Allosteric modulators of G protein-coupled receptors, including opioid receptors, have been proposed as possible therapeutic agents with enhanced selectivity. BMS-986122 is a positive allosteric modulator (PAM) of the -opioid receptor (-OR). BMS-986187 is a structurally distinct PAM for the -opioid receptor (-OR) that has been reported to exhibit 100-fold selectivity in promoting -OR over-OR agonism. We used ligand binding and second-messenger assays to show that BMS-986187 is an effective PAM at the -OR and at the-opioid receptor (-OR), but it is ineffective at the nociceptin receptor. The affinity of BMS-986187 for -ORs and-ORs is approximately 20- to 30-fold higher than for -ORs, determined using an allosteric ternary complex model. Moreover, we provide evidence, using a silent allosteric modulator as an allosteric antagonist, that BMS-986187 and BMS-986122 bind to a similar region on all three traditional opioid receptor types (-OR, -OR, and-OR). In contrast to the dogma surrounding allosteric modulators, the results indicate a possible conserved allosteric binding site across the opioid receptor family that can accommodate structurally diverse molecules. These findings have implications for the development of selective allosteric modulators.
Background and Purpose: The δ-opioid receptor is an emerging target for the management of chronic pain and depression. Biased signalling, the preferential activation of one signalling pathway over another downstream of δ-receptors, may generate better therapeutic profiles. BMS 986187 is a positive allosteric modulator of δ-receptors. Here, we ask if BMS 986187 can directly activate the receptor from an allosteric site, without an orthosteric ligand, and if a signalling bias is generated. Experimental Approach: We used several clonal cell lines expressing δ-receptors, to assess effects of BMS 986187 on events downstream of δ-receptors by measuring G-protein activation, β-arrestin 2 recruitment, receptor phosphorylation, loss of surface receptor expression, ERK1/ERK2 phosphorylation, and receptor desensitization. Key Results: BMS 986187 is a G protein biased allosteric agonist, relative to βarrestin 2 recruitment. Despite showing direct and potent G protein activation, BMS 986187 has a low potency to recruit β-arrestin 2. This appears to reflect the inability of BMS 986187 to elicit any significant receptor phosphorylation, consistent with low receptor internalization and a slower onset of desensitization, compared with the full agonist SNC80. Conclusions and Implications: This is the first evidence of biased agonism mediated through direct binding to an allosteric site on an opioid receptor, without a ligand at the orthosteric site. Our data suggest that agonists targeting δ-receptors, or indeed any GPCR, through allosteric sites may be a novel way to promote signalling bias and thereby potentially produce a more specific pharmacology than can be observed by activation via the orthosteric site. 1 | INTRODUCTION Chronic pain and depression are two of the most common medical ailments experienced worldwide and are often co-morbid. For example, an estimated 25% of the United States population (75 million people) experience moderate-to-severe chronic pain (Reinke, 2014), whereas an estimated 15-20% experience depression (Kessler & Bromet, 2013). Opioid analgesics that target the μ-opioid receptor are the most widely prescribed drugs for both chronic and acute pain but suffer from serious side effects including respiratory depression and abuse liability (McNicol et al., 2017; Przewłocki & Przewłocka, 2001). Treatments for depression are varied, but under the best circumstances, only an estimated 50% of patients show full remission (Rush et al., 2006). Mounting evidence suggests that agonists targeting the δ-opioid receptor, a GPCR, are effective in preclinical models of chronic pain and depression and could provide for new therapies
A new approach for the separation of 6-aminoquinolyl-carbamyl (AQC)-derivatized amino acids has been proposed. The chromatography used ion-pairing mechanism to increase the method selectivity. Mobile phase was based on triethylamine buffer containing N,N-dimethyloctylamine as a modifier. A number of factors, buffer composition and pH, counterion concentration, temperature and acetonitrile gradient profile, were optimized to achieve final chromatographic conditions. With the presented analytical method, the separation and identification of 34 AQC-amino acids and amino compounds present in human plasma is possible. The results of validation proved the applicability of the method for quantification of 27 amino acids in biological samples. The ultrafiltration proposed as deproteinization procedure gave repeatable and reliable results for the amino acids under investigation. This method introduced in routine testing can be a suitable tool for amino acid profiling in plasma including all aspects of clinical application.
Opioids alleviate pain, but adverse effects severely limit their usefulness. To solve this problem, biased ligands favoring 1 signaling pathway downstream of the μ-opioid receptor over another are being developed. In the target article, the authors synthesize compounds that preferentially activate G-protein or β-arrestin signaling. They find that increased bias towards G-protein signaling produces better antinociception with minimal side effects in mice models. G-protein–biased opioids may provide a safer treatment strategy.
ID 23842 Poster Board 565Mu-opioid receptor agonists are the gold standard for pain treatment. While efficacious for moderate to severe pain, these drugs are associated with serious adverse effects including abuse liability and respiratory depression. One emerging strategy for safer pain treatments is the development of positive allosteric modulators (PAMs) of the mu-opioid receptor (mu-PAMs). Mu-PAMs can enhance analgesia by increasing the activity of endogenous neurotransmitters released during pain states. Because mu-PAMs do not activate MOR in the absence of an orthosteric agonist and the promoting effects on endogenous opioids are localized and temporal, this allosteric strategy holds the promise of analgesia with fewer side effects. Previous work has identified one molecule, BMS-986122 (Figure 1), that enhances the antinociceptive effects of endogenous opioids with a reduced ability to cause constipation, respiratory depression, and reward. However, BMS-986122 is weak with an affinity of 5mM for the mu-opioid receptor, and we do not know how the compound interacts with the receptor. Hence, the presented research aimed to explore structure-activity relationships (SAR) of the BMS986122 scaffold for the design of more potent compounds. Derivatives of BMS-986122 were synthesized and fully characterized (purity >95%). The potency and efficacy of the compounds were determined by their ability to shift the dose-response curve of the standard mu-opioid agonist DAMGO using b-arrestin recruitment and GTPg 35 S binding assays. By altering the substitution patterns on the aromatic rings and replacing the heterocyclic core (Figure 1) we were able to identify compounds that had no effect on their own but shifted the concentration-response curve for DAMGO up to 35-fold. There was only a minor difference between individual enantiomers (*). Our study has generated a SAR framework of BMS986122, which will facilitate the understanding of the allosteric pharmacophore and the design of better mu-PAMs.Support/Funding Information: R37 DA039997 Figure 1. Structure of BMS986122.
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