A novel electrophilic high affinity irreversible probe for the cannabinoid receptor

Morse, Kelley L.; Fournier, Donna J.; Li, Xiuyan; Grzybowska, Jolanta; Makriyannis, Alexandros

doi:10.1016/0024-3205(95)00176-7

Cited by 34 publications

(46 citation statements)

References 10 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We have reported the development of cannabinergic covalent probes and their potential application in GPCR binding site structural studies (Charalambous et al, 1992;Guo et al, 1994;Morse et al, 1995;Picone et al, 2002). The work described here clearly supports the role of the CWXP motif, which is found in Rhodopsin as well as other family 1A GPCRs, located in TMH6 of the CB1 receptor and its involvement in ligand recognition and binding.…”

supporting

confidence: 69%

“…This moiety will selectively react with more reactive nucleophiles, such as the thiol group of cysteine, but not with the hydroxyl of serine side chains or water. Preliminary studies with AM841 (R. P. Picone, D. J. Fournier, and A. Makriyannis, unpublished observations), as well as our earlier generations of C-3 side-chain functionalized isothiocyanates, demonstrated significant efficacy at irreversibly occupying CB1 binding sites in rat brain synaptosomes (Guo et al, 1994;Morse et al, 1995;Picone et al, 2002). This binding was not recoverable with sedimentation washing, which is widely accepted as evidence of irreversible occupation with nonradiolabeled spontaneously reacting isothiocyanate ligands (Guo et al, 1994;Morse et al, 1995;Zhu et al, 1996;Picone et al, 2002;Tahtaoui et al, 2003).…”

Section: Discussionmentioning

confidence: 91%

“…One of our most successful designs encompasses a classical cannabinoid scaffold functionalized in the terminal position of the C-3 alkyl side chain (Charalambous et al, 1992;Guo et al, 1994;Morse et al, 1995;Picone et al, 2002). Together, these covalent probes have been very effective in irreversibly and selectively binding to CB1 in a highly specific manner.…”

Section: Discussionmentioning

confidence: 99%

“…Affinity Labeling. Labeling methods were adapted from our previously published procedures with a few modifications (Guo et al, 1994;Morse et al, 1995). Membranes containing CB1 receptor were allowed to equilibrate with AM841 for 1 h in a 30°C shaking water bath, after which suspensions were rapidly sedimented (27,200g) at 30°C.…”

Section: Methodsmentioning

confidence: 99%

See 3 more Smart Citations

(-)-7′-Isothiocyanato-11-hydroxy-1′,1′-dimethylheptylhexahydrocannabinol (AM841), a High-Affinity Electrophilic Ligand, Interacts Covalently with a Cysteine in Helix Six and Activates the CB1 Cannabinoid Receptor

Picone

Khanolkar²,

Xu³

et al. 2005

Mol Pharmacol

Self Cite

174

View full text Add to dashboard Cite

The CB1 cannabinoid receptor has been shown to play important physiological roles in the central nervous system, as well as peripherally, and is a target for development of therapeutic medications. To gain insight on the ligand binding site(s) and structural features of activation, we designed and synthesized (Ϫ)-7Ј-isothiocyanato-11-hydroxy-1Ј,1Ј-dimethylheptylhexahydrocannabinol (AM841), a classical cannabinoid affinity label that incorporates an isothiocyanate substituent as an electrophilic reactive group capable of interacting irreversibly with a suitably located and properly oriented nucleophilic amino acid residue at or near the binding site. To obtain evidence for the site of covalent attachment of AM841, C6.47, identified in part by interactive ligand docking, was mutated to serine, alanine, and leucine to reduce or eliminate the nucleophilic character. Wild-type (WT) and mutant CB1 receptors were evaluated for their abilities to recognize a series of cannabinergic ligands. Each bound comparably to WT, excluding C6.47L, which displayed a reduced affinity for It is noteworthy that AM841 was shown to bind irreversibly to WT CB1 but exhibited no covalent attachment with the mutants and behaved as an agonist suggesting irreversible attachment to C6.47 maintains CB1 in its active state. The evidence presented identifies C6.47 as the site of covalent bond formation with AM841 and combined with the binding data fully supports the molecular modeling. These studies present the first report of tandem applications of affinity labeling, site-directed mutagenesis, and interactive ligand docking for CB1.The CB1 and CB2 cannabinoid receptors are relatively new members in the G-protein-coupled receptor (GPCR) superfamily. They have been shown to play important physiological roles and represent targets for development of therapeutic medications. From a pharmacological standpoint, agonist activation of both receptors results in the release of G␣ iproteins, causing a concomitant reduction in intracellular This work has been supported by National Institute on Drug Abuse grants DA05955 (to R.P.P.) DA00355 (to A.D.K.), DA09158, DA03801, DA07215, DA07312 (to A.M.), DA03934, DA00489 (to P.H.R.), DA05274, and DA09978 (to M.E.A.).

show abstract

supporting

confidence: 69%

Section: Discussionmentioning

confidence: 91%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

(-)-7′-Isothiocyanato-11-hydroxy-1′,1′-dimethylheptylhexahydrocannabinol (AM841), a High-Affinity Electrophilic Ligand, Interacts Covalently with a Cysteine in Helix Six and Activates the CB1 Cannabinoid Receptor

Picone

Khanolkar²,

Xu³

et al. 2005

Mol Pharmacol

Self Cite

174

View full text Add to dashboard Cite

show abstract

“…AM4113, AM251, and SR141716A were tested for binding to the CB1 receptor using a rat brain membrane preparation, and for CB2 receptor binding using HEK293 cell membranes expressing hCB2 membranes, as previously described using [ 3 H]CP-55,940 (Morse et al, 1995;Lan et al, 1999;Makriyannis et al, 2005;McLaughlin et al, 2006). The concentrated stocks (10 mM) were diluted into TME buffer (50 mM Tris-HCl, 3 mM MgCl 2 , 100 mM NaCl, 0.2 mM EDTA, pH 7.4) with 0.1% BSA, and transferred to 96-well plates containing [ 3 H]CP-55,940 (specific activity 128 Ci/mmol; NIDA) at a final concentration of 0.76 nM.…”

Section: Methodsmentioning

confidence: 99%

The Novel Cannabinoid CB1 Receptor Neutral Antagonist AM4113 Suppresses Food Intake and Food-Reinforced Behavior but Does not Induce Signs of Nausea in Rats

Sink

McLaughlin

Wood

et al. 2007

Neuropsychopharmacol

143

193

View full text Add to dashboard Cite

Drugs that interfere with cannabinoid CB1 transmission suppress various food-motivated behaviors, and it has been suggested that such drugs could be useful as appetite suppressants. Biochemical studies indicate that most of these drugs assessed thus far have been CB1 inverse agonists, and although they have been shown to suppress food intake, they also appear to induce nausea and malaise. The present studies were undertaken to characterize the behavioral effects of AM4113, which is a CB1 neutral antagonist, and to examine whether this drug can reduce food-reinforced behaviors and feeding on diets with varying macronutrient compositions. Biochemical data demonstrated that AM4113 binds to CB1 receptors, but does not show inverse agonist properties (ie no effects on cyclic-AMP production). In tests of spontaneous locomotion and analgesia, AM4113 reversed the effects of the CB1 agonist AM411. AM4113 suppressed food-reinforced operant responding with rats responding on fixed ratio (FR) 1 and 5 schedules of reinforcement in a dosedependent manner, and also suppressed feeding on high-fat, high-carbohydrate, and lab chow diets. However, in the same dose range that suppressed feeding, AM4113 did not induce conditioned gaping, which is a sign of nausea and food-related malaise in rats. These results suggest that AM4113 may decrease appetite by blocking endogenous cannabinoid tone, and that this drug may be less associated with nausea than CB1 inverse agonists. Neuropsychopharmacology (2008) 33, 946-955;

show abstract

A Late‐Stage Aryl C−H Olefination Strategy and Its Application Towards Global Proteome Profiling of Δ⁸‐Tetrahydrocannabinol

Lim

Tang

et al. 2023

Chemistry A European J

View full text Add to dashboard Cite

Drugs and bioactive natural products exert their pharmacological effects by engaging numerous cellular targets in our body. Identification of these protein targets is essential for understanding the mechanism‐of‐action of these compounds, thus contributing to improved drug design in drug discovery programs. Termed “in situ drug profiling”, a common strategy for studying these bioactive compounds centralized on the covalent capture of protein targets along with a reporter tag to facilitate downstream proteomic analyses. Though highly successful, such reliance on innate electrophilic traps to facilitate covalent capture restricted its applications to covalent acting compounds. Late‐stage C−H functionalization (LSF) may resolve this by substituting biologically inert C−H bonds with desired electrophilic groups. Herein, we demonstrated this concept by arming a diverse range of electron‐rich aromatic drugs and natural products with α,β‐unsaturated esters, via late‐stage C−H olefination with an arylthio‐based carboxylic acid ligand developed by Ibanez and co‐workers. We also showed that covalent probes generated from this LSF approach could be applied for “in situ drug profiling” of Δ8‐THC, as exemplified by the successful target engagement of α‐4 db, a Δ8‐THC‐based probe, to its native target hCB2R. In combination with AfBP 7, a photoaffinity‐based derivative of Δ8‐THC, we identified several novel putative targets that could account for some of the effects in THC consumption. We anticipate our C−H LSF strategy to be widely adopted for future studies of non‐covalent drugs.

show abstract

A novel electrophilic high affinity irreversible probe for the cannabinoid receptor

Cited by 34 publications

References 10 publications

(-)-7′-Isothiocyanato-11-hydroxy-1′,1′-dimethylheptylhexahydrocannabinol (AM841), a High-Affinity Electrophilic Ligand, Interacts Covalently with a Cysteine in Helix Six and Activates the CB1 Cannabinoid Receptor

(-)-7′-Isothiocyanato-11-hydroxy-1′,1′-dimethylheptylhexahydrocannabinol (AM841), a High-Affinity Electrophilic Ligand, Interacts Covalently with a Cysteine in Helix Six and Activates the CB1 Cannabinoid Receptor

The Novel Cannabinoid CB1 Receptor Neutral Antagonist AM4113 Suppresses Food Intake and Food-Reinforced Behavior but Does not Induce Signs of Nausea in Rats

A Late‐Stage Aryl C−H Olefination Strategy and Its Application Towards Global Proteome Profiling of Δ⁸‐Tetrahydrocannabinol

Contact Info

Product

Resources

About

A novel electrophilic high affinity irreversible probe for the cannabinoid receptor

Cited by 34 publications

References 10 publications

(-)-7′-Isothiocyanato-11-hydroxy-1′,1′-dimethylheptylhexahydrocannabinol (AM841), a High-Affinity Electrophilic Ligand, Interacts Covalently with a Cysteine in Helix Six and Activates the CB1 Cannabinoid Receptor

(-)-7′-Isothiocyanato-11-hydroxy-1′,1′-dimethylheptylhexahydrocannabinol (AM841), a High-Affinity Electrophilic Ligand, Interacts Covalently with a Cysteine in Helix Six and Activates the CB1 Cannabinoid Receptor

The Novel Cannabinoid CB1 Receptor Neutral Antagonist AM4113 Suppresses Food Intake and Food-Reinforced Behavior but Does not Induce Signs of Nausea in Rats

A Late‐Stage Aryl C−H Olefination Strategy and Its Application Towards Global Proteome Profiling of Δ8‐Tetrahydrocannabinol

Contact Info

Product

Resources

About

A Late‐Stage Aryl C−H Olefination Strategy and Its Application Towards Global Proteome Profiling of Δ⁸‐Tetrahydrocannabinol