Direct inhibition of microglial activation by a µ receptor selective agonist alleviates inflammatory-induced pain hypersensitivity.

Opioids are widely used in the treatment of moderate and severe pain. Nociceptive stimulation has been reported to potentially promote microglial activation and neuroinflammation, which also causes chronic pain sensitization. The aim of this study was to demonstrate whether the novel µ receptor agonist MEL-0614 could inhibit activated microglia directly and the associated signaling pathway. Mice were administered lipopolysaccharide and formalin to induce allodynia. Von Frey test was used to detect the anti-allodynia effect of MEL-0614 before and after LPS and formalin injection. In the spinal cord, the levels of proinflammatory cytokines and microglial activation were determined after MEL-0614 administration. BV2 and primary microglia were cultured to further explore the effect of MEL-0614 on LPS-induced microglial activation and key signaling pathways involved. MEL-0614 partially prevented and reversed allodynia induced by LPS and formalin in vivo, which was not inhibited by the µ receptor antagonist CTAP. Minocycline was effective in reversing the established allodynia. MEL-0614 also downregulated the activation of microglia and related proinflammatory cytokines in the spinal cord. Additionally, in BV2 and primary microglia, MEL-0614 inhibited the LPS-induced upregulation of proinflammatory factors, which was unaffected by CTAP. The NLR family pyrin domain containing 3 (NLRP3) related signaling pathway may be involved in the interaction between MEL-0614 and microglia. The opioid agonist MEL-0614 inhibited the activation of microglia and the subsequent upregulation of proinflammatory factors both in vivo and in vitro. Notably, this effect is partially mediated by the µ receptor.

Repeated Endomorphin Analogue MEL-0614 Reduces Tolerance and Improves Chronic Postoperative Pain without Modulating the P2X7R Signaling Pathway.

The clinical treatment of chronic postoperative pain (CPSP) remains challenging. The side effects of chronic morphine treatment limit its clinical application. MEL-0614, a novel endomorphin analogue that is highly selective and agonistic for µ opioid receptor (MOR), produces a more powerful analgesic effect than that of morphine. In this study, we explored the difference in antinociceptive tolerance and related mechanisms between MEL-0614 and morphine in CPSP induced in a skin/muscle incision and retraction (SMIR) mice model. We found that acute administration of MEL-0614 (1, 3, 5, and 10 nmol, i.t.) produced a dose-dependent analgesic effect that was superior to that of morphine in the SMIR mice model. Long-term MEL-0614 treatment (10 nmol, i.t.) did not induce tolerance compared with morphine. Notably, tolerance induced by morphine could be greatly prevented and/or inhibited via cross-administration or coadministration between MEL-0614 and morphine. In addition, MEL-0614 accelerated the recovery of postoperative pain, whereas morphine aggravated postoperative pain and prolonged its recovery time regardless of preoperative or postoperative treatment. In addition, MEL-0614 did not activate microglia and the P2X7R signaling pathway and showed reduced expression iba1 and P2X7R compared with that observed after morphine administration. Release of inflammatory factors was induced by continued administration of morphine during SMIR surgery, but MEL-0614 did not promote the activation of inflammatory factors. Our results showed that MEL-0614 has superior analgesic effects in CPSP and leads to tolerance to a lesser degree than morphine. Further, MEL-0614 may be used as a promising treatment option for the long-term treatment in CPSP.

Design, synthesis, and biological activity of new endomorphin analogs with multi-site modifications.

Endomorphin (EM)-1 and EM-2 are the most effective endogenous analgesics with efficient separation of analgesia from the risk of adverse effects. Poor metabolic stability and ineffective analgesia after peripheral administration were detrimental for the use of EMs as novel clinical analgesics. Therefore, here, we aimed to establish new EM analogs via introducing different bifunctional d-amino acids at position 2 of [(2-furyl)Map4]EMs. The combination of [(2-furyl)Map4]EMs with D-Arg2 or D-Cit2 yielded analogs with enhanced binding affinity to the µ-opioid receptor (MOR) and increased stability against enzymatic degradation (t1/2 > 300 min). However, the agonistic activities of these analogs toward MOR were slightly reduced. Similar to morphine, peripheral administration of the analog [D-Cit2, (2-furyl)Map4]EM-1 (10) significantly inhibited the pain behavior of mice in multiple pain models. In addition, this EM-1 analog was associated with reduced tolerance, less effect on gastrointestinal mobility, and no significant motor impairment. Compared to natural EMs, the EM analogs synthesized herein had enhanced metabolic stability, bioavailability, and analgesic properties.

MEL endomorphins act as potent inflammatory analgesics with the inhibition of activated non-neuronal cells and modulation of pro-inflammatory cytokines.

Effective treatment of inflammatory pain is a major clinical concern for both patients and physicians. Traditional analgesics such as morphine and coxibs are not effective in all patients and have various unwanted side effects. Accumulating evidence has suggested that endomorphins (EMs), particularly EM-1, possess potent anti-inflammatory effects. However, poor bioavailability and low resistance to enzymatic degradation impede their direct application in the treatment of inflammation. A series of novel peptides based on the structure of EM-1, with lower undesired effects than their parent compounds, called MEL-EMs were discovered and synthetized in our preceding studies. Here, we selected two (MEL-0614 and MEL-N1606) to further investigate their anti-inflammatory effects. This work showed that MEL analogs exerted potent analgesic effects with the inhibition of activated glial cells and macrophages in a CFA-induced inflammatory pain model. Furthermore, multiple-dose administration of MEL analogs did not prolong CFA-induced chronic inflammatory pain, in contrast to morphine. Together, our findings revealed that MEL analogs may serve as effective candidates for chronic inflammation treatment.