Metabolic Changes of Drugs

Books: 1. Wilson and Gisvold’s Textbook of Organic Medicinal and Pharmaceutical Chemistry 11th ed. Lippincott, Williams & Wilkins ed.

2. Foye’s Principles of Medicinal Chemistry

Introductory Concepts

■ Biochemically speaking: Metabolism means Catabolism (breaking down of substances) + Anabolism (building up or synthesis of substances)

■ But when we speak about drug metabolism, it is only catabolism

■ That is drug metabolism is the break down of drug molecules

■ So what is building the drug molecules? We use the word “synthesis”, then

■ Drugs are synthesized in laboratory and thus is not an endogenous event

■ Lipid soluble drugs require more metabolisms to become polar, ionizable and easily excretable which involve both phase I and phase II mechanisms.

What Roles are Played by Drug Metabolism?

■ One of four pharmacokinetic parameters, i.e., absorption, distribution, metabolism and excretion (ADME)

■ Elimination of Drugs: Metabolism and excretion together are elimination

■ Excretion physically removes drugs from the body

The major excretory organ is the kidney. The kidney is very good at excreting polar and ionized drugs without any major metabolism. The kidney is unable to excrete drugs with high LWPC

■ In general, by metabolism drugs become more polar, ionizable and thus more water soluble to enhance elimination

■ It also effect deactivation and thus detoxication or detoxification

■ Many drugs are metabolically activated (Prodrugs)

■ Sometimes drugs become more toxic and carcinogenic

Metabolite activity

Examples and notes

Inactive (detoxification)

Similar activity to the drug

Different activity

Toxic metabolites

N

NO

Ph

Cl

CH3

N

NO

Ph

Cl

CH3

OHN

HN

O

Ph

Cl

Diazepam(Sustained anxiolytic action)

Hydroxylation

Temazepam(Short duration)

Oxazepam(short duration)

N-Demethylation OH

N

CONHNHCH

CH3

CH3

N

CONHNH2

Ipronazid(Antidepressant)

N-Dealkylation

Isoniazid(Antituberculosis)

NCOCH3

HO

OC2H5

NHCOCH3

OC2H5

NH2

OC2H5N-Hydroxyphenacetin

(Hepatotoxic)Phenacetin(Analgesic)

Phenetidine

Substances responsible for methemoglobinamia

Other substances responsible for hepatotoxicity

OH

Phenol

Phenol sulphokinase

3'-Phosphoadenosine-5'-phosphosulfate (PAPS)

OS

O

O OH

Phenyl hydrogen sulfate

Routes that result in the formation of inactive metabolites are often referred to as detoxification.

The metabolite may exhibit either a different potency or duration of action or both to the original drug.

Stereochemistry of Drug Metabolism

O

CH2COCH3

PhH

OH

O

O

CH2COCH3

PhH

OH

O O

H2C

HPh

OH

O

OH

CH3H

O

CH2COCH3

HPh

OH

O

O

H2C

HPh

OH

O

OH

HCH3

S-(-)-Warfarin

S-6-Hydroxywarfarin

R-(+)-Warfarin

Major routeMinor route

R,S-(+)-alcohol derivative R,R-(+)-alcohol derivative

HO

CH3

HCOOH

COOH

HCH3

Metabolism

R-(-)-Ibuprofen(inactive) S-(+)-Ibuprofen

(active)

Sites of Drug Metabolism

Liver: Major site, well organized with all enzyme systems

The first-pass effect

Following drugs are metabolized extensively by first-pass effect: Isoproterenol, Lidocaine Meperidine, Morphine, Pentazocine, Propoxyphene, Propranolol, Nitroglycerin, Salicylamide

Intestinal Mucosa: The extra-hepatic metabolism, contains CYP3A4 isozyme

Isoproterenol exhibit considerable sulphate conjugation in GI tract

Levodopa, chlorpromazine and diethylstilbestrol are also reportedly metabolized in GI tract

Esterases and lipases present in the intestine may be particularly important carrying out hydrolysis of many ester prodrugs

Bacterial flora present in the intestine and colon reduce many azo and nitro drugs (e.g., sulfasalazine)

Intestinal -glucuronidase can hydrolyze glucuronide conjugates excreted in the bile, thereby liberating the free drug or its metabolite for possible reabsorption (enterohepatic circulation or recycling)

Enzymes Involved in Drug Metabolism

Cytochrome P450 system: localized in the smooth endoplasmic reticulum.

Cytochrome P450 is a Pigment that, with CO bound to the reduced form, absorbs maximally at 450nm

Cytochromes are hemoproteins (heme-thiolate) that function to pass electrons by reversibly changing the oxidation state of the Fe in heme between the 2+ and 3+ state and serves as an electron acceptor–donor

P450 is not a singular hemoprotein but rather a family of related hemoproteins. Over 1000 have been identified in nature with ~50 functionally active in humans with broad substrate specificity

CYP450, Hepatic microsomal flavin containing monooxygenases (MFMO or FMO) Monoamine Oxidase (MAO) and Hydrolases

Simplified apoprotein portion

Heme portion with activated Oxygen

N N

NN

CH3

HOOC

HOOCCH3 CH3

CH2

CH3

CH2

Fe+3

L

OH R

Substrate binding site

Cytochrome P450: Naming

■ Before we had a thorough understanding of this enzyme system, the CYP450 enzymes were named based on their catalytic activity toward a specific substrate, e.g., aminopyrine N-demethylase now known as CYP2E1

■ Currently, all P450’s are named by starting with “CYP” (CYtochrome P450, N1, L, N2 - the first number is the family (>40% homology), the letter is the subfamily (> 55% homology), and the second number is the isoform. The majority of drug metabolism is by ~10 isoforms of the CYP1, CYP2 and CYP3 families in humans

■ Major human forms of P450: Quantitatively, in the liver the percentages of total P450 protein are: CYP3A4 – 28%, CYP2Cx – 20%, CYP1A2 – 12%, CYP2E1 – 6%, CYP2A6 – 4%, CYP2D6 – 4%

■ By number of drugs metabolized the percentages are: CYP3A4 – 35%, CYP2D6 – 20%, CYP2C8 and CYP2C9 – 17%, CYP2C18 and CYP2C19 - 8% CYP 1A1 and CYP1A2 -10%, CYP2E1 – 4%, CYP2B6 – 3%

CYP family

Main functions

CYP1 Xenobiotic metabolism

CYP2 Xenobiotic metabolism, Arachidonic acid metabolism

CYP3 Xenobiotic and steroid metabolism

CYP7 Cholesterol 7α-hydroxylation

CYP11 Cholesterol side-chain cleavage, Steroid 11β –hydroxylation, Aldosterone synthesis

CYP17 Steroid 17α-hydroxylation

CYP19 Androgen aromatization

CYP21 Steroid 21-hydroxylation

CYP24 Steroid 24-hydroxylation

CYP27 Steroid 27-hydroxylation

Few Important CYP450 Isozymes

EC Recommended name Family/gene

1.3.3.9 * secologanin synthase CYP72A1

1.14.13.11 * trans-cinnamate 4-monooxygenase CYP73

1.14.13.12 * benzoate 4-monooxygenase CYP53

1.14.13.13 * calcidiol 1-monooxygenase CYP27

1.14.13.15 * cholestanetriol 26-monooxygenase CYP27

1.14.13.17 * -monooxygenase CYP7

1.14.13.21 * flavonoid 3'-monooxygenase CYP75

1.14.13.28 * 3,9-dihydroxypterocarpan 6a-monooxygenase CYP93A1

1.14.13.30 * leukotriene-B4 20-monooxygenase CYP4F

1.14.13.37 * methyltetrahydroprotoberberine 14-monooxygenase CYP93A1

1.14.13.41 * tyrosine N-monooxygenase CYP79

The drug interactions depend upon:

a)the isoform(s) required by the drug in question,

b)the isoforms altered by concomitant therapy,

c)the type of enzyme alteration (induction or inhibition).

Drug Interactions & Metabolism

General Metabolic Pathways

Glucuronic acid conjugation Sulfate Conjugation Glycine and other AA Glutathion or mercapturic acid Acetylation Methylation

Reduction Aldehydes and ketones Nitro and azo Miscellaneous

Oxidation Aromatic moieties Olefins Benzylic & allylic C atoms

and -C of C=O and C=N At aliphatic and alicyclic C C-Heteroatom system

C-N (N-dealkylation, N-oxide formation, N-hydroxylation)

C-O (O-dealkylation)C-S (S-dealkylation, S-oxidation,

desulfuration) Oxidation of alcohols and

aldehydes Miscellaneous

Phase II - Conjugation

Phase I - Functionalization

Drug Metabolism

Hydrolytic Reactions Esters and amides Epoxides and arene oxides

by epoxide hydrase

Tetrahydrocannabinol (1-THC) Metabolism

The metabolite is polar, ionisable and hydrophilic

O C5H11

OH

CH3

H3C

CH3O C5H11

OH

CH2OH

H3C

CH3O C5H11

OH

COOH

H3C

CH3

O C5H11

OR

COOR

H3C

CH3

OCOO-

OHOH

HOH

1

7

2

345

6

1-THC 7-Hydroxy-1-THC 1-THC-7-oic Acid

Glucuronide conjugate at eitherCOOH or phenolic OH group

Where R =

-Glucuronyl moiety

Oxidative Reactions

OH O

C C

OC C

C H

C OH

O CO P

S CS P S CH3

SH, S CH3

O

R O CH3

R OH

R N H

R N

R N CH2R

R N

R N OH

R NH

O

CHRO"Activated Oxigen"

[FeO]3+

Arene OxidesArenols

Epoxides

Benzylic, allylic aliphatic C

Hydroxylation

Miscellaneous Oxidations +

Desulfuration S-Dealkylation and S-Oxidation

O-Dealkylation N-HydroxylationN-Dealkyaltion and Oxidative DeaminationN-Oxide Formation

■ Hydroxylation is the primary reaction mediated by CYP450

■ Hydroxylation can be followed by non-CYP450 reactions including conjugation or oxidation to ketones or aldehydes, with aldehydes getting further oxidized to acids

■ Hydroxylation of the carbon α to heteroatoms often lead to cleavage of the carbon – heteroatom bond; seen especially with N, O and S, results in N–, S– or O–dealkylation.

■ Must have an available hydrogen on atom that gets hydroxylated, this is important!!!

Aromatic Hydroxylation

■ Mixed function oxidation of arenes to arenols via an epoxide intermediate arene oxide

■ Major route of metabolism for drugs with phenyl ring

■ Occurs primarily at para position

■ Substituents attached to aromatic ring influence the hydroxylation

■ Activated rings (with electron-rich substituents) are more susceptible while deactivated (with electron withdrawing groups, e.g., Cl, N+R3, COOH, SO2NHR) are generally slow or resistant to hydroxylation

R1 R1

OH

R1

O

R1

OH

OH

R1

SGlutathione

R1

Macromolecule

Spontaneous

Epoxide hydrolase

Glutathione

Macromolecule

R1

OH

OH

Aromatase

CYP450

OH

OH

Epoxide Hydrase

N

N

O

H

H

O N

N

O

H

H

O

CYP2C19

HO

H

CH3

CH3

OH

NO

CH3

H

H

N

C CH

OH

HO

Phenytoin p-hydroxyphenytoinAmphetamine

Propranolol17--Ethinylestradiol

O

CH3

O O

ONa

Ca+2

HN

O

H3C

CH3 F

C

N

C OOH

HO O

2

Warfarin sodium

Atorvastatin

CH3

O

O

N

N

Phenylbutazone

Cl

Cl

HN

HN

N H3C

O

O

H3C

N S

OH

O

ClonidineProbenecid

Antihypertensive drug clonidine undergo little aromatic hydroxylation and the uricosuric agent probenecid has not been reported to undergo any aromatic hydroxylation

Diazepam Chlorpromazine

CH3

Cl

O

N

N

Cl

CH3

CH3NN

SPreferentially the more electron rich ring is hydroxylated

NIH Shift: Novel Intramolecular Hydride shift named after National Institute of Health where the process was discovered. This is most important detoxification reaction for arene oxides

R

O

SpontaneousRearrangement

R

-O H

H+

NIH Shift

R

O

HH

R

OH

Arenol

Arene Oxide

Oxidation of olefinic bonds (also called alkenes)

EpoxideAlkene trans dihydrodiol derivative

Epoxide hydrolaseO OHOH

■ The second step may not occur if the epoxide is stable, usually it is more stable than arene oxide

■ May be spontaneous and result in alkylation of endogenous molecules

■ Susceptable to enzymatic hydration by epoxide hydrase to form trans-1,2-dihydrodiols (also called 1,2-diols or 1,2-dihydroxy compounds)

■ Terminal alkenes may form alkylating agents following this pathway

NH2O

N

NH2O

N

NH2O

N

Epoxide hydrolaseCYP3A4

O HO OH

Carbamazepine Carbamazepine 10,11 epoxide Carbamazepine trans 10,11 diol

(Active) (Active & Toxic) (Inactive)

Q. Any similarities or dissimilarities with aromatic – NIH Shift, Conjugation

with macromolecules?

Benzylic Carbon Hydroxylation■ Hydroxylate a carbon attached to a phenol group (aromatic ring)

■ R1 and R2 can produce steric hindrance as they get larger and more branched

■ So a methyl group is most likely to hydroxylate

■ Primary alcohol metabolites are often oxidized further to aldehyde and carboxylic acids and secondary alcohols are converted to ketones by soluble alcohol and aldehyde dehydrogenase

Dicarboxylic acid is the major metabolite

ONa

O

CH3

H3C

O

N

Tolmetin sodium

CR1

R2

H CR1

R2

OH

Tolbutamide Metabolism

OOO

CH3NH

NH

S

H3C

OOO

CH3NH

NH

S

C

CYP2C9

HOH

H

Oxidation at Allylic Carbon Atoms

C C CC R3R1

R2 R4

C C CC R3R1

R2 R4

OHHHH H H

HH

O C5H11

OH

CH3

H3C

CH3

O C5H11

OH

CH2OH

H3C

CH3O C5H11

OH

CH3

H3C

CH3

HO

O C5H11

OH

CH3

H3C

CH3

HO

1-THC

12

345

6

77

7-Hydroxy-1-THC 6-Hydroxy-1-THC 6-Hydroxy-1-THC

+ +

N

NHO

H3CO

H2C

H

N

NHO

H3CO

H2C

OH

Quinine

1

23

3-Hydroxyquinine

O

O

O

CH3

CH3

2' 3' O

O

O

CH3

CH3

O

O

O

CH3

CH3

OH O

O-Glucuronide Cojugate

Hexabarbital 3'-Hydroxyhexabarbital 3'-Oxohexabarbital

Pentazocine

Hydroxylation at C to C=O and C=N

The benzodiazepines are classic examples with both functionalities

The sedative hypnotic glutethimide possesses C to carbonyl function

R C C R'

O H

H

R C C R'

O H

OH

N

N

CH3 O

Cl

3

N

N

CH3 O

Cl

OHN

HN

O

Cl

OHN-demethylation

N

N

(CH3CH2)2NCH2CH2 O

Cl N

N

CH3 O

O2N

3 3

Diazepam (3S) N-Methyloxazepamor 3-Hydroxydiazepam

Oxazepam

F

Flurazepam Nimetazepam

NH

C6H5

CH2CH3

OO NH

C6H5

CH2CH3

OO

HO

1

34 4

Glutethemide 4-Hydroxyglutethemide

Aliphatic hydroxylation

■ Catalyzes hydroxylation of the ω and ω-1 carbons in aliphatic chains

■ Generally need three or more unbranched carbons

C C CR1

C C CR1

OH

C C CR1 OH

H

H H

H

H

H

H

H

H H

H

H

H

H

H

H

H

H

H

N

N

H

H

O

O

O

N

N

H

H

O

O

OOH

CYP450

OH

O

CH3

CH3H3C

OH

O

CH3

CH3H3C

OH

CYP450

Pentobarbital Metabolism

Ibuprofen Metabolism OH

O

CH3

CH3HOOC

+

Alicyclic (nonaromatic ring) Hydroxylation

Acetohexamide Metabolism

■ Cyclohexyl group is commonly present in many drug molecules

■ The mixed function oxydase tend to hydroxylate at the 3 or 4 position of the ring

■ Due to steric factors if position 4 is substituted it is harder to hydroxylate the molecules

H3C

O

OOO

NH

NH

S

H3C

O

OOO

NH

NH

SCYP450

OH

Oxidation Involving Carbon-Heteroatom Systems

■ C-N, C-O and occasionally C-S

■ Two basic types of biotransformation processes:

1. Hydroxylation of -C attached directly to the heteroatom (N,O,S). The resulting intermediate is often unstable and decomposes with the cleavage of the C-X bond:

Oxidative N-, O-, and S-dealkylation as well as oxidative deamination reaction fall under this category

2. Hydroxylation or oxidation of heteroatom (N, S only, e.g., N-hydroxylation, N-oxide formation, sulfoxide and sulfone formation)

■ Metabolism of some N containing compounds are complicated by the fact that C or N hydroxylated products may undergo secondary reactions to form other, more complex metabolic products (e.g., oxime, nitrone, nitroso, imino)

R X C

H

R X C

O

H

R XH

O

+

Usually Unstable

C-N systems

■ Aliphatic (1o, 2o, 3o,) and alicyclic (2o and 3o) amines; Aromatic and heterocyclic nitrogen compounds; Amides

■ Enzymes:

1. CYP mixed-function oxidases: -C hydroxylation and N-oxidation

2. Amine oxidases or N-oxidases (non-CYP, NADPH dependent flavoprotein and require O): N-oxidation

R1 N C

H

R1 N C

O

H

R1 NH +

O

Carbinolamine

R2 R2 R2

3o or 2o amine 2o or 1o amine

C

H

NH2

C

O

NH2

H

NH3+O

Carbinolamine1o amine Carbonyl Ammonia

■ 3o Aliphatic and alicyclic amines are metabolized by oxidative N-dealkylation (CYP)

■ Aliphatic 1o, 2o amines are susceptible to oxidative deamination, N-dealkylation and N-oxidation reactions

■ Aromatic amines undergoes similar group of reactions as aliphatic amines, i.e., both N-dealkylation and N-oxidation

N-Dealkylation (Deamination)

C N

H

R2

R1 C

R2

R1R3

R4

O + HN R3

R4

C N

OH

R2

R1 R3

R4

CYP450 Spontaneous

NCH3

CH3

N NCH2

CH3

N N

CH3

NCYP2C19 Spontaneous

OH

H

■ Deamination and N-dealkylation differ only in the point of reference; If the drug is R1 or R2 then it is a deamination reaction and If the drug is R3 or R4 then it is an N-dealkylation

■ In general, least sterically hindered carbon () will be hydroxylated first, then the next, etc. Thus the more substituent on this C, the slower it proceeds; branching on the adjacent carbon slows it down, i.e. R1, R2 = H is fastest.

■ Any group containing an -H may be removed, e.g., allyl, benzyl. Quaternary carbon cannot be removed as contain no -H

■ The more substituents placed on the nitrogen the slower it proceeds (steric hindrance)

■ The larger the substituents are the slower it proceeds (e.g. methyl vs. ethyl). In general, small alkyl groups like Me, Et and i–Pro are rapidly removed; branching on these substituents slows it down even more

Imipramine N-Dealkylation

Alicyclic Amines Often Generate Lactams

N

N

CH3 N

N

CH3

OH

N

N

CH3

O

Nicotine Carbinolamine Cotinine

CH3

N

CH3

N O

Cyproheptadine Lactum metabolite

NH

O

H3C NH

O

OHH3C

C6H52

1

Phenmetrazine Carbinolamineintermediate

3

C6H5

NH

O

OH3C

C6H5

3-Oxophenmetrazine

COOCH3

HN

HydrolysisCOOH

HN

COOH

HN

OMethylphenidate Ritalinic Acid 6-Oxoritalinic Acid

CH3

CH3

CH3

CH3

O

N

HN

N

CH3

H3C

CH3

CH3

N

O

NH2C

ON

CH3

H3C

CH3

CH3

CH3N

O

CH3

CH3

N

Br

NN

CH3

CH3

3oAmine drugs

DisopyramideLidocaine Tamoxifen

Diphenhydramine

Cl

CH3

CH3NN

S

Chlorpromazine Benzphetamine Brompheniramine

O

N

CH3

CH3O

HO OH

N

O

CH3

H

CH3

O

CH3

NAlicyclic Amine drugs

Meperidine Morphine Dextromethorphan

2o & 1o Amines

Generally, dealkylation of secondary amines occurs before deamination. The rate of deamination is easily influenced by steric factors both on the -C and on the N; so it is easier to deaminate a primary amine but much harder for a tertiary amine.

CH3

HNCH3

CH3

NH2

CH3

O

CH2

ONH3

Methampetamine Ampetamine Phenylacetone

Cl

NHCH3

O

Cl

NH2

O

Ketamine Norketamine

Exceptions: Some 2o and 3o amines can undergo deamination directly without dealkylation.

Propranolol

O

HN CH3

CH3

OH

Direct OxidativeDeamination

O

HN CH3

CH3

OHO

OH

O

HN CH3

CH3

OHO

NH2

CH3H3C

OH

O H OH2N

Carbinolamine

O

H

O

NH3

Oxidative DeaminationThrough Primary Amine

AldehydeMetabolite

Primary Amine Metabolite(Desisopropyl Propranolol)

N-Oxidation

N NH H H OH

N O

1 aromatic amine Hydroxylamine Nitroso

R C N

H

H

R C N

H

H

H

H

H

OHR C N

H

H

R C N

H

H

O

O

1 amine Hydroxylamine Nitroso Nitro

O

R C N

H

H

R C N

H

H

CH3

H

CH3

OHR C N

H

H

2 amine Hydroxylamine Nitrone

CH2

O

R C N

H

H

R C N

H

H

CH3

CH3

CH3

CH3

3 amine N-Oxide

O

Aromatic amines

1° amines

2° amines

3° amines

Cl

H3C

CH3

H

H

N

Cl

H3C

CH3

OH

H

NCYP450

■ The attack is on the unbonded electrons so 3o amines can be oxidized

■ Generally, only occurs if nothing else can happen, so it is a rare reaction

■ Performed by both amine oxidases and hepatic MFO’s

■ Good examples would include amines attached to quaternary carbons since they cannot be deaminated

H3C

CH3

H

H

N

NH2

PhentermineAmantadine

Chlorphentermine N-Hydroxylation Hydroxylamine

Nitroso

Nitro

Amides

C-N bond cleavage via -C hydroxylation (formation of carbinolamide) and N-hydroxylation reactions

N

N

CH3 O

C6H5

Cl N

N

H2CO

C6H5

ClN

HN

O

C6H5

Cl

OH

N

N(CH3CH2)2NCH2CH2 O

Cl

Diazepam Carbinolamide Desmethyldiazepam

F

Flurazepam

OH

CH2

O

HN

N

O

O O

R1

R2

CH3

Hexobarbital: R1 = , R2 = CH3

Mephobarbital: R1 = C6H5, R2 = CH2CH3

Cl

SO2NHCNHCH2CH2CH3

O

Chlorpropamide

Oxidation involving C-O System (O-Dealkylation)

C O R3 HO R3+

H

R1

R2

C O R3

OH

R1

R2

CYP450 SpontaneousR1 C

R2

O

■ Converts an ether to an alcohol plus a ketone or aldehyde

■ Steric hindrance discussion similar to N-dealkylation

O

O

O

NH2

NH2

N

N

CH3

H3C

H3C

O

O

O

NH2

NH2

N

N

CH2

H3C

H3C

OH

O

O

NH2

NH2

N

NH3C

H3C

OH

Trimethoprim O-Dealkylation

CH3O

CH3

O

HN

OH

OH3C O

CH3

Cl

O

N

O

O

N

O

ON

NH2

N

NH3C

H3CH3C

O

H

CH3

CH3

OH

NO

O

N

CH3

O OH

CH3

Codeine Phenacetin Indomethacin

PrazosinMetoprolol

H3C C OH H3C C OH

OH

H3C C O

H H

H

H

CYP450 Spontaneous

■ One exception that appears to be a form of O-dealkylation is the oxidation of ethanol by CYP2E1

■ In this case R3 is hydrogen instead of carbon to form the terminal alcohol rather than an ether

■ The enzyme involved is CYP2E1 and has been historically referred to as the Microsomal Ethanol Oxidizing System (MEOS)

Oxidation involving C-S System

■ S-Dealkylation

■ Desulfuration

■ S-Oxidation

C S R3R1 C SR1 C OR1 HS R3+

R2R2

OHH

R2

R3

CYP450 Spontaneous

Steric hindrance discussion similar to N-dealkylation

R1 C R2

S

R1 C R2

O

R1 S R2 R1 S R2

O

R1 S R2

O

OSulfoxide Sulfone

N

N

SCH3

NH

N

6-(Methylthio)-purine

N

N

SCH2

NH

N

OH

N

N

SH

NH

N

CH2

O

6-Mercaptopurine

H

HS

O

O

SN

NH3C

NO2

OH3C

H3C O

SP

O

CF3

S

COOH

CH2C6H5

H

H

O

O

SN

N

NO2

OH3C

H3C O

OP

O

H

H

O

O

ON

N

Methitural

Parathione Paraoxone

2-Benzylthio-4-trifluoromethyl benzoic acid

Thiopental Pentobarbital

CH3S

CH3

NN

S

CH3S

CH3

NN

S

CH3S

CH3

NN

S

CH3S

CH3

NN

S

CH3S

CH3

NN

S

OO

O

O O O

Thioridazine

Ring Sulfoxide Ring Sulfone

Mesoridazine Sulforidazine

Oxidative Dehalogenation

R C

H

Cl

Cl

R C

OH

Cl

Cl

R CO

Cl

R CO

OH+

H Cl

+H2O

CYP450

H Cl

+

Spontaneous

■ Requires two halogens on carbon

■ With three there is no hydrogen available to replace

■ With one, the reaction generally won’t proceed

■ The intermediate acyl halide is very reactive

O2N

OH

OH

NHCOCCl

O

HCl

O2N

OH

OH

NHCOC

O

OH

O2N

OH

OH

NHCOCCl2OHO2N

OH

OH

NHCOCHCl2

ChloramphenicolOxamyl Chloride

Derivative

Oxamic Acid Derivative

Tissue Nucleophiles

Covalent Binding(Toxicity)

Q. What is Gray Baby Syndrome?

C

H3C

NH N

S

HN

HN

CH3

N

NC

H3C

NH N

S

HN

HN

CH3

N

NO

MFMO

Cimetidine MFMO S-Oxidation

Hepatic Microsomal Flavin Containing Monooxygenases (MFMO or FMO)

■ Oxidize S and N functional groups

■ Mechanism is different but end products are similar to those produced by S and N oxidation by CYP450

■ FMO’s do not work on primary amines

■ FMO’s will not oxidize substrates with more than a single charge

■ FMO’s will not oxidize polyvalent substrates

Q. What is the difference with MFO?

Non-Microsomal Oxidation Reactions

■ Monoamine oxidase (outer membrane of mitochondria, flavin containing enzyme )

■ Dehydrogenases (cytoplasm)

■ Purine oxidation (Xanthene oxidase)

C N HR1

R2

H

R3

CR1

R2

O + H N H

R3

Monoamine oxidase

■ Two MAOs have been identified: MAO–A and MAO–B. Equal amounts are found in the liver, but the brain contains primarily MAO–B; MAO–A is found in the adrenergic nerve endings

■ MAO–A shows preference for serotonin, catecholamines, and other monoamines with phenolic aromatic rings and MAO–B prefers non–phenolic amines

■ Metabolizes 1° and 2° amines; N must be attached to α-carbon; both C & N must have at least one replaceable H atom. 2° amines are metabolized by MAO if the substituent is a methyl group

■ –Phenylisopropylamines such as amphetamine and ephedrine are not metabolized by MAOs but are potent inhibitors of MAOs

O

HN N

NH

NHO

O

HNN

N NH

O

HN N

NH

NHO

OH

O

HNHN

NH

NHO

O

Hypoxanthine Xanthine Uric acid(hydroxy tautomer)

Uric acid(keto tautomer)

Xanthineoxidase

Xanthineoxidase

Purine oxidation

C OHR1

R2

HCR1

R2

O

R1 C O

OH

R1 C O

H

H3C

H2C

OH H3C

HC

O H3CC

O

OHAlcohol

DehydrogenaseAldehyde

DehydrogenaseEthanol Metabolism

Alcohol dehydrogenase Aldehyde dehydrogenase

Metabolizes 1° and 2° alcohols and aldehydes containing at least one “H” attached to -C; 1° alcohols typically go to the aldehyde then acid; 2° alcohols are converted to ketone, which cannot be further converted to the acid. The aldehyde is converted back to an alcohol by alcohol (keto) reductases (reversible), however, it goes forward as the aldehyde is converted to carboxylic acid; 3° alcohols and phenolic alcohols cannot be oxidized by this enzyme; No “H” attached to adjacent carbon

Molybdenum Containing

Reductive Reactions

■ Bioreduction of C=O (aldehyde and keton) generates alcohol (aldehyde → 1o alcohol; ketone → 2o alcohol)

■ Nitro and azo reductions lead to amino derivatives

■ Reduction of N-oxides to their corresponding 3o amines and reduction of sulfoxides to sulfides are less frequent

■ Reductive cleavage of disulfide (-S-S-) linkages and reduction of C=C are minor pathways in drug metabolism

■ Reductive dehalogenation is a minor reaction primarily differ from oxidative dehalogenation is that the adjacent carbon does not have to have a replaceable hydrogen and generally removes one halogen from a group of two or three

Reduction of Aldehydes & Ketones

■ C=O moiety, esp. the ketone, is frequently encountered in drugs and additionally, ketones and aldehydes arise from deamination

Ketones tend to be converted to alcohols which can then be glucuronidated. Aldehydes can also be converted to alcohols, but have the additional pathway of oxidation to carboxylic acids

■ Reduction of ketones often leads to the creation of an asymmetric center and thus two stereoisomeric alcohols are possible

■ Reduction of , –unsaturated ketones found in steroidal drugs results not only in the reduction of the ketone but also of the C=C

■ Aldo–keto oxidoreductases carry out bioreductions of aldehydes and ketones. Alcohol dehydrogenase is a NAD+ dependent oxidoreductase that oxidizes alcohols but in the presence of NADH or NADPH, the same enzyme can reduce carbonyl compounds to alcohols

R C O

H

R C

H

OH

H

Aldehyde 1 alcohol

R C O

R2

R1 C

R2

OH

H

Ketone 2 alcohol

R1

CR2

O

N

R

HH

H2N

OH+

R1

CR2

HO H

+N+

R

H2N

O

Ketone Chiral AlcoholRed Nicotinamide moietyof NADPH or NADH

Ox Nicotinamide moiety

of NADP+ or NAD+

OH

H3C

O CH3

H3C

H2N

O

O

OH

O

OH

OH

O

O

CH2

HO

OH

O

N

O HO

OH

O

N

O

O

OH

O

H2C

O

CH3

H

C6H5

R (+)-Warfarin

O

OH

O

H2C CH3

H

C6H5

HO H

O

OH

O

H2C CH3

H

C6H5

H OH

R,S (+)-Warfarin R,R (+)-Warfarin

+

Naloxone Daunomycin Naltrexone

OH

C

O

CH3

CH

OH

C

CH3

CH

H

H

HO

HNorethindrone 3,5-Tetrahydronorethindrone

H2

CCH

CH3

NH2

Amphetamine

H2

CC

CH3

O

H2

CCH

CH3

OH

Phenylacetone 1-Phenyl-2-propanol

CC CH3

NHCH3

OHH H C

CCH3

O

OHH

CCH

CH3

OH

OHH

(-)-Ephedrine 1-Hydroxy-1-phenyl-propane-2-one

1-Phenyl-1,2-propandiol

Reduction of Nitro & Azo Compounds

N NR

Azido

NH2R

Amine

NH + N N

N2

N N R2R1 R1 NH2 H2N R2+

Azo Two 1 amines

HNR1

Hydrazo

HN R2

R C N

H

H

R C N

H

H

H

H

H

OHR C N

H

H

R C N

H

H

O

O

1 amineHydroxylamineNitrosoNitro

O

R1 and R2 are almost always aromatic

Usually only seen when the NO2 functional group is attached directly to an aromatic ring and are rare

Nitro reduction is carried out by NADPH-dependent microsomal and soluble nitroreductases (hepatic)

NADPH dependent multicomponent hepatic microsomal reductase system reduces the azo

Bacterial reductases in intestine can reduce both nitro and azo

Cl

HO

O2N N

N

O

O

NNaNN

O2NOS

NH

O O

N

N

O

HO

OHN

SNH2

O O

N

NH2

N

H2NS

NH2

O O

H2N NH2

NH2

H2N

+

Prontosil Sulfanilamide 1,2,3-Triaminobenzene

Clonazepam Sulfasalazine Dantrolene

X

Reduction of Sulfur Containing Compounds

Sulfoxide reduction (Cannot reduce a sulfone) R1 S R2

O

R1 S R2 R1 S R2

O

O

R1 S S R1 SHR2 HS R2+Disulfide reduction

H3C

H3CCH3

S

S

CH3NSS

N H3C

H3CS

SHN

DisulfiramN,N-Diethylthiocarbamic

Acid

O

H3C

OH

OF

CH3

S

H

Sulindac

SulfoneSulfoxide

Hydrolytic Reactions

■ Enzymes: Non-microsomal hydrolases; however, amide hydrolysis appears to be mediated by liver microsomal amidases, esterases, and deacylases

■ Electrophilicity of the carbonyl carbon, Nature of the heteroatom, substituents on the carbonyl carbon, and substituents on the heteroatom influnce the rate of hydrolysis

■ In addition, Nucleophilicity of attacking species, Electronic charge, and Nature of nucleophile and its steric factors also influence the rate of hydrolysis

R1 R2 Name Susceptibility to Hydrolysis

C O Ester Highest

C S Thioester

O O Carbonate

C N Amide

O N Carbamate

N N Ureide Lowest

Table: Naming carbonyl - heteroatom groups

Hydrolyzes (adds water to) esters and amides and their isosteres; the OH from water ends up on the carboxylic acid (or its isostere) and the H in the hydroxy or amine

R1 C R2

O

+

R1 C

O

O R2 R1 C

O

OH HO R2

R1 C

OHN R2 R1 C

O

OH H2N R2

O C O R2R1

O

HO C O R2R1

O

OH HO C OHR2

O

HO O C O O HH

+++

Carbonate Carbonic acid derivative Carbonic acid

O C NR1

O

HO C NR1

O

OH HO C OH

O

HN O C O O HH

+++

Carbamate Carbamic acid derivativeCarbonic acid

R2

R3

R2

R3

R2

R3

N C N

O

HO C N

O

NH HO C OH

O

HN O C O O HH

+++

Urea derivative Carbamic acid derivativeCarbonic acid

R3

R4

R3

R4

R2

R3

R1

R2

R1

R2

R1 CHN N

OR2

R3

R1 C OH

O

H2N NR2

R3

+

Hydrazide Hydrazine

Ester hydrolysis

Amide hydrolysis (slower)

Carbonate hydrolysis

Carbamate hydrolysis

Urea hydrolysis

Hydrazide hydrolysis

The Reactions

Drug Examples

H3COO

O

N

CH3

O

Cocaine

OHO

O

N

CH3

O

H3COO

N

CH3

HO+

Benzoylecgonine Methylecgonine

H3C O

O

O

OH

H3C OOH

O

OH

OH+

Aspirin Salicylic Acid

CH3

CH3N

H2N

O

O

CH3

CH3N

H2N

O

HN

Procainamide

Procaine

H2N

O

OH

Slow Hydrolysis

Rapid Hydrolysis

OH

OH3C O

CH3

Cl

O

N

Indomethacin

CH3

CH3

CH3

CH3

O

N

HN

Lidocaine

O

O

N

O

ON

NH2

N

NH3C

H3C

Prazosin

Stereoselectivity of Hydrolysis

Etomidate (Amidate, hypnotic): R-(+)-isomer is more rapidly hydrolyzed, but S-(-)-isomer is more rapidly hydroxylated.

N

N COOH

Ph

H CH3

N

N COOEt

Ph

H CH3

P450

N

N COOEt

Ph

HO CH3

N

NH

COOEtPh

O

CH3

Etomidate

R(+)-Isomer

esterase

S(-)-Isomer+

The Concept of Prodrugs and Antedrugs

M D M D M D M D

D D ID

activation

Prodrug

inactivation

Antedrug

= Barrier & ID = inactive drug, D = active drug, M = modifier

M M M

(I) Prodrug: Need metabolic activation

(II) Antedrug: Active drug that is quickly inactivated thereby minimizing systemic effects

Prodrugs and Related Terms

■ Albert in 1958 coined the term prodrug to refer a pharmacologically inactive compound that is metabolically activated in the mammalian system

■ Hard Drugs are not susceptible to metabolic or chemical transformation, have high lipid solubility and thus accumulation or high water solubility

Celecoxib: t1/2 10-12 h in humans t1/2 ca. 680 h (Liver toxicity)

■ Soft drugs are active compounds that after exerting its action undergo inactivation to give a nontoxic product. Indeed soft drugs are a group of modified compounds that are also designed to delivery the drugs in to the brain (the chemical delivery system). Bodor coined the term.

O

ONH2S

CH3

F3C NN O

ONH2S

Cl

F3C NN

Basic Concepts of Prodrugs

■ Carrier-linked prodrugs: a pro-moiety is attached, which is not necessary for activity but may impart some desired property to the drug, such as increased lipid or water solubility, or site-directed delivery

■ Advantages may include:

1. increased absorption

2. alleviation of pain at the site of injection if the agent is given parenterally

3. elimination of an unpleasant taste associated with the drug

4. decreased toxicity

5. decreased metabolic inactivation

6. increased chemical stability

7. prolonged or shortened action

■ Bioprecursor prodrugs contain no pro-moiety but rather rely on metabolism to introduce the functionality necessary to create an active species

ONa

O

O

O2N

Cl

Cl

O

HN

OH

O

CH3O2N

Cl

Cl

O

HN

OH

O

O

H3C

CH3

N

S

O

OHO

OH

CH3

HN

Cl

O

CH3

O (CH2)14CH3

Prodrug: Chloramphenicol Hemisuccinate Na Salt

■ Inactive as it is and activated by hydrolysis by plasma esterases to chloramphenicol/ prednisolon

■ Increased water solubility for parenteral administration, which otherwise would precipitate and cause pain by damaging surrounding tissues

Prodrug: Chloramphenicol Palmitate

Prodrug: Clindamycin Palmitate

■ Inactive as it is; activated by hydrolysis by intestinal esterases to chloramphenicol/ clindamycin

■ Minimize their bitter taste and improve their palatability in pediatric liquid suspensions

O

HO OH

OO

O

O

O-Na+

Prodrug: Prednisolon Hemisuccinate Sodium Salt

O

O O

COONa

CH3

CH3

ON

SNH

Prodrug: Carbenicillin Indanyl Ester

■ Inactive as it is and activated by hydrolysis by plasma esterases to carbenicillin

■ Lipophilic indanyl ester furnish improved oral bioavailability

O

H3C

OH

OF

CH3

S

H

H3C

OH

OF

CH3

S

H

O

H3C

OH

OF

CH3

S

H

O

Sulfide(Active)

Sulindac(Inactive)

Sulfone(Inactive)

Prodrugs of Functional Groups

Carboxylic acids and alcohols: Most common

Amines and azo linkages: Not been used much

Carbonyl compounds: Not found to be used widely

Carboxylic Acids and Alcohols

Converted to ester prodrugs which are often hydrolyzed to active drug by different types of esterase enzymes:

Ester hydrolase

Lipase

Cholesterol esterase

Acetylcholinesterase

Carboxypeptidase

Cholinesterase

Microflora in the gut

Manipulation of steric and electronic properties of promoiety allows control of rate and extent of hydrolysis

O

ODrug Promoiety

ODrug Promoiety

or

O

Esterase

O

OHDrug Promoiety+ HO

OHDrug Promoiety

O

HO+

CH3

HOH

NH+O

O

O

O

H3C CH3

CH3

H3C CH3CH3

Esterase CH3

HOH

NH+HO

HO

O

H3C CH3

CH3

OH

Pivalic Acid

Epineprine

Dipivefrin

Advantage of Prodrug Formation I: Increased absorption of hydrophilic drugs by making less hydrophilic or more lipophilic

Prodrug of Epinephrine: Dipivefrin

More lipophilic, thus achieve higher intraocular concentration

Hydrolysis occur in cornea, conjunctiva, and aqueous humor after ophthalmic application

Advantage of Prodrug Formation II: Masking unpleasant taste

Chloramphenicol palmitate and Clindamycin palmitate has already been shown. Other drugs include

ON

CH3

CH3

OO

H2N

NS

CH3O

CH3

OO

H CH3

H3CN

O

CH3

O

O CH3

O

O

OCH3

CH3OH

O

CH3

CH3

HO

H3C

HOH3C

O

CH3

OH

H3C

CH3

O

OCH3

N-Acetyl sulfisoxazole

Erythromycin estolate

CH3

H3CN

O

CH3

O

O CH3

O

O

OCH3

O

O

CH3

H3CH3C

OH3C

O

O

H3C

CH3

OCH3

OCH3

O

H3C

O

Troleandomycin

Not all carboxylic esters hydrolyzed in vivo where double ester approach is used

N

HN

SR1

O

O

CH3

CH3

COOR2

N

HN SR1

O

OCOOR2

R3

Esterase

(R2 = Ethyl, Propyl, Butyl, Phenyl)

Penicillin Esters

(R2 = Ethyl, Propyl, Butyl, Phenyl)

Cephalosporin Esters

Esterase

No Reaction

No Reaction

N

HN S

O

O

NN

S

H2N

OCH3

OCH3

O O O

CH3

CH3

O

OCH3

CH3

N

HN S

O

O

NN

S

H2N

OCH3

OCH3

O O O

CH3

CH3

H HOCH3

CH3

N

HN S

O

O

NN

S

H2N

OCH3

OCH3

O OH

OH3C

H3C

Esterase

Cefpodoxime Proxetil(Prodrug)

+ CO2 +

+

Advantage of prodrug formation III: Increase hydrophilicity and thus water solubility to apply parenterally or also orally when compounds are too lipophilic to formulate in liquid dosage form

O P

O

O-Na+

OH

Drug

Drug

O

CH2

CH2

O

O-Na+O OHDrug

OHDrug HO P

O

O-Na+

OH

O

CH2

CH2

O

O-Na+HO

Succinates

Phosphates

+

+

O

-O

O

O

Drug

Drug OH + O

O

O

Rapid and thus the prodrug is unstable

H3C

CH3N

S

O

OHO

OH

CH3

HN

Cl

O

CH3H

P OHOO-

H2O H3PO4

H3C

CH3

N

S

O

OHHO

OH

CH3

HN

Cl

O

CH3

Clindamycin Phosphate

Phosphatase

Clindamycin

Chemical Delivery System

HO

HO CH2CH

NH2

COOH

BBB; Active transport to CNS

HO

HO CH2CH2NH2

HO

HO CH2CH

NH2

COOH

L-DopaDopamine (Active)

by L-Amino acid delivery system

The site specific delivery of drugs is an important way of increasing drug’s therapeutic index. The knowledge of prodrug and drug metabolism is used to concentrate drugs at its target site thus minimizing the systemic toxicity.

Why

?

I stopped taking medicineas I prefer original disease

to side effects!!

Because,Vioxx’ll treat pain but who’ll treat

vioxx??

Antedrugs (Soft Drugs)

Safety-Based Drug Withdrawals from U.S. Market (2006-2007)

Drugs Therapeutic activity Date approved

Date withdrawn

Primary health risk

Vioxx (Rofecoxib)

Antiinflammatory (COX-2 inhibitor)

05/99 09/04 Myocardial Infarction etc.

Ximelagatran (Exanta) Anticoagulant 2006 Hepatotoxicity

Tegaserod (Zelnorm)

IBS, constipation 2002 2007 Cardiovascular ischemic events

Aprotinin (Trasylol)

Induce bleeding during sergery)

1960s 2007 Ischemic colitis and Severe constipation

It takes about 10-15 years

$897 millions to $1.7 billions

Overall attrition rate 10,000:1

To bring a drug from concept to market

Because of unintended systemic actions in most therapeutic classes of drugs

Why the Adverse Drug Reactions Occur?

What is Antedrug?

An active synthetic drug which is inactivated by a metabolic process upon entry into the systemic circulation.

Therefore, a true antedrug acts only locally.

True Antedrug

Partial Antedrug

Inactive Metabolite

Less active metabolite

Lee HJ and Soliman MRI (1982). Science, 215, 989.

O

OH

OH

O

OH

OCOR

O

OH

OH

O

OH

OH

O

OH

CO2R

OH

O

OH

CO2-

(IA = Inactive Compound, A = Active Compound)

Antedrug

Prodrug

Hydrocortisone

IA A

A IA

Chemical Approaches

1) The Carboxylic Esters and Amides

2) 20-Thioester Derivatives

3) -Butyrolactone Derivatives

O

CH3

HOCH3

CO2RO

OH

H

HH

O

CH3

HOCH3

COOHO

OH

H

HH

hydrolysis in plasma

Inactivation of Steroid 21-ate Esters in Bood Plasma.

Stable locally and active Inactive

O

CH3

HOCH3

O

H

HF

OCOCH3

CO2MeOH

plasma

t1/2 6.3 min

O

CH3

HOCH3

O

H

HF

OH

CO2MeOH

plasma

t1/2 90 min

O

CH3

HOCH3

O

H

HF

OH

COOHOH

O

CH3

HOCH3

H

HF

MeOCOEt

O

F

SCH2F

O

CH3

HOCH3

H

HF

MeOCOEt

O

F

OH

Liver

InactiveInactivation of Fluticasone Propianate

Advantages of Antedrugs

Localization of the drug effects

Elimination of toxic metabolites, increasing the therapeutic index

Avoidance of pharmacologically active metabolites that can lead to long-term effects

Elimination of drug interactions resulting from metabolite inhibition of enzymes

Simplification of PK problems caused by multiple active species

M.O.F.Khan*, K.K.Park, H.J.Lee. Antedrugs: An Approach to Safer Drugs. Curr. Med. Chem., 12(19), 2227-2239, 2005.

Phase II: Drug Conjugation

Attachment of small polar endogenous molecules such as glucuronic acid, sulfate and amino acids to Phase I metabolites or parent drugs

Products are more water-soluble and easily excretable

Attenuate pharmacological activity and thus toxicity

Trapping highly electrophilic molecules with endogenous nucleophiles such as glutathione prevent damage to important macromolecules (DNA, RNA, proteins)

Regarded as true detoxifying pathway (with few exceptions)

In general, appropriate transferase enzymes activate the transferring group (glucuronate, sulphate, methyl, acetyl) in a coenzyme form

Glucuronidation is the most common conjugation pathway

The coenzyme, UDP glucuronic acid is synthesized from the corresponding phosphate

UDP-glucuronic acid contains D-glucuronic acid in the -configuration at the anomeric center, but glucuronate conjugates are -glycoside, meaning inversion of stereochemistry is involved in the glucuronidation

Glucuronides are highly hydrophilic and water soluble

UDP glucuronosyltransferase is closely associated with Cyp450 so that Phase I products of drugs are efficiently conjugated

Four general classes of glucuronides: O-, N-, S-, and C-

Neonates have undeveloped liver UDP-glucuronosyltransferase activity, and may exhibit metabolic problem. For example, chloramphenicol (Chloroptic) leads neonates to “gray baby syndrome”

Neonatal jaundice may be attributable to their inability to conjugate bilirubin with glucuronic acid

Glucuronic Acid Conjugation

Formation of Glucuronide Conjugate

OHOHO

HOOPO3

2-

HOOHO

HOHO

O

HOUTP PPi

Phosphorylase

-D-Glucose-1-phosphate

P

O

O

O-

P

O

O-

O

ON

HO OH

NH

O

O

OHOOC

HOHO

HOO P

O

O

O-

P

O

O-

OO N

HO OH

NH

O

O

2NAD+

RXH

UDPOHOHO

HOXR

HO

UDP-Glucuronyl-transferase(microsomal)

-D-Glucuronide

UDPG

Uridine-5'-diphospho--D-Glucose (UDPG)

O

Types of Compounds Forming Glucuronides

TYPE EXAMPLES

O-Glucuronide

Phenols

Alcohols

Enols

N-hydroxyamines/amides

Acetaminophen morphine

Chloramphenicol Propranolol

Hydroxycoumarine

N-hydroxydapsone N-Hydroxy-2-acetylaminoflourene

OH

CH3

O

HN

HO OH

N

O

CH3

O2N

Cl

Cl

O

HN

OH

OH

H

CH3

CH3

OH

NO

O O

OH

SO2

H2N NHOH NCH3

OH

Aryl acids

Arylalkyl acids

OH

COOH

O

OH

O

CH3

N

HN

NH2

O2N

Salicylic acid

Fenoprofen

N-Glucuronides

Arylamines7-Amino-5-nitroindazole

AlkylaminesN

H

CH3

N

Desipramine

AmidesH3C

ONH2

NH2

H3C O

O

O

Meprobamate

Sulfonamides

OO

H2N

NH

S

CH3

CH3

NO

Sulfisoxazole

3o AminesCH3N

Cyproheptadine

S-Glucuronides

SulfhydrylCH3

HSN

N

H3C

H3CS

SHN

Methimazole

Carbodithioic acid

Disulfirum (reduced form)

CH3

O

O

N

NC-Glucuronides

Phenylbutazone

Sulfate Conjugation

Occurs less frequently than does glucuronidation presumably due to fewer number of inorganic sulfates in mammals and fewer number of functional groups (phenols, alcohols, arylamines and N-hydroxy compounds)

Three enzyme-catalyzed reactions are involved in sulfate conjugation

S

O

O

-O O-

ATP PPi

Mg+2

ATP sulfurylaseSulfate

OOP

O

O-

OS

O

O

-OAd

HO OH

Adenosine-5'-phosphosulfate (APS)

Mg+2

APS phosphokinase

OOP

O

O-

OS

O

O

-OAd

-2O3PO OH

ATP ADP

3'-phosphoadenosine-5'-phosphosulfate (PAPS)

PAP

Sulfotransferase

RXH

S

O

O

-O XR

Sulfateconjugate

(soluble)

Sulfation of Drugs

COOHH3C

H

H

N

HO

HOHO CH3

HOCH3

CH3

HOH

N

Phenolic sulfation predominates

Phenolic O-glucuonidation competes favorably with sulfation due to limited sulfate availability

Sulfate conjugates can be hydrolyzed back to the parent compound by various sulfatases

Sulfoconjugation plays an important role in the hepatotoxicity and carcinogenecity of N-hydroxyarylamides

In infants and young children where glucuronyltransferase activity is not well developed, have predominating O-sulfate conjugation

Examples include: -methyldopa, albuterol, terbutaline, acetaminophen, phenacetin

-Methyldopa

CH3

CH3

CH3

OH

HOH

NHO

Albuterol Terbutaline

Possible Mechanism of Phenacetin Toxicity

Electrophilic nitreneum

Amino Acid Conjugation

The first mammalian drug metabolite isolated, hippuric acid, was the product of glycine conjugation of benzoic acid

Amino acid conjugation of a variety of caroxylic acids, such as aromatic, arylacetic, and heterocyclic carboxylic acids leads to amide bond formation

Glycine conjugates are the most common

Taurine, arginine, asparagine, histidine, lysine, glutamate, aspartate, alanine, and serine conjugates have also been found

COH

R O

Benzoic Acid, R = HSalicylic Acid, R = OH

CONHCH2COH

R O O

Hippuric Acid, R = HSalicyluric Acid, R = OH

Mechanism of Amino Acid conjugation

An Acyl-CoA Intermediate

Glycine Conjugate R = HGlutamine Conjugate R = CH2CH2CONH2

Drug-COOH

Brompheniramine Metabolism

Br

NN

CH3

CH3

P450

Br

NNH

CH3

P450

Br

NNH2

P450

Br

N CHO

Br

NN

CH3

CH3

Br

NO

HN COOH

Br

N COOH

Brompheniramine

Aldehyde dehydrogenase

Glycine N-acyltransferase

Carboxylic Acid metabolite

Brompheniramine N-oxide Glycine conjugate

Glutathione Conjugation

Glutathione is a tripeptide (Glu-Cys-Gly) – found virtually in all mammalian tissues

Its thiol functions as scavenger of harmful electrophilic parent drugs or their metabolites

Examples include SN2 reaction, SNAr reaction, and Michael addition

NH

HN

NH2

OHS

O

O

HO

O

OH

NH

HN

NH2

OS

O

O

OH

O

HO

S

HN

NH

NH2

O

O

O

OH

O

HO

Glutathione reduced form (GSH) Glutathione oxidized form (GSSG)

SN2 Examples

R X Y

CH3O2SOOSO2CH3

ONO2

O

HONO2

NO2

R X SG

ONO2

O

HONO2

SG

SGCH3O2SO

ONO2

OH

HONO2

S+ G

GSH

Glutathione-S-Transferase

+ Y- SN2 X = C, O, S; Y = leaving group or epoxideA.

-SG

Busulfan

1.

-SG -SG + GSSG2.

Nitroglycerine

OO

OCH3

CH3

ON

O

O

Naproxcinod

SNAr Examples

Z

X

N

N

N

NH

H3C

N

N

+N

S

O

O-

Z

SG

N

N

N

NH

H3C

N

N

+N

S

O-

O-

SG

H3C

N

N

NO2

SGN

N

N

NH

SH

GSHSNRrB.

1.

Azathioprine

-SG

+

1-Methyl-4-nitro-5-(S-glutathionyl)

imidazole6-Mercaptopurine

Michael Addition

C. Z-SG

H+

ZSG Michael Addition

HO OH

N

O

CH3

HO O

N

O

CH3

O OH

N

O

CH3

-SG

HO OH

N

O

CH3

SG

-SG

HO OH

N

O

CH3

GS

Mercapturic Acid Conjugates

NH

HN

NH2

OS

O

O

HO

O

OH

Drug Amino Acid(AA)

-Glutamyltranspeptidase

-Glutamyl-AA

NH2

HN

S

O

O

HO

Drug

Glutathione Conjugate

Glycine

CysteinylGlycinase

NH2HO

S

O

Drug

S-substitutedCysteineDerivative

AcetylCoA CoASH

NH

H2N

S

O

Drug

CH3

O

Mercapturicacid conjugate

Acetyl ConjugationAcetyl Conjugation

Metabolism for drugs containing a primary amino group, (aliphatic and aromatic amines), amino acids, sulfonamides, hydrazines, and hydrazides

The function of acetylation is to deactivate the drug, although N-acetylprocainamide is as potent as the parent antiarrhythmic drug procainamide (Procanbid) or more toxic than the parent drug, e.g., N-acetylisoniazid

Acetylation is two-step, covalent catalytic process involving N-acetyl transferase

H3C SCoA

O CoASH

H3C X

O

H2N R

X-

H3C

O

NHRX-

N-Acetylation of amines

Genetic polymorphism in N-acetyltransferase activityMultiple NAT2 alleles (NAT2*5, *6, *7, and *14) have substantially decreased acetylation activity and are common in Caucasians and populations of African descent. In these groups, most individuals carry at least one copy of a slow acetylator allele, and less than 10% are homozygous for the wild type (fast acetylator) trait. The ratio of NAT2 activity is 7 in Caucasians to 18 in the Chinese population.

Example of Acetylated Drugs

O

CH3

CH3

ONH NH2

O

OHSHO

Cilastatin

NHHN

SH3C

HO

COOHO

N

Imipenem

Fatty Acid and Cholesterol Conjugation

Hydroxyl-containing drugs can undergo conjugation with a wide range of endogenous fatty acids such as saturated acids from C10 to C16 and unsaturated acids such as oleic and linoleic acids

Cholesterol ester metabolites have been detected for drugs containing either an ester or a carboxylic acid

HO

O

OO

O

NCl

ClOH

Cl O (CH2)10 COOH

Prednimustine

Fatty AcidCholesterol

Methyl Conjugation Minor conjugation pathway, important in biosynthesis of epinephrine

and melatonin; in the catabolism of norepinephrine, dopamine, serotonin, and histamine; and in modulating the activities of macromolecules (proteins and nucleic acids)

Except for the formation of quarternary ammonium salts, methylation of an amine reduces the polarity and hydrophilicity of the substrates

A variety of methyl transferase, such as COMT (catechol O-methyl transferase), phenol-O-methyltransferase, N-methyl transferase, S-methyltransferase etc are responsible for catalyzing the transfer of methyl group from SAM to RXH

H3CS

H2N COOH

S+

H2N COOH

O

HO OH

AdCH3

HX-RCH3-X-R

S

H2N COOH

O

HO OH

AdMethionine

adenosyltransferase

Methyltransferase+

Mthetionine

S-Adenosylmethionine

Mechanism of methyl conjugation

ATP PPi + Pi

Case Study

Case 2. Imagine yourself as a drug information specialist at a poison control center. A technician from the coroner’s office is investigating a case and requires assistance in identifying the possible sources of benzodiazepines (BZDs) in the toxicology profile of a particular corpse. The technician has identified four distinct BZDs in this blood sample. She believes that the major component is diazepam (1) (72% of the identified BZDs) and that the remaining three components are metabolites (NOTE: the assay identifies only active compounds).

CH3

Cl

O

N

N

Cl

O

N

HN

CH3

Cl

O

N

N

Cl

O

N

HN

OH

OH

1

2

3

4

Q. What are the three structures of potential ACTIVE metabolites for diazepam?

http://www-home.cr.duq.edu/~harrold/basic_concepts_index.html

Assignment: Due by this Friday

Study Guide

1. What Roles are Played by Drug Metabolism? Know with structural examples

2. Role of stereochemistry in metabolism of drugs with example of warfarin, ibuprofen and itomidate

3. What is first pass effect; enterohepatic circulation? Why and how they occur? Drug examples

4. Metabolisms in the intestinal mucosa

5. CYP450, Hepatic microsomal flavin containing monooxygenases (MFMO or FMO) Monoamine Oxidase (MAO) and Hydrolases. Drugs metabolised by these enzymes and the active sites of these enzymes. Types of metabolic reaction catalyzed by these enzymes

6. Specific CYP enzymes with the number of drugs they metabolize

7. Few CYP family with their main functions

8. Drug interaction basics related to metabolic enzymes

Study Guide Cont.

9. Mechanism and routes of aromatic hydroxylation. The effects of electron donating and withdrawing groups in aromatic hydroxylation. Drug examples. What is NIH shift?

10.Oxidation of olefins. Role of epoxide hydrolase. Can olefenic epoxide be converted to alcohol as in aromatic epoxide by NIH shift?

11.What type of C in a drug molecule can not be hydroxylated?

12.What is allylic and benzylic hydroxylation? Show drug examples.

13.Show the drug examples where hydroxylation occur on Cα to C=O and C=N bonds

14.Show the drug examples where hydroxylation occur at aliphatic and alicyclic carbon atoms. Which carbons are more easily hydroxylated?

15.What is N-oxidatin and N-dealkylation. What enzymes are involved? How do you differentiate between N-dealkylation and deamination. Drug examples. What types of drugs generates lactams instead of causing dealkylation?

16.What is the difference between mixed function oxidases and amine oxidases?

Study Guide Cont.

17.What is the difference between ethanol oxidation and O-dealkylation?

18.What is S-dealkylation, desulfuration and S-oxidation? Drug examples.

19.How does steric factors influence S- O- and N-dealkylations?

20.Oxidative dehalogenation with special example of chloramphenicol. Why chloramphenicol cause toxicity to the babies?

21.What is MFMO and its active site? What types of functional groups are metabolized by this enzyme? Drug examples.

22.MAO, dehydrogenases, xanthene oxidases and their functions with drug examples. Difference between MAO-A and MAO-B.

23.Alcohol and aldehyde dehydrogenases, the coenzymes and the types of drugs they work on.

24.Azo and nitro reductases, their coenzymes and the drugs they act on.

Study Guide Cont.

25.Different types of hydrolytic enzymes. Compare rate of hydrolysis of esters, amides, carbonates and carbamates.

26.What are prodrugs and antedrugs? What are the advantages? Examples.

27.What are different types of conjugation reactions?

28.The enzymes and substrates involved in glucuronidation, and sulfate conjugation.

29.Why acetaminophen is toxic to neonates? Mechanism of phenacetin and acetaminophen toxicity.

30.What types of drugs or metabolites may form glycin conjugates?

31.What are different mechanisms involved in glutathione conjugation? What is mercapturic acid conjugate? Mercapturic acid conjugate of acetaminophen is a sign of its toxicity – why?

32.Mechanism of acetylation. What is slow and fast acetylator?

33.What is COMT? What coenzymes is involved in its action? What types of drugs and/or neurotransmitters are metabolized by COMT?