Improving Anti-Neurodegenerative Benefits of Acetylcholinesterase Inhibitors in Alzheimer's Disease: Are Irreversible Inhibitors the Future?

Abstract

Decades of research have produced no effective method to prevent, delay the onset, or slow the progression of Alzheimer's disease (AD). In contrast to these failures, acetylcholinesterase (AChE, EC 3.1.1.7) inhibitors slow the clinical progression of the disease and randomized, placebo-controlled trials in prodromal and mild to moderate AD patients have shown AChE inhibitor anti-neurodegenerative benefits in the cortex, hippocampus, and basal forebrain. CNS neurodegeneration and atrophy are now recognized as biomarkers of AD according to the National Institute on Aging-Alzheimer's Association (NIA-AA) criteria and recent evidence shows that these markers are among the earliest signs of prodromal AD, before the appearance of amyloid. The current AChE inhibitors (donepezil, rivastigmine, and galantamine) have short-acting mechanisms of action that result in dose-limiting toxicity and inadequate efficacy. Irreversible AChE inhibitors, with a long-acting mechanism of action, are inherently CNS selective and can more than double CNS AChE inhibition possible with short-acting inhibitors. Irreversible AChE inhibitors open the door to high-level CNS AChE inhibition and improved anti-neurodegenerative benefits that may be an important part of future treatments to more effectively prevent, delay the onset, or slow the progression of AD.

Keywords: Alzheimer’s disease; acetylcholinesterase; acetylcholinesterase inhibitor; atrophy; butyrylcholinesterase; donepezil; galantamine; methanesulfonyl fluoride; metrifonate; rivastigmine.

Figure 1

Panel (A) shows the normal ephemeral (microseconds) covalent acetyl-enzyme complex that is formed as an intermediate step in the hydrolysis of acetylcholine (shown). Panel (B) shows a schematic of a competitive inhibitor binding reversibly (spanning) the catalytic site representing donepezil or galantamine (note that the competitive inhibitor does NOT form a covalent bond with the serine sidechain OH required for acetylcholine hydrolysis). Panel (C) shows a longer-lasting covalent bond (signified by heavier red bars) formed between pseudo-irreversible inhibitors (spontaneously hydrolyzed with a half-time of hours) and the enzyme. The schematic box in Panel (C) represents the corresponding carbamoyl- or phosphoryl-enzyme covalent binding, respectively, for the case of rivastigmine, metrifonate, or DDVP, wherein the specific molecular structure of each pseudo-irreversible inhibitor intermediate not shown. Panel (D) shows an example of the irreversible sulfonyl-enzyme covalent complex (no spontaneous hydrolysis, no recovery) that permanently excludes acetylcholine binding and hydrolysis. The specific sulfonyl-enzyme covalent complex shown in Panel (D) is that formed during methanesulfonyl fluoride inhibition.

Figure 2

A computational model of the expected accumulated AChE inhibition in the CNS (upper solid line) versus peripheral tissues (lower dotted line) during three weeks of daily doses of an irreversible inhibitor (e.g., methanesulfonyl fluoride, MSF), computed as producing an equal 10% inhibition of currently active AChE in both CNS and peripheral tissues with each dose. The saw-tooth appearance of the lines shows the increment of inhibition (upward points) added with each dose. The downward slope between doses is the decrease in inhibition produced by new synthesis of the enzyme in the dose-to-dose interval. MSF disappears rapidly from blood, within a few hours, producing the pulsatile inhibition shown above. These pharmacological calculations (repeated dosing with recovery between doses) predict the accumulated effects occurring over 21 days [128]. The separation between the levels of CNS versus peripheral tissue accumulated AChE inhibition caused by differences in enzyme recovery rates, as shown above, does not occur with short-acting competitive or pseudo-irreversible inhibitors [69]. (Modified from Journal of Alzheimer’s Disease, 55, Cholinesterase Inhibitor Therapy in Alzheimer’s Disease: The Limits and Tolerability of Irreversible CNS-Selective Acetylcholinesterase, 1285–1294 (2017), with permission of IOS Press.The publication is available at IOS Press through http://dx.doi:10.3233/JAD-160733).The validity of the pharmacodynamics shown in Figure 2 was tested in an experiment in which rats were treated with methanesulfonyl fluoride (MSF), an irreversible AChE inhibitor, in accordance with the 21 day protocol modeled in Figure 2. In this experiment, which is explained in detail elsewhere [128], rats were sacrificed at the end of 21 days of treatment with MSF. As modeled by the computations, CNS AChE was inhibited much more (~75%) than AChE in peripheral tissues (<25% AChE), all without observable signs of toxicity (Figure 3). Seventy-five percent CNS AChE inhibition is at the upper end of the expected therapeutic window for AD and <25% is well below the beginning of toxicity from peripheral tissues [69,87,88,89]. Similarly, rats aged 24 months were pretreated with MSF in a computationally based 4 week protocol designed to produce ~50% CNS AChE inhibition, actually showed in 56% inhibition ex vivo, and such MSF pretreatment enhanced memory function in the aged animals to that equal to young animals [129]. The ability to produce highly selective CNS AChE inhibition without peripheral toxicity has been further confirmed in monkeys (Macaca fascicularis) treated with escalating doses of MSF over 3 months, ending with ten weeks of continuous MSF treatment at 5 times the human clinical dose. Cortical biopsies confirmed ~80% and ~45% cortical AChE and BChE inhibition, respectively, with no gastrointestinal toxicity, no neuropathy, nor any other troublesome effects [69].

Figure 3

Accumulated AChE inhibition in four rat tissues after three weeks of repeated doses of 0.3 mg/kg MSF (IM) given three times per week to approximate the smaller daily dose shown in Figure 2. The animals were sacrificed 24 h after the last injection and smooth muscle (ileum), skeletal muscle (pectoral), cardiac muscle (heart), and whole brain were assayed for AChE inhibition, compared to untreated controls. CNS is significantly more inhibited than peripheral tissues (**p < 0.01), but peripheral tissues are not different from each other. Error bars show SEM [128]. (From British Journal of Clinical Pharmacology, 75, A Randomized Phase 1 Study of Methanesulfonyl Fluoride, an Irreversible Cholinesterase Inhibitor, for the Treatment of Alzheimer’s Disease, 1231-1239 (2013), with permission Wiley Press).

Figure 4

Comparison of brain AChE inhibition produced by reversible inhibitors (donepezil, rivastigmine, and galantamine) to an irreversible inhibitior (methanesulfonyl fluoride). The reversible AChE inhibitors, because of peripheral toxicity, cannot be tolerated by patients at doses that produce more than about 25%–35% AChE inhibition in the brain [82,83,84,85,86]. In contrast, an irreversible AChE inhibitor, because of inherent selectivity for inhibiting brain AChE and the absence of peripheral toxicity, can be administered at doses that produce ~66% brain AChE inhibition [69,126], a level that is within the therapeutic window [87,88,89] and is associated with strong cognitive improvement [126].