Radiofluorination of non-activated aromatic prosthetic groups for synthesis and evaluation of fluorine-18 labelled ghrelin(1–8) analogues

Childs, Marina D.; Yu, Lihai; Kovacs, Michael S.; Luyt, Leonard G.

doi:10.1039/d1ob01023a

Cited by 1 publication

(6 citation statements)

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“…The binding affinities of all analogues toward the GHSR were evaluated in a competitive radioligand-displacement binding assay against [ 125 I]-ghrelin(1–28) using HEK293 cells transiently transfected with GHSR-eYFP (Table ). As controls, acylated human ghrelin(1–28) and peptide 1 were evaluated and found to possess binding affinity values of 2.48 and 0.09 nM, respectively, which are in agreement with published literature values. , Additionally, all analogues were evaluated in vitro for proteolytic stability in human serum and human liver S9 fraction, the results of which are summarized in Table .…”

Section: Resultsmentioning

confidence: 99%

“… Stability of peptides (A) 1 and (B) 2 in human serum over 24 h. (Part A is adapted from adapted from ref ( 39 ) with permission from The Royal Society of Chemistry) …”

Section: Resultsmentioning

confidence: 99%

“…As controls, acylated human ghrelin(1–28) and peptide 1 were evaluated and found to possess binding affinity values of 2.48 and 0.09 nM, respectively, which are in agreement with published literature values. 36 , 39 Additionally, all analogues were evaluated in vitro for proteolytic stability in human serum and human liver S9 fraction, the results of which are summarized in Table 1 .…”

Section: Resultsmentioning

confidence: 99%

“…One of the drawbacks of using peptides as pharmaceutical agents is their tendency toward rapid blood clearance due to low proteolytic stability. 9 The in vitro stability of peptide 1 in human serum was recently reported, revealing substantial proteolytic degradation and a half-life of 4.7 h. 39 In the present study, as an additional comparison, a control sequence of ghrelin(1−8), with minimal modification compared to the native sequence, was also evaluated by the same method. The peptide [Dpr 3 (n-octanoyl)]ghrelin(1−8) (2) (Figure 1) was synthesized, and its serum stability was analyzed.…”

mentioning

confidence: 99%

“…Subsequent in vivo evaluation of [ 18 F]1 in normal mice revealed rapid blood clearance and high tracer uptake in the liver and intestines. 39 A possible explanation for this observation is poor metabolic stability of the imaging probe. While peptide 1 offers substantially improved binding affinity over natural ghrelin, there has been little investigation into the proteolytic stability of this analogue.…”

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confidence: 99%

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Discovery of Ghrelin(1–8) Analogues with Improved Stability and Functional Activity for PET Imaging

Childs

Chandrabalan

Hodgson

et al. 2023

ACS Pharmacol. Transl. Sci.

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The highest affinity ghrelin-based analogue for fluorine-18 positron emission tomography, [Inp1,Dpr3(6-FN),1Nal4,Thr8]ghrelin(1–8) amide (1), has remarkable subnanomolar receptor affinity (IC50 = 0.11 nM) toward the growth hormone secretagogue receptor 1a (GHSR). However, initial in vivo PET imaging and biodistribution of [18F]1 in mice demonstrated an unfavorable pharmacokinetic profile with rapid clearance and accumulation in liver and intestinal tissue, prompting concerns about the metabolic stability of this probe. The aims of the present study were to examine the proteolytic stability of ghrelin analogue 1 in the presence of blood and liver enzymes, structurally modify the peptide to improve stability without impeding the strong binding affinity, and measure the presently unknown functional activity of ghrelin(1–8) analogues. The in vitro stability and metabolite formation of 1 in human serum and liver S9 fraction revealed a metabolic soft spot between amino acids Leu5 and Ser6 in the peptide sequence. A focused library of ghrelin(1–8) analogues was synthesized and evaluated in a structure–activity–stability relationship study to further understand the structural importance of the residues at these positions in the context of stability and receptor affinity. The critical nature of l-stereochemistry at position 5 was identified and substitution of Ser6 with l-2,3-diaminopropionic acid led to a novel ligand with substantially improved in vitro stability while maintaining subnanomolar GHSR affinity. Despite the highly modified nature of these analogues compared to human ghrelin, ghrelin(1–8) analogues were found to recruit all G protein subtypes (Gαq/11/13/i1/oB) known to associate with GHSR as well as β-arrestins with low micromolar to nanomolar potencies. The study of these analogues demonstrates the ability to balance desirable ligand properties, including affinity, stability, and potency to produce well-rounded candidate molecules for further in vivo evaluation.

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Section: Resultsmentioning

confidence: 99%

“… Stability of peptides (A) 1 and (B) 2 in human serum over 24 h. (Part A is adapted from adapted from ref ( 39 ) with permission from The Royal Society of Chemistry) …”

Section: Resultsmentioning

confidence: 99%