Probing opioid receptor–ligand interactions by employment of indolizidin‐9‐one amino acid as a constrained Gly²‐Gly³ surrogate in a leucine‐enkephalin mimic

Abstract

The relationship between the conformation and biological activity of Leu-enkephalin was studied using (2S,6R,8S)-9-oxo-8-N-(Boc)amino-1-azabicyclo[4.3.0]nonane-2-carboxylic acid [(2S,6R,8S)-1, I(9)AA] as a constrained Gly(2)-Gly(3) dipeptide surrogate. [I(9)AA](2,3)-Leu-enkephalin 12 was assembled using solid-phase peptide synthesis on Merrifield resin with TBTU as the coupling reagent. The in vitro assays indicated that [I(9)AA](2,3)-Leu-enkephalin 12 exhibited affinities for the mu- and delta-opioid receptors that were three orders of magnitude lower than that of Leu-enkephalin, as well as partial agonist character for both receptors. In in vivo assays for spinal analgesia, the indolizidinone analog 12 showed significantly enhanced duration of action, indicating an increased metabolic stability. Conformational analysis was performed using NMR and CD spectroscopy. The amide temperature coefficients and 3J(NH-CalphaH) coupling constants for 12 could not support a hydrogen-bonded beta-turn structure; however, its CD spectrum indicated a turn conformation. Incorporation of indolizidinone amino acid 1 into Leu-enkephalin thus provided additional support for the importance of a turn conformation for the biological activity of the native peptide.

Conformational constraint in protein ligand design and the inconsistency of binding entropy

“…[8][9][10][11][12] This is closely aligned with another advantage, the potential for establishing the bioactive conformation of the unconstrained parent compound. [13][14][15] In the case of peptide and peptidomimetic structure, additional reported benefits of adding organic constraints or effecting cyclization include increased resistance to proteases and in vivo metabolic stability, [16][17][18][19][20][21][22][23] as well as improved cell membrane permeability [24][25][26][27][28][29] (although the latter may be a contingent rather than a necessary property of cyclic peptides 30,31 ). It is the prospect of increased affinity, however, that has attracted the most research attention over the years.…”

Synthesis of 1‐(m‐Hydroxybenzyl)‐Substituted 1,2,3,4‐Tetrahydroisoquinoline‐3‐carboxylic Acid Derivatives as Opioid Peptide Mimetics − Unexpected Amide Bond Cleavages under Mild Conditions

“…Proposal for the mechanism involved in the cleavage of the TicϪPhe amide bond antagonist (Kb ϭ 1.3 µm). [14] The in vivo analgesic effect was tested in the tail withdrawal test after intrathecal injection in rats. At a dose of 40 µg per rat, full analgesia was reached at 15 min after injection, and lasted for more than 2 h. In comparison to intrathecally administered morphine, compound 15 is about six times less potent.…”

“…At a dose of 40 µg per rat, full analgesia was reached at 15 min after injection, and lasted for more than 2 h. In comparison to intrathecally administered morphine, compound 15 is about six times less potent. [14] …”

“…[13] Antagonist properties were determined in the GTPγS assay on human δ-receptor transfected C6 glioma cells, while the in vivo analgesia was determined in rats after intrathecal injection. [14] (1R,3S)-1-(m-Benzyloxybenzyl)-2-pivaloyl-1,2,3,4-tetrahydroisoquinoline-3-carboxylic Acid (5): tBuLi (1.7  solution in pentane, 7.8 mL, 11.49 mmol) was added under Ar atmosphere at Ϫ78°C to a solution of -2-pivaloyl-1,2,3,4-tetrahydroisquinoline-3-carboxylic acid [5] (4, 1.50 g, 5.75 mmol) and DMPU (1.4 mL, 11.49 mmol) in anhydrous THF (40 mL). After the mixture had been stirred for 1.5 h, a solution of m-benzyloxybenzyl bromide [7] (3.18 g, 11.49 mmol, 2 equiv.)…”

Parallel synthesis and biological evaluation of different sizes of bicyclo[2,3]‐Leu‐enkephalin analogues

“…In order to further understand the relationship between their conformations and their bioactivities, constrained bicyclic dipeptides have been inserted into Leu-enkephalin by conventional synthetic strategies. [22][23][24] However, these solution-phase synthesized bicyclic-Leu-enkephalin analogues are limited in numbers, and the available isomers show limited biological activity. The synthetic methods did not give all possible conformations, due to the lack of a robust methodology for varying the size and stereochemistry of the mimetics.…”