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. 1999 Apr;126(7):1575-84.
doi: 10.1038/sj.bjp.0702469.

An investigation into the structural determinants of cannabinoid receptor ligand efficacy

G Griffin  1 , E J Wray, W K Rorrer, P J Crocker, W J Ryan, B Saha, R K Razdan, B R Martin, M E Abood
Affiliations

An investigation into the structural determinants of cannabinoid receptor ligand efficacy

G Griffin et al. Br J Pharmacol. 1999 Apr.

Abstract

1. A number of side-chain analogues of delta8-THC were tested in GTPgammaS binding assay in rat cerebellar membranes. O-1125, a saturated side-chain compound stimulated GTPgammaS binding with an Emax of 165.0%, and an EC50 of 17.4 nM. 2. O-1236, O-1237 and O-1238, three-enyl derivatives containing a cis carbon-carbon double bond in the side-chain, stimulated GTPgammaS binding, acting as partial agonists with Emax values ranging from 51.3-87.5% and EC50 values between 4.4 and 29.7 nM. 3. The stimulatory effects of O-1125, O-1236, O-1237 and O-1238 on GTPgammaS binding were antagonized by the CB1 receptor antagonist SR 141716A. The K(B) values obtained ranged from 0.11-0.21 mM, suggesting an action at CB1 receptors. 4. Five-ynyl derivatives (O-584, O-806, O-823, O-1176 and O-1184), each containing a carbon-carbon triple bond in the side-chain, did not stimulate GTPgammaS binding and were tested as potential cannabinoid receptor antagonists. 5. Each -ynyl compound antagonized the stimulatory effects of four cannabinoid receptor agonists on GTPgammaS binding. The K(B) values obtained, all found to be in the nanomolar range, did not differ between agonists or from cerebellar binding affinity. 6. In conclusion, alterations of the side-chain of the classical cannabinoid structure may exert a large influence on affinity and efficacy at the CB1 receptor. 7. Furthermore, this study confirms the ability of the GTPgammaS binding assay to assess discrete differences in ligand efficacies which potentially may not be observed using alternative functional assays, thus providing a unique tool for the assessment of the molecular mechanisms underlying ligand efficacies.

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Figures

Figure 1
Figure 1
Chemical structures.
Figure 2
Figure 2
Effect of O-1236, O-1237, O-1238 and O-1125 on [35S]-GTPγS binding. (A) Concentration-response curves of O-1125 and O-1238 constructed in the presence of GDP (100 μM). (B) Concentration-response curves of O-1236 and O-1237 conducted in the presence of GDP (10 μM). Data represent percentage stimulation over basal levels. Results are presented as means±s.e.mean for n=3–4 experiments.
Figure 3
Figure 3
Effect of O-584, at a concentration of 100 nM on the mean concentration-response curves of WIN 55212-2, HU-210, O-689 and CP 55,940. Data represent percentage stimulation over basal levels. Results are presented as means±s.e.mean for n=3–5 experiments.
Figure 4
Figure 4
Effect of O-823, at concentrations of 30, 100 and 300 nM on the mean concentration-response curves of WIN 55212-2, HU-210, O-689 and CP 55,940. Data represent percentage stimulation over basal levels. Results are presented as means±s.e.mean for n=4 experiments.
Figure 5
Figure 5
Effect of O-1184, at a concentration of 100 nM on the mean concentration-response curves of WIN 55212-2, HU-210, O-689 and CP 55,940. Data represent percentage stimulation over basal levels. Results are presented as means±s.e.mean for n=4–8 experiments.
Figure 6
Figure 6
Displacement of bound [3H]-SR 141716A from cerebellar membranes by O-1184 and O-584 in the presence of binding buffer A or GTPγS assay buffer. The data are presented as percentage of displacement of specific binding; 0.35 nM [3H]-SR 141716A was the concentration of radioligand used. Non-specific binding was measured in the presence of SR 1417167A (1 μM). Data points are the means±s.e.mean of three experiments performed in triplicate.
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