Aryl-Cyclohexanone as a potential CB2 agonist: in vitro and in silico evidence in inflammatory modulation

Inflammation is an evolutionarily conserved adaptive process essential for host defense against harmful stimuli, aimed at restoring homeostasis. However, exacerbated or dysregulated responses are associated with the progression of several autoimmune and chronic inflammatory diseases, representing a significant clinical challenge. The endocannabinoid system has been described as an important immunomodulatory pathway in which synthetic cannabinoids exert immunosuppressive effects, providing novel therapeutic targets. Aryl-Cyclohexanone (AD) has previously demonstrated anti-inflammatory properties in both in vitro and in vivo models.

Herein we aimed to investigate its immunomodulatory profile in vitro, using lipopolysaccharide (LPS)-induced inflammation in murine macrophages (J774), and its potential agonist action on CB2 receptor using human macrophages (THP-1), as well as confirm the specific and stable connection with cannabinoid receptor type 2 (CB2) through in silico docking and molecular dynamics analyses.

In J774 macrophages, AD treatment preserved cell viability, reduced nitric oxide metabolites production, normalized apoptotic events, and enhanced phagocytosis. Additionally, decreased Toll-like receptor 4 and increased mannose receptor (CD206) expressions were observed, along with a significant reduction in pro-inflammatory cytokines production (IL-12p70, TNF-α, IFN-γ, MCP-1, and IL-6). In THP-1 macrophages, the compound maintained its anti-inflammatory activity only in the absence of the selective CB2 inverse agonist (SR144528), preserving cell viability and reducing nitric oxide metabolites and pro-inflammatory cytokine production. These findings indicate that its immunomodulatory effect is directly associated with CB2 receptor interaction. Complementary silico analyses confirmed a specific and stable interaction with CB2 receptor, supporting a relevant agonistic activity observed in in vitro experiments.

The results demonstrate that AD exerts significant anti-inflammatory and immunomodulatory effects, associated with CB2 receptor agonism, highlighting its potential as a promising candidate for developing therapeutic strategies for handling inflammatory diseases.