Motifs in Natural Products as Useful Scaffolds to Obtain Novel Benzo[ d]imidazole-Based Cannabinoid Type 2 (CB2) Receptor Agonists

Abstract

The endocannabinoid system (ECS) constitutes a broad-spectrum modulator of homeostasis in mammals, providing therapeutic opportunities for several pathologies. Its two main receptors, cannabinoid type 1 (CB1) and type 2 (CB2) receptors, mediate anti-inflammatory responses; however, their differing patterns of expression make the development of CB2-selective ligands therapeutically more attractive. The benzo[d]imidazole ring is considered to be a privileged scaffold in drug discovery and has demonstrated its versatility in the development of molecules with varied pharmacologic properties. On the other hand, the main psychoactive component of Cannabis sativa, delta-9-tetrahydrocannabinol (THC), can be structurally described as an aliphatic terpenoid motif fused to an aromatic polyphenolic (resorcinol) structure. Inspired by the structure of this phytocannabinoid, we combined different natural product motifs with a benzo[d]imidazole scaffold to obtain a new library of compounds targeting the CB2 receptor. Here, we synthesized 26 new compounds, out of which 15 presented CB2 binding and 3 showed potent agonist activity. SAR analysis indicated that the presence of bulky aliphatic or aromatic natural product motifs at position 2 of the benzo[d]imidazoles ring linked by an electronegative atom is essential for receptor recognition, while substituents with moderate bulkiness at position 1 of the heterocyclic core also participate in receptor recognition. Compounds 5, 6, and 16 were further characterized through in vitro cAMP functional assay, showing potent EC50 values between 20 and 3 nM, and compound 6 presented a significant difference between the EC50 of pharmacologic activity (3.36 nM) and IC50 of toxicity (30-38 µM).

Keywords: CB2 agonists; benzimidazoles; cannabinoid receptor; natural products; synthesis.

Figure 1

THC and a representative compound from the synthesized series as a combination of a cyclic terpenoid and an aromatic core.

Scheme 1

Synthetic route for 2-alkoxybenzimidazole derivatives. a. R¹X, NaH, DMF; b. R²OH, NaH, DMF.

Scheme 2

Synthetic route for 2-aryloxybenzimidazole derivatives. a. R¹X, TBAB, K₂CO₃, DMF; b. m-CPBA, DCM; c. Cs₂CO₃, DMF, resorcinol, or 1,1-dimethylheptylresorcinol.

Scheme 3

Synthetic route for compounds 1–7. a. TsCl, DABCO, DCM; b. 2-mercaptobenzo[d]imidazole, TBAB, K₂CO₃, DMF; c. R¹X, TBAB, K₂CO₃, DMF.

Scheme 4

Synthetic route for compounds 13–15 and 18–20. a. 1-adamantanol, TFA; b. R¹X, TBAB, K₂CO₃, DMF; c. anisyl chloride, TBAB, K₂CO₃, DMF.

Figure 2

Bar graph of single dose (10 µM) inhibition of [³H]WIN55212-2 specific binding by the reported compounds.

Figure 3

Concentration–response curve of compounds 5, 6, and 16 for concentration-dependent inhibition of forskolin-stimulated cAMP accumulation in recombinant CHO cells expressing hCB2 receptors. Data for compounds 19 and 22 could not be determined. WIN55212−2 was assessed in parallel for all assays; data presented as normalized response to 100 nM WIN55212−2 maximum response.

Figure 4

Concentration–response curves of compounds on [3H]-CP55940 determined as described in Methods. Points indicate means ± SD of 3 independent experiments run in duplicate.

Figure 5

Binding modes of reported compounds docked in CB2 receptor orthosteric pocket. (A) Superposition of compounds 5, 6, 16 with WIN55212-2; (B) binding interactions established by compound 5; (C) compound 6; and (D) compound 16 within the orthosteric pocket of CB2 receptor. Transmembrane domains I–VII.

Figure 6

Dose–response curve of neutral red uptake assay of compound 6 in two different cell lines: HEK-293 and MCF-7. Data presented as normalized response relative to control (DMSO). Experiments were performed in triplicate.