Synthesis and opioid activity of conformationally constrained dynorphin A analogues. 1. Conformational constraint in...

A constrained analogue of the opioid peptide dynorphin A (Dyn A) cyclized in the "message" sequence was designed which may be compatible with the helical conformation proposed by Schwyzer (Biochemistry 1986, 25, 4281-4286) as the conformation Dyn A adopts at kappa opioid receptors. On the basis of molecular modeling with AMBER, we prepared the lactam cyclo-[D-Asp2,Dap5]Dyn A-(1-13)NH2 (1; Dap = alpha, beta-diaminopropionic acid) containing a four-atom bridge between positions 2 and 5 as a possible constraint compatible with an alpha-helix, along with the homologues with five-(2) and six-atom (3) bridges containing Dab (alpha, gamma-diaminobutyric acid) and Orn, respectively, in position 5. All of the cyclic peptide analogues exhibited high binding affinity for both kappa and mu receptors and high potency in the guinea pig ileum (GPI) assay. As ring size increased, a trend in receptor selectivity from slightly kappa selective (compound 1) to nonselective for kappa vs mu (compound 2) to slightly mu selective (compound 3) was observed in the radioligand binding assays. The results in the GPI for antagonism of these peptides by naloxone paralleled the results of the binding assays and indicated that compound 1 preferentially interacted with kappa receptors in this tissue. Novel byproducts were also obtained from the cyclization reactions with HBTU (2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate) and characterized as [D-Asp2,X(Tmg)5]Dyn A-(1-13)NH2 (where X = Dap, Dab, or Orn and Tmg = tetramethylguanidinium). All of the Tmg linear byproducts bound with high affinity to kappa and mu receptors and also exhibited potent agonist activity in the GPI. Circular dichroism spectra of compound 1 and the parent peptide Dyn A-(1-13)NH2 determined in 80% trifluoroethanol at 5 degrees C were consistent with some alpha-helical content in the peptides; comparison of the delta epsilon at 222 nm suggested that compound 1 possessed slightly higher helical content than Dyn A-(1-13)NH2 under these experimental conditions. The cyclic Dyn A analogues 1-3 described here represent the first Dyn A analogues constrained in the "message" sequence with demonstrated high affinity and potency at kappa receptors.