Interaction of tachykinins with their receptors studied with cyclic analogues of substance P and neurokinin B.

Abstract

The activities of two groups of cyclic agonists of substance P (SP) have been studied. The disulfide bridge constraints have been designed on the basis of conformational studies on SP and physalaemin indicating an a-helical structure for the core of these two tachykinins (group I) and a folding of the C-terminal carboxamide towards the side chains of the glutamines 5 and 6 (group II). Only peptides simulating the a-helix present substantial potencies. [Cys3,6JSP is as active as SP in inhibiting '25I-labeled Bolton and Hunter SP-specific binding on rat brain synaptosomes and on dog carotid bioassay, two assays specific for the neurokinin 1 receptor. Moreover, [Cys3,6]SP is as potent as neurokinin B in inhibiting 125I-labeled Bolton and Hunter eledoisin-specific binding on rat cortical synaptosomes as well as in stimulating rat portal vein, two tests specific for the neurokinin 3 receptor. Interestingly, in contrast to neurokinin B, [Cys3,6JSP is a weak agonist of the neurokinin 2 receptor subtype, as evidenced by its binding potency in inhibiting 3H-labeled neurokinin A-specific binding on rat duodenum and in inducing the contractions of the rabbit pulmonary artery, a neurokinin 2-type bioassay. To increase the specificity of the cyclic analogue [Cys3"6]SP positions 8 and 9 were modified. [Cys3'6,Tyr8,Ala9SP is slightly less selective than SP for the neurokinin 1 receptor subtype. [Cys2,5]neurokinin B constitutes a selective cyclic agonist for the neurokinin 3 receptor. The very weak potencies of the peptides from group II indicate that a certain degree of flexibility in the C-terminal moiety is required. Collectively, these results suggest that the neurokinin 1 and neurokinin 3 tachykinin receptors may recognize a similar three-dimensional structure of the core of the tachykinins. Different orientations of the common Cterminal tripeptide may be related to the selectivity for the different receptor subtypes.Conformational analyses of tachykinins (Table 1) were first done on the substance P(5-11) fragment because an earlier report indicated a crucial role of the C-terminal hexapeptides of substance P (SP) and eledoisin (ELE) for spasmogenic activity on the guinea pig ileum (1). Conformational energy calculations have been conducted using both SP and SP(5-11) (2-4). The most stable predicted conformations exhibit a turn around either the Gly-9 and Phe-8 or Phe-7 and Phe-8 residues leading to a spatial proximity of the C-terminal methionine and of the glutamine side chains (2-7). However, we have demonstrated the existence in the rat brain (8, 9) and spinal cord (10) of a specific binding site for SP that requires the whole sequence of SP for full binding potency. This result was further confirmed by several groups (11-13). This observation led us to study the conformational behavior of the undecapeptide in different solvents using NMR and CD spectroscopies. The main features of the conformational model deduced from this study were the flexibility of the

Receptors for Substance P and Related Neurokinins

“…In general, neurokinins, tachykinins and a few analogues, some of them selective for one or the other receptor type, have been used. Pos itive and significant correlations between binding and activity were reported by Laufer et al [1986]; Dion et al [ 1987b]; Ploux et al [1987] and Bergstrom et al [1987a], The comparison performed by Dion et al [1987b] is summarized in table 5. In this study, the binding of BH-SP to rat cerebral cortex synaptosomes was correlated with the biological activity measured in our labora tory on the DCA and the GPI (NK| receptor systems) and on the RPA (NK2).…”

“…Two different neurokinin-binding sites, corre sponding to NK| and NK3, have been identi fied in brain homogenates of various mam mals by several workers [Viger et al, 1983;Torrens et al, 1984Torrens et al, , 1985Cascieri and Liang, 1984;Laufer et al, 1986], while an NK2 site described by Quirion and Dam [1985] appears to be controversial [Ploux et al, 1987;Bergstrom et al, 1987b], NK2 sites have been described in several peripheral tis sues by Buck et al [1984]; Burcher et al, [1986] ; Lee et al [1986], and most recently by Bergstrom et al [1987a]. In general, BH tachykinins have been used in studies pub lished before 1987; more recently tritiated neurokinins have also been tested [Ploux et al, 1987;Bergstrom et al, 1987a, b].…”

“…In another study, Ploux et al [1987] have used a series of cyclic analogues of SP to establish positive and significant correla tions between the binding affinities mea sured with labeled neurokinins (125I-BH-SP, 3H-NKA and l25I-BH-eledoisin) to rat cere bral cortex synaptosomes and the biological activities respectively on the DCA, RPA and RPV.…”

“…New terms how ever should describe biological entities iden tified and characterized in pharmacological and biochemical studies. In this line of thought, a few years ago we proposed the term NK-P to describe the receptor which mediates the relaxation of the dog carotid artery (DCA) in response to SP and related neurokinins, the term NK-A to indicate receptors for NKA which are present in the rabbit pulmonary artery (RPA) [D'Orleans-Juste et al, 1986], the rat vas deferens [Holzer-Petsche et al, 1985; Tousignant et al, 1987] and in other tissues, for instance the human bronchus [Advenier et al, 1987]; the term NK-B to indicate the receptor for NKB which is to be found in the rat portal vein (RPV) [Mastrangelo et al, 1987;Regoli et al, 1987b;Iversen et al, 1987], In recent studies, positive significant correlations have been found when plotting the relative affinities of various neurokinins as measured in biological assays against the affinities evaluated by the binding [Dion et al, 1987b;Ploux et al, 1987], We have therefore suggested ] to abandon the terms SP-P and SP-E and re place them by NK-P, NK-A and NK-B, which indicate definite pharmacological and biochemical entities. At the Substance P Symposium in Montreal, in July 1986, the terms NKj, NK, and NK3 were proposed to indicate receptors for SP, NKA and NKB, respectively (table 1).…”

Synthesis and chemical stability of a disulfide bond in a model cyclic pentapeptide: Cyclo(1,4)‐Cys‐Gly‐Phe‐Cys‐Gly‐OH

“…Presumably, cyclic peptide interaction with endopeptidases is restricted because of the conformational rigidity of the backbone 15,16. Furthermore, the probability of determining the cyclic peptide conformation in solution is higher than that of the linear peptide because of the limited conformers in cyclic peptides 17,18. The solution conformation has been used to determine the bioactive conformation of the peptide and to design a more selective peptide or peptidomimetics.…”