Download Drug Metabolism

Pharmacology: Drug Metabolism (Lash)
INTRODUCTION:

Drug Metabolism: chemical alteration of xenobiotics in the body
RAR of chemical bonds
Incorporation or loss of atoms or molecules
A combination of these reactions

Xenobiotic: any compound normally foreign to living systems
Xenobiotic transformation results from:
o Enzymatic processes
o Non-enzymatic processes (ie. acid hydrolysis)
o Rearrangements (ie. of unstable metabolites)

Metabolism:
Of drugs/xenobiotics that are analogues of physiological substances: often metabolized by specific enzymes
normally responsible for metabolizing the physiological compounds
Of drugs/xenobiotics that have no endogenous counterparts: metabolized by enzyme systems that exhibit a
broad range of specificity (many appear to have evolved for this purpose)

End Result of Metabolism:
Facilitates Elimination: generally converts xenobiotics to more POLAR, HYDROPHILIC compounds that are more
readily excreted
Pharmacological Inactivation: may also occur, but NOT ALWAYS
SITES OF DRUG BIOTRANSFORMATION:

Liver: principal organ of drug metabolism (although almost all tissues have some capacity for this process)

Portals of Entry: many have significant capacity for drug metabolism
Examples:
o Lung (inhalation)
o GI tract (ingestion)
o Skin (dermal absorption)
o Nasal mucosa (inhalation)
These sites may have enzymes that exhibit higher specific activities than the liver: but total capacity for
metabolism may be less than the liver

Kidneys: also involved in drug metabolism (not just excretion)

First Pass Effect:
Orally administered drug  Absorbed from SI  Enters portal system  Travels to liver  Metabolized before
reaching circulation
o Other contributors to first-pass effect:

Extensive metabolism of some drugs in intestinal mucosa itself

Metabolism by intestinal microorganisms
Greatly restricts the oral bioavilability of some drugs

Subcellular Localization of Drug Metabolizing Enzymes:
Smooth endoplasmic reticulum (microsomal fraction)
Cytoplasm
Mitochondria
OVERVIEW OF PATHWAYS OF DRUG METABOLISM:

Two Major Categories:
Phase I: oxidation, reduction or hydrolysis reactions
o Convert parent compound to more polar metabolite
o May make the compound more readily excreted
o May reveal a functional group that can serve as a site for additional metabolism
Phase II: conjugation of a small, endogenous substrate molecule with functional groups already present on drug
or added/revealed by phase I metabolism
Important Point: either one of the above phases may occur first
PHASE I REACTIONS- MONOOXYGENASE SYSTEM (MIXED-FUNCTION OXIDASE):

Cytochrome P450 Monooxygenase System:
Overall Reaction: Drug + NADPH + O2  Oxidized Drug + NADP+ + H2O
Basics:
o Flavoprotein (NADPH cytochrome P450 oxidoreductase) + a hemoprotein terminal oxidase (cyt. P450)
o Requires NADPH (source of reducing equivalents)
o P450s located in two different compartments (SER and mitochondria) and have 2 different electron
transfer chains in these compartments
o Identified by distinct optical spectrum due to thiol ligand to heme moiety

Type I Ligands: include many drugs and environmental chemicals
 Bind hydrophobic site on the protein in close enough proximity to the heme iron to
all both perturbation of the absorption spectrum and interaction with molecular
oxygen
 Vast majority of these ligands are SUBSRATES

Type II Ligands: interact directly with heme iron of P450
 Associated with organic compounds having nitrogen atoms with sp2 or sp3
nonbonded electrons that are sterically accessible
 Vast majority of these ligands are INHIBITORS
o Important Difference Spectra:

Oxidized P450: most important difference spectra is type I

Reduced P450: most important difference spectra are carbon monoxide spectrum and type
III spectrum
 CO: characteristic Amax of 450nm
 Type III: two pH-dependent peaks at 430 and 455nm
Components Required for P450 System:
o NADPH
o NADPH-cytochrome P450 reductase (flavoprotein)
o Cytochrome P450
o Lipid environment
Cytochrome P450 Catalytic Cycle:
o Drug adds first to cytochrome P450 (Fe3+)
o Addition of electron from NADPH (Fe2+)
o Addition of O2 (Fe2+−O)
o Addition of another electron from NADPH (Fe2+−O2(-))
o Addition of 2 protons (H+) and release of water (Fe3+−O)
o Transfer of O to drug and release of hydoxylated drug (d-OH)  regeneration of P450 (Fe3+)
Cytochrome P450 Nomenclature:
o Example of CYP2C11

CYP: cytochrome P450

2: gene family

C: gene subfamily

11: individual gene
o Pseudogene: P450 name followed by a P
Cytochrome P450 in Humans:
o 18 Families and 44 Subfamilies:

CYPs for Drug Metabolism:
 CYP1 (3 subfamilies, 3 genes, 3 pseudogenes)
 CYP2 (13 subfamilies, 16 genes, 16 pseudogenes)
o Also involved in steroid metabolism
 CYP3 (1 subfamily, 4 genes, 2 pseudogenes)

Many CYPs with Other Functions Pertaining to Endogenous Compounds:
 Synthesis and elimination of cholesterol
 Regulation of blood homeostasis
 Steroid metabolism
 Arachidonic acid metabolism

Specific CYPs to Know:
 CYP3A4: involved in biotransformation of a majority of all drugs
o Expressed at significant levels (especially extrahepatically)
o
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
Now known to be a significant contributing factor to poor bioavailability
of many drugs- participates in metabolism in the GI tract
o Humans have 57 sequenced CYP genes and 58 psuedogenes

Pseudogene: defective gene that does not produce a functional protein
 Relics of gene duplications where one of the copies has degenerated and lost its
function
Cytochrome P450 exhibit a broad substrate specificity and are capable of catalyzing lots of reactions
o Drugs can also be substrates of MORE THAN 1 CYP
Reactions Catalyzed by Cytochrome P450s:
o Hydroxylations:

RH  ROH
 Aliphatic (reaction on aliphatic side chain)
 Aromatic (reaction on ring structure)
o Epoxidation:

R2C=C2R  epoxide (can occur at C=C)
o Dealkylation and Deamination:

Deamination: N-CH3  NH
o N-Oxidation:

Hydroxylamines: amide/primary or secondary amine (R2NH  R2NOH)

N-oxides: tertiary amines (R3N  R3N-O)
o Sulfur Oxidation:

R2S  R2S=O
o Dehalogenation:

RX  ROH (halogen replaced by OH)
o Desulfuration:

RSH (thiol)  ROH (hydroxyl)
Flavin-Containing Monooxygenase (FMO, Ziegler’s Enzyme):
Basics:
o Require NO additional proteins (ie. no reductase required)
o Do require NADPH
o Catalyze same overall monooxygenase reaction as cytochrome P450s
o Located in the SER
Also exhibit a broad specificity (like P450s), but do have some preference:
o Prefer oxidation of secondary and tertiary amines to hydroxyl amines and N-oxides
o Also catalyze S-oxidation
Also represent a multiple gene family: however, only 5 gene products (compared to >600 CYP gene products)
known in mammals
Reactions Catalyzed by FMOs:
o Tertiary amines  N-oxides (N-oxidation)
o Secondary amines (ie. N-Acylarylamines)  hydroxylamines
o Oxidation of hydrazines followed by removal of H2O and N2  reactive intermediates
o 2 Thiols  Disulfide bond (*ONLY IN FMOs*)
o Sulfur oxidation to form sulfides
Catalytic Cycle:
o Major Difference from CYPs: drug does not add first!
o Basic Cycle:

NADPH + H+ adds to FMO-FAD  FMO-FADH2 (NADP+)

O2 adds to give FMO-FADOOH (NADP+)

Drug (X) adds and is oxidized and released (XO)  FMO-FADHOH (NADP+)

NADP+ + H2O release, regenerating FMO-FAD
PHASE II REACTIONS:

Basics:
Require a parent drug or phase I metabolite with suitable functional groups (OH, NH2, SH) to undergo
conjugation with endogenous substances
Drug conjugates are more polar, less lipid-soluble, and more readily excreted
o Can often be less biologically active (but not always)
-



Involve High-Energy Intermediates and Specific Enzymes (Transferases):
o UDP-glucoronosyl transferase (SER)
o N-acetyl transferase (in this lecture, we are talking about the ones in the SER/microsome)
o Glutathione S-transferase (cytoplasm, SER)
o Acyl-CoA glycine transferase (cytoplasm)
o Sulfotransferase (cytoplasm)
o Transmethylase (cytoplasm)
Glucoronidation Pathways (3 Steps):
UTP + G1P  UDPG + Pyrophosphate (UDPG pyrophosphorylase)
o UDPG= uridine diphosphate glucose
UDPG + 2 NAD+ + H2O  UDPGA + 2 NADH + 2H+ (UDPG dehydrogenase)
o UDPGA= uridine diphosphate glucuronic acid (performs conjugation reaction)
O-Glucuronide Formation:
o –OH + UDPGA  O-glucuronide (UDP glucuronosyl transferase)
N-Glucuronide Formation:
o –SH + UDPGA  S-glucuronide (UDP glucuronosyl transferase)
Other Molecules Formed Via this Pathway:
o S-glucuronides
o C-glucuronides
o O-glycosides
Sulfate Conugation Pathway (3 Steps):
ATP + SO4  APS + pyrophosphate
o APS= adenosine-5’-phosphosulfate
APS + ATP  PAPS + ADP
o PAPS= 3’-Phosphoadenosine-5’-phosphosulfate (performs conjugation reactions)
Conjugation reactions occur at:
o –OH groups
o –NH groups
Glutathione Conjugation (Mercapturate) Pathway:
R-X + GSH  R-SG
o Loses glutamate (γ-glutamyl transpeptidase)
o Loses glycine (cysteinyl glycine dipeptidase) to form a cysteine conjugate
Cysteine conjugate  Mercapturate (readily excreted)
o Utilizes N-Acetyltransferases in microsome/SER
Conjugation reactions can occur at:
o Halogen side groups
o C=C
o Epoxides
FACTORS AFFECTING DRUG METABOLISM:

General:
Possible Factors: genetic, environmental and physiological factors
Most Important:
o Genetically determined polymorphisms in drug oxidations and conjugations
o Concomitant use of other drugs
o Exposure to environmental pollutants and industrial chemicals
o Disease state
o Age
Responsible for:
o Decreased efficacy
o Prolonged pharmacological effects
o Increased toxicity

Induction:
Increased synthesis of de novo cytochrome P450 protein
Leads to an increased rate of biotransformation and decreased availability of parent drug OR increased toxicity if
metabolized to a reactive species
Some drugs can induce their own metabolism as well (ie. carbamazepine)
-


Generally specific for a given P450 family, although within a family structurally diverse chemicals can have similar
effects
o Induction of CYP1A: exposure to polycyclic aromatic hydrocarbons from industrial pollutants, cigarette
smoke, and charbroiled meats (induction both in liver and extrahepatically)
o Induction of CYP3A4: glucocorticoids, anticonvulsants
o Induction of CYP2E1: isoniazid, acetone, chronic EtOH consumption
Many inducers of P450s also induce enzymes involved in phase II biotransformation
Inhibition:
Results in elevated levels of parent drug, prolonged pharmacological effects, and increased incidence of druginduced toxicity
Competition between 2 or more drugs for the active site of an enzyme may lead to decrease in metabolism of
one agent
Examples:
o Inhibition of CYP2D6: by quinidine
o Inhibition of oxidative drug metabolism:

Cimetidine and ketoconazole (form a tight complex with heme iron of cytochrome P450)

Macrolide Abx (metabolite of these compounds binds heme)

Suicide inactivators of P450 that result in destruction of heme
 Secobarbital
 Synthetic steroids (norethindrone and ethinyl estradiol)
o Inhibition of phase II enzymes: depletion of necessary co-factors
Genetic Polymorphisms:
Genetic differences in the ability of individuals to metabolize a drug through a given pathway
Can be rapid or slow metabolizers
Impaired metabolism through a given pathway associated with increased incidence of adverse effects in slow
metabolizers
All deficiencies of drug metabolism inherited as autosomal recessive traits