PL-265

Ocular pain is a core symptom of inflammatory or traumatic disorders affecting the anterior segment. To date, the management of chronic ocular pain remains a therapeutic challenge in ophthalmology. The main endogenous opioids (enkephalins) play a key role in pain control but exhibit only transient analgesic effects due to their rapid degradation. The aim of this study was to explore the antinociceptive and anti-inflammatory effects of topical administration of PL265 (a dual enkephalinase inhibitor) on murine models of corneal pain. On healthy corneas, chronic PL265 topical administration did not alter corneal integrity nor modify corneal mechanical and chemical sensitivity. Then, on murine models of corneal pain, we showed that repeated instillations of PL265 (10 mM) significantly reduced corneal mechanical and chemical hypersensitivity. PL265-induced corneal analgesia was completely antagonized by naloxone methiodide, demonstrating that PL265 antinociceptive effects were mediated by peripheral corneal opioid receptors. Moreover, flow cytometry (quantification of CD11b+ cells) and in vivo confocal microscopy analysis revealed that instillations of PL265 significantly decreased corneal inflammation in a corneal inflammatory pain model. Chronic PL265 topical administration also decreased Iba1 and neuronal injury marker (ATF3) staining in the nucleus of primary sensory neurons of ipsilateral trigeminal ganglion. These results open a new avenue for ocular pain treatment based on the enhancement of endogenous opioid peptides' analgesic effects in tissues of the anterior segment of the eye. Dual enkephalinase inhibitor PL265 seems to be a promising topical treatment for safe and effective alleviation of ocular pain and inflammation.

The first line pharmacological treatment of cancer pain is morphine and surrogates but a significant pain relief and a reduction of the side-effects of these compounds makes it necessary to combine them with other drugs acting on different targets. The aim of this study was to measure the antinociceptive effect on cancer-induced bone pain resulting from the association of the endogenous opioids enkephalin and non-opioid analgesic drugs. For this purpose, PL265 a new orally active single dual inhibitor of the two degrading enkephalins enzymes, neprilysin (NEP) and aminopeptidase N (APN) was used. It strictly increased the levels of enkephalin at their sites of releases. The selected non-opioid compounds are: gabapentin, A-317491 (P2X3receptor antagonist), ACEA (CB1 receptor antagonist), AM1241 (CB2 receptor antagonist), JWH-133 (CB2 receptor antagonist), URB937 (FAAH inhibitor), and NAV26 (Nav1.7 channel blocker).

Experiments. Experiments were performed in 5–6 weeks old (26–33g weight) C57BL/6 mice. Cell culture and cell inoculation. B16-F10 melanoma cells were cultured and when preconfluent, treated and detached. Finally related cells were resuspended to obtain a concentration of 2×106cells/100μL. Then 105cells were injected into the right tibial medullar cavity. Control mice were treated by killed cells by freezing.Behavioural studies. Thermal withdrawal latencies were measured on a unilatered hot plate (UHP) maintained at 49±0.2 °C.Mechanical threshold values were obtained by performing the von Frey test using the “up and down” method. To evaluate the nature (additive or synergistic) of the interactions between PL265 and different drugs, an isobolographic analysis following the method described by Tallarida was performed.

The results demonstrate the ability of PL265, a DENKI that prevents the degradation of endogenous ENKs, to counteract cancer-induced bone thermal hyperalgesia in mice, by exclusively stimulating peripheral opioid receptors as demonstrated by used of an opioid antagonist unable to enter the brain. The development of such DENKIs, endowed with druggable pharmacokinetic characteristics, such as good absorption by oral route, can be considered as an important step in the development of much needed novel antihyperalgesic drugs. Furthermore, all the tested combinations resulted in synergistic antihyperalgesic effects. As shown here, the greatest synergistic antinociceptive effect (doses could be lowered by 70%) was produced by the combination of PL265 with the P2X3receptor antagonist (A-317491), cannabinoid CB1 receptor agonist (exogenous, ACEA and endogenous URB937-protected-AEA) and Nav1.7 blocker (NAV26) whose mechanism of action involves the direct activation of the enkephalinergic system.

These multi-target-based antinociceptive strategies using combinations of non-opioid drugs with dual inhibitors of enkephalin degrading enzymes may bring therapeutic advantages in terms of efficacy and safety by allowing the reduction of doses of one of the compounds or of both, which is of the utmost interest in the chronic treatment of cancer pain.

This article presents synergistic antinociceptive effect produced by the combination of PL265 with non-opioid analgesic drugs acting via unrelated mechanisms. These multi-target-based antinociceptive strategies may bring therapeutic advantages by allowing the reduction of doses, which is of great interest in the chronic treatment of cancer pain.