Xenobiotic Biotransformation Basic Concepts
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Transcript of Xenobiotic Biotransformation Basic Concepts
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Copyright 2008, The Johns Hopkins University and Michael A. Trush. All rights reserved. Use of these materialspermitted only in accordance with license rights granted. Materials provided AS IS; no representations orwarranties provided. User assumes all responsibility for use, and all liability related thereto, and must independently
review all materials for accuracy and efficacy. May contain materials owned by others. User is responsible forobtaining permissions for use from third parties as needed.
This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike License. Your use of thismaterial constitutes acceptance of that license and the conditions of use of materials on this site.
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Section A
Biotransformation: Basic Concepts
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Renal Excretion of Chemicals
Afferent arteriole
Efferent arteriole
Glomerulus
Bowmans capsule
Proximal tubule Water soluble
Lipid soluble
Filtered drugPassive
reabsorption
Excretion and/or further passive reabsorption
Unltered drug
Activesecretion
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Biotransformation of Xenobiotics
Biological basis for xenobiotic metabolism:
! To convert lipid-soluble, non-polar, non-
excretable forms of chemicals to water-soluble, polar forms that are excretable in
bile and urine
Continued
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Biotransformation Reactions
! Phase I Reactions
Enzymatic reactions that addor
exposefunctional groups to xenobioticssuch as -OH, -SH, -NH2
or COOH
Functional groups are analogous tohaving a trailer hitch on a vehicle
Continued
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Biotransformation Reactions
! Phase II Reactions
Enzymatic reactions that result in the
conjugation of large water-soluble,charged (polar)biomolecules to
xenobiotics
For these reactions to occur, afunctional group must be present oneither the parent compound or itsPhase I product
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Foreign Chemical(xenobiotic)
lipophilic not charged
not water soluble poorly excretable
TRUCK
The Truck-Hitch-Trailer Analogy to Xenobiotic Biotransformation
Photo by John Pittman. Creative Commons BY-NC-SA.
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Foreign Chemical(xenobiotic)
lipophilic not charged
not water soluble poorly excretable
TRUCK HITCH
Phase 1 enzymesadd or expose a
functional group
still lipophilic
possibly reactive
poorly water soluble poorly excretable
catalyzed by P450s
The Truck-Hitch-Trailer Analogy to Xenobiotic Biotransformation
Photo by John Pittman. Creative Commons BY-NC-SA.
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Foreign Chemical(xenobiotic)
lipophilic not charged
not water soluble poorly excretable
TRUCKHITCH
Phase 1 enzymesadd or expose a
functional group
still lipophilic
possibly reactive
poorly water soluble poorly excretable
catalyzed by P450s
Phase 2 enzymesconjugate (transfer)
endogenous molecules*to the functional group
TRAILER
not lipophilic
usuallynot reactive
water soluble products excretable
catalyzed by transferases
*sugars, amino acids, sulfates, acetyl groups
The Truck-Hitch-Trailer Analogy to Xenobiotic Biotransformation
Photo by John Pittman. Creative Commons BY-NC-SA.
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Section B
Biotransformation: Enzymes
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Organ and Cellular Location of
Biotransformation Enzymes! Organs involved in biotransformation
Liver
Lung
Kidney
Intestine
Enterocytes
Gut flora (contribute to entero-hepaticcirculation)
Skin
Gonads
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Biotransformation Enzyme-
Containing Cells in Various Organs
Organ Cell s)
Liver Parenchymal cells (hepatocytes)
Kidney Proximal tubular cells (S3 segment)
Lung Clara cells, Type II alveolar cells
Intestine Mucosa lining cells
Skin Epithelial cellsTestes Seminiferous tubules, Sertoliscells
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Nature of the Xenobiotic
Metabolizing Enzyme System! Phase I metabolism
Small molecular weight changes like
hydroxylation, reduction, hydrolysis, etc.
In general, Phase I metabolismprepares the xenobiotic for subsequent
Phase II reactions
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Cytochrome P450
Characteristics! Can metabolize many xenobiotics (broad
substrate specificity)
!
Can catalyze many types of reactions! Is widely distributed among tissues, and
tissue distribution can be quite varied
Continued
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Multiple Forms of P450Major Mammalian Cytochrome P450 Gene Families
Continued
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Multiple Forms of P450Major Mammalian Cytochrome P450 Gene Families
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Nature of the Xenobiotic
Metabolizing Enzyme System! Phase II metabolism
Involves the complexing or conjugation
of xenobiotics with relatively large andhighly water-soluble adducts to form
glucuronides, sulfates, and glutathioneadducts
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Nature of Systems Involved in
Phase II MetabolismFour primary enzymes:
1.
Glucuronosyltransferaseglucuronic acid
2.
Sulfotransferasesulfate3.
Glutathione-S-transferaseglutathione(GSH)
4.
Acetyltransferaseacetyl
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Phase II Reactions
! Many of the characteristics describedearlier for cytochrome P450 also apply tothese Phase II enzymes
! However, cytochrome P450 is localized incellular membraness, whereas Phase IIenzymes are, for the most part, in the
cytoplasm (water portion) of cells
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Phase II Enzymes: ExamplesGlucuronidation and Sulfation of a Hydroxyl Group
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Glutathione-S-TransferaseStructure of Reduced Glutathione (MW 307)
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Section C
Factors Affecting Biotransformation
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Factors that Affect
Xenobiotic Biotransformation
Likewise, even within a species,including man, there are differences
The basis of such differences is oftengenetic (polymorphisms)
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Examples of Factors that Affect
Xenobiotic Biotransformation! Species, strain, and genetic variation
- Hexobarbital
Aflatoxin B1
Benzo[a]pyrene 7,8 dihydrodiol
Isoniazid
!
Age! Diet
! Exposure to other chemicals
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Species Differences in the Duration ofAction and Metabolism of Hexobarbital
Species Duration of
ction
(Sleeping time, min)
Relative Enzyme
ctivity
(!g hexobarbitolmetabolized/gm/liver/hr)
Mouse 12 8 598 184
Rabbit 49 12 196 28
Rat 90 15 134 51Dog 315 105 36 30
Source: G.P. Quinn, et.al. Species, Strain and Sex Differences in the Metabolism of
Hexobarbital, Aminopyrine and Aniline. Biochem. Pharmacol. 1:152, 1958
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Species, Strain,
and Genetic Variation
Continued
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A B D E F G H I J
1000
2000
3000
4000
C A B D E F G H I JC
His+revertants/mg
Biopsy samples
Metabolism of Aflatoxin B1
and Benzo[a]pyrene
Inter-individual differences in the metabolism of aflatoxin B1 (A)and benzo[a]pyrene 7,8-dihydrodiol (B) to mutagens( assessed
by Ames test) by microsomes from samples of human liver
obtained during abdominal surgery
A B
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The Bimodal Distribution of Patients intoThose who Rapidly Inactivate Isoniazid and
Those who Slowly Metabolize It50
40
30
20
10
0 1 2 3 4 5 6 7 8 9 10 11 12
Plasma Isoniazid ( g/mL)
NumberofPati
ents
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Age as Affecting Xenobiotic
Biotransformation
Schematic representation of the ontogeny ofhepatic drug metabolic activity
Birth Puberty Adult Elderly (>65 yrs)
Age
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Exposure to Other Chemicals
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Section D
Induction of
Biotransformation Enzymes
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Induction of Xenobiotic
Metabolizing Systems1.
Many chemicals can induce the synthesisof the enzymes involved in Phase I and IIxenobiotic metabolism and include
chemicals found in the environment, thediet, and cigarette smoke
2.
Inducers often exhibit specificity for the
enzymes which they induce
Continued
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Induction of Xenobiotic
Metabolizing Systems3.
Depending on the inducer, fairly highdose levels or repeated dosing may berequired; on the other hand, TCDD
(dioxin) is effective as an inducer at1 microgram/kg in some species
Continued
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Induction of Xenobiotic
Metabolizing Systems4.
Studies have demonstrated that acluster of genes referred to as the
Ah locuscontrols the induction of
xenobiotic enzyme activities by polycyclicaromatic compounds and TCDD
Continued
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Induction of Xenobiotic
Metabolizing Systems5.
Such toxic responses as cancer, chemical-induced cataracts, aplastic anemia, andfetal toxicity have been demonstrated to
be affected by this cluster of genes
6.
Evidence exists for theAh locusin man
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Characteristics of the Hepatic Effects ofPhenobarbital and Polycyclic Aromatic Hydrocarbons
haracteristics Phenoba rbital
Polycyclic
Hydrocarbons
Onset of effects 812 hours 36 hours
Time of maximum effect 35 days 2448 hours
Persistence of induction 57 days 512 days
Liver enlargement Marked Slight
Protein synthesis Large increase Small increase
Phospholipid synthesis Marked increase No effect
Liver blood flow Increase No effect
Biliary flow Increase No effectGlucuronidation Increase Small increase
Glutathione conjugation Small increase Small increase
Epoxide hydrolase Increase Small increase
Cytosolic receptor None identified Identified
Continued
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TheAh Receptor
!Ah receptor = Arylhydrocarbon receptor
! Examples = 3-methylcholanthrene
benzo[a]pyrene!Also called TCDD receptoror dioxin
receptor
Continued
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Bioactivation as a Basis for
Chemical Toxicity! One of the possible results of the
interaction of a xenobiotic with enzymesystems is the biotransformation of that
compound to a chemically reactiveintermediate (i.e. Bioactivation)
! The reaction of either this initial reactive
metabolite or secondary reactiveproducts with target molecules bringsabout changes in cellular function (the
Molecular Targets Concept)
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Proposed Relationship Between Biotransformation,Bioactivation, and Toxicity of a Xenobiotic
49Adapted from Casarett & Doulls Toxicology. 4
th
Edition.
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Metabolism and Bioactivationof Benzo[a]pyrene
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Chemical Nature ofReactive Intermediates
! ElectrophilesForm covalent (irreversible)bonds with cellular nucleophiles such asGSH, proteins and DNA
! Free RadicalsOdd or unpaired electron
Can act as electrophiles
Can abstract hydrogen from targetmolecules, such as lipids or nucleic acids
Can activate molecular oxygen
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Acetominophen is good exampleof a xenobiotic whose toxicity is
due to bioactivation to anelectrophile
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Relationship between hepatic glutathione levels and
covalent binding of acetaminophen to targetnucleophiles(proteins)
Bioactivation of Acetaminophen
100
80
60
40
20
20 40 60 100 200 400 600 10000
1
2
CovalentB
inding(molecu
les/mgprotein
)
InitialGlutathionein
Liver(%)
Dose of Acetaminophen (mg/kg)
Covalent binding
Glutathione
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Human paraquat exposure canresult in lung toxicity due to its
accumulation in lung cells andredox cycling
! The structure of the herbicide paraquat (A)and the polyamines putrescine (B) and
spermine (C)
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Redox Cycling of Xenobiotics
! Redox cyclingof xenobiotics initially resultsin the formation of a form of active oxygencalled superoxide(O2
)
!
Through a series of non-enzymatic oftenmetal catalyzed, reactions other forms ofreactive oxygen are formed
These include hydrogen peroxide(H2O2), the hydroxyl radical ( OH) andsinglet oxygen (1O2)
Continued
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Redox Cycling of Xenobiotics
! This results in an oxidative stressin cellsand the subsequent modification of criticalbiomolecules leading to cellular toxicity
!
In this situation what is the active formthat causes toxicity?