Opioid compounds with mixed mu agonist/delta antagonist properties are expected to be analgesics with low propensity to produce tolerance and dependence. In an effort to strengthen the mu agonist component of the mixed mu agonist/delta antagonist H-Tyr-Tic-Phe-Phe-NH(2) (TIPP-NH(2)), analogues containing structurally modified tyrosine residues in place of Tyr(1) were synthesized. Among the prepared compounds, H-Dmt-Tic-Phe-Phe-NH(2) (DIPP-NH(2); Dmt = 2',6'-dimethyltyrosine) and H-Dmt-TicPsi[CH(2)NH]Phe-Phe-NH(2) (DIPP-NH(2)[Psi]) retained a mixed mu agonist/delta antagonist profile, as determined in the guinea pig ileum and mouse vas deferens assays, whereas H-Tmt-Tic-Phe-Phe-NH(2) (Tmt = N,2',6'-trimethyltyrosine) was a partial mu agonist/delta antagonist and H-Tmt-TicPsi[CH(2)NH]Phe-Phe-NH(2) was a mu antagonist/delta antagonist. DIPP-NH(2)[Psi] showed binding affinities in the subnanomolar range for both mu and delta receptors in the rat brain membrane binding assays, thus representing the first example of a balanced mu agonist/delta antagonist with high potency. In the rat tail flick test, DIPP-NH(2)[Psi] given icv produced a potent analgesic effect (ED(50) = 0.04 microg), being about 3 times more potent than morphine (ED(50) = 0.11 microg). It produced less acute tolerance than morphine but still a certain level of chronic tolerance. Unlike morphine, DIPP-NH(2)[Psi] produced no physical dependence whatsoever upon chronic administration at high doses (up to 4.5 microg/h) over a 7-day period. In conclusion, DIPP-NH(2)[Psi] fulfills to a large extent the expectations based on the mixed mu agonist/delta antagonist concept with regard to analgesic activity and the development of tolerance and dependence.

Opioid peptide analogs consisting entirely of aromatic amino acid residues and contining conformationally restricted phenylalanine derivatives in position 2 of the peptide sequence were synthesized and pharmacologically characterized in vitro. Whereas the existence of at least three major opioid receptor classes (u, 6, and K) is now well-established, the development ofpotent opioid agonists and antagonists with high specificity for each receptor type and of ligands with receptor-specific agonist/antagonist properties continues to be an important goal in opioid pharmacology. The fact that A and 8 opioid receptors differ from one another in their conformational requirements for peptide ligands was first established through comparison of the receptor binding profiles of a cyclic enkephalin analog and its linear correlate (1). This observation led to the realization that conformational restriction of peptides either locally through incorporation of backbone or side-chain conformational constraints at a specific amino acid residue or more globally through peptide cyclizations may often result in improved receptor selectivity. The use of this strategy resulted in a number of conformationally restricted opioid peptide analogs with agonist properties that showed high preference for either 1L or 6 receptors (for a review, see ref.2). It has often been speculated but never demonstrated unambiguously that conformational restriction of peptides in some cases might also reduce or even totally abolish their intrinsic activity ("efficacy") and, thus, may produce partial agonists or antagonists. No examples of opioid peptide analogs with significant antagonist properties as a consequence of conformational restriction have been reported to date. The only opioid-peptide-derived antagonists with reasonable potency described so far were obtained through diallylation of the N-terminal amino group. An enkephalin analog of this type, NN-diallyl-Tyr-Aib-Aib-Phe-Leu-OH (ICI 174,864; Aib = aminoisobutyric acid) (3), has been useful as a 6-selective antagonist.In this paper we report that the tetrapeptide amide H-Tyr-D-Phe-Phe-Phe-NH2 (la) is a potent p-selective opioid agonist. This compound consists entirely of aromatic amino acids that can be conformationally restricted in a number of interesting ways. We show that substitution of the D and L isomers of the conformationally restricted phenylalanine analogs Na-methylphenylalanine (NMePhe) and tetrahydro-3-isoquinoline carboxylic acid (Tic) (Fig. 1) for D-Phe2 in peptide la produced astonishing changes in receptor affinities and intrinsic activities. Most importantly, these structureactivity studies defined a class of potent and selective 6 antagonists, characterized by the N-terminal sequence H-Tyr-Tic-Phe-. MATERIALS AND METHODSPeptide Synthesis. Peptide analogs 1-7 were synthesized by the usual solid-phase technique with N"-t-butyloxycarbonylprotected amino acids and with benzotriazol-1-yl-oxytris-(dimethylamino)phosphonium hexafluorophosphate as coupling agent as described el...

No abstract

A topochemical model to explain the bioactivity of morphiceptin (Tyr1-Pro2-Phe3-Pro4-NH2) was developed by taking account of accessible conformations around rotatable bonds which define relative spatial arrangements of opioid pharmacophores, the amine and phenolic groups of tyrosine and the aromatic ring of phenylalanine, necessary for receptor recognition. For this purpose, 1H-NMR measurements and computer simulations were extensively carried out on 10 stereoisomeric analogs related to morphiceptin: Tyr-Pro-(L and D)-Phe- (L and D)-Pro-NH2; Tyr-Pro-(L and D)-(NMe)Phe-(L and D)-Pro-NH2; Tyr-(NMe)Ala-Phe-D-Pro-NH2; and Tyr-Ala-Phe-D-Pro-NH2. These analogs are structurally close to one another but display various opiate potencies from highly active to inactive. The conformation of each rotatable bond has been specifically identified by measuring accessible space for the analogs, in which the difference in composition is observed in the specific site affecting only the conformation around the target bond. The most interesting characteristic of the model is a requirement of a cis amide bond linking residues 1 and 2. The model also requires the side chains in a trans conformation (chi 1 = 180 degrees) for the Tyr and Phe residues. The distances between the three pharmacophores, d1 (Tyr N to Tyr OH), d2 (Tyr N to the center of the aromatic ring of the third residue), and d3 (Tyr OH to the center of the aromatic ring of the third residue), were found to be approximately 8, approximately 7, and approximately 11-13 A, respectively. This model should aid in pharmaceutical design of peptide and nonpeptide ligands with opioid potencies.

According to the membrane compartment concept the receptor specificity of ligands is based not only on ligand-receptor complementarity but also on specific ligand-membrane interactions. Elaboration of this concept for opioid peptide-receptor interactions had led to the assumption that mu- and delta-receptors are located in anionic and cationic membrane compartments, respectively, and to the prediction that positively charged opioid receptor ligands should display mu-receptor selectivity. To assess the validity of this model, we synthesized a series of dermorphin analogues carrying a net positive charge and tested them in mu- and delta-receptor representative binding assays and bioassays. Some but not all of the prepared compounds showed the receptor-selectivity profile expected on the basis of the membrane compartment concept. In particular, gradual augmentation of the positive charge from 1+ to 3+ in a series of dermorphin-(1-4) tetrapeptide analogues produced an enhancement of mu-receptor affinity and a progressive decrease in delta-receptor affinity, resulting in increasingly higher mu-receptor selectivity. The most selective compound was [D-Arg2,Lys4]dermorphin-(1-4)-amide (DALDA), showing a selectivity ratio (Ki delta/Ki mu = 11,400) more than 10 times higher than that of DAGO (Ki delta /Ki mu = 1050) and, thus, displaying unprecedented mu-receptor specificity. Because of its high positive charge (3+), DALDA may be particularly useful as a very specific agonist for studying peripheral mu-receptor interactions.