Techn.
Format: PDF
Kopierschutz: 1 - PDF Watermark

E-Book, Englisch, Band Volume 493, 645 Seiten, Web PDF

Reihe: Methods in Enzymology

There are numerous excellent reviews on fragment-based drug discovery (FBDD), but there are to date no hand-holding guides or protocols with which one can embark on this orthogonal approach to complement traditional high throughput screening methodologies. This Methods in Enzymology volume offers the tools, practical approaches, and hit-to-lead examples on how to conduct FBDD screens. The chapters in this volume cover methods that have proven to be successful in generating leads from fragments, including chapters on how to apply computational techniques, nuclear magnetic resonance, surface plasma resonance, thermal shift and binding assays, protein crystallography, and medicinal chemistry in FBDD. Also elaborated by experienced researchers in FBDD are sample preparations of fragments, proteins, and GPCR as well as examples of how to generate leads from hits. - Offers the tools, practical approaches, and hit-to-lead examples on how to conduct FBDD screens - The chapters in this volume cover methods that have proven to be successful in generating leads from fragments, including chapters on how to apply computational techniques, nuclear magnetic resonance, surface plasma resonance, thermal shift and binding assays, protein crystallography, and medicinal chemistry in FBDD

Autoren/Hrsg.

Inhaltsverzeichnis

1;Front Cover;1
2;Fragment-Based Drug Design: Tools, Practical Approaches, and Examples;4
3;Copyright Page;5
4;Contents;6
5;Contributors;14
6;Preface;22
7;Contents of previous volume;24
8;Section One: Tools;54
8.1;Chapter 1: Designing a Diverse High-Quality Library for Crystallography-Based FBDD Screening;56
8.1.1;1. Introduction;57
8.1.2;2. Library Requirements for Different Screening Methods;59
8.1.3;3. Library Design for X-Ray Screening;61
8.1.4;4. Implementation;66
8.1.5;5. Conclusions;70
8.1.6;References;72
8.2;Chapter 2: Preparation of Protein Samples for NMR Structure, Function, and Small-Molecule Screening Studies;74
8.2.1;1. Introduction;76
8.2.2;2. Bioinformatics Infrastructure and Target Curation;77
8.2.3;3. Ligation-Independent High-Throughput Cloning and Analytical Scale-Expression Screening;82
8.2.4;4. Midi-Scale Protein Expression and Purification ;91
8.2.5;5. Preparative-Scale Fermentation;98
8.2.6;6. Preparative-Scale Purification;100
8.2.7;7. Salvage Strategies ;102
8.2.8;8. E. coli Single Protein Production System for Isotopic Enrichment;108
8.2.9;9. Conclusions;109
8.2.10;Acknowledgments;110
8.2.11;References;110
8.3;Chapter 3: Key Factors for Successful Generation of Protein-Fragment Structures: Requirement on Protein, Crystals, and Technology;114
8.3.1;1. Introduction;115
8.3.2;2. General Target Properties and Protein Sources in Protein-Fragment Crystallography;116
8.3.3;3. Fragment Properties and the Crystallization Setup;119
8.3.4;4. Quality of Crystallographic Data;129
8.3.5;5. Application of the Free Mounting System and the Picodropper Technology to Improve Ligand Occupancy;131
8.3.6;6. Conclusions;135
8.3.7;Acknowledgments;135
8.3.8;References;135
8.4;Chapter 4: Predicting the Success of Fragment Screening by X-Ray Crystallography;144
8.4.1;1. Introduction;145
8.4.2;2. Fragment Screening and Druggability;146
8.4.3;3. Fragment Screening Campaigns in This Review;148
8.4.4;4. Crystallization Conditions of Fragment Targets;151
8.4.5;5. Solvent Content and Solvent Channels;152
8.4.6;6. Pocket Predictions for Fragment Screening Targets;154
8.4.7;7. Summary;163
8.4.8;Acknowledgments;164
8.4.9;References;164
8.5;Chapter 5: Fragment Screening of Stabilized G-Protein-Coupled Receptors Using Biophysical Methods;168
8.5.1;1. Introduction;169
8.5.2;2. Case Study 1: Biacore Fragment Screen of Adenosine A2A StaR;170
8.5.3;3. Case Study 2: TINS NMR Fragment Screen of beta1-Adrenergic StaR;177
8.5.4;4. Discussion;186
8.5.5;Acknowledgments;188
8.5.6;References;188
8.6;Chapter 6: Using Computational Techniques in Fragment-Based Drug Discovery;190
8.6.1;1. Introduction;191
8.6.2;2. Fragment Library Design;192
8.6.3;3. In Silico Fragment Screening;196
8.6.4;4. Hit Triage;201
8.6.5;5. Hit Follow-Up;203
8.6.6;6. Iteration;205
8.6.7;7. Conclusion;205
8.6.8;References;206
9;Section Two: Practical Approaches;210
9.1;Chapter 7: How to Avoid Rediscovering the Known;212
9.1.1;1. Why FBDD?;213
9.1.2;2. Types of Screen;214
9.1.3;3. Size of a Fragment;215
9.1.4;4. Choice of Fragments;216
9.1.5;5. Fragment Progression;218
9.1.6;6. Conclusion;219
9.1.7;References;220
9.2;Chapter 8: From Experimental Design to Validated Hits: A Comprehensive Walk-Through of Fragment Lead Identification Using Surface Plasmon Resonance;222
9.2.1;1. Introduction: Biophysical Principles of Surface Plasmon Resonance;223
9.2.2;2. Preparing the Instrument;225
9.2.3;3. Surface Preparation;227
9.2.4;4. Target Immobilization;227
9.2.5;5. Buffer and Compound Preparation;234
9.2.6;6. Assay Development;236
9.2.7;7. Aligning the SPR Assay with Crystallography;239
9.2.8;8. Pilot Screening;240
9.2.9;9. Setting Up the Fragment Screen;240
9.2.10;10. Executing the Screen;250
9.2.11;11. Primary Screen Data Reduction;252
9.2.12;12. Data Quality Control and Extraction of the Equilibrium Binding Level;253
9.2.13;13. Scaling and Normalization of Primary Screening Data;255
9.2.14;14. Primary Screen Active Selection;258
9.2.15;15. Collecting Dose-Response Hit Confirmation Data;259
9.2.16;16. Dose-Response Data Reduction and Quality Control;261
9.2.17;17. Global Analysis for KD Determination;262
9.2.18;18. Conclusion;269
9.2.19;Acknowledgments;269
9.2.20;References;270
9.3;Chapter 9: Practical Aspects of NMR-Based Fragment Screening;272
9.3.1;1. Introduction;273
9.3.2;2. Constructing the Fragment Library;274
9.3.3;3. Developing the Screen;280
9.3.4;4. After the Screen;285
9.3.5;5. Conclusions;288
9.3.6;Acknowledgments;289
9.3.7;References;289
9.4;Chapter 10: Binding Site Identification and Structure Determination of Protein-Ligand Complexes by NMR: A Semiautomated Approach;294
9.4.1;1. Introduction;295
9.4.2;2. Automated and Semiautomated Chemical Shift Assignment ;296
9.4.3;3. Identification of Ligand Binding Sites by Chemical Shift Mapping ;303
9.4.4;4. 3D Structure Determination by NMR;311
9.4.5;5. Conclusion;321
9.4.6;References;321
9.5;Chapter 11: Protein Thermal Shifts to Identify Low Molecular Weight Fragments;330
9.5.1;1. Introduction;331
9.5.2;2. Thermal Shift Assays;332
9.5.3;3. Binding Affinity in Thermal Shifts;335
9.5.4;4. Typical Thermal Shift Assay Development;342
9.5.5;5. Dynamic Range of Thermal Shift Assays and Guidelines For a "Significant" Binding Event ;346
9.5.6;Acknowledgment;349
9.5.7;References;349
9.6;Chapter 12: HTS Reporter Displacement Assay for Fragment Screening and Fragment Evolution Toward Leads with Optimized Binding Kinetics, Binding Selectivity, and Thermodynamic Signature;352
9.6.1;1. Introduction;353
9.6.2;2. The Reporter Displacement Assay;354
9.6.3;3. Residence Time and Kinetic Selectivity in Fragment Evolution;358
9.6.4;4. High-Throughput Thermodynamics in Fragment Evolution;364
9.6.5;5. Conclusion;372
9.6.6;Acknowledgments;372
9.6.7;References;372
9.7;Chapter 13: Fragment Screening Purely with Protein Crystallography;374
9.7.1;1. Introduction;375
9.7.2;2. The Primary Library Screen;382
9.7.3;3. The Secondary Library Screen;401
9.7.4;4. Why Screening Purely with X-Ray Structures Works;406
9.7.5;Acknowledgments;407
9.7.6;References;407
9.8;Chapter 14: Computational Approach to De Novo Discovery of Fragment Binding for Novel Protein States;410
9.8.1;1. Introduction;411
9.8.2;2. Protein Modeling;412
9.8.3;3. Fragment Binding in Protein Model: Methods for Free Energy Calculation;416
9.8.4;4. Protein Binding Site Characterization Via Fragment Simulations ;421
9.8.5;5. Fragment-Based Design;428
9.8.6;6. Future Directions;429
9.8.7;Acknowledgments;432
9.8.8;References;432
10;Section Three: Examples;434
10.1;Chapter 15: Lead Generation and Examples: Opinion Regarding How toFollow Up Hits;436
10.1.1;1. Introduction;437
10.1.2;2. Ligand Efficiency;439
10.1.3;3. Four Different Approaches for Converting Fragment Hits to Leads;444
10.1.4;4. Following Up on Hits: Anchor-Based Drug Discovery;451
10.1.5;5. Conclusions;469
10.1.6;References;470
10.2;Chapter 16: Medicinal Chemistry Inspired Fragment-Based Drug Discovery;474
10.2.1;1. Introduction;475
10.2.2;2. Medicinal Chemistry Engagement in Fragment-Based Drug Design;480
10.2.3;3. Case Studies-MACS2b and Ketohexokinase;482
10.2.4;4. Conclusion;497
10.2.5;Acknowledgment;497
10.2.6;References;497
10.3;Chapter 17: Effective Progression of Nuclear Magnetic Resonance-Detected Fragment Hits;500
10.3.1;1. Introduction;501
10.3.2;2. How to Plan for a Successful NMR-Based FBDD Campaign?;501
10.3.3;3. How to Prioritize NMR-Detected Fragment Hits for Lead Generation?;508
10.3.4;4. How to Progress NMR-Detected Fragment Hits into Leads?;510
10.3.5;5. In-house Example of a Successful FBDD Campaign;511
10.3.6;Acknowledgments;519
10.3.7;References;519
10.4;Chapter 18: Advancing Fragment Binders to Lead-Like Compounds Using Ligand and Protein-Based NMR Spectroscopy;522
10.4.1;1. Introduction;523
10.4.2;2. Strategies for Defining Hits;524
10.4.3;3. The Fragment Library and Protein Production;525
10.4.4;4. NMR Follow-Up and Fragment Hit-To-Lead;526
10.4.5;5. Characterizing Binding Modes and Co-Structure Information through Docking;529
10.4.6;6. The Process in an Example;530
10.4.7;7. Summary and Conclusions;537
10.4.8;Acknowledgment;537
10.4.9;References;537
10.5;Chapter 19: Electron Density Guided Fragment-Based Drug Design-A Lead Generation Example;540
10.5.1;1. Introduction;541
10.5.2;2. Electron Density Guided FBDD;541
10.5.3;3. Ketohexokinase;544
10.5.4;4. Ketohexokinase FBDD;546
10.5.5;5. First View of the Solution Activity of the Arylamide Lead ;559
10.5.6;Acknowledgments;559
10.5.7;References;560
10.6;Chapter 20: Experiences in Fragment-Based Lead Discovery;562
10.6.1;1. Introduction;563
10.6.2;2. Maintaining and Enhancing a Fragment Library;565
10.6.3;3. Issues with Different Methods for Fragment Screening;566
10.6.4;4. Hit Rates for Different Classes of Target;574
10.6.5;5. Success Stories in Fragment Evolution;576
10.6.6;6. Thoughts on How to Decide Which Fragments to Evolve;579
10.6.7;7. Final Comments;581
10.6.8;Acknowledgments;582
10.6.9;References;582
10.7;Chapter 21: Fragment Screening of Infectious Disease Targets in a Structural Genomics Environment;586
10.7.1;1. Introduction;587
10.7.2;2. Methods;591
10.7.3;3. Case Studies;596
10.7.4;4. Conclusions;602
10.7.5;Acknowledgments;603
10.7.6;References;603
11;Author Index;610
12;Subject Index;632
13;Color Plates;646