Identification of New Lupane-Type Triterpenoids as Inverse Agonists of RAR-Related Orphan Receptor Gamma (RORγ)

Patrik F Schwarz, Alexander F Perhal, Famke Guder, Jorge Enrique Hernández González, Kerrin Janssen, Ece Sağıroğlu, Ammar Tahir, Johannes Kirchmair, Natacha Rochel, Verena M Dirsch, Ya Chen

Abstract:

Targeting retinoic acid-related orphan receptor γ (RORγ) with inverse agonists presents a promising therapeutic strategy for treating autoimmune diseases, including psoriasis, rheumatoid arthritis, and multiple sclerosis. Through structure-based virtual screening, we identified a lupane-type pentacyclic triterpenoid, (2Z)-2-(2-furanylmethylene)-3-oxolup-20(29)-en-28-oic acid (
15), as a new inverse agonist of RORγ. The compound exhibited IC
50 values of 0.4 μM and 0.9 μM in Gal4-RORγ and full-length RORγ luciferase assays, respectively. Compound
15 showed improved potency and efficacy compared to a structurally related known inverse agonist, betulinic acid. Among the four additional analogues tested (
15.1-
15.4), two (
15.2 and
15.3) also demonstrated RORγ inverse agonist activity with low micromolar IC
50 values in Gal4-RORγ luciferase assay. Real-time quantitative polymerase chain reaction experiments confirmed that compounds
15,
15.2, and
15.3 downregulated RORγ target genes. Thermal shift assays showed that both betulinic acid and
15 stabilized the RORγ ligand-binding domain. Molecular docking and structure-activity relationship analysis revealed distinct binding modes within the RORγ ligand-binding domains, further supported by site-directed mutagenesis. These findings expand the repertoire of RORγ inverse agonists based on the pentacyclic triterpenoid scaffolds.

External IDs:doi:10.1021/acs.jnatprod.5c00416