Tetrahydroquinoline-Isoxazole/Isoxazoline Hybrid Compounds as Potential Cholinesterases Inhibitors: Synthesis, Enzyme Inhibition Assays, and Molecular Modeling Studies

Abstract

A series of 44 hybrid compounds that included in their structure tetrahydroquinoline (THQ) and isoxazole/isoxazoline moieties were synthesized through the 1,3-dipolar cycloaddition reaction (1,3-DC) from the corresponding N-allyl/propargyl THQs, previously obtained via cationic Povarov reaction. In vitro cholinergic enzymes inhibition potential of all compounds was tested. Enzyme inhibition assays showed that some hybrids exhibited significant potency to inhibit acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Especially, the hybrid compound 5n presented the more effective inhibition against AChE (4.24 µM) with an acceptable selectivity index versus BChE (SI: 5.19), while compound 6aa exhibited the greatest inhibition activity on BChE (3.97 µM) and a significant selectivity index against AChE (SI: 0.04). Kinetic studies were carried out for compounds with greater inhibitory activity of cholinesterases. Structure-activity relationships of the molecular hybrids were analyzed, through computational models using a molecular cross-docking algorithm and Molecular Mechanics/Generalized Born Surface Area (MM/GBSA) binding free energy approach, which indicated a good correlation between the experimental inhibition values and the predicted free binding energy.

Keywords: Alzheimer’s disease; cholinesterase inhibitors; cross-docking and MM/GBSA free binding energy; hybrid compounds.

Figure 1

Hybrid compounds from commercial drugs tested on cholinergic enzymes.

Scheme 1

Synthesis of new THQ–isoxazoline (5a-p) and THQ–isoxazole (6a-ab) hybrid compounds.

Figure 2

The Lineweaver–Burk plot of AChE/BChE (0.02 U) with acetylthiocholine/butyrylthiocholine substrate in the absence and presence of inhibitor 5n (a) and 6aa (b), respectively.

Figure 3

Predicted binding modes for studied compounds within the AChE active site. The most populated pose cluster is shown in (a), and the identified intermolecular interactions are depicted in (b). Compounds are shown in ball-and-stick representation with cyan carbons. Relevant amino acids are shown in tubes in the following coloring code: catalytic triad in yellow, anionic site in magenta, acyl pocket in green, and peripheral anionic site in orange. Secondary protein structure is depicted as white ribbons. (c). Predicted binding mode for galantamine (shown with carbons in plum) within AChE active site.

Figure 4

Scatter plot of the experimental activities (IC₅₀) converted to free binding energies (ΔG_expt) vs. predicted free binding energies (ΔG_pred). Solid line is shown for the linear regression fit (R² = 0.83).