Structure–Activity Study of Ghrelin(1–8) Resulting in High Affinity Fluorine-Bearing Ligands for the Ghrelin Receptor

Charron, Carlie L.; Hou, Jinqiang; McFarland, Mark S.; Dhanvantari, Savita; Kovacs, Michael S.; Luyt, Leonard G.

doi:10.1021/acs.jmedchem.7b00164

Cited by 23 publications

(56 citation statements)

References 36 publications

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“…This suggests that LEAP2 and ghrelin share a common ligand-binding pocket on GHSR1a. Previous studies demonstrated that the N-terminal fragment of ghrelin is essential for the binding and activation of GHSR1a [25,26]. The present study suggested that the N-terminal fragment of LEAP2 is important for binding to this receptor.…”

Section: Discussionsupporting

confidence: 65%

Identifying the binding mechanism of LEAP2 to receptor GHSR1a

Wang

Shao

et al. 2019

The FEBS Journal

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Liver‐expressed antimicrobial peptide 2 (LEAP2) is a highly conserved secretory peptide first isolated in 2003. However, its exact biological functions remained elusive until a recent study identified it as an endogenous antagonist for the growth hormone secretagogue receptor (GHSR1a), a G protein‐coupled receptor for which the gastric peptide ghrelin is the endogenous agonist. By tuning the ghrelin–GHSR1a system, LEAP2 has an important function in energy metabolism. In the present study, we first demonstrated that LEAP2 and ghrelin actually bound to GHSR1a in a competitive manner, rather than in a non‐competitive manner as previously reported, by binding assays and activation assays. Subsequently, we demonstrated that the antagonistic function of LEAP2 was drastically affected by the manner of its addition. LEAP2 primarily affected the maximal activation effect when added before ghrelin, whereas it primarily affected half‐maximal effective concentration when added at the same time as ghrelin. Thus, LEAP2 behaved as a competitive antagonist if added at the same time as the agonist and a non‐competitive antagonist if added before the agonist. This unusual property of LEAP2 might be caused by its slow dissociation from receptor GHSR1a. We also found that the N‐terminal fragment of LEAP2 was important for receptor binding. Our present study revealed an antagonistic mechanism for LEAP2, and will facilitate the design of novel antagonists for receptor GHSR1a in future studies.

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Section: Discussionsupporting

confidence: 65%

Identifying the binding mechanism of LEAP2 to receptor GHSR1a

Wang

Shao

et al. 2019

The FEBS Journal

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“…These results correlate to those of the CD spectra, as the binding improves with increased α‐helicity. This relates to earlier findings that have noted that the first 5–8 amino acids are critical for receptor binding . Therefore modifying these positions to incorporate a lactam bridge would interfere with receptor recognition.…”

Section: Resultssupporting

confidence: 56%

“…We propose that an improved ghrelin imaging probe, for fluorescence imaging of ovarian cancer cells or tissue samples through the use of chemi‐staining or confocal microscopy, can be developed through the creation of stapled ghrelin analogues. Previous studies have shown that truncated fluorescently labelled ghrelin analogs, where the fluorescent dye is attached through a Lys ɛ‐amine of a modified ghrelin(1–19), provides a fluorescent probe with comparable GHS‐R1a affinity to that of natural ghrelin(1–28) and was demonstrated to bind to GHS‐R1a in cardiac tissues . This imaging probe was also used to distinguish between prostate cancer and benign hyperplasia in ex vivo prostate tissue and for in vivo biodistribution analysis .…”

Section: Resultsmentioning

confidence: 99%

“…Competitive ligand binding assays were performed in triplicate using HEK293/GHS‐R1a cells and [ 125 I]Ghrelin as the competitive ligand, as previously described . Receptor binding affinities were calculated as IC 50 values as previously described …”

Section: Methodsmentioning

confidence: 99%

“…The first five amino acids have been shown to be key in binding to GHS‐R1a, as ghrelin(1–5) still retains its affinity to the receptor . Recently, ghrelin derivatives have been developed for the purpose of molecular imaging and were determined to have similar GHS‐R1a affinity as that of the endogenous ligand, even when truncated to eight amino acids length . This makes it an ideal candidate for modification for the purpose of imaging GHSR using fluorescence microscopy .…”

Section: Introductionmentioning

confidence: 99%

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Stapled ghrelin peptides as fluorescent imaging probes

et al. 2018

Self Cite

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Fluorescently labelled ghrelin is an effective imaging probe for ex vivo biopsy analysis, in vivo distribution studies, and cell‐based analyses. The objective of our study was to improve the receptor affinity and stability of this ghrelin probe through cyclization, thereby providing a chemical probe with advantages in specificity and sensitivity as compared to immunohistochemical approaches. Truncation of ghrelin to its first 20 essential binding amino acids simplifies chemical synthesis, but reduces the α‐helical content of the peptide, which is important for receptor recognition. To overcome this limitation, we used a “staple scan” to synthesize stable α‐helical cyclic ghrelin(1‐20) analogues using a lactam bridge in either the i, i + 4 or i, i + 7 position. Stapling improved helicity in every case when compared to the linear sequence; however, the binding affinity to the receptor was dependent on the staple position. The peptide with the greatest improvement resulted in a [θ]222/[θ]208 ratio of 0.84, and an IC50 of 7.85 nM. The lead analogue was fluorescently labeled on the C‐terminal lysine of the peptide and microscopy experiments confirmed receptor binding in cells expressing GHS‐R1a. We postulate that the lead stapled peptide can be used as a cancer cell‐specific fluorescent stain with potential research and clinical applications.

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Lymph Node Dissection in Testicular Cancer: The State of the Art and Future Perspectives

Małkiewicz,

Świrkosz,

Lewandowski

et al. 2024

Curr Oncol Rep

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Purpose of Review This narrative review provides a comprehensive overview of the evolving role of retroperitoneal lymph node dissection (RPLND) in the management of testicular cancer (TC). It explores the significance of RPLND as both a diagnostic and therapeutic tool, highlighting its contribution to accurate staging, its impact on oncological outcomes, and its influence on subsequent treatment decisions. Recent Findings RPLND serves as an essential diagnostic procedure, aiding in the precise assessment of lymph node involvement and guiding personalized treatment strategies. It has demonstrated therapeutic value, particularly in patients with specific risk factors and disease stages, contributing to improved oncological outcomes and survival rates. Recent studies have emphasized the importance of meticulous patient selection and nerve-sparing techniques to mitigate complications while optimizing outcomes. Additionally, modern imaging and surgical approaches have expanded the potential applications of RPLND. Summary In the context of TC management, RPLND remains a valuable and evolving tool. Its dual role in staging and therapy underscores its relevance in contemporary urological practice. This review highlights the critical role of RPLND in enhancing patient care and shaping treatment strategies, emphasizing the need for further research to refine patient selection and surgical techniques.

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Structure–Activity Study of Ghrelin(1–8) Resulting in High Affinity Fluorine-Bearing Ligands for the Ghrelin Receptor

Cited by 23 publications

References 36 publications

Identifying the binding mechanism of LEAP2 to receptor GHSR1a

Identifying the binding mechanism of LEAP2 to receptor GHSR1a

Stapled ghrelin peptides as fluorescent imaging probes

Lymph Node Dissection in Testicular Cancer: The State of the Art and Future Perspectives

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