RORα: A Promising Target for Drug Development in Cancer and Immune Disorders

RORα is a tumor suppressor that exhibits reduced expression in various tumor cell lines and tissues. Researchers have been striving to discover potent, selective, and chemically synthesized ligands for RORα, as it holds great promise as a target for circadian rhythm, cancer, and metabolic and immune diseases. However, there are currently no drugs that specifically target RORα on the market. Pharmaceutical companies are exploring the use of natural products as selective agonists of RORα. For instance, GENFIT and BICOLL are developing neoruscogenin, the first potent bioavailable natural product, as a therapeutic drug for autoimmune disorders. Developing drugs that target RORα would be a significant achievement in treating related diseases.

We performed reverse docking using AlphaFold DB and Schrödinger's small molecule drug discovery platform. The highly accurate model acquired by AlphaFold2 will significantly influence biology and enable structure-based drug design. CSL, a natural active product, has anti-cancer angiogenesis and anti-rheumatoid effects and is worthy of attention. Based on the results of reverse docking, we focused on RORα and RORγ. We demonstrated the agonistic effect of CSLs on RORα and the inhibitory effect of CSLs on RORγ, which explains their anti-tumor and anti-inflammatory efficacy.

We explored the conformational features of RORα and RORγ and the effects of CSLs on protein conformational binding using various methods, including MD simulation, radius of rotation, calculation of binding free energy, hydrogen bond formation, and ligand-residue interaction network profiles. The results showed that both systems were relatively stable throughout the simulation, and hydrophobic stacking significantly contributed to the binding free energy due to the large number of aromatic and hydrophobic rings in the active site residues. The energy decomposition analysis revealed that electrostatic interactions were potentially critical binding forces, while van der Waals forces played a more prominent role in the system. The amino acids GLN19 and ARG97 in the RORα binding site and GLN25, LbleEU26, ARG103, and ARG106 in the RORγ binding site exhibited high hydrogen bond occupancy and played a crucial role in protein stability, as verified by our point mutation and dual luciferase reporter experiments.

These findings offer opportunities for designing natural modulators of RORs as potential therapeutic agents in metabolic and immune disorders. Based on our recent reverse docking study of CSLs, we found that they were natural RORα agonists with a weak inverse agonistic effect on RORγ. By calculating the binding energy of protein ligands for stable trajectories, CSLs exhibited the ability to bind directly to RORs-LBD with high affinity, showing lower binding energy than endogenous ligands. This lays a foundation for further studies on RORs and small molecule ligands to better understand the physiological functions of RORs.