The Melanocortin System: Structure Activity Relationships of Alpha-N-Methylated MT-II Analogues and Mutation Studies of Human Melanocortin Receptor Subtypes 1 and 4
AuthorDedek, Matthew Milan
Keywordsstructure activity relationship
melanocortin 1 receptor
melanocortin 4 receptor
AdvisorHruby, Victor J.
Committee ChairHruby, Victor J.
MetadataShow full item record
PublisherThe University of Arizona.
RightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
AbstractThe melanocortin system regulates various physiological processes including feeding behavior, sexual function, skin pigmentation and photoprotection via five G-protein coupled receptors and several endogenous ligands. There is a need for selective and potent ligands to the human melanocortin receptors (hMCRs) that can chemically resolve these various functions. This thesis presents three studies aimed at refining the understanding of the structural differences between binding pockets of the hMCR subtypes. In the first study α-N-methylated analogues of the non-selective agonist, MT-II, are evaluated for their in vitro function. This study produced the most potent hMC1R selective agonist to date. The following two studies examine the effects of mutations on the biological activity of melanocortin receptor subtypes 1 and 4. Much of the mutation study data is preliminary and requires a demonstration of reproducibility.
Degree ProgramMedical Pharmacology
Degree GrantorUniversity of Arizona
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Synthesis of Novel Amino Acids and Use of Peptides & Peptidomimetics Containing Unnatural Amino Acids for the Development of Selective Melanocortin Peptide Antagonists and for the Study of Melanocortin Receptor SignalingHruby, Victor J; Qu, Hongchang; Mash, Eugene A.; Pyun, Jeffrey; Walker, F. Ann (The University of Arizona., 2007)Unnatural amino acids are indispensible tools, not only for the elucidation of molecular mechanisms during the study of the complicated biological system, but also for the development of novel peptide and protein drugs with better efficacy and lower toxicity. Beta-substituted gamma,delta-unsaturated amino acids have been shown to be an important type of novel amino acid because of the terminal double bond which can be converted to many other functionalities. The methodology for the synthesis of syn-beta-substituted gamma,delta-unsaturated amino acids has been developed. However, there is no satisfactory general method for the synthesis of anti-beta-substituted gamma,delta-unsaturated amino acids. Therefore, a general methodology was developed by using the Eschenmoser-Claisen rearrangement for the synthesis of both racemic and optically active anti-beta-substituted gamma,delta-unsaturated amino acids. This rearrangement is highly diastereoselective and good asymmetric induction was obtained with a relatively small C2-symmetric chiral auxiliary (2R,5R)-dimethylpyrrolidine. In an effort to design peptide antagonists that are selective for human melanocortin 4 receptor, highly constrained trans and cis 4-guanidinium proline derivatives were synthesized and incorporated in various melanotropin analogues designed to mimic the endogenous hMC1,4R selective antagonist hASIP (Agouti Signaling Protein) central loop. Biological assays show that some of these analogues are highly selective for hMC1R and/or hMC4R with partial agonist or antagonist activities due to a new beta-turn structure induced by the presence of the constrained amino acids. According to molecular modeling studies, the lowest energy conformations of these selective analogues resemble the NMR solution structure of the hASIP central loop. Therefore, a new template was developed for the rational design of novel selective melanotropin analogues that may have therapeutic potential. To further understand the molecular mechanisms of hMC4R signaling upon agonist activation, an hMC4R selective nonpeptide agonist THIQ and its fluorescent dye labeled derivatives were needed to compare to peptide agonist MTII with regard to receptor phosphorylation, internalization, etc. An improved synthetic method was developed for the efficient synthesis of THIQ. A method for the synthesis of TRITC labeled THIQ derivatives was also developed.
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Investigation of the Melanocortin Receptor Biased Signaling and the Peptide Drug Design for Melanocortin and Opioid Receptor SystemsHruby, Victor; Liu, Zekun; Mash, Eugene; Montfort, William; Miesfeld, Roger (The University of Arizona., 2022)Melanocortin receptors are important GPCRs controlling different biological functions. As cell membrane integrated receptors, melanocortin receptors have complicated 7-transmembrane domains and delicate internal structures. As membrane protein, melanocortin receptors are highly mobile and their conformations are subjected to change upon ligands binding and induction. The phenomenon of GPCRs’ potential to have different states/conformations is the functional basis of multiple signaling pathways. Ligand types determines the GPCR conformations when they bind and GPCR conformations determine which signaling protein can be recruited, further determining which pathway to initiate. This is the simple accepted mechanism of GPCR biased signaling. In the previous decades, we achieved tremendous milestones in making ligands receptor selective, for example, making MC1R selective agonists, making MC4R selective agonists and so on. However, the receptor subtype selectivity does not mean the ligand is necessary to inflict a single biological signaling outcome: The ligand can be a balanced ligand on this GPCR subtype and activate multiple pathways that this receptor controls. In this respect, making the GPCR ligand go biased signaling is important for the ligand’s anticipated drug effect and for the avoidance of side effects as well. In this dissertation, the author Zekun Liu first uses melanocortin receptor 4(MC4R) as an example to elucidate the mechanism of biased signaling and its influence in MCR peptide drug development. Meanwhile, to test the hypothesis of the relation between receptor structure and function, plasmon waveguide resonance (PWR) research was done. Apart from the biased signaling study, this dissertation also involves the peptide drug development for MC4R and MC5R in introduction of halogenation, beta homo amino acids and N-methylations. Besides the traditional single MCR ligand design, there is also the development of novel conjugation of MC1R agonist and melanoma killing drug as a novel macromolecule to target and kill melanoma. And the thinking of pharmacophore conjugation is also applied to opioid receptor research where Mu and Delta opioid receptors can form a dimer and control specific feedback regulation. In conclusion, this dissertation describes GPCR biased signaling (using MC4R as an example), a couple cases of MCR drug developments, opioid drug developments and DM1 drug development, with the application of multiple scientific approaches including molecular docking, molecular dynamics simulation, solid phase peptide synthesis, PWR, NMR, radiation based cAMP and binding assay for GPCR signaling, and so on. Hopefully, the thinking of the author can produce some inspiration for future peptide research.