Celastrol Derivatives: A Reverse Docking Study Reveals Potential Anti-inflammatory and Anti-tumor Targets

Celastrol (CSL) is a pentacyclic triterpenoid terpene extracted from the root bark of Tripterygium wilfordii. It has shown remarkable efficacy in anti-cancer and cerebral neurology; however, its toxicity limits its clinical use. To overcome this issue, we have obtained a series of derivatives through biotransformation in the early stage, which have reduced toxicity and increased efficacy.

We used 23391 proteins of Homo sapiens from AlphaFold DB and the Schrödinger platform Small Molecule Drug Discovery for reverse docking with the basic scaffold of CSL to discover the pharmacological activity of its derivatives. Based on the drug target database, we selected the RORα and RORγ of Retinoic Acid Related-Orphan Receptors (RORs) family as targets for our study. We filtered the series of compounds through QikProp, docked them for dynamics simulation and molecular mechanics-generalized born surface area (MMGBSA) binding energy calculations. We also performed fluorescence polarization assay (FP assay), luciferase reporter gene assay, and the CCK8 assay.

In summary, we performed reverse docking of CSLs to find its key targets RORα and RORγ to explain its anti-inflammatory and anti-tumor effects. We found binding sites GLN19, ARG97, ARG100 for RORα-Ligand binding domain (LBD) and GLN25, LEU26, ARG103, ARG106 for RORγ-LBD, and screened for the highest affinity derivatives. The luciferase reporter gene assay showed that 2 μM 18-OH-CSL and 28-OH-CSL had the strongest agonistic effect on RORα-LBD, and 12-OH-CSL and a-ring aromatized 15-OH-CSL had the strongest inhibitory effect on RORγ-LBD. Meanwhile, the positive and negative coulomb energy in MMGBSA supported these experimental results.