Anti-inflammatory Activity of Indole and Amide Derivatives of Ursolic Acid: Design, Synthesis, and Docking Studies

Ursolic acid (3-hydroxy-urs-12-ene-28-oic acid, UA) is one of the most notable pentacyclic triterpene which we have isolated from the biomass marc of the lavender plant Lavandula angustifolia. Then, UA was structurally modified by an introduction of indole ring at the C-3 position and amide group at the C-17 position with the aim to enhance its pharmacological potential. Firstly, we investigated the cytotoxic potential of all the synthesized derivatives in RAW 264.7 cells. The results of the study suggested that UA and its derivatives were non-cytotoxic to macrophages up to concentrations of 10.0 µM. In the present study we used Lipopolysaccharide (LPS), a potent activator of immune cells including macrophages consequently activating the inflammatory cascade. The nitric oxide (NO) inhibition was assessed for potential of these derivatives in LPS induced RAW 264.7 macrophages wherein the results demonstrated that among all, indole derivatives significantly reduced the levels of inorganic nitric oxide in cell culture supernatant in comparison to the parent molecule (UA) and as well as amide derivatives. Furthermore, compound 3 and 6 were evaluated for its cytokine inhibitory potential wherein, the results demonstrated that compound 3 and 6 significantly reduced the LPS-induced upregulation of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β and PGE 2) and further upregulated the levels of anti-inflammatory cytokine (IL-10) in comparison to the UA in RAW 264.7 cells. The effect of these derivatives was also access on principal inflammatory mediators including iNOS, COX-2 and ROS. The results showed a reduction in the protein expression of iNOS and COX-2 indicating their potential role in dampening inflammation. Furthermore, ROS analysis demonstrated that both compound 3 and 6 significantly inhibited the LPS-induced reactive oxygen species. Additionally, molecular docking studies were performed to study the interaction of these hits with NF-κB protein. Since, it is a critical transcription factor that plays a central role in the regulation of the immune and inflammatory responses. The results of molecular docking demonstrated that these potent compounds might have worked through NF-κB signalling pathway, which might be the mechanism of action.