Exploring Structure-Activity Relationship in Tacrine-Squaramide Derivatives as Potent Cholinesterase Inhibitors

Abstract

Tacrine was the first drug to be approved for Alzheimer's disease (AD) treatment, acting as a cholinesterase inhibitor. The neuropathological hallmarks of AD are amyloid-rich senile plaques, neurofibrillary tangles, and neuronal degeneration. The portfolio of currently approved drugs for AD includes acetylcholinesterase inhibitors (AChEIs) and N-methyl-d-aspartate (NMDA) receptor antagonist. Squaric acid is a versatile structural scaffold capable to be easily transformed into amide-bearing compounds that feature both hydrogen bond donor and acceptor groups with the possibility to create multiple interactions with complementary sites. Considering the relatively simple synthesis approach and other interesting properties (rigidity, aromatic character, H-bond formation) of squaramide motif, we combined this scaffold with different tacrine-based derivatives. In this study, we developed 21 novel dimers amalgamating squaric acid with either tacrine, 6-chlorotacrine or 7-methoxytacrine representing various AChEIs. All new derivatives were evaluated for their anti-cholinesterase activities, cytotoxicity using HepG2 cell line and screened to predict their ability to cross the blood-brain barrier. In this contribution, we also report in silico studies of the most potent AChE and BChE inhibitors in the active site of these enzymes.

Keywords: 6-chlorotacrine; 7-methoxytacrine; Alzheimer’s disease; bis(7)-tacrine; cholinesterases; in silico; in vitro; squaramides; tacrine.

Figure 1

Schematic drawing for the design of tacrine-squaramide homodimers.

Scheme 1

Synthetic procedure for tacrine-squaramide derivatives 3a–3g, 4a–4g and 5a–5g.

Figure 2

Steady state inhibition of cholinesterase substrate hydrolysis by compound 4b (hAChE) and 3e (hBChE) at different concentrations. Lineweaver−Burk plots of initial velocity at increasing substrate concentrations (AChE: 0.1563–1.250 mM; BChE: 2.5–20.0 mM) are presented. Lines were derived from a linear regression of the data points.

Figure 3

Docking results for the tacrine-squaramide hybrids 4b and 3e within hAChE (A,B) and hBChE (C,D) active sites, respectively (PDB IDs: 4EY7, 4BDS). A and C—Superimposed analogues 4b and 3e (orange and purple carbon atoms, respectively) as 3D figures; B and D—2D figures 4b and 3e, respectively. Generally to A and C—important amino acid residues involved in the ligand-enzyme interactions are displayed as green and blue carbon atoms, respectively; catalytic triad residues are shown in yellow, the rest of the enzyme is represented as a light-grey cartoon. Figures A and C were generated with PyMol 1.5.0.4 (The PyMOL Molecular Graphics System, Version 1.5.0.4 Schrödinger, LLC, Mannheim, Germany), Figures B and D were created with Discovery Studio 2016 Client software.