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[125I][Tyr3]octreotide labels human somatostatin sst2 and sst5 receptors
Abstract:
Human somatostatin (somatotropin release inhibiting factor = SRIF) receptor subtypes sst2 and sst5 were stably expressed in Chinese hamster lung fibroblast (CCL39) cells. [125I][Tyr3]octreotide labelled with high affinity and in a saturable manner both sst2 (pKd = 9.89+/-0.02, Bmax = 210+/-10 fmol/mg, n = 3) and sst5 sites (pKd = 9.64+/-0.04, Bmax = 920+/-170 fmol/mg, n = 3). The pharmacological profile of sst2 sites established in CCL39 cells using SRIF and various peptide analogues was very similar to that described previously in CHO cells and in human cortex: SRIF14 = SRIF28 > or = seglitide > BIM 23014 = RC 160 > octreotide > CGP 23996 > or = L362,855 > BIM 23052 > L361,301 = cortistatin14 > BIM 23030 > BIM 23056 > cycloantagonist SA. However, peptides classically perceived as sst2 receptor selective (e.g., seglitide, octreotide, vapreotide) showed also high affinity for human sst5 receptors labelled with [125I][Tyr3]octreotide: SRIF28 > seglitide > SRIF14 > L361,301 = octreotide > cortistatin14 = BIM 23014 = BIM 23052 > L362,855 = RC160 > CGP 23996 > BIM 23056 > cycloantagonist SA > BIM 23030. Further radioligand binding studies were performed with [Leu8,D-Trp22,125I-Tyr25]SRIF28 ([125I]LTT-SRIF28) and [125I]CGP 23996. At sst2 receptors, Bmax values determined with [125I][Tyr3]octreotide, [125I]LTT-SRIF28 and [125I]CGP 23996 were in the same range (180-370 fmol/mg). 5'-Guanylyl-imidodiphosphate (GppNHp) displaced all three radioligands to the same extent (85%) and the pharmacological profiles were superimposable. By contrast, at sst5 receptors Bmax values were very different: [125I][Tyr3]octreotide (920 fmol/mg), [125I]CGP 23996 (3530 fmol/mg) and [125I]LTT-SRIF28 (6950 fmol/mg). GppNHp affected [125I][Tyr3]octreotide more than [125I]CGP 23996 binding, whereas [125I]LTT-SRIF28 was much less affected. In addition, the affinity values determined in competition experiments at sst5 receptors, varied markedly; whereas SRIF14, cortistatin14 and SRIF28 showed 2-, 4- and 8-fold differences in affinity at sst5 receptors labelled with [125I][Tyr3]octreotide and [125I]LTT-SRIF28 compounds such as RC160, L363,301, L362,855, octreotide or CGP 23996 showed between 42- and 123-fold lower affinity when sst5 sites were labelled with [125I]LTT-SRIF28. The present data suggest caution to be used when comparing affinity profiles determined in binding studies using different radioligands. In addition, the present results suggest that effects produced by octreotide and related short chain SRIF analogues on hormone release, modulation of tumour growth and central effects may be mediated by either sst2 and/or sst5 receptors.
Insights
This study reveals that common somatostatin (SRIF) receptor ligands bind to both sst2 and sst5 subtypes. Caution is advised when comparing binding affinities using different radioligands for these somatostatin receptors.
Area of Science:
- Endocrinology
- Pharmacology
- Molecular Biology
Background:
- Human somatostatin (SRIF) receptor subtypes sst2 and sst5 play crucial roles in various physiological processes.
- Understanding the binding characteristics of SRIF receptor ligands is essential for developing targeted therapies.
Purpose of the Study:
- To characterize the binding affinity and pharmacological profiles of SRIF receptor subtypes sst2 and sst5.
- To investigate the influence of different radioligands on binding affinity measurements.
- To clarify the receptor mediation of SRIF analogue effects.
Main Methods:
- Stable expression of human sst2 and sst5 receptors in Chinese hamster lung fibroblast (CCL39) cells.
- Radioligand binding assays using [125I][Tyr3]octreotide, [125I]LTT-SRIF28, and [125I]CGP 23996.
- Competition binding experiments with various SRIF analogues.
Main Results:
- Both sst2 and sst5 receptors exhibited high affinity for [125I][Tyr3]octreotide.
- Pharmacological profiles of sst2 sites were consistent across different radioligands.
- Significant variations in Bmax values and affinity profiles were observed for sst5 receptors depending on the radioligand used.
- Peptides considered sst2-selective also showed high affinity for sst5 receptors.
Conclusions:
- Comparing somatostatin receptor affinity profiles requires careful consideration of the radioligand employed.
- The effects of octreotide and related SRIF analogues may be mediated by both sst2 and sst5 receptors.
- These findings have implications for the interpretation of SRIF receptor binding studies and drug development.