An additional design feature, taking advantage of the neighboring unoccupied anionic site of the poisoned enzyme, is to factor a cationic head into the antidote drug molecule. It is to be separated from the "reactivator" portion so as to "fit" and position the nucleophilic end of the drug for maximum effectiveness. One such drug is pralidoxime (Protopam chlorideR, 2-PAM), which is easily synthesized from pyridine-2-carboxaldehyde (Eq. 8.7). Figure 8-12 schematically represents such a reactivation of the poisoned enzyme. The mechanism of reactivation may be viewed as a concerted action. The covalent phosphorous-oxygen bond is broken by the oxime oxygen of 2-PAM, which has been weakened by simultaneous hydrogen bonding to the serine residue oxygen atom. The quaternized pyridine nitrogen atom of 2-PAM, by binding to the anionic site of the enzyme, can be presumed to aid in the specificity and effectiveness of the displacement reaction.12

Pralidoxime Chloride

Enzyme regeneration is 105 times faster than it is with 2HN-OH alone. Under actual poisoning conditions treatment includes 2-PAM as well as heroic doses of atropine (up to 50 mg in 24 hours) to counteract the muscarinic intoxication symptoms. Intravenous barbiturates to stop convulsions may also be utilized.

12 Oximes can exist in two stereochemical forms, syn and antl, which roughly correspond to cis and trans in olefins; pralidoxime is in the anti configuration.

Other agents useful as cholinesterase reactivators include nicotinhydroxamic acid methoiodide and the bis-compound obidoxime (not available in the United States). Toxic overdoses of carbamate AChE inhibitors are not reversible with pralidoxime-type antidotes.13 In fact, because of some anti-AChE activity of their own, they would actually aggravate the toxicity of physostigmine and neostigmine overdoses.


Nicotinhydroxamic Acid 2 2

methiodide Obidoxime Chloride

The rate at which AChE reactivators function (or for that matter spontaneous reactivation by in vitro or in vivo hydrolysis) depends very much on the size and degree of branching of the alkyl groups R. If the groups are small, such as methyl or ethyl, then active enzyme regeneration may be a matter of hours. When the bulkier and branched isopropyl group is reached (e.g., isofluorophate), no measurable spontaneous in vivo reactivation is likely to occur. Steric hindrance due to "bulk" is the apparently significant factor here.

A somewhat related concept is the decreased ability of reactivators to regenerate the free enzyme from its poisoned serine-organophosphate ester linkage as a function of time, a phenomenon called aging. Here too the effect is related to structural variations of the alkyl or alkoxy groups bonded to the phosphorous atom. This effect was noted to be more pronounced, (i.e., arising sooner) with esters of secondary (and tertiary) alcohols. It then became apparent that "aging" actually involved the loss, by hydrolysis, of an alkoxy group from the phosphorylated enzyme, at a rate proportional to its degree of branching. Thus a 2-propoxy group from an isofluorophate-inactivated enzyme would "leave" more readily than would a methoxy from malathion or metrifonate (Table 8-6) (Eq. 8.8). One plausible explanation for the decreased ability of the antidote to reactivate the "aged" poisoned enzyme is that nucleophilic approach to the phosphate ester is now electrostatically repulsed by the negative oxyanion.


Poisoned AChE "Aged"AChE (8-8)

8.6. Memory and Alzheimer's Disease

Declining memory and cognition has long been associated with aging. Recent memory is particularly affected. With increasing public awareness of Alzheimer's disease and other

13 Several texts erroneously state that they are (Bowman and Rand, 1980).

senile dementias, the tempo of research has greatly accelerated to find causes and discover drugs to treat and, ideally, to reverse the pathology. The tools for such studies have generally improved over the past decade.

The search for an underlying chemical basis has concentrated on central cholinergic involvement. It has been known for some time that muscarinic—cholinergic antagonists capable of crossing the blood-brain barrier (BBB) can impair cognitive functions and produce a temporary state of amnesia. The belladonna alkaloids in the form of extracts of the various Solanaceae plants have been used as mind-altering potions (and poisons) for millennia. Therapeutic doses, or overdoses, of these drugs today [e.g., atropine (AT), Scopolamine (SP)] can produce numerous adverse effects including blurred vision, slurred speech, drowsiness, impaired motor function, confusion, disorientation, memory impairment, and hallucinations—all ascribable to central effects. The significant relationship of cortical cholinergic innervation to memory has been established by showing that drugs capable of stimulating central cholinergic functions can reverse recent memory loss and other performance deficits induced by anticholinergics. Experiments with aged and young monkeys demonstrated more consistent improvements with young animals when physostigmine was used.


2-(N-acetamido) pyrrolidones

Cholinomimetics that can increase central cholinergic activity have produced some improvements in Alzheimer's disease; AChE inhibitors, particularly physostigmine, have been most studied. A study with an experimental potent, centrally acting anticholinesterase compound l,2,3,4-tetrahydro-9-aminoacridine (THA, Tacrine) involving Alzheimer's patients over a year-long period showed encouraging improvement without some side effects.

Other approaches that are not related to the cholinergic system that are used to treat learning, memory, and cognitive impairment irrespective of disease type are with so-called nootropics (Schindler et al., 1984). These are compounds without either sedative or stimulatory properties that may positively affect blood flow and metabolism in the brain. They tend to share the ability to protect (rats) against chemically induced disruptions of memory deficits and learned avoidance behavior. Among the promising compounds are several 2-(N-acetamido)-pyrrolidones.

Potential modulators of memory storage are also under investigation. A diversity of substances appear to facilitate memory including Substance P, naloxone, amphetamine, and vasopressin.

8.7. Cholinergic Blocking Agents

Drugs capable of blocking, or otherwise antagonizing, the effects of ACh at muscarinic cholinergic receptors and at nicotinic receptors of the NMJ of skeletal muscle will now be

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