PnPP‐19, a spider toxin peptide, induces peripheral antinociception through opioid and cannabinoid receptors and inhibition of neutral endopeptidase

• Published in: British journal of pharmacology Vol. 173; no. 9; pp. 1491 - 1501
• Main Authors: Freitas, A C N, Pacheco, D F, Machado, M F M, Carmona, A K, Duarte, I D G, Lima, M E
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01-05-2016; John Wiley and Sons Inc
• Subjects: Analgesics, Opioid - pharmacology; Animals; Dose-Response Relationship, Drug; Enzyme Inhibitors - pharmacology; Male; Neprilysin - antagonists & inhibitors; Neprilysin - metabolism; Peptides - pharmacology; Rats; Rats, Wistar; Receptor, Cannabinoid, CB1 - antagonists & inhibitors; Receptors, Opioid - metabolism; Research Paper; Research Papers; Spider Venoms - chemistry; Structure-Activity Relationship
• ISSN: 0007-1188; 1476-5381
• DOI: 10.1111/bph.13448

Background and Purpose
The synthetic peptide PnPP‐19 has been studied as a new drug candidate to treat erectile dysfunction. However, PnTx2–6, the spider toxin from which the peptide was designed, induces hyperalgesia. Therefore, we intended to investigate the role of PnPP‐19 in the nociceptive pathway.
Experimental Approach
Nociceptive thresholds were measured by paw pressure test. PnPP‐19 was administered intraplantarly alone or with selective cannabinoid or opioid receptor antagonists. The hydrolysis of PnPP‐19 by neutral endopeptidase (NEP) (EC 3.4.24.11), an enzyme that cleaves enkephalin, was monitored by HPLC and the cleavage sites were deduced by LC–MS. Inhibition by PnPP‐19 and Leu‐enkephalin of NEP enzyme activity was determined spectrofluorimetrically.
Key Results
PnPP‐19 (5, 10 and 20 μg per paw) induced peripheral antinociception in rats. Specific antagonists of μ opioid receptors (clocinnamox), δ opioid receptors (naltrindole) and CB1 receptors (AM251) partly inhibited the antinociceptive effect of PnPP‐19. Inhibition of fatty acid amide hydrolase by MAFP or of anandamide uptake by VDM11 enhanced PnPP‐19‐induced antinociception. NEP cleaved PnPP‐19 only after a long incubation, and Ki values of 35.6 ± 1.4 and 14.6  ± 0.44 μmol·L−1 were determined for PnPP‐19 and Leu‐enkephalin respectively as inhibitors of NEP activity.
Conclusions and Implications
Antinociception induced by PnPP‐19 appears to involve the inhibition of NEP and activation of CB1, μ and δ opioid receptors. Our data provide a greater understanding of the antinociceptive effects of PnPP‐19. This peptide could be useful as a new antinociceptive drug candidate.