Expanding the Library of 1,2,4-Oxadiazole Derivatives: Discovery of New Farnesoid X Receptor (FXR) Antagonists/Pregnane X Receptor (PXR) Agonists

Abstract

Compounds featuring a 1,2,4-oxadiazole core have been recently identified as a new chemotype of farnesoid X receptor (FXR) antagonists. With the aim to expand this class of compounds and to understand the building blocks necessary to maintain the antagonistic activity, we describe herein the synthesis, the pharmacological evaluation, and the in vitro pharmacokinetic properties of a novel series of 1,2,4-oxadiazole derivatives decorated on the nitrogen of the piperidine ring with different N-alkyl and N-aryl side chains. In vitro pharmacological evaluation showed compounds 5 and 11 as the first examples of nonsteroidal dual FXR/Pregnane X receptor (PXR) modulators. In HepG2 cells, these compounds modulated PXR- and FXR-regulated genes, resulting in interesting leads in the treatment of inflammatory disorders. Moreover, molecular docking studies supported the experimental results, disclosing the ligand binding mode and allowing rationalization of the activities of compounds 5 and 11.

Keywords: 1,2,4-oxadiazole; farnesoid X receptor antagonist; inflammatory disorders; pregnane X receptor agonist.

Figure 1

Library of 1,2,4-oxadiazole derivatives generated in this study.

Scheme 1

Reagents and conditions: (a) NH₂OH HCl, K₂CO₃ in CH₃OH, reflux; (b) N-Boc-Inp-OH, DIPEA, HBTU in DMF dry, 80 °C, 80% yield; (c) TFA:CH₂Cl₂ 1:1, 2 h, quantitative yield; (d) bromoethane, 1-bromopropane, 1-bromobutane, 1-bromopentane, 1-bromohexane, 1-bromoheptane, 2-bromopropane, 2-bromobutane, methyl 3-(bromomethyl)benzoate or methyl 4-(bromomethyl)benzoate, DIPEA, in CH₃CN dry, 60 °C, overnight, the yield of each compound is: 77% for 2, 87% for 3, 72% for 4 and 5, 63% for 6, 82% for 7, 50% for 8, 72% for 9 and 80% for 12 and 13 (e) DIBAL-H (1 M in toluene) in THF dry, 0 °C, 48 h, and 75 and 92% yields for compounds 10 and 11.

Figure 2

Panels (A–C) Effects of compounds 5 and 11 on the relative mRNA expression of FXR genes target in HepG2 cells left untreated, treated with 6-ECDCA (10 μM), compounds 5 or 11 (10 μM), or a combination of 6-ECDCA plus 5 or 11. Results are expressed as mean ± SEM. (D–F) Effects of compounds 5 and 11 on the relative expression of PXR and inflammatory cytokines in Human-CaCo2 cells left untreated, treated with TNFα (100 ng/mL) alone, or in combination with 5 or 11 (10 μM). Results are expressed as mean ± SEM. * p < 0.05 versus not treated cells (NT).

Figure 3

Binding modes of (A) compound 5 (in yellow sticks) in the X-ray structure of PXR-LBD (PDB ID 7AXE) and (B) compound 11 (in light green sticks) were observed in the X-ray structure of PXR-LBD (PDB ID 3HVL) [34]. PXR is shown as tan cartoon, while the interacting residues of the receptors are shown in tan sticks. Hydrogens are omitted for clarity. Hydrogen bonds are shown as dashed black lines.

Figure 4

Binding modes of (A) compound 5 (in yellow sticks) and (B) compound 11 (in light green sticks) in the X-ray structure of FXR-LBD (PDB ID 4OIV) [35]. FXR is shown as a gray cartoon, while the interacting residues of the receptors are shown as gray sticks. Hydrogens are omitted for clarity. Hydrogen bonds are shown as dashed black lines. The bridging water molecules are reported as red sticks with explicit hydrogens.