Discovery of Farnesoid X Receptor Antagonists Based on a Library of Oleanolic Acid 3-O-Esters through Diverse Substituent Design and Molecular Docking Methods

The pentacyclic triterpene oleanolic acid (OA, 1) with known farnesoid X receptor (FXR) modulatory activity was modified at its C-3 position to find new FXR-interacting agents. A diverse substitution library of OA derivatives was constructed in silico through a 2D fingerprint similarity cluster strategy. With further docking analysis, four top-scored OA 3-O-ester derivatives were selected for synthesis. The bioassay results indicated that all four compounds 3 inhibited chenodeoxycholic acid (CDCA)-induced FXR transactivation in a concentration-dependent mode. Among them 3b and 3d are more active than the parent compound OA. A molecular simulation study was performed to attempt to explain the structure-activity relationship (SAR) and the antagonistic action. To the best of our knowledge, this is the first report on semi-synthetic pentacyclic triterpenoids with FXR-modulatory activities.


Synthesis of compound 4
To a stirred solution of 1a (200 mg, 0.44 mmol) in a mixed solvent of dioxane (2 ml) and water (2 ml) was added K 2 CO 3 (121 mg, 0.88 mmol) and tetrabutylammonium bromide (112 mg, 0.35 mmol). After benzyl chloride (100 μl, 0.88 mmol) was added dropwise, the mixture was heated to 80ºC and maintained for 3 h. The excess K 2 CO 3 was filtered off. The filtrate was concentrated under reduced pressure. And the residue was purified through a silica chromatography column (petroleum ether : acetone 10 : 1) to afford the product (4) as white solid (198 mg, 83.5 %).

Synthesis of compound 7
To a stirred solution of 6 (20 mg, 0.17 mmol) in dry DMF (0.2 ml) was added N-chlorosuccinimide (NCS) (5 mg, 0.037 mmol). The mixture was heated to 40ºC, and two drops of 2M HCl (aq.) was added. After 10 min, the color of the solution turned to light blue. Then the reaction was moved to room temperature. After another portion of NCS (12 mg, 0.084 mmol) was added, the mixture was stirred at room temperature for 2 h. Then water (2 ml) was added, and the aqueous layer was extracted by diethyl ether (5 ml × 3). The organic layer was collected, dried over Na 2 SO 4 and concentrated. And the residue was purified through a silica chromatography column (petroleum ether : EtOAc 15 : 1) to afford the product (7)

Synthesis of compound 8
To a stirred solution of 7 (10 mg, 0.06 mmol) in dichloromethane (DCM) (0.2 ml) was added propargyl alcohol (5 μl, 0.09 mmol) and Et 3 N (13 μl, 0.09 mmol) at room temperature. Then, the mixture was heated to 50 ºC for 2 h. After the removal of the solvent, the residue was purified through a silica chromatography column (petroleum ether : EtOAc 3 : 1) to afford the product (8) as white solid (10 mg

Synthesis of compound 2a
To a stirred solution of 8 (12 mg, 0.07 mmol) in acetone (0.4 ml) was added Jones' reagent (50 μl) at room temperature. Then, the mixture was stirred for 3 h. Then water (2 ml) was added, and the aqueous layer was extracted by diethyl ether (5 ml × 3). The organic layer was collected, washed with water (5 ml × 3), dried over Na 2 SO 4 and concentrated. The residue was recrystallized in CHCl 3 to afford the product (2a) as white solid (12 mg Synthesis of compound 9 2a (13.8 mg, 0.072 mmol) was stirred with SOCl 2 (1 ml) at 80ºC for 6 h. Then the excess SOCl 2 was evaporated under reduced pressure. And the residue was used directly for reactions below without any further purification.

Synthesis of compound 2d
After the mixture of 14 (164 μl, 2 mmol), hydroquinone (5 mg, 0.05 mmol) and 15 (0.2 ml, 2.3 mmol) was stirred at 100ºC for 10 h, another portion of 15 (0.34 ml, 4 mmol) was added, and the reaction mixture was heated to 140 ºC. After another 20 h, the reaction was stopped and concentrated. The residue was recrystallized in toluene to afford product (