The Natural Product Magnolol as a Lead Structure for the Development of Potent Cannabinoid Receptor Agonists

<div><p>Magnolol (4-allyl-2-(5-allyl-2-hydroxyphenyl)phenol), the main bioactive constituent of the medicinal plant <i>Magnolia officinalis</i>, and its main metabolite tetrahydromagnolol were recently found to activate cannabinoid (CB) receptors. We now investigated the structure-activity relationships of (tetrahydro)magnolol analogs with variations of the alkyl chains and the phenolic groups and could considerably improve potency. Among the most potent compounds were the dual CB₁/CB₂ full agonist 2-(2-methoxy-5-propyl-phenyl)-4-hexylphenol (<b>61a</b>, <i>K</i>_i CB₁∶0.00957 µM; <i>K</i>_i CB₂∶0.0238 µM), and the CB₂-selective partial agonist 2-(2-hydroxy-5-propylphenyl)-4-pentylphenol (<b>60</b>, <i>K</i>_i CB₁∶0.362 µM; <i>K</i>_i CB₂∶0.0371 µM), which showed high selectivity versus GPR18 and GPR55. Compound <b>61b</b>, an isomer of <b>61a</b>, was the most potent GPR55 antagonist with an IC₅₀ value of 3.25 µM but was non-selective. The relatively simple structures, which possess no stereocenters, are easily accessible in a four- to five-step synthetic procedure from common starting materials. The central reaction step is the well-elaborated Suzuki-Miyaura cross-coupling reaction, which is suitable for a combinatorial chemistry approach. The scaffold is versatile and may be fine-tuned to obtain a broad range of receptor affinities, selectivities and efficacies.</p></div

Radioligand binding results of key compounds 61, 61a, 61b.

<p>Concentration-dependent inhibition of specific [³H]CP55,940 binding by <b>61</b> (▪) at membrane preparations of CHO cells expressing (A) human CB₁, or (B) human CB₂ receptors, respectively (<i>K</i>_i CB₁∶0.145 µM, CB₂∶0.0294 µM). The biphenol <b>61</b> is substituted with two alkyl residues, a propyl residue at one side and a hexyl chain at the other side of the biphenylic core. Each alkyl side chain is located in the <i>para</i>-position of one of the phenolic hydroxyl groups. Substitution of the hydroxyl group in the <i>para</i>-position of the propyl residue (<b>61a</b> (▾)) resulted in a remarkable increase in (A) CB₁ receptor affinity (<i>K</i>_i: 0.00957 µM), while (B) CB₂ receptor affinity was barely affected (<i>K</i>_i: 0.0238 µM) compared to the parent compound <b>61</b>. An introduction of a methoxy group in <i>para</i>-position of the hexyl side chain (<b>61b</b> (•)) had different effects: (A) <b>61b</b> displayed a moderately decreased CB₁ receptor affinity (<i>K</i>_i: 0.313 µM) and (B) a drastical loss in CB₂ receptor affinity (<i>K</i>_i: 0.281 µM) compared to <b>61</b>. Data points represent means ± SEM of three independent experiments, performed in duplicates.</p

Synthesis of magnolol derivatives and analogs.

<p>(a) Pd(PPh₃)₄, Na₂CO₃, toluene, EtOH, H₂O, 100°C, 18h; (b) CH₂Cl₂, BBr₃, −78°C to rt.</p

Synthesis of intermeditates.

<p>(a) Br₂, NaHCO₃, CHCl₃, 0°C; (b) three steps, (1) <i>n</i>-butyllithium, Et₂O, −78°C; (2) B(OCH₃)₃, Et₂O, −78°C to rt; (3) HCl, Et₂O; (c) CH₃I, NaOH, benzyl-tri-<i>n</i>-butylammonium bromide, CH₂Cl₂ : H₂O (1∶ 1), 12 h, rt.</p

Potencies and Efficacies of Magnolol Derivatives and Analogs at human Cannabinoid Receptor Subtypes.^a

<p>Potencies and Efficacies of Magnolol Derivatives and Analogs at human Cannabinoid Receptor Subtypes.^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077739#nt101" target="_blank">a</a>}</p

Structural comparison of Δ⁹-THC (1), the synthetic CP55,940 (4), tetrahydromagnolol (12).

<p>Structural comparison of Δ⁹-THC (1), the synthetic CP55,940 (4), tetrahydromagnolol (12).</p

Structures of selected cannabinoid receptor ligands.

<p>Structures of selected cannabinoid receptor ligands.</p

Activities of Magnolol Derivatives and Standard Compounds at human GPR18 and GPR55.^a

<p>Activities of Magnolol Derivatives and Standard Compounds at human GPR18 and GPR55.^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0077739#nt104" target="_blank">a</a>}</p

Effects of 61, 61a and 61b on forskolin(10 µM)-induced cAMP production.

<p>CHO cells expressing (A) human CB₁, or (B) human CB₂ receptors. The maximal effect of the full agonist CP55,940 is represented by the green triangle symbol (▾). Data points represent means ± SEMs of three independent experiments, performed in duplicates.</p

Schematic illustration of microscale reaction of NTPDases at capillary inlet

<p><b>Copyright information:</b></p><p>Taken from "A capillary electrophoresis method for the characterization of ecto-nucleoside triphosphate diphosphohydrolases (NTPDases) and the analysis of inhibitors by in-capillary enzymatic microreaction"</p><p></p><p>Purinergic Signalling 2005;1(4):349-358.</p><p>Published online Jan 2005</p><p>PMCID:PMC2096555.</p><p></p> 1. Injection of a sample of 4 nl of 320 µM of ATP (substrate) in reaction buffer containing UMP (20 µM) as an internal standard in the absence or presence of test compound (potential inhibitors) (0.3 p.s.i., 5 s); 2. Injection of enzyme (0.3 p.s.i., 5 s); 3. Injection of 320 µM of ATP (substrate) in reaction buffer containing UMP (20 µM) as an internal standard in the absence or presence of test compound (0.3 p.s.i., 5 s); 4. Overlayed plugs are then allowed to stand during a predetermined period of 5 min; 5. Subsequently a −60 µA current is applied and the reaction products migrate to the detector. Electrophoresis conditions were as described in the experimental section