Homology models of the cannabinoid CB1 and CB2 receptors. A docking analysis study

doi:10.1016/j.ejmech.2004.10.002

European Journal of Medicinal Chemistry

Volume 40, Issue 1, January 2005, Pages 75-83

https://doi.org/10.1016/j.ejmech.2004.10.002 Get rights and content

Abstract

The 3D models of both CB1 and CB2 human receptors have been established by homology modeling using as template the X-ray structure of bovine Rhodopsin (code pdb: 1F88) a G-protein-coupled receptor (GPCR). A recursive approach comprising sequence alignment and model building was used to build both models, followed by the refinement of non-conserved regions. The cannabinoid system has been studied by means of docking techniques, using the 3D models of both CB1 and CB2 and well known reference inverse agonist/antagonist compounds. An approach based on the flexibility of the structures has been used to model the receptor–ligand complexes. The structural effects of ligand binding were studied and analyzed on the basis of hydrogen bond interactions, and binding energy calculations. Potential interaction sites of the receptor were determined from analysis of the difference accessible surface area (DASA) study of the protein with and without ligand.

Introduction

Cannabinoid receptors interact with cannabinoid drugs including the classical cannabinoids such as Δ⁹-tetrahydrocannabinol (Δ⁹-THC) that is the main psychotropic constituent of cannabis, their synthetic analogs and with the endogenous cannabinoids [1], [2], [3], [4]. The potential therapeutic applications claimed for cannabinoid receptor agonists include attenuation of nausea and vomiting in cancer chemotherapy, management of glaucoma, the suppression of muscle spasticity/spasm associated with multiple sclerosis and spinal cord injury, disorders associated with Alzheimer’s disease [5], [6] and therapeutical effects of analgesia [7], [8], [9], [10], [11]. CB1 receptor antagonists/inverse agonists have potential therapeutic application as appetite suppressants [12], [13], [14] and in the management of schizophrenia [15]. The side effects accompanying these therapeutic responses include alterations in cognition and memory, dysphoria, euphoria, and sedation [16].

There are two types of cannabinoid receptors that have so far been identified, CB1 cloned in 1990 [17], and CB2 cloned in 1993 [18]. The CB1 cannabinoid receptor has been cloned from rat [17], mouse [19], and human [20], [21] tissues and shows 97–99% amino acid sequence identity across species and it is found primarily in brain and neuronal tissue, whereas CB2 is found mainly in immune cells where they may mediate an immunosuppresant effect. The CB2 cannabinoid receptor shows 44% identity with the CB1 cannabinoid receptor [18]. At present, there is some preliminary pharmacological evidence that supports the existence of additional types or subtypes of cannabinoid receptors [22], [23], [24].

Both cannabinoid receptor types belong to the large family of G-protein-coupled receptors (GPCRs) [25] controlling a wide variety of signal transduction. In addition, CB1 receptors are also coupled through G-proteins to several types of calcium and potassium channels. GPCRs are a large superfamily, which are integral membrane proteins that are characterized by seven hydrophobic transmembrane helices (TMH). Therefore, the knowledge of the structural features of cannabinoid receptors is of the utmost importance for the understanding of their function and for their use for drug design. For theses purposes, in the last decade, the structures used as template in the molecular modeling studies of GPCRs have been based on the structure of bacteriorhodopsine (BR) [26] or in low-resolution crystal structures of GPCRs. The ground state of Rhodopsin was determined at 2.8 Å resolution by X-ray crystallography [27] and NMR [28], [29]. This experimental result has allowed to undertake the modeling of the GPCRs with greater reliability. Therefore, we have used this structure as suitable template to build by homology modeling 3D models of both CB1 and CB2 receptors.

Identification of the binding site and binding conformations of cannabinoid ligands within the CB receptors is of great interest for the understanding of principles that account for the interactions between the ligand and the amino acid residues and for the design of new ligands. Site-specific mutation studies on the Rhodopsin subfamily of receptors suggest that the ligand binding site is localized within the TM core region in the crevice formed by TM3, TM5, TM6, and TM7 [27], [30].

Ligand design represents an integral approach useful to provide structural information not available from the experiment methods. Docking studies have been used to study the binding orientations and prediction of binding affinities.

Classical cannabinoids are tricyclic terpenoid derivatives bearing a benzopyran moiety. This class includes the natural product (–)Δ⁹-THC and other pharmacologically active constituents of the plant Cannabis sativa. A second class of cannabimimetics was developed at Pfizer which includes bicyclic (e.g. CP55940) and tricyclic (e.g. CP55244) analogs lacking the pyran rings of classical cannabinoids (Fig. 1). The diarylpyrazoles are another type of cannabinoid analogs, one of them is SR141716A developed by Sanofi which is currently in clinical phase for the treatment of obesity and addictions. The other diarylpyrazol is SR144528 developed by Sanofi which is a potent and selective CB2 receptor ligand. A second chemical class of cannabinergics are the aminoalkylindoles that were developed at Sterling Winthrop as potential non-steroidal anti-inflamamatory agents. WIN55212-2 is a potent CB1 and CB2 agonist with high stereoselectivity and a slight preference for CB2. AM-630 is the first CB2-selective antagonist derived from this class of compounds [1] (Fig. 2).

In the present study, a homology model of the CB1 and CB2 receptor was constructed using the X-ray crystal structure of bovine Rhodopsin as the structural template. A molecular docking approach with FlexiDock [31] was employed to identify the binding site in CB1 and CB2 receptors from representative inverse agonist or antagonist ligands (Fig. 2).

Section snippets

Homology model of the CB1 and CB2 receptor

The modeling process by homology consists in several steps essential for obtaining a correct sequence alignment of the target sequences CB1 and CB2 with the homologous visual Rhodopsin (Rho) in the inactivated state (code PDB: 1F88) used as basic structure. The approach followed was developed and improved by Burke et al. [32]. The sequential alignment of Rhodopsin and the cannabinoid receptor CB1 and CB2 was performed using the sequence alignment program CLUSTALW [33] followed by a manual

Homology modeling

Multiple Sequence Alignment. Sequentially, CB1 and CB2 share 21% and 20% of identity with visual Rhodopsin (code pdb: 1F88) and only 44% between CB1 and CB2. Fig. 3 shows a schematic representation of the cannabinoid receptors. Residues highly conserved in the GPCR family are shown in Table 1.

The sequential alignment of Rhodopsin and cannabinoid receptors is shown in Fig. 4. Most of the key residues characteristic of GPCRs are conserved in CB1 and CB2. The major sequence differences between

Conclusion

In conclusion, we have provided, the 3D models of the cannabinoid receptors CB1 and CB2 based on the highest resolution structure of a GPCR (1F88). The differences and analogies of both receptors have also been studied.

A model of the ligand–protein complexes are described by means of docking studies. The structural effects of ligand binding have been analyzed on the basis of hydrogen bond interactions, aromatic interactions and binding energy interactions in final complexes from manual docking

Acknowledgements

The authors thank Emilia Bayo for her help in the transcription of this manuscript. This project is supported by the Ministerio of Ciencia y Tecnologia, Spain (SAF 2000-0114-C02-01) and Comunidad de Madrid (08.5/0050/2003 1).

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Original articleHomology models of the cannabinoid CB1 and CB2 receptors. A docking analysis study

Abstract

Introduction

Section snippets

Homology model of the CB1 and CB2 receptor

Homology modeling

Conclusion

Acknowledgements

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Prog. Neurobiol.

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Eur. J. Pharmacol

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J. Mol. Biol.

J. Mol. Biol.

J. Mol. Biol.

Eur. J. Pharmacol.

J. Biol. Chem.

Pharmacol. Rev.

Mini Rev. Med. Chem.

J. Neurosci.

Int. J. Geriatr. Psychiatry

Curr. Opin. Invest. Drugs

Curr. Opin. Invest. Drugs

Expert Opin. Invest. Drugs

Pharmacol. Ther

J. Physiol. Regul. Integr. Comp. Physiol.

Behav. Pharmacol.

Nature

Nature

DNA Seq.

Nucleic Acids Res.

Biochem. J.

Neurochemistry

Neuroscience

Science

Biochemistry

Biochemistry

Biochemistry

Original article
Homology models of the cannabinoid CB1 and CB2 receptors. A docking analysis study