In vitro functional assays as a tool to study new synthetic opioids at the μ-opioid receptor: Potential, pitfalls and progress

Vandeputte, Marthe M.; Vasudevan, Lakshmi; Stove, Christophe

doi:10.1016/j.pharmthera.2022.108161

Cited by 14 publications

(10 citation statements)

References 159 publications

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“…. This highlights the importance of carrying out SAR evaluations within a specific assay in a specific time period and emphasizes that one should be cautious when aiming at directly comparing the results obtained with different functional assays or over a prolonged period of time, as also reported elsewhere …”

Section: Resultsmentioning

confidence: 60%

“…5 This highlights the importance of carrying out SAR evaluations within a specific assay in a specific time period and emphasizes that one should be cautious when aiming at directly comparing the results obtained with different functional assays or over a prolonged period of time, as also reported elsewhere. 49 ■ CONCLUSIONS This study reports on the in vitro CB 1 activation potential of a panel of recently emerged and newly synthesized cumylderived indole-and indazole-3-carboxamide SCRAs by applying different assays monitoring the same or different signaling events. This is the first work describing the in vitro CB 1 activity of a systematic panel of SCRAs related to the recently detected CUMYL-CBMICA and CUMYL-CBMINA-CA.…”

Section: ■ Introductionmentioning

confidence: 99%

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Comprehensive Characterization of a Systematic Library of Alkyl and Alicyclic Synthetic Cannabinoids Related to CUMYL-PICA, CUMYL-BUTICA, CUMYL-CBMICA, and CUMYL-PINACA

Janssens

Ametovski

Sparkes

et al. 2022

ACS Chem. Neurosci.

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Over 200 synthetic cannabinoid receptor agonists (SCRAs) have been identified as new psychoactive substances. Effective monitoring and characterization of SCRAs are hindered by the rapid pace of structural evolution. Ahead of possible appearance on the illicit drug market, new SCRAs were synthesized to complete a systematic library of cumyl-indole-(e.g., CUMYL-CPrMICA, CUMYL-CPMICA) and cumyl-indazole-carboxamides (e.g., CUMYL-CPrMINACA, CUMYL-CPMINACA), encompassing butyl, pentyl, cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl, and cyclohexylmethyl tails. Comprehensive pharmacological characterization was performed with three assay formats, monitoring the recruitment of either wild-type or C-terminally truncated (βarr2d366) β-arrestin2 to the activated cannabinoid 1 receptor (CB 1 ) or monitoring G βγ -mediated membrane hyperpolarization. Altered compound characterization was observed when comparing derived potency (EC 50 ) and efficacy (E max ) values from both assays monitoring the same or a different signaling event, whereas ranges and ranking orders were similar. Structure−activity relationships (SAR) were assessed in threefold, resulting in the identification of the pendant tail as a critical pharmacophore, with the optimal chain length for CB 1 activation approximating an npentyl (e.g., cyclopentylmethyl or cyclohexylmethyl tail). The activity of the SCRAs encompassing cyclic tails decreased with decreasing number of carbons forming the cyclic moiety, with CUMYL-CPrMICA showing the least CB 1 activity in all assay formats. The SARs were rationalized via molecular docking, demonstrating the importance of the optimal steric contribution of the hydrophobic tail. While SAR conclusions remained largely unchanged, the differential compound characterization by both similar and different assay designs emphasizes the importance of detailing specific assay characteristics to allow adequate interpretation of potencies and efficacies.

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Section: Resultsmentioning

confidence: 60%

Section: ■ Introductionmentioning

confidence: 99%

Comprehensive Characterization of a Systematic Library of Alkyl and Alicyclic Synthetic Cannabinoids Related to CUMYL-PICA, CUMYL-BUTICA, CUMYL-CBMICA, and CUMYL-PINACA

Janssens

Ametovski

Sparkes

et al. 2022

ACS Chem. Neurosci.

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“…Raw data analysis was performed as described previously, 20 and GraphPad Prism 9 (San Diego, CA, USA) was used to generate concentration–response curves via three‐parameter logistic regression. In line with previous studies, 19,21 data were normalized to the maximum response of hydromorphone (100%). Normalized area under the curve (AUC) values from the different experiments were combined to obtain final concentration–response curves, including calculation of potency (EC 50 ) and efficacy ( E max , relative to hydromorphone) values.…”

Section: Methodsmentioning

confidence: 99%

In vitro μ‐opioid receptor activation potential of U10 and β‐U10, positional isomers of the synthetic opioid naphthyl U‐47700

Vandeputte

Stove

2023

Drug Testing and Analysis

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New synthetic opioids (NSOs) with diverse chemical structures continue to appear on recreational drug markets worldwide. U‐type opioids have become one of the largest groups of non‐fentanyl‐related NSOs. Starting in 2020, a previously unreported U‐compound coined “β‐U10” (2‐naphthyl U‐47700; N‐[2‐(dimethylamino)cyclohexyl]‐N‐methylnaphthalene‐2‐carboxamide) was identified in Australia and the United States. β‐U10 is a positional isomer of α‐U10 (1‐naphthyl U‐47700), more commonly known as “U10.” Here, the first comparative in vitro pharmacological characterization of naphthyl U‐47700 (U10 and β‐U10), together with the structural analogue U‐47700 and fentanyl, is reported. Application of a cell‐based μ‐opioid receptor (MOR) activation (β‐arrestin 2 recruitment) assay demonstrated β‐U10 (EC50 = 348 nM; Emax = 150% vs. hydromorphone) to be less potent than U‐47700 (EC50 = 116 nM; Emax = 154%) and fentanyl (EC50 = 9.35 nM; Emax = 146%) but considerably more active than the α‐isomer (EC50 value in the μM range). For the latter, maximum receptor activation could not be reached at 100 μM. The difference in MOR activation potential for U10 and β‐U10 stresses the importance of (analytical) differentiation between closely related analytes. The emergence of β‐U10 on the recreational drug market is an example of the continuing emergence of non‐fentanyl‐related NSOs and further emphasizes the need to closely monitor fluctuations in the drug supply.

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“…Nevertheless, the use of opioids can give rise to undesirable side effects, including fatigue, nausea, and dizziness . Furthermore, high doses of opioids can result in respiratory depression, a potentially fatal adverse side effect. , During the 1980s, the prescription of opioids for noncancer pain and chronic conditions increased in parallel with the production of pharmaceutical opioids in the United States . By 2010, a second wave of the opioid crisis began in the United States, which was linked to an increase in availability and demand for heroin .…”

Section: Introductionmentioning

confidence: 99%

In Vitro Functional Profiling of Fentanyl and Nitazene Analogs at the μ-Opioid Receptor Reveals High Efficacy for Gi Protein Signaling

Tsai,

Chen,

Baumann

et al. 2024

ACS Chem. Neurosci.

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Novel synthetic opioids (NSOs), including both fentanyl and non-fentanyl analogs that act as μ-opioid receptor (MOR) agonists, are associated with serious intoxication and fatal overdose. Previous studies proposed that G-protein-biased MOR agonists are safer pain medications, while other evidence indicates that low intrinsic efficacy at MOR better explains the reduced opioid side effects. Here, we characterized the in vitro functional profiles of various NSOs at the MOR using adenylate cyclase inhibition and β-arrestin2 recruitment assays, in conjunction with the application of the receptor depletion approach. By fitting the concentration−response data to the operational model of agonism, we deduced the intrinsic efficacy and affinity for each opioid in the Gi protein signaling and β-arrestin2 recruitment pathways. Compared to the reference agonist [D-Ala 2 ,N-MePhe 4 ,Gly-ol 5 ]enkephalin, we found that several fentanyl analogs were more efficacious at inhibiting cAMP production, whereas all fentanyl analogs were less efficacious at recruiting β-arrestin2. In contrast, the non-fentanyl 2-benzylbenzimidazole (i.e., nitazene) analogs were highly efficacious and potent in both the cAMP and β-arrestin2 assays. Our findings suggest that the high intrinsic efficacy of the NSOs in Gi protein signaling is a common property that may underlie their high risk of intoxication and overdose, highlighting the limitation of using in vitro functional bias to predict the adverse effects of opioids. In addition, the extremely high potency of many NSOs now infiltrating illicit drug markets further contributes to the danger posed to public health.

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In vitro functional assays as a tool to study new synthetic opioids at the μ-opioid receptor: Potential, pitfalls and progress

Cited by 14 publications

References 159 publications

Comprehensive Characterization of a Systematic Library of Alkyl and Alicyclic Synthetic Cannabinoids Related to CUMYL-PICA, CUMYL-BUTICA, CUMYL-CBMICA, and CUMYL-PINACA

Comprehensive Characterization of a Systematic Library of Alkyl and Alicyclic Synthetic Cannabinoids Related to CUMYL-PICA, CUMYL-BUTICA, CUMYL-CBMICA, and CUMYL-PINACA

In vitro μ‐opioid receptor activation potential of U10 and β‐U10, positional isomers of the synthetic opioid naphthyl U‐47700

In Vitro Functional Profiling of Fentanyl and Nitazene Analogs at the μ-Opioid Receptor Reveals High Efficacy for Gi Protein Signaling

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