Fragment-based Drug Discovery

Daniel A. Erlanson is the co-founder and President of Carmot Therapeutics, Inc., which is developing fragment-based approaches to address unmet needs in drug discovery. Prior to Carmot, Dr. Erlanson worked in medicinal chemistry and technology development at Sunesis Pharmaceuticals, which he joined at the company's inception. Before Sunesis, he was an NIH postdoctoral fellow with Dr. James A. Wells at Genentech. Dr. Erlanson earned his Ph.D. in chemistry from Harvard University in the laboratory of Gregory L. Verdine and his BA in chemistry from Carleton College. He edits a blog devoted to fragment-based drug discovery, Practical Fragments. Wolfgang Jahnke is a Director and Leading Scientist at the Novartis Institutes for Biomedical Research in Basel, Switzerland. His major interests are Structural Biophysics and Fragment-based Drug Discovery. He has received several honors, among them the Industrial Investigator Award from the Swiss Chemical Society, and several Novartis-internal Awards. Dr. Jahnke received his PhD from the TU Munchen, working with Horst Kessler on the development and application of novel NMR methods. Prior to joining Novartis, he worked with Peter Wright at the Scripps Research Institute in La Jolla.

• Contributors XVPreface XXIA Personal Foreword XXIIIPart I The Concept of Fragment-based Drug Discovery 11 The Role of Fragment-based Discovery in Lead Finding 3Roderick E. Hubbard1.1 Introduction 31.2 What is FBLD? 41.3 FBLD: Current Practice 51.3.1 Using Fragments: Conventional Targets 51.3.2 Using Fragments: Unconventional Targets 131.4 What do Fragments Bring to Lead Discovery? 141.5 How did We Get Here? 161.5.1 Evolution of the Early Ideas and History 161.5.2 What has Changed Since the First Book was Published in 2006? 161.6 Evolution of the Methods and Their Application Since 2005 191.6.1 Developments in Fragment Libraries 211.6.2 Fragment Hit Rate and Druggability 221.6.3 Developments in Fragment Screening 231.6.4 Ways of Evolving Fragments 231.6.5 Integrating Fragments Alongside Other Lead-Finding Strategies 231.6.6 Fragments Can be Selective 241.6.7 Fragment Binding Modes 251.6.8 Fragments, Chemical Space, and Novelty 271.7 Current Application and Impact 271.8 Future Opportunities 28References 292 Selecting the Right Targets for Fragment-Based Drug Discovery 37Thomas G. Davies, Harren Jhoti, Puja Pathuri, and Glyn Williams2.1 Introduction 372.2 Properties of Targets and Binding Sites 392.3 Assessing Druggability 412.4 Properties of Ligands and Drugs 422.5 Case Studies 432.5.1 Case Study 1: Inhibitors of Apoptosis Proteins (IAPs) 442.5.2 Case Study 2: HCV-NS3 462.5.3 Case Study 3: PKM2 472.5.4 Case Study 4: Soluble Adenylate Cyclase 492.6 Conclusions 50References 513 Enumeration of Chemical Fragment Space 57Jean-Louis Reymond, Ricardo Visini, and Mahendra Awale3.1 Introduction 573.2 The Enumeration of Chemical Space 583.2.1 Counting and Sampling Approaches 583.2.2 Enumeration of the Chemical Universe Database GDB 583.2.3 GDB Contents 593.3 Using and Understanding GDB 613.3.1 Drug Discovery 613.3.2 The MQN System 623.3.3 Other Fingerprints 633.4 Fragments from GDB 653.4.1 Fragment Replacement 653.4.2 Shape Diversity of GDB Fragments 663.4.3 Aromatic Fragments from GDB 683.5 Conclusions and Outlook 68Acknowledgment 69References 694 Ligand Efficiency Metrics and their Use in Fragment Optimizations 75György G. Ferenczy and György M. Keserû4.1 Introduction 754.2 Ligand Efficiency 754.3 Binding Thermodynamics and Efficiency Indices 784.4 Enthalpic Efficiency Indices 814.5 Lipophilic Efficiency Indices 834.6 Application of Efficiency Indices in Fragment-Based Drug Discovery Programs 884.7 Conclusions 94References 95Part II Methods and Approaches for Fragment-based Drug Discovery 995 Strategies for Fragment Library Design 101Justin Bower, Angelo Pugliese, and Martin Drysdale5.1 Introduction 1015.2 Aims 1025.3 Progress 1025.3.1 BDDP Fragment Library Design: Maximizing Diversity 1035.3.2 Assessing Three-Dimensionality 1035.3.3 3DFrag Consortium 1045.3.4 Commercial Fragment Space Analysis 1055.3.5 BDDP Fragment Library Design 1085.3.6 Fragment Complexity 1115.3.6.1 Diversity-Oriented Synthesis-Derived Fragment-Like Molecules 1135.4 Future Plans 1145.5 Summary 1165.6 Key Achievements 116References 1166 The Synthesis of Biophysical Methods In Support of Robust Fragment-Based Lead Discovery 119Ben J. Davis and Anthony M. Giannetti6.1 Introduction 1196.2 Fragment-Based Lead Discovery on a Difficult Kinase 1216.3 Application of Orthogonal Biophysical Methods to Identify and Overcome an Unusual Ligand: Protein Interaction 1276.4 Direct Comparison of Orthogonal Screening Methods Against a Well-Characterized Protein System 1316.5 Conclusions 135References 1367 Differential Scanning Fluorimetry as Part of a Biophysical Screening Cascade 139Duncan E. Scott, Christina Spry, and Chris Abell7.1 Introduction 1397.2 Theory 1407.2.1 Equilbria are Temperature Dependent 1407.2.2 Thermodynamics of Protein Unfolding 1427.2.3 Exact Mathematical Solutions to Ligand-Induced Thermal Shifts 1437.2.4 Ligand Binding and Protein Unfolding Thermodynamics Contribute to the Magnitude of Thermal Shifts 1457.2.5 Ligand Concentration and the Magnitude of Thermal Shifts 1477.2.6 Models of Protein Unfolding Equilibria and Ligand Binding 1487.2.7 Negative Thermal Shifts and General Confusions 1507.2.8 Lessons Learnt from Theoretical Analysis of DSF 1517.3 Practical Considerations for Applying DSF in Fragment-Based Approaches 1527.4 Application of DSF to Fragment-Based Drug Discovery 1547.4.1 DSF as a Primary Enrichment Technique 1547.4.2 DSF Compared with Other Hit Identification Techniques 1597.4.3 Pursuing Destabilizing Fragment Hits 1667.4.4 Lessons Learnt from Literature Examples of DSF in Fragment-Based Drug Discovery 1687.5 Concluding Remarks 169Acknowledgments 169References 1708 Emerging Technologies for Fragment Screening 173Sten Ohlson and Minh-Dao Duong-Thi8.1 Introduction 1738.2 Emerging Technologies 1758.2.1 Weak Affinity Chromatography 1758.2.1.1 Introduction 1758.2.1.2 Theory 1778.2.1.3 Fragment Screening 1798.2.2 Mass Spectrometry 1858.2.2.1 Introduction 1858.2.2.2 Theory 1868.2.2.3 Applications 1868.2.3 Microscale Thermophoresis 1878.2.3.1 Introduction 1878.2.3.2 Theory 1898.2.3.3 Applications 1898.3 Conclusions 189Acknowledgments 191References 1919 Computational Methods to Support Fragment-based Drug Discovery 197Laurie E. Grove, Sandor Vajda, and Dima Kozakov9.1 Computational Aspects of FBDD 1979.2 Detection of Ligand Binding Sites and Binding Hot Spots 1989.2.1 Geometry-based Methods 1999.2.2 Energy-based Methods 2019.2.3 Evolutionary and Structure-based Methods 2029.2.4 Combination Methods 2029.3 Assessment of Druggability 2039.4 Generation of Fragment Libraries 2059.4.1 Known Drugs 2069.4.2 Natural Compounds 2079.4.3 Novel Scaffolds 2089.5 Docking Fragments and Scoring 2099.5.1 Challenges of Fragment Docking 2099.5.2 Examples of Fragment Docking 2109.6 Expansion of Fragments 2129.7 Outlook 214References 21410 Making FBDD Work in Academia 223Stacie L. Bulfer, Frantz Jean-Francois, and Michelle R. Arkin10.1 Introduction 22310.2 How Academic and Industry Drug Discovery Efforts Differ 22510.3 The Making of a Good Academic FBDD Project 22610.4 FBDD Techniques Currently Used in Academia 22810.4.1 Nuclear Magnetic Resonance 22910.4.2 X-Ray Crystallography 23010.4.3 Surface Plasmon Resonance/Biolayer Interferometry 23110.4.4 Differential Scanning Fluorimetry 23210.4.5 Isothermal Titration Calorimetry 23210.4.6 Virtual Screening 23210.4.7 Mass Spectrometry 23310.4.7.1 Native MS 23310.4.7.2 Site-Directed Disulfide Trapping (Tethering) 23410.4.8 High-Concentration Bioassays 23410.5 Project Structures for Doing FBDD in Academia 23510.5.1 Targeting p97: A Chemical Biology Consortium Project 23510.5.2 Targeting Caspase-6: An Academic–Industry Partnership 23610.6 Conclusions and Perspectives 239References 24011 Site-Directed Fragment Discovery for Allostery 247T. Justin Rettenmaier, Sean A. Hudson, and James A. Wells11.1 Introduction 24711.2 Caspases 24911.2.1 Tethered Allosteric Inhibitors of Executioner Caspases-3 and -7 24911.2.2 Tethering Inflammatory Caspase-1 25011.2.3 Tethered Allosteric Inhibitors of Caspase-5 25111.2.4 General Allosteric Regulation at the Caspase Dimer Interface 25211.2.5 Using Disulfide Fragments as “Chemi-Locks” to Generate Conformation-Specific Antibodies 25311.3 Tethering K-Ras(G12C) 25411.4 The Master Transcriptional Coactivator CREB Binding Protein 25611.4.1 Tethering to Find Stabilizers of the KIX Domain of CBP 25611.4.2 Dissecting the Allosteric Coupling between Binding Sites on KIX 25711.4.3 Rapid Identification of pKID-Competitive Fragments for KIX 25811.5 Tethering Against the PIF Pocket of Phosphoinositide-Dependent Kinase 1 (PDK1) 25911.6 Tethering Against GPCRs: Complement 5A Receptor 26111.7 Conclusions and Future Directions 263References 26412 Fragment Screening in Complex Systems 267Miles Congreve and John A. Christopher12.1 Introduction 26712.2 Fragment Screening and Detection of Fragment Hits 26812.2.1 Fragment Screening Using NMR Techniques 27012.2.2 Fragment Screening Using Surface Plasmon Resonance 27112.2.3 Fragment Screening Using Capillary Electrophoresis 27212.2.4 Fragment Screening Using Radioligand and Fluorescence-Based Binding Assays 27312.2.5 Ion Channel Fragment Screening 27512.3 Validating Fragment Hits 27612.4 Fragment to Hit 27912.4.1 Fragment Evolution 28012.4.2 Fragment Linking 28112.5 Fragment to Lead Approaches 28112.5.1 Fragment Evolution 28212.5.2 Fragment Linking 28412.6 Perspective and Conclusions 285Acknowledgments 287References 28713 Protein-Templated Fragment Ligation Methods: Emerging Technologies in Fragment-Based Drug Discovery 293Mike Jaegle, Eric Nawrotzky, Ee Lin Wong, Christoph Arkona, and Jörg Rademann13.1 Introduction: Challenges and Visions in Fragment-Based Drug Discovery 29313.2 Target-Guided Fragment Ligation: Concepts and Definitions 29413.3 Reversible Fragment Ligation 29513.3.1 Dynamic Reversible Fragment Ligation Strategies 29513.3.2 Chemical Reactions Used in Dynamic Fragment Ligations 29613.3.3 Detection Strategies in Dynamic Fragment Ligations 29913.3.4 Applications of Dynamic Fragment Ligations in FBDD 30113.4 Irreversible Fragment Ligation 31113.4.1 Irreversible Fragment Ligation Strategies: Pros and Cons 31113.4.2 Detection in Irreversible Fragment Ligation 31113.4.3 Applications of Irreversible Fragment Ligations in FBDD 31313.5 Fragment Ligations Involving Covalent Reactions with Proteins 31613.6 Conclusions and Future Outlook: How Far did We Get and What will be Possible? 319References 320Part III Successes from Fragment-based Drug Discovery 32714 BACE Inhibitors 329Daniel F. Wyss, Jared N. Cumming, Corey O. Strickland, and Andrew W. Stamford14.1 Introduction 32914.2 FBDD Efforts on BACE1 33314.2.1 Fragment Hit Identification, Validation, and Expansion 33314.2.2 Fragment Optimization 33314.2.3 From a Key Pharmacophore to Clinical Candidates 34014.3 Conclusions 346References 34615 Epigenetics and Fragment-Based Drug Discovery 355Aman Iqbal and Peter J. Brown15.1 Introduction 35515.2 Epigenetic Families and Drug Targets 35715.3 Epigenetics Drug Discovery Approaches and Challenges 35815.4 FBDD Case Studies 35915.4.1 BRD4 (Bromodomain) 36015.4.2 EP300 (Bromodomain) 36315.4.3 ATAD2 (Bromodomain) 36415.4.4 BAZ2B (Bromodomain) 36415.4.5 SIRT2 (Histone Deacetylase) 36515.4.6 Next-Generation Epigenetic Targets: The “Royal Family” and Histone Demethylases 36615.5 Conclusions 367Abbreviations 368References 36816 Discovery of Inhibitors of Protein–Protein Interactions Using Fragment-Based Methods 371Feng Wang and Stephen W. Fesik16.1 Introduction 37116.2 Fragment-Based Strategies for Targeting PPIs 37216.2.1 Fragment Library Construction 37216.2.2 NMR-Based Fragment Screening Methods 37316.2.3 Structure Determination of Complexes 37416.2.4 Structure-Guided Hit-to-Lead Optimization 37516.3 Recent Examples from Our Laboratory 37616.3.1 Discovery of RPA Inhibitors 37716.3.2 Discovery of Potent Mcl-1 Inhibitors 37816.3.3 Discovery of Small Molecules that Bind to K-Ras 37916.4 Summary and Conclusions 382Acknowledgments 383References 38417 Fragment-Based Discovery of Inhibitors of Lactate Dehydrogenase A 391Alexander L. Breeze, Richard A. Ward, and Jon Winter17.1 Aerobic Glycolysis, Lactate Metabolism, and Cancer 39117.2 Lactate Dehydrogenase as a Cancer Target 39217.3 “Ligandability” Characteristics of the Cofactor and Substrate Binding Sites in LDHA 39417.4 Previously Reported LDH Inhibitors 39517.5 Fragment-Based Approach to LDHA Inhibition at AstraZeneca 39817.5.1 High-Throughput Screening Against LDHA 39817.5.2 Rationale and Strategy for Exploration of Fragment-Based Approaches 39917.5.3 Development of Our Biophysical and Structural Biology Platform 40017.5.4 Elaboration of Adenine Pocket Fragments 40417.5.5 Screening for Fragments Binding in the Substrate and Nicotinamide Pockets 40517.5.6 Reaching out Across the Void 40717.5.7 Fragment Linking and Optimization 40817.6 Fragment-Based LDHA Inhibitors from Other Groups 41017.6.1 Nottingham 41017.6.2 Ariad 41317.7 Conclusions and Future Perspectives 417References 41918 FBDD Applications to Kinase Drug Hunting 425Gordon Saxty18.1 Introduction 42518.2 Virtual Screening and X-ray for PI3K 42618.3 High-Concentration Screening and X-ray for Rock1/2 42718.4 Surface Plasmon Resonance for MAP4K4 42818.5 Weak Affinity Chromatography for GAK 42918.6 X-ray for CDK 4/6 43018.7 High-Concentration Screening, Thermal Shift, and X-ray for CHK2 43218.8 Virtual Screening and Computational Modeling for AMPK 43318.9 High-Concentration Screening, NMR, and X-ray FBDD for PDK1 43418.10 Tethering Mass Spectometry and X-ray for PDK1 43518.11 NMR and X-ray Case Study for Abl (Allosteric) 43618.12 Review of Current Kinase IND’s and Conclusions 437References 44219 An Integrated Approach for Fragment-Based Lead Discovery: Virtual, NMR, and High-Throughput Screening Combined with Structure-Guided Design. Application to the Aspartyl Protease Renin 447 Simon Rüdisser, Eric Vangrevelinghe, and Jürgen Maibaum19.1 Introduction 44719.2 Renin as a Drug Target 44919.3 The Catalytic Mechanism of Renin 45119.4 Virtual Screening 45219.5 Fragment-Based Lead Finding Applied to Renin and Other Aspartyl Proteases 45519.6 Renin Fragment Library Design 46419.7 Fragment Screening by NMR T1ρ Ligand Observation 46919.8 X-Ray Crystallography 47319.9 Renin Fragment Hit-to-Lead Evolution 47519.10 Integration of Fragment Hits and HTS Hits 47619.11 Conclusions 479References 480Index 487