Isosorbide-2-benzyl carbamate-5-benzoate is a highly potent and selective BuChE inhibitor. Meanwhile, isosorbide-2-aspirinate-5-salicylate is a highly effective aspirin prodrug that relies on the salicylate portion to interact productively with human BuChE. By integrating the salicylate group into the carbamate design, we have produced isosorbide-2-benzyl carbamate-5-salicylate, an inhibitor of high potency (150 pM) and selectivity for human BuChE over AChE (666000) and CES2 (23000). Modeling and mutant studies indicate that it achieves its exceptional potency because of an interaction with the polar D70/Y332 cluster in the PAS of BuChE in addition to pseudosubstrate interactions with the active site.
Isosorbide-2-carbamate-5-esters are highly potent and selective butyrylcholinesterase inhibitors with potential utility in the treatment of Alzheimer's Disease (AD). They are stable in human plasma but in mouse plasma they undergo hydrolysis at the 5-ester group potentially attenuating in vivo potency. In this paper we explore the role of the 5-position in modulating potency. The focus of the study was to increase metabolic stability while preserving potency and selectivity. Dicarbamates and 5-keto derivatives were markedly less potent than the ester class. The 2-benzylcarbamate-5-benzyl ether was found to be potent (IC(50) 52 nM) and stable in the presence of mouse plasma and liver homogenate. The compound produces sustained moderate inhibition of mouse butyrylcholinesterase at 1mg/kg, IP.
We report herein that a variety of isosorbide di-esters, previously reported to be novel substrates for butyrylcholinesterase (BuChE, EC 3.1.1.8), are in fact inhibitors of the homologous enzyme acetylcholinesterase (AChE), with IC(50) values in the micromolar range. In vitro studies show that they are mixed inhibitors of the enzyme, and thus the ternary enzyme-inhibitor-substrate complex can form in acetylcholinesterase. This is rationalised by molecular modelling which shows that the compounds bind in the mid-gorge area. In this position, simultaneous substrate binding might be possible, but the hydrolysis of this substrate is prevented. The di-esters dock within the butyrylcholinesterase gorge in a very different manner, with the ester sidechain at the 5-position occupying the acyl pocket at residues Leu286 and Val288, and the 2-ester binding to Trp82. The carbonyl group of the 2-ester is susceptible to nucleophilic attack by Ser198 of the catalytic triad. The larger residues of the acyl pocket in acetylcholinesterase prevent binding in this manner. The results complement each other and explain the differing behaviours of the esters in the cholinesterase enzymes. These findings may prove very significant for future work.
Title: Pharmacological effects of a novel isosorbide-based butyrylcholinesterase inhibitor Khan D, Gilmer JF, Carolan CG, Gaynor JM, Ryder SA Ref: Chemico-Biological Interactions, 175:231, 2008 : PubMed
Isosorbide-2-benzylcarbamate-5-benzoate, a novel butyrylcholinesterase inhibitor, shows interspecies variation in its inhibitory activity (IC(50) of 4.3 nM for human plasma butyrylcholinesterase, but 1.09 microM for mouse plasma butyrylcholinesterase). Stability studies revealed that this drug is resistant to hydrolysis by human plasma (no degradation in 1 h). However, it was found to undergo rapid degradation when incubated with mouse plasma or mouse liver homogenate, yielding benzyl carbamate and benzoic acid. The addition of the carboxylesterase inhibitor bis-(4-nitrophenyl) phosphate (BNPP) inhibited the degradation of the novel drug, indicating that it may be a substrate for both butyrylcholinesterase and carboxylesterase. The absence of carboxylesterase from human plasma explains the drug's stability in this medium. In vivo, pharmacodynamic studies on single doses of 1 mg/kg to naive male C57BL/6 mice revealed maximal plasma butyrylcholinesterase inhibition 20 min after intraperitoneal administration (approximately 60% inhibition) and 1 h after administration by gavage (approximately 45% inhibition). While this plasma butyrylcholinesterase inhibition was short-lived, the drug also penetrated the blood-brain barrier resulting in a slight (10-15%) but persistent (> or =72 h) reduction in brain butyrylcholinesterase activity.
Title: Synthesis and structure activity relationships (SAR) of a new class of potent and selective butyrylcholinesterase inhibitors Gaynor JM, Dillon GP, Reidy S, Gilmer JF Ref: Chemico-Biological Interactions, 157-158:380, 2005 : PubMed
Reported here is the synthesis and SAR of novel group of highly potent and selective inhibitors of human plasma butyrylcholinesterase (BuChE; EC 3.1.1.8). The design is based on the discovery that isosorbide 2-esters are hydrolysed by BuChE at exceptionally rapid rates. Two families of carbamates were synthesised in which the vulnerable 2-ester was replaced with a carbamate or reversed carabamate. Several compounds in one of the families are among the most potent and selective BuChE inhibitors reported.
Butyrylcholinesterase [EC 3.1.1.8] present widely in mammalian tissue does not have a precisely defined biological function or known endogenous substrate. However, it plays an important role in the detoxification of certain xenobiotics and is an established vector for the systemic liberation of other drugs from their prodrugs. While investigating a series of isosorbide-based prodrugs, we discovered that BuChE catalyses the hydrolysis of esters of the simple sugar isosorbide with unusually rapidity and in some cases with remarkable regioselectivity. In this study, a series of isosorbide esters were synthesised and their rates of hydrolysis measured by HPLC following incubation in diluted plasma solution. In general, little hydrolysis of the 5-ester group could be observed but the 2-ester group was usually hydrolysed very rapidly and the hydrolysis rate exhibited an unusual dependence on the identity of the 5-group. The results indicate that while the 5-ester group is not itself hydrolysed it is important for productive binding in isosorbide diesters.
