Lecture 32/33 Flashcards
Anticholinoesterases - Feng
1
Q
AChEI
A
- Indirect acting cholinomimetics
- Increase ACh by decreasing AChE hydrolysis
2
Q
AChE + Hydrolysis
A
- ACh = ester
- ACh signaling is terminated by catalytic ester hydrolysis by AChE
- AChE cleaves 10,000 ACh per second, VERY efficient, reaching almost diffusion reaction rates
3
Q
Electrophiles + Nucleophiles
A
- Electrophiles love electrons (electron deficient)
- Cleavage of esters from nucleophilic attack
- Nucleophile has lone pair of electrons that donate and form new bond to electrophile
4
Q
Acid-Catalyzed Hydrolysis
A
- Ester reversibly goes between being a weak and strong electrophile by transferring partial charges
- Hydrolysis in increased by the transforming into a strong electrophile which activates the carbonyl carbon
5
Q
Base-Catalyzed Hydrolysis
A
-Accelerated by a strong nucleophile like -OH
6
Q
Catalytic Triad - AChE
A
- serine hydrolase has triad: Glu-His-Ser
- Serine acts as the nucleophile
- Histadine residue increases the nucleophilicity by removing proton from serine
- Glutamate interacts with imidazole to orient
7
Q
Hydrolysis Steps
A
- Nucleophilic attack of carbonyl carbon by deprotonated serine (base-catalyzed), acetylated AChE = inactive
- Regeneration - assisted by histadine group (acid-catalyzed), crucial step for AChEI, when acetylated with carbamyl or phosphate the enzyme is more stable when inactive and hydrolysis take a lot longer
8
Q
AChE + Acid/Base Catalysis
A
- Base-catalyzed hydrolysis portion: -OH from serine in nucleophilic attack
- Acid-catalyzed hydrolysis portion - imidazole proton protonating carbonyl O
- Makes this enzyme super efficient with fast rates by utilized both aspects
9
Q
Ester AChE Inhibitors
A
- Classic
- Carbamates or phosphoesters
- Compete for site of activation
- Close in structural and chemical properties to ACh
- Inhibits AChE so that ACh can’t bind and it returns to the receptor to activate it
- Same effect as cholinergic agonist
10
Q
Reversible Inhibitor Types
A
- Substrates that slow hydrolytic regeneration (carbamates)
2. Binds with AChE with greater affinity but does NOT react as substrate (edrophonium)
11
Q
Carbamyl Inhibitors
A
- Esters of carbamic acid
- Interacts with AChE as a substrate but increases the time to regenerate enzyme (Ex: insecticide)
- Minutes to regenerate, but the enzyme will work again (reversible)
- Form covalent bond with Serine-O
- Slower regeneration rate since carbonyl C of carbamate is less electrophilic than acetate (water is less attracted to it)
12
Q
Electronic Factors
A
-Carbonyl groups are stabilized by delocalizing nitrogen lone pair onto the carbonyl
13
Q
Aryl Carbamate
A
- Better than alkyl since aryl is a better leaving group
- Phenoxide anion drives reaction to completion with serine
- Therefore, carbamylates AChE quickly to inhibit it
14
Q
Physostigmine: SAR
A
-Tertiary amine
-Natural product from African calabar bean
-Activity varies based on pH (more activity at lower pH), contributes that the positive charge is important for activity
1. Carbamate = essential (equiv. to ester)
2 Aromatic ring = important (good leaving group)
3. Pyrrolidine N = important, ionized in blood, equiv. to quart. N of ACh
15
Q
Physostigmine Analogs
A
- Miotine
2. Neostigmine
16
Q
Miotine
A
- Simplified analog of Physostigmine
- Susceptible to hydrolysis
- Crosses BBB as free base
- CNS effects
17
Q
Neostigmine
A
- Fully ionized
- Doesn’t cross BBB
- No CNS effects
- Constant inhibition activity regardless of pH
- More stable to hydrolysis by adding extra N-Me group
- Carbamylates AChE
18
Q
Neostigmine SARs
A
- Carbamate with extra N-Me
- Benzene ring
- Quart. Ammonium
19
Q
Edrophonium
A
- Short acting inhibitor that binds to ionic site but NOT esterase site of AChE
- simple alcohol with quart. ammonium group, binds electrostatically by hydrogen bonds
- Edrophonium doesn’t make covalent bonds with AChE
- Enzyme inhibiting complex is short lived and therefore short acting
20
Q
Alzheimer’s + AChEI
A
-Important because there is decreased activity of chol. neurons, therefore increasing ACh level in brain
-This combats the loss of ACh in brain from death of chol. neurons
EX: Rivastigmine (Exelon) - carbamate-derived AChEI, analog of Physostigmine
Donepezil (Aricept) - untypical AChEI that reacts with active and peripheral site
21
Q
Non-Classic AChEI
A
-Binds to AChE with higher affinity than ACh but doesn’t react with enzyme as substrate
EX: Tacrine and Velnacrine
22
Q
Tacrine
A
- Approved by FDA for Alzheimer’s
- D/C in 2013
- Tertiary amines
- Well absorbed
- Used systemically
- Will cross BBB and enter CNS
23
Q
Irreversible AChEI
A
- Phosphoesters - pesticides
- Organo-phosphates undergo initial binding and hydrolysis by the enzyme, resulting in phosphorylated active site
- Covalent phosphorous-enzyme bonds are VERY stable and therefore VERY slow to hydrolyze (hours)
- Much longer duration of action which is considered “irreversible” since new enzyme synthesis if required to recover enzyme function
24
Q
Aging of AChEI + ChE Reaction
A
- Aging is from dealkylation from partial hydrolysis of phosphate =O to -OH
- Results are poor electrophiles
- These poor electrophiles don’t allow for hydrolysis to activate or reaction with antidote to occur
- Phosphorous atom of anionic P is less electrophilic
- Explains why these were used for insecticides and in chemical warfare (their use here pushed for the research of antidotes)
25
Warfare Use of Organo-Phosphorous Compounds
- Deadly agent in warfare (Iraq 1980-88) and terrorism (Tokyo Subway Attack 1995)
- Agent irreversibly inhibits AChE, "permanent" activation of chol. receptors by agent which leads to asphyxia due to inability to control pulmonary and diaphragm muscles which causes death
- AChE disabled for hours to days
26
DIFD
- Agent used in warfare
- Irreversible phosphorylation
- F = good leaving group
- Aging process occurs
- P-O bond is very stable (Ser-O-P)
27
Organo-Phosphorous Antidote
- Needs to cleave P-O bond
- AChE disabled, regeneration via antidote only occurs BEFORE aging
- Aged AChE is permanently disabled
28
Antidote Design
- Strong nucleophile is required to cleave P-O bond
- Needs to be suitable to cleave phosphate esters
- Water is too weak
- Hydroxylamine is a stronger nucleophile but is too toxic for clinical use
- Therefore, need to increase its selectivity by increasing its binding interactions
29
PAM
- Pyridine Aldoxime Methiodide
- Oxime = chemical compound belonging to imines
- When R1 = organix side chain and R2 = H, then its an oxime
- Quat. N is added to bind to anionic region of receptor
- Side chain designed to place hydroxylamine moiety in correct position relative to P-Serine bond
- Pralidoxime is a million times more effective than hydroxylamine
30
PAM Nuc- Site
- Oximes = used to reactivate phosphorylated AChE
- 2-PAM has nucleophilic site (N-OH) that interacts with phosphorylated site on AChE
- High degree of selectivity and strong binding affinity for AChE (due to positive charge) and carrier hydroxylamine-like nucleophile close to P-Serine
31
PAM Mechanism
- Oxime nucleophilically attacks the phosphorous atom
- Frees serine from phosphate group
- 1st line of defense against nerve gas since it reactivates ACh
32
Limits of Pralidoxime
- Only available agent to be determined as clinically effective as antidote for phosphate ester AChE poisoning
- Can inhibit AChE at doses higher than optimal dose
- Quat. ammonium can't cross BBB
- Aging decreases 2-PAM and other oxime reactivators' effectiveness
33
ProPAM
- Prodrug of 2-PAM
- Passes BBB as free base
- Oxidized in CNS to 2-PAM by riboflavin
- Then reactivated phosphorylated AChE in CNS
34
Medicinal Organophosphates
- Phospholine Iodide - used for glaucoma
- Quat. N added to increase binding (more selective, lower doses, safer)
- Utilizes similar concept to PAM
35
Additional Functional Groups
- Added on in components of nerve gas and medications
- Adding Quat. N adds extra ionic binding
- Stronger binding to AChE
- Insecticides like this are toxic and must be handled with extreme caution
36
Parathion --> Paraoxon (Insecticides)
1. Thiophosphate insecticides are lipid soluble and rapidly absorbed
2. Compounds have little AChEI activity since P -> S isn't as nucleophilic as P -> O
3. Rapidly bioactivated by desulfization by microsomal oxidation in insects to afford oxo derivatives to phosphate esters that are more potent
- Insects have more oxidizing enzymes (like P450s) than we do
37
Parathion in Mammals
- Low AChEI activity
- Metabolized into an inactive metabolite and is excreted
- Poor detoxified though due to lack of carboxylate ester, therefore still dangerous to humans
38
Malathion --> Malaoxon
- Detoxified in birds and humans
- Less toxic overall as insecticide
- Carboxylate ester bond is the site of detoxification
- Metabolized to a less toxic carboxylic acid metabolite which is rapidly excreted in urine as carboxylate anions in humans
- Lower esterase activity in insects, leading to their death
39
Aging + Malathion
- With at least one phosphoester group, aging occurs in insects
- Doesn't happen in birds and humans since it is rapidly metabolized at carboxylate ester site
- Two ester types in malathion: phosphoester and carboxylate ester
