metabolism...mechanism based inhibitors (suicide inhib) fe(ii) heme in cyp sub families ex. alkynes...
TRANSCRIPT
Metabolism
•Chemical transformaion of xenobiotics
•Occurs in mostly in liver (enzymatic prosesses)
•Convertion into more hydrophil. subst. - excretion urine
•May convert procarciogenics into cytotox., muthagenic compounds
•Different persons may have differences in methabolism (genetic diff., physiol. factors)
•Methabolism of one xenobiotic may influence metab. of amother
Xenobiotics
•Drugs
•Other foreign non-essential compounds
Metabolism in non-hepatic tissue
•Intestine mucosa
•Kidney
•Lung
•Bacteria in GI-tract
First-pass metabolism:
Xenobiotic metabolized before reaching
general circulation
First-pass metabolism:
Xenobiotic metabolized before reaching general circulation
A) Metab. lungs (inhaled subst)
Intestine mucosa, GI bacteria
B)
GI Liver
General Circulation
Absorb under tung
Absorbfrom rectum
Pathways of metabolism
Phase 1: Biotransformation
Attachment of new functional groups, transformation of exist. funct. groups
oxidation, reduction, hydroxylation, hydrolysis etc.
Phase 2: Conjugation.
Masking of an exist. funct. group by for instance
acetylation, glycosylation, attachment amino acid etc
More hydrophilic drug
Renal excretion
O OHHO
NCH3
Morphine
Demethylation
Phase 1 O OHHO
NH
Conjugationglucuronic acid
Phase 2 O OHO
NH
O OHCO2H
OHHO
Phase 1
Metabolism by cytochrome P450 enzyme systhem (CYP450)
•Located in endoplasmatic reticulum (liver and other cells)
•Electron transport systhem - oxidation, monooxygenase
•Heme protein + flavoprotein
•Capablee of oxidation - many differen xenobiotics
N
NN
N
Fe3+
S
Cystein
HOH
CHEMICAL REVIEWSVolume 104, Issue 9 (September 8, 2004)
3947-3980 Mechanism of Oxidation Reactions Catalyzed by Cytochrome P450 Enzymes
Bernard Meunier, Samuël P. de Visser, and Sason Shaik
<http://dx.doi.org/10.1021/cr020443g>http://dx.doi.org/10.1021/cr020443g
Heme protein
Fe3+
Heme protein
Fe2+FMN
FMNH2
Flavoprotein
Flavoprotein
ox. formred. form
red. form ox. form
NAD
NADH
red. form
ox. form
Substrate-red. form
Substrate-ox.form
N
N
N
NHH3C
H3C
O
O
H OH
OH
OHH
H
OPO3 2-
Flavin mononucleotide
FMN
N
HN
NH
NHH3C
H3C
O
O
H OH
OH
OHH
H
OPO3 2-
FMNH2
N
N N
N
NH2
O
OHHO
OP
O
O
OH
P
OO
OHHO
N
O
HO
O
NH2
NAD
N
N N
N
NH2
O
OHHO
OP
O
O
OH
P
OO
OHHO
N
O
HO
O
NH2
NADH
H H
Nicotinamid adenine dinucleotide
CYP450 families and sub-families
Family 1:
CYP1A1
Aromatic hydrocarbon hydroxylase, metabol. PAH etc.
OOH
OH
More hydrophilicnot tox.Nu
OH
Nu
CYP1A2
Ox of arylamines, nitrosamines, aromatic hydrocarbons
Family 2:
CYP2A6
CYP2B6
CYP2C
CYP2D6: Often enantioselective, lipophil. amines
CYP2E1: Halogenated hydrocarbons, other org solvents
Family 3:
CYP3A4
CYP450 / Mechanisms of metobolic transformations
CYP450
Fe3+
CYP450
Fe3+
CYP450
Fe2+
R-H
CYP450
Fe2+
R-H
O2
CYP450
Fe3+
R-H
O2
R-H
e
O2
NADPH
CYP450
Fe3+
R-H
O2
2 H
H2O
CYP450
Fe5+
R-H
O
CYP450
Fe4+
R
OH
R-O-H
R-HCYP450
Fe3+
R
OH
H
H=
+
H2O
Hydroxylation of alkane
Dehydrogenation of alkane
R-CH3
CYP450R-CH2OH
Other enzymesR-CO2H
Fe5+O
Alkene, aromatic!-complex
Fe4+O
Radical"-complex
Fe3+
H
OH
Allylic alchohol
Fe3+O
R
R
O
R
R
O
1,2 migration
Proposed mech. ox of alkenes
alkynes
O
H2O R
O
OH
R
R
Fe3+
Fe3+ O
R
Fe3+ O
RR
O
Proposed mech. ox of aromatics
Fe5+ O
Fe4+ O
Fe3+ O
Fe3+
O
RR
R R
RR
R R
R=H
Fe3+
Fe3+
O HH
taut
HO
Fe5+O Fe4+
OH
X
R
H
X
R
X
R
Fe3+
X
R
OH
i.e. hemiacetal
O+ HX-R
Fe4+OH
Fe3+
X
R
H O
i.e. N oxide
Proposed mech. react. on heteroatom cont. compounds
N, O, S dealkylation
cleav. of small alkylgroups (Me)
Dehalogenation: HX + carbonyl comp.
Proposed mech. react. on heteroatom cont. compounds
Fe5+O Fe4+
OH
N
R
H
N
R
N
R
Fe4+OH
Fe3+
N
R
H O
N
R''
R
R'
O
Stable if R, R' R''!H
N
H
H
R'
O
N
OH
H
hydroxylamine
N-oxide
N
H
R
R'
O
Rearrang.N
OH
R
R'
hemiaminal
O
NR
R'
H
N
O
H
nitroso comp.
sulfide ox, see FMO
CYP450 Induction / inhibition by xenobiotics
Xenobiotics may enhance metabol. of them selvs as well as other comp. taken at the same time
Induce transcript CYP450 mRNA - Synth. CYP450 enzymes (enzyme induction)
•Drugs
•Ethanol
•Organic solvents
•Components in cig. smoke
•
NH
OH
St. Johns Worth
(Johannesurt, prikkperikum)
OH
HO
O OH
R
O OHOH
HO
R=H: Hypericin
R=OH Pseudohypericin
CYP450 Inhibitors
Reversible CYP enzyme inhibitors: Several drugs
ex. antimycotic azoles
Squalene
Squalene epoksidase
O
Squalene epoksid-syklase
HO
Lanosterol
Lanosterol14a-demetylase
Antimycotic azoles
HO
Ergosterol
HO
H
H
HO
Kolesterol
H
H
H
Antimycoticallylamines
Cell wall component fungi
NN
Cl
KlotrimazoleCanesten etcCYP450:
CYP450 Inhibitors
Complexation inhibitors
ex. metabolites from alkylamines
R-NMe2
AlkylamineNo inhib. activity
CYP450
R-N=O
NitrosoalkaneIrreversible complexation with Fe(II) heme in CYP sub families
Mechanism based inhibitors (suicide inhib)
ex. alkynes
CYP450
FeO3+ O
NH2-EnzymeR
NH
O
Enzyme
H2OR
O
OH
R
R
R
HO
H
HH
OH
Ethynyl estradiol
Phase 1 react. not involving CYP450
microsome: Artefactual spherical particle, not present in the living cell,
derived from pieces of the endoplasmic reticulum present in homogenates of tissues or cells:
microsomes sediment from such homogenates when centrifuged at 100 000 g and higher:
the microsomal fraction obtained in this way is often used as a source of mono-oxygenase enzymes.
Other microsomal enzymes
Azoreductase
Nitroreductase
Flavinmonooxygenase-FMO (N and S-ox.)
Peroxidases
N
N NH2
S
O
O
H2N
H2N
NH2S
O
O
H2N
Prontocil Sulfanilamide
R NO2 R NH2
Flavinmonooxygenase-FMO
Cont. FAD
N
N
N
NHH3C
H3C
O
O
H OH
OH
OHH
H
OPO3 2-
Flavin mononucleotide
FMN
N
HN
NH
NHH3C
H3C
O
O
H OH
OH
OHH
H
OPO3 2-
FMNH2
N
N
N
NHH3C
H3C
O
O
H OH
OH
OHH
H
O P O
O
OH
O
OH
O O
HO OH
N
N
N
N
NH2
FAD
N
HN
N
NHH3C
H3C
O
O
RFADH2
Flavin adenine dinucleotide
N
N
N
NHH3C
H3C
O
O
H OH
OH
OHH
H
OH
Riboflavin(Vit B2)
N
N N
N
NH2
O
ORHO
OP
O
O
OH
P
OO
OHHO
N
O
HO
O
NH2
N
N N
N
NH2
O
ORHO
OP
O
O
OH
P
OO
OHHO
N
O
HO
O
NH2
NADH
H H
Nicotinamid adenine dinucleotide
NADR=H
R=Phosphate: NADP+, NADPH
N
N N
N
NH2
O
OHHO
HO
Adenine(Vit. B4)
N
CO2H
Nicotinic acid / Niacin(Vit. B3)
Flavinmonooxygenase-FMO
N
N
N
NHH3C
H3C
O
O
R
H2O
N
HN
N
NHH3C
H3C
O
O
R
OH
N
HN
N
NHH3C
H3C
O
O
R
O
OH
FAD
Substrate Substrate-O
N
HN
NH
NHH3C
H3C
O
O
R
FADH2
O2
NADPH
NADP+
RO
O
H
NuPeroxide
Peracid
O Nu
N-oxide
Sulfoxide
+ ROH
•Amine: ox. to N-oxide / hydroxylamine
•Sulfide: ox to sulfoxide , furter to sulfone
•Thiol:
Ox of soft Nu
R-SH R-S-S-R R-S-S-R
O
Non-microsomal enzymes
•Enzymes in mitokondria
•Enzymes in soubile tissue fractions
R-OH
prim or sec.
Alchohol dehydrogenase(NAD cont.)
aldehyde / ketoneR
O
H
Aldehyde dehydrogenase
R
O
OH
Alcohol dehydrogenase Aldehyde dehydrogenaseCH3OH HCO2HCH2O
Slow
EtOH cometitive substr.
rel. fast
Tox. effects:AcidosisBlindness
HN
NH
NH
HN
O
O
NH
Tetrahydrofolate
NH
O CO2H
CO2H
HN
NH
NH
HN
O
O
N
NH
O CO2H
CO2H
OH
HCO2H
Dietary folic acid
10-Formyl-THF dehydrogenase
CO2
Non-microsomal enzymes (Phase 1)
Molybdenum Hydroxylases
•Aldehyde oxidase
•Xantine oxidase
•Xantine dehydrogenase
Xanthine oxidase
Electron transfer: FAD - Fe2S2ˇI - Fe2S2ˇII - Moco - Substrate
Active form
Cont. Mo in cat. site
Cont FAD and 2 Fe/s clusters
Use H2O not O2
•Aldehyde oxidase
R H
O
R OH
O
N
Aza heterocycle
N OH NH
O
N
N N
N
H2N
AcO
AcO
FamciclovirAntiviral (Herpes etc)Pro-drug
Aldehyde oxidaseEsterase / hydrolysis HN
N N
N
H2N
HO
HO
O
Penciclovir
•Xantine oxidase
•Xantine dehydrogenase
(requires NAD+)
xanthine oxidoreductase
HN
N
O
NH
N
Hypoxanthine
HN
NH
O
NH
N
Xanthine
O
HN
NH
O
NH
HN
Uric acid
O
O
HN
N
O
NH
N
Allopurinol
HN
NH
O
NH
N
AlloxanthineInhib. enzyme
O
Treatment of gout (podagra)
Non-microsomal enzymes (Phase 1)
Oxidative deamination of amines
•Monoamine oxidase (MAO)
•Diamine oxidase (DAO)
R-CH2-NH2 + O2 [R-CH=NH] R-CHO + NH4+
NH
NH2
HO
Serotonin
5-Hydroksotryptamin (5-HT)
MAO
NH
O
HO
Serotonine:
•Neurotransmittor; temp. control, mood
•Depresion: Low serotonine activity
•MAO Inhibitors - Older antidepresants
(low selectivity)
NNH
O O
Cl
Moklobemid
Aurorix® Moklobemid®
Other MAO substrates:
HN
OH
HO
HOH
Noradrenaline
NH2HO
HO
Dopamine
Not MAO substrates (subst at !-C):
NH2
(S)-amfetamin
Low dopamine conc. ! Parkinston
Active transport re-uptake transmittor
(not Acetylcholine)
Non-selective monoamine re-uptake inhib.
Tricyclic antidepressants
SSRI (selective serotonine re-uptake inhib.)
“Lykkepiller” Prozac etc (Fontex)
NH2HO
HO
NH2HO
HO
OH
DA NE
HO
NH
NH2
5-HT!
"
"-Arylaminer
Y
X
RZ
N
!"
"-Arylam
! "-Arylamin
Stor gruppe / sterisk hindingHindrer reopptak
O
F3C
NHH2N
Non-microsomal enzymes (Phase 1)
Oxidative deamination of amines
•Monoamine oxidase (MAO)
•Diamine oxidase (DAO)
Oxidize diamines, histamineNH2
HN
N
MAO like enzymes in plants
N
N NH
N
HN
BAPCYtokinin (Plant growth hormone)
N
N NH
N
NH2
O
+N
N NH
NN
N NH
N N
N NH
N
CK analogs, Anders Bråthe
Non-microsomal enzymes (Phase 1)
Miscellaneous react.
Reductions
S S RR R-SH
R-SO-R R-S-R (i.e.DMSO)
R-CO-R R-CHOH-R
R-NO-R R-NH-R
R-CH=CH-R R-CH2-CH2-R
R-OH R-H
Ar-OH Ar-H
!-Oxidation
R-CH2-CH2-CO2H R-CH2-CH2-CO-S-CoA R-CO2H
+
CHa-CO-S-CoA
Acetyl-CoA
R-CO-S-CoA
N
N N
N
NH2
O
OHPO3
OP
OP
O
HN
O OO OOH
O
HN
S
O
O
Acetyl-CoA
Nu
Hydrolysis - Esterases
O
ORR'
O
NRR'
R''
O
OHR
Esters as pro-drugs
Pathways of metabolism
Phase 1: Biotransformation
Attachment of new functional groups, transformation of exist. funct. groups
oxidation, reduction, hydroxylation, hydrolysis etc.
Phase 2: Conjugation.
Masking of an exist. funct. group by for instance
acetylation, glycosylation, attachment amino acid etc
More hydrophilic drug
Renal excretion
O OHHO
NCH3
Morphine
Demethylation
Phase 1 O OHHO
NH
Conjugationglucuronic acid
Phase 2 O OHO
NH
O OHCO2H
OHHO
Phase 2: Conjugation
Most comp. excreted as cojugates, ionic, hydrophilic groups added,
most common glucuronation
•Glucuronic acid conjugation
•Sulfate conjugation
•Conjugation with amino acids
•Acetylation
•Glutathione conjugation
•Methylation
Phase 2: Conjugation
•Glucuronic acid conjugation
Substrates:
•Alchohols
•Phenols
•Amines
•Sulfides
•Carboxylic acids
•1,3-Dicarbonyls
O
HO
OHOHOH
OH
Glucose
O
CO2H
OHOOH
OH
UDP-Glucuronate
P P O
HO OH
N
N
O
O
RX
H
O
CO2H
OHOH
OH
XR
RXH: Xenobiotic / Phase 1 metabolite
O
CO2H
OHOH
OH
XR
Kidney Urive excretion
Bile (galle)
RXH
O
CO2H
OHOH
OH
XR
IntestineRXH
Poor reabsorb.
O
CO2H
OHOH
OH
XR
Reabsorb
Entro-hepatic recycling
Important for many hormones etc
O
CO2H
OHOOH
OH
UDP-Glucuronate
P P Uridine
O
H
O
CO2H
OHOH
OH
OR
O
R
O
Relatively labile
Hydrolysis
O
H R
O
HNu Macromolecule
Nu Macromolecule
R
O
Altered protein (hapten)Unwanted immune responce / allergic react.
Acyl migr.
ex. NSAIDs
Ar-NH2
Phase 1Ar-NH-OH Ar-NH-OH-Glu
UrinepH<7
Ar-NH-OH
H2O
Ar-N
Nitrene(c.f. carbene)
Bladder cancer
Phase 2: Conjugation
•Sulfate conjugation: Phenols, (alcohols, N-compounds)
ATPO
NPO
O
OH
S
O
O
O
HO3P OH
PAPS
N
N
N
NH2HO
O
S
O
O
O
•Conjugation with amino acids (Most ofthen Gly): Carboxylic acids
R-CO2H
O
R SCoA
H2N CO2HO
R NH
CO2H
No tox. conjugates known
Phase 2: Conjugation
•Acetylation: N-compounds
O
H3C SCoA
NH N
O
•Glutathione conjugation: Electrophilic species
HS NH
HN
O CO2H
O
NH2
CO2HR-X + S NH
HN
O CO2H
O
NH2
CO2H
R
Glutathione conjugate
S OH
HN
O
O
R
Mercapturic acid der.
•Alkylhalides
•Epoxides
•Michael acceptors etc
may otherwise
alkylate biomolecules
Br
BrRSH
Br
S
S R
Nu(i.e. DNA)
HN
O
OH
Paracetamol
Phase 1
CYP450
H2O
N
O
O
HS-R
O
HN
O
SR
H
HN
O
OH
SR
H2N Liver Protein
Liver dammage
Phase 2: Conjugation
•Methylation (O and N- compd)
Prod. may be more lipophilic
React. mainly aimed at converting endogenic compouds
O.Metylation by COMT (catecol O-methyl transferase)
ATPO
N
HO OH
N
N
N
NH2HO
O
H3C
SAM (S-adenocyl methionine)
S
H3C
CO2H
H2N
HO
HO
SAM may also methylate N-comp.
HO
HO
OH
NH2
Noradrenaline(Norepinephrine)
MAO
HO
HO
OH
CHOAldehyde Dehydrogenase
HO
HO
OH
CO2H
COMT
HO
H3CO
OH
NH2
COMT
HO
H3CO
OH
CO2HMAO Aldehyde
Dehydrogenase
OH
NH2
EphedrineAdrenerg agonist