Nociceptin orphanin FQ (N/OFQ) and its receptor (NOP) are implicated in a variety of physiological processes both at central and peripheral levels. The identification of selective ligands is required to determine the potential of the NOP receptor as a target of innovative drugs. [(pF)Phe⁴,Arg¹⁴,Lys¹⁵]N/OFQ-NH₂ (UFP-102) has been recently generated in our laboratories and here we describe the in vitro pharmacological characterisation of this novel NOP ligand. Human recombinant NOP receptors expressed in Chinese hamster ovary cells (CHO_hNOP) and native animal receptors expressed in isolated tissues were used. Receptor binding, stimulated GTP[³⁵S] binding, and cAMP levels were evaluated in CHO_hNOP according to the methods described by McDonald et al., 2002 while the mouse/rat vas-deferens (m/rVD) and guinea pig ileum (gpI) were used according to Bigoni et al., 1999.

In [³H]N/OFQ binding studies UFP-102 displayed 10-fold greater affinity (pK_i 11.32±0.08) than N/OFQ (pK_i 10.01±0.04). In functional studies UFP-102 mimicked the effects of N/OFQ showing similar maximal effects but higher potencies (see table 1). In electrically stimulated mVD, rVD and gpI the effects of both N/OFQ and UFP-102 were competitively antagonised by the NOP selective antagonists UFP-101 and J-113397 with pK_B values in the range 6.91 - 7.33 and 7.75 - 8.12, respectively. Moreover the effects of N/OFQ and UFP-102 in all tissues were not affected by the universal opioid receptor antagonist naloxone (1 µM).

Table 1 Potencies of N/OFQ and UFP-102 in in vitro assays

Preparation / Assay	N/OFQ	UFP-102
CHOhNOPGTP[35S]	8.70 ± 0.07 1	0.12 ± 0.04
CHOhNOP cAMP	9.86 ± 0.13	10.17 ± 0.07
mVD	7.76 ± 0.05	9.44 ± 0.08
rVD	7.24 ± 0.06	8.57 ± 0.08
gpI	8.05 ± 0.06	9.14 ± 0.10

A separate series of experiments were performed in mVD tissues taken from wild type (NOP^+/+) and NOP receptor knockout (NOP^-/-) mice. N/OFQ and UFP-102 concentration dependently inhibited the electrically stimulated twitch in NOP^+/+ tissues (pEC₅₀ 7.62 and 9.40, respectively) while both peptides were inactive up to micromolar concentrations in mVD taken from NOP^-/- animals.

Overall we conclude that UFP-102 behaves as a highly potent and selective NOP receptor ligand and to date represents the most potent full agonist described in literature.

Bigoni R et al., (1999) Naunyn-Schmiedeberg's Arch. Pharmacol., 361, 565-568.
McDonald J et al., (2002) Eur J Pharmacol 443:1-3, 7-12.
Mogil JS & Pasternak GW (2001) Pharmacol Rev., 53, 381-415.

This work was supported by funds from the University of Ferrara, from the University of Leicester and from a collaborative IASP grant Ferrara / Leicester.