In Silico and Ex Vivo Analyses of the Inhibitory Action of the Alzheimer Drug Posiphen and Primary Metabolites with Human Acetyl- and Butyrylcholinesterase Enzymes

Abstract

Alzheimer’s disease (AD) is the most common neurodegenerative disorder worldwide. Ongoing research to develop AD treatments has characterized multiple drug targets including the cholinergic system, amyloid-β peptide, phosphorylated tau, and neuroinflammation. These systems have the potential to interact to either drive or slow AD progression. Promising agents that simultaneously impact many of these drug targets are the AD experimental drug Posiphen and its enantiomer phenserine that, currently, are separately being evaluated in clinical trials. To define the cholinergic component of these agents, the anticholinesterase activities of a ligand dataset comprising Posiphen and primary metabolites ((+)-N1-norPosiphen, (+)-N8-norPosiphen, and (+)-N1,N8-bisnorPosiphen) were characterized and compared to those of the enantiomer phenserine. The “target” dataset involved the human cholinesterase enzymes acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Binding interactions between the ligands and targets were analyzed using Autodock 4.2. The computationally determined inhibitory action of these ligands was then compared to ex vivo laboratory-measured values versus human AChE and BChE. While Posiphen lacked AChE inhibitory action, its major and minor metabolites (+)-N1-norPosiphen and (+)-N1,N8-bisnorPosiphen, respectively, possessed modest AChE inhibitory activity, and Posiphen and all metabolites lacked BChE action. Phenserine, as a positive control, demonstrated AChE-selective inhibitory action. In light of AChE inhibitory action deriving from a major and minor Posiphen metabolite, current Posiphen clinical trials in AD and related disorders should additionally evaluate AChE inhibition; particularly if Posiphen should be combined with a known anticholinesterase, since this drug class is clinically approved and the standard of care for AD subjects, and excessive AChE inhibition may impact drug tolerability.

Dual-Mode Ratiometric Electrochemical and Turn-On Fluorescent Detection of Butyrylcholinesterase Utilizing a Single Probe for the Diagnosis of Alzheimer’s Disease

“…Here, Re was selected as the substrate molecule, cyclopropionyl chloride was selected as the triggering group, and Re-BChE was synthesized via a one-step acyl chlorination reaction. The synthesized probes were characterized by 1 H, 13 C NMR and HR-MS (Figure S1, Supporting Information). To determine the feasibility of Re-BChE as a fluorescence and electrochemical probe, the reaction between Re-BChE and BChE was analyzed using high-performance liquid chromatography−mass spectrometry (HPLC−MS).…”

“…Consequently, the prepared sensing probe was utilized successfully for rapid monitoring of BChE activity in real samples with high accuracy and good recovery. The electrochemical-fluorescence method opens a new avenue for detecting a wide variety of biological enzymes for in vitro diagnostics, thereby expanding the scope of its applications in clinical medicine and bioinformatics.The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acs.analchem.3c00974.Instruments, samples preparation, 1 H NMR and13 C NMR and mass spectra of Re-BChE, HPLC/MS, FTIR, EIS, CV obtained at SiCFME/CNT/Re, scan rate., pH effect, DFT calculations, DPVs of modification steps, selectivity and competition tests, electrode reproducibility, electrode stability, and detection of BChE in serum samples (PDF)…”

“…Recent research and experimental studies have revealed that BChE is linked to drug metabolism and neurodevelopment, as well as being involved in the cholinergic neurotransmission’s co-regulation with acetylcholinesterase (AChE). The level and activity of BChE have been shown to increase in the brains of patients with progressive stages of AD, whereas the level of AChE decreases dramatically. − Therefore, BChE could be considered a potential target for increasing AChE levels and thereby enhancing cognitive functions in advanced AD patients. − In addition, recent studies have implicated BChE in a variety of other diseases, including cancers, trait anxiety, diabetes, and cardiovascular disease, making it an essential biomarker for health. ,, For the diagnosis of biological lesions, it would be highly advantageous to develop a chemical tool with high specificity and accuracy for monitoring BChE activity.…”

Carltonine-derived compounds for targeted butyrylcholinesterase inhibition

“…1 H NMR (600 MHz, CDCl 3 ) δ: 7.45-7.40 (m, 2H), 7.39-7.34 (m, 2H), 7.34-7.28 (m, 1H), 7.23-7.17 (m, 1H), 7.00-6.94 (m, 3H), 6.92-6.88 (m, 1H), 6.87-6.84 (m, 1H), 6.73-6.67 (m, 2H), 5.87 (ddt, J = 17.1 Hz, J = 10.2 Hz, J = 6.4 Hz, 1H), 5.19 (d, J = 17.1 Hz, 1H), 5.15 (d, J = 10.2 Hz, 1H), 5.01 (s, 2H), 3.64 (s, 2H), 3.48 (s, 1H), 3.16 (d, J = 6.4 Hz, 2H), 2.74-2.65 (m, 4H). 13 (9). Compound 8 (15 mg; 0.040 mmol); NaH (2 mg; 0.048 mmol); allyl bromide (5 μl; 0.052 mmol) in dry THF (2 mL).…”

“…[5][6][7][8] In a healthy human brain, AChE dominates, constituting approximately 90% of ChE activity, with BChE contributing the remainder. 9 The human BChE gene, located on the third chromosome (3q26), exhibits polyallelism and encodes a monomeric subunit comprising 574 amino acid residues with a molecular weight of approximately 65.1 kDa. 10 BChE, synthesized in the human liver, is widespread in human plasma, brain tissue, and leg muscles, among other locations.…”