Browsing by Subject "SAR"

Now showing 1 - 3 of 3
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    Discovery of small molecule modulators of protein-protein interactions by FRET-based high-throughput screening and structure-based drug design
    (2019-08) Lo, Chih Hung
    Protein–protein interactions (PPIs) are of pivotal importance in the regulation of biological systems and are consequently implicated in the development of disease states. Here, we investigated two classes of protein, including a transmembrane protein (tumor necrosis factor receptor 1 (TNFR1)) and intrinsically disordered proteins (tau and huntingtin (HTT)), which are implicated in autoimmune diseases and neurodegenerative diseases respectively. Receptor-specific inhibition of TNFR1 signaling is a highly sought after strategy for treatment of inflammatory diseases such as rheumatoid arthritis. In this study, we investigated the structure-function relationship of TNFR1 by engineering a TNFR1 fluorescence resonance energy transfer (FRET) biosensor to monitor the structural and conformational changes of the receptor. We have also shown using small-molecule tool compounds, that the disruption of receptor-receptor interactions (competitive inhibition) and perturbation of the receptor conformational dynamics (allosteric inhibition) are both feasible approaches to inhibit TNFR1 signaling. We have also made a major discovery showing that long-range structural couplings, between TNFR1 membrane distal and proximal domains, mediated through the ligand-binding loop, determine the conformational states of the receptor that act as a molecular switch in receptor function. In addition to deepening the understanding of a novel mechanism in TNF receptor activation, we have optimized a lead compound through medicinal chemistry by improving its potency by more than sixty-fold to the nanomolar range, thereby advancing therapeutic developments in these clinically important targets. The heterogeneity of tau and HTT pathology is one of the major challenges that plagues current clinical trials, hence impeding the discovery of a cure for Alzheimer’s disease (AD) and Huntington’s disease (HD). We have engineered novel FRET biosensors of these proteins to target the ensemble of heterogeneous protein oligomers or aggregates in cells. The biosensors are not only capable of monitoring oligomer conformations, but can also be used as a high-throughput screening platform. Using these technologies, we have discovered small-molecule inhibitors of tau oligomerization or HTT aggregation that rescue cell cytotoxicity with nanomolar potency.
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    A Solution Towards Weight Management: Discovery, Design, Synthesis, and Characterization of Melanocortin Ligands
    (2021-05) Koerperich, Zoe
    The melanocortin receptors are a family of rhodopsin-like (family A) G protein-coupled receptors (GPCRs) that are known to have a variety of conserved physiological functions. The following chapters of this dissertation explore four different projects designed to examine this receptor family through: 1) synthesis of novel ligands using traditional structure activity relationships (SAR), 2) use of a novel mixture based positional scanning approach for ligand discovery, 3) synthesis of endogenous ligand mimetics with established scaffolds, and 4) use of various functional, binding affinity, and proximity assays for in vivo evaluation. Chapter 1 serves as an introduction to the melanocortin receptor (MCR) family as well as ligands (endogenous and synthetic) that are well known in the field. This chapter also introduces the concept of selectivity between the five melanocortin receptors, particularly the centrally expressed melanocortin-3 receptor (MC3R) and melanocortin-4 receptor (MC4R). Chapter 2 describes all the materials and methods used in Chapters 3-6 for peptide synthesis, characterization, and pharmacological evaluation. Chapter 3 describes the exploration of single nucleotide polymorphisms in the antagonist signaling molecule Agouti-related Protein (AgRP) by incorporating them into a modified AgRP octapeptide macrocyclic scaffold. Chapter 4 investigates the roles of ligands in protein:protein interactions, specifically those of the MC4R and the kappa opioid receptor (KOR). Chapter 5 demonstrates the complexity involved in designing a selective ligand through focused tetrapeptide SAR and functional assessment in a cAMP accumulation assay. Chapter 6 outlines the discovery of a completely novel MC3R antagonist scaffold using a mixture based positional scanning approach. Lastly, Chapter 7 broadly summarizes the dissertation presented and reflects upon knowledge and experienced gained, advancements to the field, and potential future directions for the work presented in this thesis.
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    Tetrapeptide Melanocortin Agonist Ligands Exploring Selectivity of the mMC3R Using DPhe Substitutions in the Ac-His-Arg-DPhe-Tic-NH2 Scaffold
    (2018-03) Schlasner, Katherine
    There are five melanocortin receptors (MC1R-MC5R) that primarily activate the signaling pathway for adenylate cyclase. While both the MC3R and MC4R are hypothesized to be involved in regulating energy homeostasis, the MC3R functionality has been elusive to characterize due to the lack of MC3R-selective ligands. The melanocortin system may be a target for treating obesity or cachexia. When centrally delivered through intracerebroventricular administration, agonists decrease food intake, while antagonists increase food intake. Previous weight management therapies have focused on targeting the MC4R, though off-target cardiovascular effects may limit the clinical utility of these ligands. Therefore, obtaining a MC3R selective compound may allow for a weight regulation therapy that will not have the same cardiovascular liabilities. MC4R-selective ligands have been heavily investigated, however MC3R-selective ligands have largely gone unexplored. A mixture-based positional scan was conducted to generate scaffolds with MC3R selectivity. The lead compound from the study was Ac-His-Arg-(pI)DPhe-Tic-NH2, an MC3R agonist and an MC4R antagonist. The scaffold reverses the sequence of arginine and phenylalanine residues in the His-Phe-Arg-Trp conserved sequence found in endogenous melanocortin ligands1. This work presents a follow-up study investigating the position of the phenylalanine residue in efforts to create a MC3R selective agonist with decreased MC4R antagonist activity.