Muscarinic Activity and AChE Inhibitors

Drugs Affecting Cholinergic Neurotransmission

• Cholinergic neurons release acetylcholine (ACh)
• Cholinergic receptors are found in the autonomic nervous system (parasympathetic and sympathetic), somatic nervous system, and some CNS neurons.
• Cholinergic receptors are either muscarinic or nicotinic, based on their ability to bind muscarine or nicotine, respectively.

Introduction to Acetylcholine (ACh)

• Neurons releasing acetylcholine are called cholinergic.
• Cholinergic neurons are present in the autonomic (parasympathetic and sympathetic), somatic, and a few CNS neurons.
• ACh receptors are either muscarinic or nicotinic types.

Functions of Acetylcholine (ACh)

• Parasympathetic nerve impulses induce smooth muscle contraction in the GI and urinary tracts, contract the ciliary muscles of the eye and relax the blood vessels and heart.
• This function is associated with "Rest and Digest" activity.

Classes of Drugs

• Cholinomimetic (parasympathomimetic) agents stimulate the parasympathetic nervous system.
  • These include agonists of cholinergic receptors and inhibitors of acetylcholinesterase (AChE).
• Cholinolytic (parasympatholytic) agents have an affinity for cholinergic receptors but no intrinsic activity.
  • These are cholinergic antagonists.

Introduction, Continued

• ACh deficiencies can lead to several diseases; however, directly administering ACh is not an effective treatment.
• Direct ACh administration can cause non-selective effects and potentially severe adverse effects.

Indications for Cholinergic Agonists and Antagonists

• Muscarinic Cholinergic Agents (Agonists):
  • Post-surgical use to reestablish smooth muscle tone.
  • Glaucoma treatment
  • Potential cognitive disorder treatment (such as Alzheimer's disease).
• Muscarinic Antagonists (Anticholinergics):
  • Reducing smooth muscle spasms due to overstimulation (e.g., GI and urinary tract spasms).
  • Overactive bladder treatment.

Muscarinic Acetylcholine Receptor Subtypes

• Receptors vary in location and cellular response, affecting various functions like cognition, seizures, secretions, and autonomic ganglia.
• Differences exist in their ability to affect heart rate, smooth muscle, and other functions. There is a lack of highly selective antagonists, making selective treatment difficult.

Muscarinic Receptors are GPCRs

• ACh causes the dissociation of G-protein, leading to intracellular second messenger responses (e.g., cAMP, cGMP, IP3, DAG, arachidonic acid, and Ca2+).

Overview of GPCR Signaling

• The process of activation and deactivation of G proteins, involving GTP and GDP exchange.

Nicotinic Receptor

• Nicotinic receptors are present in the skeletal neuromuscular junction, adrenal medulla, and autonomic ganglia, linked to myasthenia gravis (an autoimmune disease).
• These receptors function as ligand-gated ion channels for Na+ and K+, enabling ion passage.

Acetylcholine Neurochemistry

• ACh is synthesized in cholinergic neurons by transferring acetyl from acetyl coenzyme A to choline.
• Choline is acquired from serine and from hydrolysis of ACh by AChE.
• Free ACh is rapidly hydrolyzed by AChE
• This process stops the effect of ACh.

SAR and Drug Design

• Understanding structure-activity relationships (SAR) of acetylcholine and cholinergic receptors is critical for designing synthetic drugs that mimic natural effects.
• Basic muscarinic receptor model shows needs for ester functionality and quaternary ammonium group separated by two carbons.
• Different types of bonds exist in ACh, including ester and ionic bonds.

SAR of Muscarinic Receptor Agonists

• The ester portion of ACh binds through hydrogen bonds.
• The quaternary ammonium portion binds through ionic bonds.
• At least two methyls must be bonded to the quaternary nitrogen for potency.
• A defined stereochemistry is important and following Ing's rule of five, involving the number of atoms separating important parts of the molecule.

ACh Stereochemistry

• ACh, while non-chiral, has different arrangements in 3D space caused by σ-bond rotations. Common isomers are synclinal and anticlinal. This affects interactions with receptors.

ACh Mimetics - Muscarinic Agonists

• Interactions of cholinergic agonists with muscarinic receptors trigger specific pharmacological responses related to tissue location (i.e., smooth muscle contraction, vasoconstriction, reduced heart rate).
• ACh is not an ideal therapeutic agent because it is poorly absorbed and has non-specific effects.

SAR of ACh

• Three important functional regions include acyloxy, ethylene, and quaternary ammonium groups.

SAR of ACh Quaternary Ammonium

• Muscarinic activity requires a positively charged nitrogen. Replacing methyl groups significantly affects activity, with larger alkyl groups leading to inactivity and ethyl group replacement leading to antagonistic effects.

SAR of Ethylene Bridge

• Agonist activity decreases with increasing chain length in the ethylene bridge region and Ing's Rule of Five predicts maximum potency requires no more than five atoms between the terminal N and hydrogen bond. Larger analogues tend to have antagonistic effects.

SAR of Ethylene Bridge

• Replacing hydrogen atoms with alkyl groups impacts agonist activity, with larger groups decreasing activity.
• Ing's Rule of Five pertains to the number of atoms between the nitrogen and terminal hydrogen, affecting muscarinic potency.
• Larger alkyl groups can bind to the receptor but exhibit antagonistic activity instead.

Stereochemistry and Acetyl-β-Methylcholine

• Muscarinic receptors display stereoselectivity
• S-enantiomer of methacholine is potent and equipotent to ACh, while the R-form is significantly less potent.
• AChE hydrolyzes the S-enantiomer much slower than ACh, explaining the prolonged action.
• The R-enantiomer does not get hydrolyzed by AChE and exhibits more antagonistic effects.

SAR of Acyloxy Group

• Aromatic choline esters usually act as antagonists
• Stability to hydrolysis is important
• Carbamic acid esters, e.g., Carbachol produce potent agonists of both muscarinic and nicotinic receptors.
• Carbachol is less susceptible to hydrolysis by AChE and gastric acid, which increases its absorption.

Ether and Ketone Analogues

• Choline ethyl ether exhibits muscarinic activity but is not commonly used clinically.
• Potent ketones can be designed by placing carbonyl groups in the same location as in ACh.
• The duration of action differs compared to ACh.

Practice Problem

• Ing's Rule of Five does not apply to muscarine (natural agonist).

Summary of ACh SAR

• Essential components include a nitrogen atom (positive charge), appropriate alkyl group size at the nitrogen, an oxygen atom and a two-carbon unit between the oxygen and nitrogen.
• Ing's rule should apply.

Clinically Relevant Therapeutics:

• Methacholine chloride, a racemic mixture, is clinically important.
• A selective muscarinic agonist with the S-enantiomer being more potent.
• Carbachol is a carbamate analogue of ACh with no receptor specificity and high resistance to hydrolysis.
• Pilocarpine HCl is unique muscarinic agonist. Different routes of administration are available

Clinically Relevant Therapeutics

• Bethanechol Chloride (Urecholine) is a selective muscarinic agonist which can be administered orally.
• Pilocarpine HCl has diverse uses and is a notable muscarinic agonist.

Clinically Relevant Therapeutics

• Cevimeline HCl is a non-classical muscarinic antagonist.

AChE Inhibitors (AChEls)

• AChEls (anticholinesterases) interfere with ACh breakdown in the synapse.
• Increasing ACh concentration leads to increased muscarinic and nicotinic effects.

AChE Inhibitors (AChEls)

• AChEls (anticholinesterases) are indirect cholinomimetics, not acting on the receptor itself, but increasing the synaptic concentration of ACh.
• Used in various disorders like myasthenia gravis, glaucoma, and some cognitive conditions with insecticides and chemical warfare agents as examples.

AChE Mechanism of Hydrolysis

• Hydrolysis of ACh occurs via sequential steps involving the catalytic serine residue of AChE.
• Acetylated AChE is inactive and cannot bind any more ACh molecules.

Reversible AChEls

• Type I reversible AChEls are substrates for AChE, forming acetylated enzyme intermediates.
• Aryl and alkyl carbamates are examples, and hydrolysis of the carbamate leads to regeneration of the active AChE (slower hydrolysis compared to acetylated enzyme).
• Aryl carbamates show better affinity (stronger inhibition) compared to alkyl carbamates.

Physostigmine

• Physostigmine, a tertiary amine, is more lipophilic than other AChEls.
• It readily crosses the blood-brain barrier (BBB).

Other Aryl Carbamates

• Neostigmine, also a carbamate ester and quaternary ammonium salt, is similar in structure and MOA to physostigmine.
• Others, like pyridostigmine (Mestinon) and edrophonium chloride (Enlon) vary in structure but are valuable medicinal chemicals

Irreversible AChEls

• Phosphate esters are extremely stable to hydrolysis.
• Irreversible AChE inhibitors like echothiophate iodide form very stable phosphate-enzyme complexes and are used topically but have a long-lasting effect.

Acetylcholine Antagonists - Muscarinic Antagonists

• Muscarinic antagonists have a high affinity for muscarinic receptors but do not have intrinsic activity.
• They act as competitive antagonists, thus producing the opposite effect of ACh.
• Clinically, they are used for GI and respiratory tract issues, overactive bladder and other issues

Clinically Relevant Agents (SAR)

• Atropine provided a structural model for designing muscarinic antagonists. The configuration of tropane rings places the amine and ester groups the same distance apart as in ACh.

SAR of Muscarinic Antagonists

• Substituents R1 and R2, crucial for potency, should be carbocyclic or heterocyclic rings.
• The R3 substituent influences binding strength, with hydroxyl and hydroxymethyl groups being particularly effective in enhancing binding by participating in hydrogen bonding.
• The X substituent may be an ester or an ether group.

SAR of Muscarinic Antagonists

• The nitrogen substituent is frequently a quaternary ammonium group but not always necessary.
• The distance between the amine and the substituted carbon is not critical in potency.
• Two methylene units are often present in highly potent compounds.

Final Comments Muscarinic Antagonists

• Antagonists compete with ACh at the receptor binding site.
• Size and steric factors in the R1, and R2 substituents play a key role in the efficiency of competing with ACh.
• Anticholinergic agents with quaternary ammonium groups are frequently administered orally or parenterally
• The nature of the substituent on the tertiary nitrogen greatly affects absorption rates..

This quiz explores the structural requirements for compounds to exhibit muscarinic activity and the criteria of Ing's Rule of Five. Additionally, it covers various aspects of acetylcholinesterase (AChE) inhibitors and their effectiveness in pharmacology. Test your knowledge on how different structural modifications impact muscarinic potency and enzyme interaction.