Celastrol Derivatives: Target Identification and Pharmacological Activity via Reverse Docking and Experimental Validation

Celastrol (CSL) is a pentacyclic triterpenoid extracted from the root bark of Tripterygium wilfordii. It has shown outstanding efficacy in anti-cancer and cerebral neurology, but its toxicity limits its clinical use. Through biotransformation, we have obtained a series of derivatives with reduced toxicity and increased efficacy. In this study, we used AlphaFold DB and the Schrödinger platform Small Molecule Drug Discovery to perform reverse docking of CSL derivatives with 23391 human proteins, and found that RORα and RORγ of the Retinoic Acid Related-Orphan Receptors (RORs) family were key targets. We performed QikProp filtering, docked for dynamics simulation, and performed molecular mechanics-generalized born surface area (MMGBSA) binding energy calculations to select 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, while 12-OH-CSL and a-ring aromatized 15-OH-CSL had the strongest inhibitory effect on RORγ-LBD. The positive and negative coulomb energy in MMGBSA supported these experimental results. Overall, our study provides insights into the pharmacological activity of CSL derivatives through reverse docking and target identification, and identifies potential lead compounds for further development.