Drug Metabolism & Delivery Flashcards
1
Q
Drug Metabolism
A
- most metabolic products are less pharmacologically active
- different metabolites have different effects
- drug metabolism usually involves pathways for the biosynthesis of endogenous substrates
- liver is major site of drug metabolism
- drug metabolism required to convert lipophilic compounds into hydrophilic compounds to be excreted
- if lipid soluble non polar compounds are not metabolised they remain in tissued and their effects are stronger
2
Q
Types of drugs and metabolism
A
- hydrophilic drug: renal excretion
- lipophilic drug no metabolism: excretion impossible
- lipophilic drug: conversion in liver to hydrophilic metabolite
3
Q
Pharmacokinetics
A
- absorbed, metabolised, distributed, and eliminated
- needs to be considered during drug design process
4
Q
Drug Dosage
A
- influenced by pharmacokinetics
- physical and chemical properties of a drug determine its success to reach its target
- consider: bioavailability, metabolic stability, chemical stability
5
Q
Phases of Metabolism
A
Phase 1: modification to hydrophilic compound
Phase 2: conjugation with large molecules for secretion
6
Q
Phase 1 Transformation
A
- introduce or unmask a functional group by hydrolysis or oxygenation
- metabolites afterwards are inactive and can be excreted
7
Q
Phase 2 Transformation
A
- generate highly polar derivatives for excretion
- addition of glucoronide, sulfate, acetate, amino acids
8
Q
Phase 1 Metabolism
A
Oxidation
- addition of oxygen or removal of hydrogen
- occurs in liver where oxidation takes place by cytochrome P450
- oxidised products are more polar and water soluble
Reduction
- removal of oxygen or addition of hydrogen
- less common
- uses cytochrome P450 or reductase enzymes
Hydrolysis
- reaction between compound and water to give polar metabolite
- uses amidase or esterase enzymes
9
Q
Types of oxidation
A
- hydroxylation: addition of hydroxyl group
- N dealkylation: removal of methyl to give amine
- O dealkylation: removal of methyl to give alcohol
- deamination: removal of amino to give carbonyl
- N oxidation: addition of oxygen to amine to give amide
- S oxidation: addition of oxygen to sulfate
10
Q
Types of Reduction
A
- nitro reduction to amine/hydroxylamine
- carbonyl reduction to alcohol (ketone or aldehyde to alcohol)
- opposite of oxidation reactions
- cytochrome 450 enzymes working in opposite direction
11
Q
Types of Hydrolysis
A
- ester or amide to acid and alcohol or amine
12
Q
Phase One Mixed Function Oxidase
A
- mixed function oxidase system is cytochrome P450 dependent
- oxygen and reducing system (NAPDH) required
- one atom of oxygen transferred to substrate and other undergoes two electron reduction and is converted to water
- found in ER of different tissues
13
Q
Cytochrome P450
A
- catalyses hydroxylation or epoxidation of substrates
- requires NADPH-cytochrom P450 reductase: flavoenzyme with one molecule of FAD and FMN
- coenzyme transfers electrons for oxygen reduction in P450
- membrane associated
- contains heme : iron cofactor
- molecule oxygen binds heme cofactor after reduction of Fe3+ to Fe2+ and is converted to reactive form
- reactive form used for oxygenation reactions
14
Q
P450 Structure
A
- promiscuous enzyme
- large binding active site
- narrow binding for molecular oxygen on heme face
- heme coordinated by Cysteine residues
15
Q
Iron-Oxo Species
A
- free form enzyme: ferric heme with water occupying the oxygen binding site
- substrate binding
- acceptance of electrons from reductase to reduce the iron
- molecular oxygen binds to heme forming ferrous oxy species
- electron acceptance from reductase to form ferric peroxy species (TS)
- pick up proton from water to form ferric hydro-peroxy species
- second hydrogen released water from compound
- activated compound formed with free radical in ferryl oxo species capable of attacking the bound substrate
- transfer activated oxygen to the substrate (oxidation)
- water displaces substrate to regenerate active site
16
Q
What happens if substrate binding to P450 is unfavorable ?
A
- mechanisms are present to see if the interaction is favoured
- if not the ferrous oxy species falls back to ferric heme and oxygen is dissociated
- ferryl oxo and ferric hydroperoxo species can also fall back without substrate modification
17
Q
Order of Iron-Oxo species
A
- ferric heme
- ferrous heme
- ferrous oxy
- ferric peroxy
- ferric hydroperoxo
- ferryl oxo
18
Q
Phase 2 Transformations
A
- when phase 1 products are not sufficiently hydrophilic or inactive to be eliminated the metabolites undergo phase 2
- phase 2 modifies functional groups by a conjugation reaction
- conjugate large polar molecules to water soluble products
- requires a coenzyme
- catalysed by transferases
- conjugates unable to cross membranes and are therefore biologically inactive
19
Q
Types of Phase 2 Conjugations
A
- glucuronidation: sugar moiety transferred to phase 1 metabolite groups
- sulfation: sulfotransferase
- acetylation: acetyltransferase
- glutathione conjugation: glutathione S transferase
- fatty acid conjugation
- condensation reactions
- amino acid conjugation
20
Q
Aspirin Metabolism
A
- aspirin hydrolysis to salicylic acid (liver activation)
- form ether glucuronide or sulfate, ester glucuronide, salicyloyl glycine
21
Q
Glucuronidation
A
- addition of sugar moiety
- conjugation to a-D-glucuronic acid
- most important path for drug metabolism
- products excreted in bile
- uses UDP-glucuronosyltransferase enzyme
- conjugates -OH, -COOH, -NH2, -SH, C-H
22
Q
a-D-glucoronic acid
A
- cofactors binds to gluco-transferase to help transfer the sugar
1. phosphorylase adds P to a-D-glucose 1-phosphate
2. a-D-glucose 1-UDP formed
3. 2 NADH removed via UDPG dehydrogenase
4. a-D-glucose 1-UDP-glucoronic acid
23
Q
N glucuronidation
A
- occurs with amines (aromatics)
- occurs with amides and sulfonamides
24
Q
O glucuronidation
A
- occurs by ester linkages with carboxylic acids
- occurs by ether linkages with phenols and alcohols
25
Q
Sulfation
A
- major pathway for phenols, alcohols, amines, and thiols
- sulfation can make reactive byproducts
- occurs at lower substrate concentrations
- PAPS (phosphoadenosine phosphosulfate) is the most common coenzyme in sulfotransferase reactions
- conjugates -OH, -NH2
26
Q
Acetylation
A
- aromatic amines and sulfonamides
- requires N-acetyltransferase and cofactor acetyl-CoA
- acetyl-sulfonamides are less soluble than parent compound and cause renal toxicity
- conjugates -OH, -NH2
27
Q
Fatty Acid Conjugation
A
- stearic and palmitic acids are conjugated to drug via esterification
- addition of a large hydrophobic molecule
- secreted in feces
28
Q
Amino Acid Conjugation
A
- active CoA amino acid conjugates reacting with drugs by N-acetylation
- glycine, glutamine, arginine
29
Q
Glutathione Conjugation
A
- tripeptide Gly-Cys-Glu conjugated by glutathione S transferase
- conjugated compounds subsequently attacked by g-glutamyltranspeptidase and a peptidase
- products can be further acetylated
30
Q
Drug Modifications and Metabolism
A
- can make drugs more or less resistant to metabolism by removing susceptible groups or adding them
- don’t want a drug breaking down too quickly or become toxic and long lasting
- ie. methyl easily metabolised so can be exchanged for Cl (or vice versa)
31
Q
Efflux Transporters
A
- cause multidrug resistance in cancer treatment
- transporters detoxify cells from toxic compounds (P-glycoprotein)
32
Q
Factors affecting metabolism
A
- rate and pathway of drug metabolism are affected by species, sex, age, hormones, etc
- drug metabolism is stereospecific where enantiomers act as two different xenobiotics with different metabolites and pharmacokinetics
- inactive enantiomer can produce toxic metabolites or be dangerous
33
Q
Drug Blood Availability
A
- drugs can bind to plasma proteins (causes lack of bioavailability)
- only the unbound compound is available for distribution into tissues
- acidic drugs tend to bind albumin and basic drugs bind alpha 1 acid glycoprotein
- can modify drugs to change their plasma binding and bioavailability
- % plasma binding gives information about the amount actually having an activity
34
Q
Prodrugs
A
- compounds that are inactive but are converted in the body to an active drug by metabolic enzyme (activated after phase 1 metabolism)
35
Q
Benefits of prodrugs
A
- improve membrane permeability
- esters: if carboxylic acid is important for drug binding but prevents drug from crossing membrane it can be hidden as an ester
- in the blood the ester is hydrolysed to active form
- N-methylation: amines methylated to increase hydrophobicity - membrane transport: mimic substrate to cross membrane
- extend life
- 6-mercaptopurine is an immune suppressant eliminated quickly. prodrug slowly converted allows longer activity - less toxicity
- salicylic acid contains phenolic -OH causing gastric bleeding. aspirin has an ester to mask this toxic group until it is hydrolysed
36
Q
Prodrug Definition
A
- pharmacologically inactive compound converted to active drug by metabolic biotransformation
- simple chemical derivatives requiring only one to two chemical or enzymatic transformation steps to yield the active parent drug
- prodrug to drug conversion can occur before/during/after absorption or in a specific site
37
Q
Utility of prodrugs
A
- improved ADMET properties
1. absorption: improve
2. distribution: modify to cross blood brain barrier
3. metabolism and excretion
4. toxicity: alleviate toxic burden - potentially easier to formulate or administer
38
Q
Prodrug activating enzymes
A
- mostly esterases
- found in many tissues in different cellular locations
39
Q
Biopharmaceutical Classification System
A
- characterisation of drugs based on solubility and permeability measures
1. high solubility and high permeability : water soluble
2. low solubility and high permeability : prodrug optimise
3. high solubility and low permeability : prodrug optimise
4. low solubility and low permeability : membrane target
40
Q
Types of Prodrugs
A
- bio-precursors
2. carrier linked prodrugs: bipartite, tripartite, mutual prodrugs
41
Q
Ideal Drug Carrier
A
- protect drug until site of action
- localise drug at the site of action
- release drug chemically or enzymatically
- minimise host toxicity
- biodegradable, biochemically inert, nonimmunogenic
- easily prepared and inexpensive
- chemically and biochemically stable
42
Q
Carrier linked prodrugs
A
- contains active drug linked to carrier group (generally esters or amides) that can be removed enzymatically (hydrolytic cleavage)
- esters and amides are major group of carrier linked prodrugs
- drug molecules contains alcohol/carboxylic acids are esterified
- bond to carrier group must be labile to allow active drug to be released
- carrier group non toxic and inactive
43
Q
Esterases
A
- hydrolase enzymes
- liberate drug’s active form
- esters are useful in modifying lipophilicity of drugs
- aliphatic esters generally enhance lipid solubility whereas phosphate esters improve aqueous solubility
44
Q
Prodrug Funcational Groups
A
- consider which functional groups are amenable to chemical prodrug derivatisation
- these are typically carboxylic, hydroxyl, amine, phosphate, and carbonyl groups
- prodrugs produced via modification of these groups include esters, carbonates, carbamates, amides, phosphates, and oximes
45
Q
Functional Group Modifications
A
- hydroxyl, carboxyl, and amine
- esters
- sulfenamides
- carbamates
- phosphate groups
- amidine or guanine groups
- all have esterases breaking an ester bond to release drug
* * review notes **
46
Q
Alcohols and carboxylic acids
A
- most common prodrug form are esters
- esterases are ubiquitous
- esters alter lipophilicity : any degree of hydrophobicity can be made to facilitate membrane diffusion
- challenging the predict their pharmacokinetics
- significant differences in specific esterase activities in preclinical species
47
Q
General Mechanism of Esterases
A
- contain catalytic triad: His, Ser, Asp
- ester come into binding site
- His deprotonates Ser-OH
- Ser attacks the ester bond
- covalent tetrahedral intermediate
- release of alcohol
- water enters and is deprotonated for attack on Ser
- release of second alcohol and the bioactive product
- see NOTES **
48
Q
Water Solubility
A
- alcohol containing drugs acylated with aliphatic acids to decrease water solubility or with carboxylic acids to increase water solubility
- phosphate or sulfate esters also increase water solubility
- aromatic or aliphatic functional groups increase hydrophobicity
- phosphate functional groups enhance solubility
- see NOTES **
49
Q
Prodrug Aqueous solubility
A
- prodrugs can overcome solubility limitations of poorly soluble drugs when phase 1 metabolism is low to moderate
50
Q
Improved lipophilicity
A
- prodrugs frequently mask polar and ionisable groups to improve oral drug delivery
- promotes membrane permeability and oral absorption
51
Q
Examples of Prodrugs
A
- phosphate often used to increase solubility
eg. Fosamprenavir (antiviral) containing a phosphate ester of amprenavir. Bioconversion by alkaline phosphatases to increase solubility by 10 times
52
Q
Improved parenteral administration
A
- increase water solubility by an ionizable/polar promoiety
to the parent drug - increase in solubility imparted by phosphate group is several orders of magnitude
53
Q
Improved topical administration
A
- opthalmic and dermal
- unfavorable physiocochemical properties of many drug molecules lead to poor permeation across the skin
- balance of the two solubilities improves drug permeation
54
Q
Carrier mediated absorption
A
- prodrugs targeted towards specific membrane transporters are designed to have structural features that would allow them to be taken up by one of the endogenous transporters present at intestinal epithelium
- target specific transporters for polar or charged drugs
- peptide transporters are attractive targets
55
Q
Site selective drug delivery
A
- site selectivity may be achieved in 4 ways: passive enrichment in the organ/transporter mediated delivery/selective metabolic activation through enzymes/antigen targeting
- usually targeting of the CNS, tumours, and liver targeting
56
Q
Proloinged drug action
A
- highly lipophilic prodrugs are slowly released from injection site and result in a prolonged duration of action
- rapid conversion on release
eg. conversion of nonsteroidal anti-inflammatory drug to glycine conjugate increases potency and extends concentration from 1 to 9 hours due to slow hydrolysis of the amide linkage
57
Q
Bipartite prodrug
A
- one carrier attached directly to the drug
- modified lipophilicity due to the carrier
- hydrolytic cleavage releases the drug
- carrier is degraded and released
eg. Latanoprost is used to treat glaucoma. The morelipophilic isopropyl ester prodrug is hydrolysed by corneal esterases to give the biologically active soluble acid
58
Q
Tripartite prodrug
A
- carrier connect to drug through a linker (enzyme cleaved)
- modified lipophilicity due to carrier
- hydrolytic cleavage releases the drug
- linker spontaneously cleaved
e. g Ampicillin has poor oral absorption but pivampicillin is a ester tripartite prodrug of the drug. Prodrug uses a -CH2 linker to link ampicillin and pivalic acid carrier. Ester has more lipophilicity and therefore greater bioavailability - SEE NOTES **
59
Q
Tripartite Mechanism of Activation
A
- ester hydrolysis by esterases to release carrier
- spontaneous removal of the linker and release of formaldehyde
- release of active drug
60
Q
Mutual Prodrug
A
- two synergistic drugs attached to each other
- one drug is the carrier for the other
- both are active drugs
eg. Sultamicillin is mutual tripartite prodrug of ampicillin and sulbactam - gives improved pharmacokinetic properties and both drugs in combination work better on bacteria to prevent resistance
61
Q
Bioprecursors
A
- metabolised by molecular modification into a new compound which is the active principle or which can be metabolised further to the active drug
- unlike carrier linked prodrugs that are active drugs linked to a carrier and released by hydrolysis: bioprecursors cannot be converted to the active drug by simply cleavage of a group of the prodrug
- activated in the body by oxidation or reduction (phase 1 metabolism)
eg. sulfasalazine activated by reduction of its azo bond by anaerobic bacteria in the lower bowel to mesalazine and sulfapyridine: anti-inflammatory and antibacterial agents are the product - increases the amount of bioavailable compound
62
Q
Macromolecular drug carrier systems
A
- absorption and distribution of the drug depend on physicochemical properties of the carrier
- Pros: specific site target, low toxicity
- Cons: maybe not well absorbed/immunogenic
- use of glycoproteins, lipoproteins, hormones, ADC, peptides, etc
63
Q
Prodrug Challenges
A
- complex synthesis
- controlling site and rate of bioconversion and metabolism
- interpretation of the preclinical results is complicated by species differences in prodrug bioconversion
- complex analytical profiling is needed and requires analysis of the prodrug, parent drug, and each metabolite
- toxicity of not only prodrug and drug but also the released promoieties or byproducts needs to be considered
- stability must be adequate to allow drug synthesis at scale
