Structure, Synthesis, and Reactivity: Intentional Design Approaches to Agonists for Liver Receptor Homolog-1 and Photochemical Arene Dearomatization Pubblico

Abstract

The nuclear receptor Liver Receptor Homolog-1 (LRH-1) is a high-value therapeutic target for inflammatory bowel diseases due to its ability to combat tissue damage specifically in the gastrointestinal tract. Developing activators (agonists) for LRH-1 would be highly beneficial to people with these autoimmune disorders, like ulcerative colitis, because the agonists would stimulate endogenous anti-inflammatory activity in the gut without global immunosuppression.

Unfortunately, LRH-1 is historically difficult to target with small molecules because of its spacious and largely hydrophobic ligand binding domain that prefers to bind lipids, which cannot be used as pharmaceuticals because of their poor metabolic profiles and low potency.

Here, crystallographic studies have driven the design of synthetic targets, each of which interrogates a specific question relating to LRH-1 activation by small molecules. By combining these intentional synthetic efforts with pointed biological feedback, we have successfully developed the only small molecule agonists that activate LRH-1 with nanomolar potencies. Additionally, we have developed a synthetic strategy to install phospholipid-mimicking groups that act synergistically with the core of our agonists to result in the most activating and potent LRH-1 ligands to date. These activators indeed bind to LRH-1 exactly as designed and are strikingly effective at restoring colitis-induced intestinal inflammation in a mouse model of colitis. 

Table of Contents

PART 1: RATIONAL DESIGN OF LRH-1 AGONISTS

Using LRH-1 to locally combat inflammation 1

CHAPTER 1: IDENTIFYING A TARGETING STRATEGY FOR LRH-1

1.1 Two Modes of LRH-1 Activation 5

1.2 Crystal Structures of RJW100 bound to LRH-1 8

1.3 Identifying a Synthetic Ligand Design Strategy for LRH-1 16

1.4 Supporting Information 17

CHAPTER 2: PHOSPHOLIPID-MIMICKING CHIMERAS AS LRH-1 AGONISTS

2.1 Optimizing a synthetic route to RJW100-DLPC hybrid precursors 34

2.3 Design and evaluation of an RJW100-DLPC chimera 37

2.3 Optimization of chimeras via simplification 39

2.4 Conclusions 44

2.5 Supporting Information. 44

CHAPTER 3: MECHANISM OF ACTIVATION AND THERAPEUTIC EFFECTS OF LRH-1 AGONIST 10CA

3.1 Phospholipid Signaling Axes 87

3.2 The Apparent Importance of Linker-Length on Activation 88

3.3 Crystal Structures of PL Mimetics Bound to LRH-1 90

3.4 Surface Residue Contacts Drive Binding and Activation by PL Mimetics 91

3.5 Chimeras Affect Coregulator Association Signatures and Stabilize AFS 93

3.6 Efficacy of 10CA in Murine Models of Colitis 96

3.6 Conclusions 99

3.7 Supporting Information 99

CHAPTER 4: LRH-1-RJW100 CRYSTAL STRUCTURES ENABLE A COMPREHENSIVE STRUCTURE-ACTIVITY RELATIONSHIP

4.1 Synthesizing a Library 112

4.2 The First Low Nanomolar LRH-1 Agonist 115

4.3 Contacts that Drive LRH-1 Activation by 6N 117

4.4 6N Stabilizes AFS, Strengthens Allosteric Signaling, and Promotes Coactivator Recruitment 122

4.5 Evaluation of 6N in Murine Intestinal Organoids 126

4.6 Conclusions 128

4.7 Supporting Information 130

CHAPTER 5: OPTIMIZATION OF LEAD AGONISTS

5.1 Strategy 196

5.2 Carboxylate Replacement 196

5.3 6N-10CA Hybrid 199

5.4 Supporting information 202

CHAPTER 6: DESIGNING NEW CHEMICAL TOOLS TO STUDY THE HUMAN NR5A RECEPTORS LRH-1 AND SF-1

6.1 NR5A Nuclear Receptors 217

6.1 Development of a Fluorescence Polarization Assay to Detect Binding 218

6.3 Evaluation of the fluorescent probe 6N-FAM 220

6.4 Utilization of 6N-FAM to Detect Phospholipid Binding to NR5A Receptors 222

6.5 Synthetic Agonist Binding Affinity Correlation with Stability and Activity 224

6.6 FP Assay Validation by Comparing to EMSA and SPR 225

6.7 Conclusions 227

6.8 Supporting Information 228

LRH-1: SUMMARY AND FUTURE DIRECTIONS

PART 2: ARENE DEAROMATIZATION

A Call for Improved Dearomative Technologies. 248

Principles of Photochemistry 250

CHAPTER 7: DEAROMATIZATION VIA RADICAL HYDROARYLATION

7.1 Our Aromatic Functionalization Space 254

7.2 Dearomative Hydroarylation Design Strategy 256

7.3 Key Differences and Assets to Photochemical Dearomative Hydroarylation 259

7.4 The Scope 260

7.5 Selectivity Rationale 263

7.6 Comparative synthesis of a pharmaceutical candidate 266

7.7 Conclusions 267

7.8 Supporting Information 267

CHAPTER 8: PHOTOCHEMICAL DEAROMATIZATION OF BENZENOIDS

8.1 Photochemical Activation of Monocyclic Aromatics 327

8.2 Proof-of-concept and preliminary scope 328

8.3 Proposed Mechanism 329

8.4 Probing Mechanism with Experimentation 331

8.5 Supporting Information 335

PHOTOCHEMICAL DEAROMATIZATION: SUMMARY AND FUTURE DIRECTIONS

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School	Laney Graduate School
Department	Chemistry
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Chemistry, Organic
Parola chiave	agonist nuclear receptor LRH-1 colitis dearomatization phospholipid photochemistry
Committee Chair / Thesis Advisor	Jui, Nathan, Emory University
Committee Members	Liebeskind, Lanny, Emory University Liotta, Dennis, Emory University Ortlund, Eric, Emory University

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