Download Drug Metabolism Biotransformation: the process whereby lipid

Drug Metabolism
Joseph K. Ritter, Ph.D., Assoc. Prof.
Biotransformation: the process whereby
lipid-soluble drugs are converted to more
water-soluble metabolites
a.k.a. drug metabolism
Implies a change in structure of the drug
Medical Sciences Building,
Room 531
[email protected]
828-1022
Department of Pharmacology and Toxicology
Medical College of Virginia Campus
Virginia Commonwealth University Richmond,
Virginia, USA
Two types biotransformations: Phase 1 reactions are nonsynthetic, Phase 2 reactions are synthetic
Drugs can undergo either type reaction alone or they can undergo
combinations of Phase 1 and Phase 2 reactions
Conversion to water-soluble products occurs through metabolic
transformations
Consequences of drug biotransformation
Drug
(D)
Alteration of structure results in a unique
molecule which can possess different
biological activities and pharmacokinetic
characteristics compared to the parent drug.
Active
Enzyme (E)
Substrates
(s)
MAJOR
Active
Inactive
More toxic
Active
-
Inactive
Metabolites
(DS)
MINOR
Active
Equally active
Metabolism can be a “Double edged sword”
Consequences of drug biotransformation
Metabolism promotes drug excretion
Sites of biotransformation: tissues
Occurs in many places in the body, but especially the liver,
gi tract, and kidneys.
Increased water solubility/ increased ionization,
especially by phase 2 reactions Æ decreases
reabsorption in GI tract and kidney
Other sites of metabolism include skin, lungs, brain,
placenta,
Prepares drug for secretion into bile and urine
Decreases entry of drugs into cells (effects on Vd)
Decreases entry of drugs into cells (effects on Vd)
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Sites of biotransformation: cellular
Different locations of enzymes that carry
out drug metabolizing reactions
Cytosol
Alcohol oxidation
Amide hydrolysis, sulfation,
glutathione conjugation
Plasma
In the liver, a major site of drug biotransformation is in the
membrane of the smooth endoplasmic reticulum
Ester hydrolysis
Cytosol
Homogenized SER purified by centrifugation =
“microsomes”
Smooth ER
P450 oxidation
Glucuronidation
Enzymes localized to microsomes include the major types
of Phase I and Phase 2 enzymes: the cytochrome P450s
and the UDP-glucuronosyltransferases, respectively.
SER is not the only cellular or extracellular location of drug
metabolizing enzymes
Mitochondria
Monoamine
oxidation
Steroid
oxidation
Oxidation I
Types of non-synthetic Phase 1 reactions
•
•
Oxidation – addition of oxygen, most common type of Phase 1
reaction
– Two types:
oxygen appears in drug metabolite structure
oxidation occurs followed by molecular
rearrangement
Reduction – removal of oxygen
Oxidative
dealkylation
N-Dealkylation
RNHCH3
oxidation / molecular
rearrangement
O-Dealkylation
•
RNH2 + CH 2O
Hydrolysis –splitting of an ester or amide bond by insertion of watersecond most common type of phase 1 reaction
ROCH 3
ROH + CH 2O
•
RSCH 3
Phase 1 reactions generally “functionalize” a chemical (adds
functional groups or unmasks functional groups such as –OH, COOH,
amine group, etc.
Functional groups serve as “handles” for synthetic type (Phase 2
reactions)
Alkyl group equals a saturated carbon side chain (methyl
or ethyl group)
S-Oxidation
R1
RNHOH
R1
R1
NH
Deamination
N
RCHCH 3
R2
R2
Oxygen in drug metabolite
R2
R3
R1
N
R2 N
R
NH2
Nicotine, methaqualone
R1
O
oxidation / molecular
rearrangementv
R2
Amphetamine,
diazepam.
OH
OH
Tertiary amines
S
R2
2-Acetylamino-fluorene,
acetaminophen.
Thioridazine, cimetidine,
chlorpromazine
R1
S
Oxygen in drug metabolite
Secondary amines
C CH3
R
CCH 3 + NH 3
O oxidation / molecular
rearrangementv
NH2
Desulfuration
O
Thiopental.
R1
R3
Oxygen in drug metabolite
6Methylthiopurine,
methitural.
Oxidation III
Aniline chlorphentermine
RNH 2
RSH + CH 2O
oxidation / molecular
rearrangement
Oxidation II
N-Oxidation
Primary amines
Codeine,
p-nitroanisole.
oxidation / molecular
rearrangement
S-Dealkylation
•
Morphine,
ethylmorphine,
benzphetamine,
aminopyrine,
caffeine,
theophylline.
R1
C
R2
S
C
R2
O
oxidation / molecular
rearrangementv
2
Oxidation IV
hydroxylation
Cytochrome P450
Phenobarbital
R-H Æ R-OH
Oxygen in drug metabolite
P450s are a major family of enzymes that catalyze oxidation
reactions
Often referred to as Microsomal Mixed Function Oxidases
because of the splitting of oxygen that occurs in the reaction.
NADPH + O2 + H+ + Drug Æ NADP + + H2O + Drug-OH
The ability to bind and utilize oxygen as a cosubstrate in the
reaction is due to the presence of a ferrous ion containing heme
group in the active P450 enzyme
Oxidation V
P450 I
the P450 reaction cycle
P450 collectively are able to oxidize thousands of drugs
NADP+
NADPH+
P 450
Reductase
Flavoprotein
(reduced)
1- Substrate Binding
Flavoprotein
(oxidized)
2- Substrate Reduction
2
e-
3- Substrate Oxygenation
P -4 5 0 -F e 2 +
RH
3
RH
3- Substrate Reduction
e-
P -4 5 0 -F e 3 +
O2
P -4 5 0 -F e 2 +
RH
O2
H 2O
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4- Substrate Rearrangement
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4- Product Dissociation
P -4 5 0 -F e 3 +
R-H
(parent drug)
Their versatility is due in part to the presence of multiple
families of enzymes, each family containing multiple family
members. Four P450s that are major contributors to drug
oxidation are CYP1A2 (12% of all drugs), CYP2B6 (20%),
CYP2E1 (6%), and CYP3A4 (28%). Others include CYP2A6,
CYP2C9, CYP2C19 and CYP2D6.
Versatility also due to broad and overlapping substrate
specificity of each P450.
Drug drug interactions stemming from competition between
drugs for P450 metabolism, especially CYP3A4 mediated
metabolism, is common
R-OH
(oxidized product)
Regulation of P450 oxidases and effect
on drug metabolism
The levels of certain P450s are higher in individuals who
drink alcohol, smoke, or use certain therapeutic drugs
(higher rates of metabolism Æ give more drug)
Other types of phase 1 reactions:
ester hydrolysis
Hydrolysis of Aspirin
E = plasma esterase
O
R
C
E
OR
+ H20
The levels of P450s are lower in the elderly and in infants
(lower rates of metabolism Æ give less drug)
R - COOH + ROH
acid
alcohol
O
O C CH3
COOH
aspirin
O
E
+ H20
OH
COOH
salicylic acid
+ HO C CH3
acetic acid
3
Ester hydrolysis II
H2N
O
C2H5
C
O
CH2
CH2
N
C2H5
procaine
(novocaine)
+ H20
H2N
E = plasma esterase
O
Amide hydrolysis
Short-acting local
anesthetic, injected
into tissue (local
effect),
metabolized
rapidly after
absorption into
blood
C2H5
C
O
H
+ HO
CH2
CH2
N
C2H5
para-aminobenzoic
acid (PABA)
PABA Æ allergic
reactions
diethylaminoethanol
Phase 2 reactions: glucuronidation
Amide hydrolysis III
Longer acting
local anesthetic
Anti-arrhythmic
UDPG is UDP-glucuronic acid
SA = salicylic acid
metabolite of aspir
E is glucuronosyl transferase
OH
E
COOH UDPG
SA
Acetylation is a major type of conjugation reaction for
drugs with amine groups (the antibacterial
sulfanilamide)
NH CO
+
HOOC
CH3
SO2NH2
Sulfanilamide
Conjugation with the amino acid glycine is another type
of conjugation reaction.
CH3
N - AT
OH
+ H 2N
COOH
SO2NH2
Acetic Acid
Acetylsulfanilamide
N-AT is N-Acetyltransferase
ether glucuronide of SA
10%
Phase 2 reactions: glycine conjugation
Phase 2 reactions: acetylation
NH2
OC6H9O6
COOH
SA
(salicylic acid)
CH2 COOH
glycine
N - AT
OH
CONH
CH2 COOH
salicyluric acid
75% major
metabolite of Aspirin
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Phase 2 reactions: glutathione conjugation
Phase 2 reactions: methylation
ACETAMINOPHEN
HNCOCH3
OH
HO
HO
CH3
CH CH2 NH
OMT
SAM
OH
CH3O
HO
HNCOCH3
PAPS
CH CH2 NH
SULFATE
OH
P-450 MIXED FUNCTION OXIDASE
45 - 50%
epinephrine
HNCOCH3
UDPGA
CH3
GLUCURONIDE
45 - 50%
HO-N-COCH3
metanephrine
OXIDATIVE STRESS (•OH, O 2 •–)
4 - 5%
HO
OH
NMT
HO
OH
CH3
HO
CH CH2 NH2
SAM
HO
CH CH2
NH
norepinephrine
epinephrine
POSTULATED
TOXIC
INTERMEDIATES
OH
LOW DOSE (1-2g)
Key Factor
NCOCH3
GLUTATHIONE
1+
HNCOCH3
HIGH DOSE (10-15g)
NUCLEOPHILIC CELL
MACROMOLECULES
HNCOCH3
O
O-, N-methyltransferase (OMT & NMT)
S-Adenosylmethionine (SAM)
OH
MERCAPTURIC
ACID
Effect of substrate (drug) concentration on
rate of metabolism
Reaction rate Æ
E + S ↔ ES → P
V/2
First order:
Rate of metabolism
proportional to
substrate concentration
Km
Alcoholic
N-Acetylcysteine
CELL
DEATH
Factors Influencing Drug Metabolism I
Enzyme Induction (slow) increases drug clearance
Plateau:
Zero order kinetics:
Rate of metabolism
independent of
substrate concentration
(constant)
Maximum velocity
(Vmax) P
CELL
MACROMOLECULES
GLUTATHIONE
OH
Diseases: Hyperthyroidism
Drugs [many]: PB, Rifampin, Phenytoin, etc.
Conditions: smoking, alcoholism
Higher doses of drugs are required
Only one induction period then stable level
Substrate concentration Æ
P450 induction
CYP1A2 is induced by cigarette smoking and consumption of
charbroiled foods (active constituents are 3methylcholanthrene and benzo[a]pyrene)
CYP2B6 and CYP3A4 are inducible by therapeutic drugs-phenobarbital, rifampin, and phenytoin.
Factors Influencing Drug Metabolism II
Enzyme Inhibition (fast) reduces drug clearance
Diseases: Hypothyroidism, Liver Disease
Drugs (many): Chloramphenicol, Cimetidine,
Disulfiram, Ethanol (acute), etc.
CYP2E1 is induced by isoniazid (drug for tuberculosis) and
by ethanol.
Dietary factors: e.g., Grapefruit juice
Conditions: Pregnancy, Aging, Newborn
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Factors Influencing Drug Metabolism III
Age: low metabolism in elderly and newborn
start low and go slow with drug dose
Nutrition: high metabolism with chronic intake of alcohol
and charcoal cooked food and lower with high
acute alcohol intake.
Factors Influencing Drug Metabolism IV
Genetic Variations:
Isoniazid [prophylaxis of tuberculosis] produces liver
injury in slow acetylators (variation in Nacetyltransferase gene 2 gene or NAT2).
Succinylcholine [surgical muscle relaxant] produces
prolonged respiratory depression (apnea) in
patients with abnormal plasma cholinesterase
which reduces the hydrolysis of succinylcholine.
Codeine (narcotic analgesic)-10-15% is converted to morphine
by CYP2D6-mediated O-demethylation (dealkylation)
CYP2D6 is polymorphic in the human population
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