Anandamide affinity

Lin S, Khanolkar AD, Fan P, Goutopoulos A, Qin C, Papahadjis D, Makriyannis A (1998) Novel analogues of arachidonylethanolamide (anandamide) affinities for the CBl and CB2 cannabinoid receptors and metabolic stability. J Med Chem 41 5353-5361... 

The development of SAR for endocannabinoid-derived structures has primarily focused on the anandamide skeleton (1) with a large number of publications addressing the requirements for activity and stability of this scaffold. More recently, some SAR has begun to emerge for the other end-ocannabinoids, in particular 2-AG (2). The following discussion will focus on highlighting some of the main features that contribute to affinity and/or stability each endocannabinoid will be treated separately. A number of detailed reviews on this subject have been published [142-146]. 

As outlined earlier, anandamide was the first among the endogenous cannabinoid receptor agonists to be identified. It exhibits higher binding affinity for the CBi receptor Ki — 89 nM) than for the CB2 receptor (il = 371 nM) [81]. Anandamide has typical cannabinoid activities including decreased spontaneous motor activity, immobility and production of hypothermia and analgesia [147, 148]. However, this action in vivo is of shorter duration than... 

In addition to anandamide, several other endogenous polyunsaturated fatty acid derivatives were also found to act as cannabimimetics. They are all now collectively referred to as endocannabinoids. Soon after the discovery of anandamide, two more fatty acid ethanolamides were isolated and found to bind to CB1 preparations with affinities similar to that of anandamide (anandamide CB1 binding affinity K = 39.2 nM, according to Hanus et al., 1993). These were the homo-y-linolenylethanol-amide (CB1 K[ = 53.4 nM) and 7,10,13,16-docosatetraenylethanolamide (CB1 K[ = 34.4 nM) (Fig. 2). All three V-acylethanolamide endocannabinoids were found to be CB1 agonists in the MVD test (Pertwee, 1994). 

Anandamide is inactivated in two steps, first by transport inside the cell and subsequently by intracellular enzymatic hydrolysis. The transport of anandamide inside the cell is a carrier-mediated activity, having been shown to be a saturable, time- and temperature-dependent process that involves some protein with high affinity and specificity for anandamide (Beltramo, 1997). This transport process, unlike that of classical neurotransmitters, is Na+-independent and driven only by the concentration gradient of anandamide (Piomelli, 1998). Although the anandamide transporter protein has not been cloned yet, its well characterized activity is known to be inhibited by specific transporter inhibitors. Reuptake of 2-AG is probably mediated by the same facilitating mechanism (Di Marzo, 1999a,b Piomelli, 1999). 

Beltramo M, Stella N, Calignano A, Lin S, Makriyannis A, Piomelli D. Functional role of high-affinity anandamide transport, as revealed by selective inhibition. Science 1997 277 1094-1097. 

Edgemond WS, Hlillard CJ, Falck JR, Kearn CS, Cambell WB. Fluman platelets and polymorphonuclear leukocytes synthesize oxygenated derivatives of arachidonylethanolamide (anandamide) their affinities for canna-binoid receptors and pathways of inactivation. Mol Pharmacol 1998 54 180-188. 

Its hypnotic properties were characterized. Its mechanism of action is far from being understood. Although it does not bind with high affinity to CB] or CB2 receptors, it exhibits some cannabimimetic actions, which could be explained at least in part by entourage effects. It is likely that oleamide and anandamide have common as well as distinct pathways of action. The 5-HT2A receptor appears to be a target for oleamide but the possibility of the existence of specific receptors for this compound is still open. Legget et have reported that oleamide is a full cannabi-... 

Anandamide was isolated from water-insoluble fractions of the porcine brain. It binds to CB1 with rather moderate affinity (Ki 61 nM) and a low affinity for the CB2 receptor (Ki 1930 nM). The name anandamide is based on its chemical nature (an amide) and the Sanskrit word ananda meaning bliss. The chemical structure of anandamide can be divided into two major molecular fragments a polar ethanolamido head group and a hydrophobic arachidonyl chain. The polar head group comprises a secondary amide functionality with an N-hydroxyalkyl substituent while the lipophilic fragment is a non-conjugated c/ s tetraolefinic chain and an n-pentyl chain reminiscent of the lipophilic side chain found in the classical cannabinoids. A number of anandamide analogs have been synthesized and demonstrated to have considerable selectivity for the CB1 receptor in comparison to the CB2 receptor. 

Makriannis, A, Deng, H. (University of Conneticut) Retro-anandamides, high affinity and stability cannabinoid receptor ligands, WO0128498 (2001). 

Anandamide was discovered as a high-affinity ligand for CB receptors in 1992 (Devane et al., 1992). Anandamide is produced in response to an increase in intracellular Ca2+ levels or activation of G-protein-coupled receptors (e.g., D2 receptors) (Piomelli, 2003). In mammalian tissues, anandamide is present at concentrations of 1-50 pmol/g (Fig. 4), and can be formed through three distinct biochemical pathways (1) the direct hydrolysis of NAPE by a NAPE-specific phospholipase D (NAPE-PLD) (Okamoto et al., 2005 Wang et al., 2006) (2) the hydrolysis of lyso-NAPE or glycerophospho-anandamide by a specific PLD (Leung et al., 2006 Simon and Cravatt, 2008 Sun et al., 2004) and (3) the hydrolysis of phospho-anandamide by a lipid phosphatase (Liu et al., 2006, 2007) (Fig. 1). 

Numerous cannabinoids, anandamides and heterocyclics have been compared for CBj and CB2 binding [87], Most compounds did not differ significantly in their binding values for the two receptors except the specific CBj antagonist SR141716A (see below) which binds weakly to CB2 and the indole compound JWH-015 (20) which showed a CBj/CB2 binding ratio of 27.75. Busch-Petersen etal. synthesized a series of cannabinoids in which rotation around the Cj -C2 bond was blocked [102]. The compound with the cis-heptene side-chain (21) had the highest affinity for CBj (0.89 [iM) with the widest separation between CBj and CB2 affinities. Pertwee et al. showed... 

A second antagonist, AM 630 (24b), a novel aminoalkylindole, was found to attenuate the ability of some cannabinoids to inhibit electrically-evoked twitches of the mouse isolated vas deferens [114]. AM 630 was a more potent antagonist of d9-THC than of anandamide (Kd of 14.0 and 278.8 nM, respectively). It was suggested that the receptors for which AM 630 has the highest activity may not be CB, cannabinoid receptors. This is supported by the observation that AM 630 is actually a cannabinoid agonist in the myenteric plexus - muscle preparation [115]. Yamada et al. [116] showed that isothiocyanate derivatives of pravadoline can serve as potential electrophilic affinity ligands for CB],... 

Semiplenamides A 359 to G 365, a series of new anandamide-like fatty acid amides, were isolated from a 1997 Papua New Guinea collection of the marine cyanobacterium Lyngbya semiplena, and all these new metabolites displayed toxicity in the brine shrimp model system. Only semiplenamides A 359, B 360, and G 365 showed weak affinity for the rat cannabinoid CBl receptor. Semiplenamide A 364 was also a moderate inhibitor (IC50 = 18.1 pM) of the anandamide membrane transporter (AMT). Novel 5y -aldol/dehydration methodology was developed for the stereoselective synthesis of the core ( )-a, P-unsaturated amide functionality of this class of natural product, and employed for the efficient synthesis of semiplenamide... 

The discovery of cannabinoid receptors naturally stimulated a search for an endogenous ligand with which the receptors naturally interact. Such a substance was isolated from the pig brain. It was found to be chemically different from plant cannabinoids it is a derivative of the fatty acid arachidonic acid (arachidonyl ethanolamide) related to the prostaglandins. This endogenous substance was named anandamide after the Sanskrit word for bliss, ananda. It has a high affinity for CBi receptors and has most of the actions of THC. 

The binding affinity of palmitylsulfonyl fluoride for the CBl receptor is about 10 times lower than anandamide. 

Very recently the authors developed a novel anandamide amidase inhibitor, AM374, whose potency in vitro and in neuroblastoma cells significantly exceeds that of other compounds developed to date as well as PMSF. In intact neuroblastoma cells, AM374 was found to dramatically increase the level of undegraded anandamide 5 5-fold at 10 nM. Interestingly, its affinity for the CBl receptor was approximately tenfold weaker than anandamide (Deutsch and Makriyannis, unpublished data). 

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