We hypothesized that under chronic pain conditions, up-regulated dynorphin A (Dyn A) interacts with bradykinin receptors (BRs) in the spinal cord to promote hyperalgesia through an excitatory effect, which is opposite to the well known inhibitory effect of opioid receptors. Considering the structural dissimilarity between Dyn A and endogenous BR ligands, bradykinin (BK) and kallidin (KD), this interaction could not be predicted, but allowed us to discover a potential neuroexcitatory target. Well known BR ligands, BK, DALKD, and HOE140 showed different binding profiles at rat brain BRs than that previously reported. These results suggest that neuronal BRs in the rat central nervous system (CNS) may be pharmacologically distinct from those previously defined in non-neuronal tissues. Systematic structure-activity relationship (SAR) study at the rat brain BRs was performed and as a result, a new key structural feature of Dyn A for BR recognition was identified: amphipathicity. NMR studies of two lead ligands, Dyn A-(4-11) 7 and [des-Arg7]-Dyn A-(4-11) 14, which showed the same high binding affinity, confirmed that the Arg residue in position 7, which is known to be crucial for Dyn A’s biological activity, is not necessary, and that a type I β-turn structure at the C-terminal part of both ligands plays an important role in retaining good binding affinities at the BRs. Our lead ligand 14 blocked Dyn A-(2-13) 10-induced hyperalgesic effects and motor impairment in in vivo assays using naïve rats. In a model of peripheral neuropathy, intrathecal (i.th.) administration of ligand 14 reversed thermal hyperalgesia and mechanical hypersensitivity in a dose-dependent manner in nerve-injured rats. Thus ligand 14 may inhibit abnormal pain states by blocking the neuroexcitatory effects of enhanced levels of Dyn A, which are likely to be mediated by BRs in the spinal cord.

Natural residues of the dimeric opioid peptide Biphalin were replaced by the corresponding homo-β 3 amino acids. The derivative 1 containing hβ 3 Phe in place of Phe showed good μ-and δ-receptor affinities ( ) and antinociceptive activity in vivo together with an increased enzymatic stability in human plasma.

An upregulation of the endogenous opioid, dynorphin A, in the spinal cord is seen in multiple experimental models of chronic pain. Recent findings implicate a direct excitatory action of dynorphin A at bradykinin receptors to promote hyperalgesia in nerve injured rats, and its upregulation may promote, rather than counteract, enhanced nociceptive input due to injury. Here we examined a model of inflammatory pain by unilateral injection of Complete Freund's Adjuvant (CFA) into the rat hind paw. Rats exhibited tactile hypersensitivity and thermal hyperalgesia in the inflamed paw by 6 hr after CFA injection, while a significant elevation of prodynorphin transcripts in the lumbar spinal cord was seen at day 3 but not at 6 hr. Thermal hyperalgesia at day 3, but not at 6 hr, after CFA injection was blocked by intrathecal administration of anti-dynorphin antiserum or by bradykinin receptor antagonists. The antihyperalgesic effect of the latter was not due to de novo production of bradykinin or upregulation of spinal bradykinin receptors. These data suggest that elevated spinal dynorphin upon peripheral inflammation mediates chronic inflammatory hyperalgesia. The antihyperalgesic effect of bradykinin receptor antagonists requires the presence of upregulated spinal dynorphin but not of de novo production of bradykinin, supporting our hypothesis that pathological levels of dynorphin may activate spinal bradykinin receptors to mediate inflammatory hyperalgesia.Perspective-This study shows that chronic peripheral inflammation induces a significant upregulation of the endogenous opioid peptide dynorphin. Elevated levels of spinal dynorphin and activation of spinal bradykinin receptors are essential to maintain inflammatory hyperalgesia. The results suggest that blockade of spinal bradykinin receptors may have therapeutic potential in chronic inflammatory pain.

Newly designed bivalent ligands—opioid agonist/NK1-antagonists have been synthesized. The synthesis of new starting materials—carboxy-derivatives of Fentanyl (1a–1c) was developed. These products have been transformed to ‘isoimidium perchlorates’ (2a–c). The new isoimidium perchlorates have been successfully implemented in nucleophilic addition reactions, with L-tryptophan 3,5-bis(trifluoromethyl)benzyl ester to give the target compounds—amides (3a–c). Perchlorates (2a–c) successfully undergo reactions with other nucleophiles such as alcohols, amines or hydrazines. The obtained compound 3b exhibited μ-opioid agonist activity and NK1-antagonist activity and may serve as a useful lead compound for the further design of a new series of opioid agonist/NK1-antagonist compounds.

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