Indole- and Pyrazole-Glycyrrhetinic Acid Derivatives as PTP1B Inhibitors: Synthesis, In Vitro and In Silico Studies.

Ledy De-la-Cruz-Martínez,Juan Manuel Germán-Acacio,Constanza Duran-Becerra,Martin González-Andrade,Olivia Soria-Arteche,Francisco Cortés-Benítez,Juan Francisco Palacios-Espinosa,J. Martin Torres-Valencia,José C. Páez-Franco,Julio Espinosa-Chávez,Jaime Pérez-Villanueva

doi:10.3390/molecules26144375

Abstract

Regulating insulin and leptin levels using a protein tyrosine phosphatase 1B (PTP1B) inhibitor is an attractive strategy to treat diabetes and obesity. Glycyrrhetinic acid (GA), a triterpenoid, may weakly inhibit this enzyme. Nonetheless, semisynthetic derivatives of GA have not been developed as PTP1B inhibitors to date. Herein we describe the synthesis and evaluation of two series of indole- and N-phenylpyrazole-GA derivatives (4a–f and 5a–f). We measured their inhibitory activity and enzyme kinetics against PTP1B using p-nitrophenylphosphate (pNPP) assay. GA derivatives bearing substituted indoles or N-phenylpyrazoles fused to their A-ring showed a 50% inhibitory concentration for PTP1B in a range from 2.5 to 10.1 µM. The trifluoromethyl derivative of indole-GA (4f) exhibited non-competitive inhibition of PTP1B as well as higher potency (IC50 = 2.5 µM) than that of positive controls ursolic acid (IC50 = 5.6 µM), claramine (IC50 = 13.7 µM) and suramin (IC50 = 4.1 µM). Finally, docking and molecular dynamics simulations provided the theoretical basis for the favorable activity of the designed compounds.

Highlights

For many years, protein tyrosine phosphatase 1B (PTP1B) has been known for its role in the negative regulation of the insulin and leptin signaling pathway [1,2,3,4]
A Using the X-ray structure of PTP1B (PDB ID: 1C83); b Using the X-ray structure of PTP1B (PDB ID: 1T49); c Binding energy values retrieved from Autodock; d Binding energy values retrieved from Autodock Vina; e CHEMPLP score values retrieved from GOLD; f GoldScore values retrieved from GOLD
We evaluated Root Mean Square Deviation (RMSD) fluctuations of the Cα carbons to know the stability of each protein–ligand complex

Summary

Introduction

Protein tyrosine phosphatase 1B (PTP1B) has been known for its role in the negative regulation of the insulin and leptin signaling pathway [1,2,3,4]. We synthesized two series of GA derivatives (Figure 1): series one regroups six A-ring-fused indoles, whereas series two regroups six A-ring-fused N-phenylpyrazoles It is well-known that the catalytic site of PTP1B is a positively-charged pocket formed by side chains of Tyr, Arg, Asp, Lys120, Phe182, Ile219, Asp181 and Gln262 [44], whereas the allosteric binding site has a hydrophobic pocket formed by side chains of Leu192, Asn193, Phe196 Glu276, Lys279, and Phe280 [45]. UA is a natural ursane-type pentacyclic triterpenoid that has been identified as a potent PTP1B inhibitor This compound enhances insulin receptor signaling and stimulates glucose uptake in vitro [46]. Reagents and conditions: (a) CrO3/H2SO4, acetone 0 °C, overnight; (b) ethyl formate, NaH, dioxane, 40 °C, 1 h 100 °C, 4 h; (c) Ar-NHNH2 hydrochloride, AcOH, reflux, 2 h; (d) Ar-NHNH2 hydrochloride, EtOH, 70 °C, 1 h

NMR Characterization of Compounds 4f and 5f

PTP1B Inhibitory Activity of Indole- and Pyrazole-GA Derivatives

Enzymatic Kinetic Studies

Enzymatic 4Kfinetic Studies

Molecular Docking Studies

Molecular Dynamics Simulations

General

Synthesis

Synthesis of N-Phenylpyrazole Derivatives 5a–f

PTP1B Inhibitory Assay

Enzyme Kinetics

In Silico Studies