3/4/2015

CHEM3407 & BPHM3015

Medicinal Chemistry

Hongzhe SUN
Chemistry & Pharmacy
HKU
2nd Semester, 2015

CHEM3407/3410 Medicinal Chemistry

8. PHARMACOKINETICS
Frequently encountered problem:
A drug containing an ionized carboxylate group shows good activity against its
target in vitro tests. When in vivo tests were carried out, the drug showed poor
activity when it was administrated orally but good activity when it was
administrated by intravenous injection

Frequently asked question:

why does this drug behave differently?
Frequently expected consequence:
- frustrate synthetic chemists and molecular biologists
- re-examine drug target? design? optimization?
Frequently used solution:
- investigate the pharmacological properties of drug so as
to improve the access of drug to the target

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CHEM3407/3410 Medicinal Chemistry

8. PHARMACOKINETICS (())
Pharmacodynamics - a study of what a drug does to the body,
whereas Pharmacokinetics - a study of what our body does to a drug.
factors affecting a drug will reach the target
may explain why active drugs in vitro but inactive in vivo
may explain why the most potent drug may be useless clinically
how the level of a drug in the blood is affected by the factors:

Factors to consider (ADME)

Drug Absorption
Drug Distribution
Drug Metabolism
Drug Excretion

CHEM3407/3410 Medicinal Chemistry

Lead Optimization- ADME

how much the drug enters the

blood from tablets or capsules in
the stomach and small intestine.

how the drug travels in

the blood and how it goes
into and comes out of
other areas of the body.
(protein-bound?) or free drug left?

how the body chemically

changes a drug in the
intestines and liver.

how the body gets the drug out by passing the drug into
the urine (via the kidneys) or stool (via the liver).

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Drugs Journey in ADME (oral drug)

brain

mouth

(blood-brain barrier)

stomach

bloodstream
small intestine
(circulation)
large intestine
(excretion)

kidney
liver
(metabolism)

(organ)
drug target
(receptors,
proteins, nucleic
acids, lipids
carbohydrates
of cancer cells,
virus, fungi
bacteria)

A 5th process, Liberation, also plays an important role.

Liberation- the process of release of a drug from the formulation.
L ADME is sometimes used in place of ADME

Absorption Solubility
Oral route is preferred for drug administration.
Injection and infusion (i.v.) formulations are required for
immobilized patients in hospitals.
Preferred schedule is once a day dosing (QD dosing) without food
effect.
Drug absorption takes place in the small intestine (6 m. long) due
to the large surface area (460 m2) created by the villus and
microvillus structure.
Absorption depends on aqueous solubility, permeability across
intestinal cell membrane and efflux pump in intestinal cells.
Thermodynamic solubility (from solid) and kinetic solubility (from
organic solution).

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Absorption Solubility
Single dose of drug

should dissolve in 250 mL of aqueous solution

(pH 1 7.5). human gut volume ~1100 mL
10 mg dose: solubility >100 M. 100 mg dose: solubility >1 mM.

Poor solubility if kinetic solubility < 50 M.

Aqueous solubility can be improved by introducing polar groups
such as hydroxyl and sulfone or ionizable centers such as basic
amines.
Solubility can be improved by introducing out-of-plane
substitutions which can decrease the lattice energy of crystalline
solid.
Solubility can be improved by formulation of salts. Amorphous
solids have much higher solubility than crystalline solids. Spray
dried dispersions.

Improve Aqueous Solubility

ClogP -0.1
Solubility < 10 ug/ml

ClogP 0.7
Solubility < 1 mg/ml

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Oral Adsorption - Permeability

3 mechanisms for absorption across intestinal cell membrane.
Passive diffusion across lipophilic membrane down a concentration
gradient. e.g. gases, low MW (<150 Da) neutral compounds, alcohol,
pesticides,
Facilitated diffusion down a concentration gradient with help from
proteins in the membrane. e.g. fructose.
Active transport of molecules with the help from proteins in the
membrane and extra energy, often against concentration gradient. e.g.
Na+, K+.
A drug is desired to be absorbed by passive diffusion.
Passive diffusion rate decreases with increasing molecular size.

Oral Adsorption - Permeability

Passive diffusion rate increases with increasing lipid solubility of
the molecule.
Charged molecules have negligible passive diffusion across cell
membrane.
Intestinal pH ranges from pH 5 to pH 8. Acidic compounds with
pKa > 4 and basic cmpd with pKa(H+) < 9 should have sufficient
neutral fraction.
Permeability decreases with increasing polarity, especially H-bond
donors.
Permeability is measured by assays using monolayer Caco2 (colon
cancer) cells, MDCK (Madin Darby Canine Kidney) epithelial
cells or artificial memb.
Poor permeability if Caco2 permeability < 30 nm/s.
Bidirectional Caco2 permeability assay (from apical side and
basolateral side)

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Absorption - logP
An oral drug cannot be too hydrophilic (low permeability) or too hydrophobic
(low aqueous solution).
LogP measures hydrophobicity of a molecule in its neutral form where
P is the octanol/water partition coefficient.
LogP > 5, too hydrophobic, poor aqueous solubility.

LogP < 1, too hydrophilic, poor permeability.

ClogP - calculated logP obtained by fragment-based prediction.
ClogP changes relative to Hydrogen (H):
Methyl +0.5, Fluorine +0.1, Chloro +0.5, Bromo +1.0, Hydroxyl -1.0
Amine -1.0, Ether -1.0, Ethyl +1.0, Iso-Propyl +1.5, Cyclopropyl +1.0
Tert-Butyl +2.0, Trifluoromethyl +1.0, Phenyl +2.0, Cyclohexyl +2.5
Amide -1.5, Ketone -0.5, Ester -0.5, Acid -0.3, Urea -1.0, Nitrile -0.5
Carbamate -1.0, Sulfonamide -1.5, Sulfone -1.5

Absorption - logP

ClogP changes relative to Benzene:

Cyclohexane +1.0, Pyridine -1.5, Thiophene -0.3, Furan -0.8
Pyrrole -1.5, N-Me Pyrrole -1.0, Oxazole -2.0
LogD is used for ionizable compounds where D is the
octanol/water distribution coefficient at a specific pH which
account for the fraction of the highly polar ionized form.
LogD difference between the neutral and fully ionized form, i.e.
~3 units.
LogD7.4 between 1 and 3 is optimal for absorption.

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Improve Permeability

Blood Level of A Drug over Time method dependence

maximum blood level concentration and time taken to reach it
concentration decreases with time (drug metabolism and excretion)

drug conc. in blood (mg/L)

Injection
Inhalation
Ingestion

Dermal
Time (hr)

- measure blood level of a drug? plasma HPLC-MS or UV detector

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Blood Level of A Drug over Time some parameters

Cmax maximum conc. of drug in blood
(represent bioavailability and the side effects)

drug conc. in blood (mg/L)

AUC Area Under the Curve

= C dt

(represent overall drug exposure, i.e. integral of the curve)

Cmin minimum or trough

conc. represent effectiveness

Time (hr)

Pharmacokinetics
Pharmacokinetics study the absorption of drug, distribution of drug
throughout the fluid and tissue of the body, metabolic
transformation, elimination of the parent compound and its
metabolites.

Pharmacokinetics data are obtained by dosing animal p.o. and i.v.

followed by measuring the plasma concentration of the compound
at different time point.

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Pharmacokinetics
AUC (nMh) - Area Under the Curve in the plasma conc. vs. time graph.

For 10 mpk p.o., it is desirable to achieve AUC > 1500 nMh.

Cmax (nM) - Maximum plasma concentration in the study.
tmax(h) - Time to reach maximum plasma concentration in the study.
F(%) - Bioavailability = AUCoral / AUCi.v. Desired F > 20 %.
t1/2(h) - Half-life of the compound in the i.v. study. t1/2 = ln2 x Vd/CL =

0.69 Vd/CL

MRT (h) - Mean Residence Time is the mean time that the compound

resides in the body in the i.v. study.

MRT = Vd / CL = t1/2 /ln2 = 1.44 t1/2

CL (mL/min/kg) - Clearance is the volume of plasma which is clear of

the comounpd per unit time in the i.v. study. CL = kel x Vd

Pharmacokinetics
CL_int (uL/min/Mcell) - Intrinsic clearance using hepatocytes.
CL_int >15, high CL.
kel (/min) - Elimination rate constant is the fraction of compound
eliminated per unit time.
Vd (L/kg) - Volume of distribution is an hypothetical term (dose
/plasma conc. at t0) for estimating degree of distribution in the
body.
Vd ~ 0.05 L/kg, compound remained in plasma due to plasma
protein binding.
Vd ~ 0.2 L/kg, compound distributed in extracellular fluid, cannot
cross cell membrane.
Vd ~ 0.55 L/kg, compound distributed in intracelluar fluid (total
body water).

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Plasma Protein Binding

Binding to plasma protein decreases the free drug concentration.
High plasma protein binding (> 99 %) prevents distribution of drug to

target tissue.
High plasma protein binding increases hepatic cycle rate and shorten

half-life.
High plasma protein binding is reflected in low Vd (0.05-0.1 L/kg).
3 major plasma proteins.

Human serum albumin (HSA) - binds hydrophobic and acidic

compounds.
-Acid glycoprotein AGP - binds basic compounds.
Globulin lipoprotein - binds non-polar compounds.

Plasma Protein Binding

Human serum albumin is a 66.5 kDa protein and the most abundant

protein in plasma ~40 mg/mL (~ 0.6 mM). HSA concentration is much

higher than drug conc. and drug target conc. Free drug conc. is
independent of drug conc. and target affinity.
In vitro assays conducted in the presence of HSA can predict in vivo

activity.
-Acid glycoprotein (AGP) is a 43 kDa protein with plasma conc. ~2

mg/mL (~ 50 M).

When drug conc. exceeds AGP conc., free drug conc. increases
dramatically.
Polar groups, such as hydroxyl and sulfone, decreases plasma protein

binding.

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Human serum albumin (HSA)

Domain III

Domain I
Cys34
helix 3
N-terminus

Domain II

Note: tri-iodobenzoic acid (TIB)

Nat. Struct. Biol. 1998, 5, 827
(PDBID: 1bke)

Single-chain protein,
MW 66.5 kDa (585 aa)
ca. 0.65 mM (40 mg/ml)
Functions: Bind and transport
various endogenous small molecules,
fatty acids; drugs; xenobiotics; metal
ions (e.g. Zn2+)

DRUG ABSORPTION
orally taken drugs must cross the gut wall to reach blood supply

most orally active drugs pass through the cells lining the gut wall
thus, drugs are required to cross two fatty cell membranes
balance of hydrophilic/hydrophobic character is required
polar drugs can be administered by injection
orally active drugs usually obey Lipinskis Rule of Five

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DRUG ADME - ABSORPTION

Drug-like molecules: -good oral bioavailability.
- good permeability and water solubility.
Lipinskis Rule of Five (formulated in 1997)
- orally active drugs generally show a balance of hydrophilic /
hydrophobic properties and obey the following rules:
Molecular weight < 500
HBD groups 5
HBA groups 10
calculated logP +5 (ClogP 5)
(P partition coefficient =

Concentration of drug in n-octanol

Concentration of drug in water

high P means high drug hydroprobicity

but not foolproof - several exceptions!

DRUG ADME - ABSORPTION

Exceptions
- small polar molecules (MW <200) that cross the gut wall through
small pores between cells (squeezing process)
- polar molecules pass through the membrane by transport proteins
i.e. amino acids, nucleic acid bases and some drugs (e.g. lisinopril,
methotrexate, erthromycin)
NH2

HO

N
N
H
O

CO2H

Lisinopril
(antihypertensive)

methotrexate
(anticancer)

erthromycin
(antibiotic)

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DRUG ADME - ABSORPTION

Vebers parameters More flexible, less likely an orally active drug
molecular flexibility is important to drug absorption
too many rotatable bonds is bad for absorption
the polar surface of the molecule plays a role
molecular weight is not a factor
Number of rotatable bonds 10
Total no. of HBDs and HBAs 12
or
Number of rotatable bonds 10
Polar surface area < 140 Angstroms2

- drugs that are usually poorly absorbed injection methods

DRUG ADME - DISTRIBUTION

once across the gut wall, drug enters blood vessels
cells lining blood vessels are loose fitting
no need to cross cell membranes
drug can quickly cross blood vessel walls through pores between
cells
drugs absorbed orally are first taken to the liver
modification of the drug is possible by enzymes in the liver - drug
metabolism
fraction of the absorbed drug is often deactivated by drug
metabolism in the liver before distribution occurs round the body first pass effect
drug is distributed evenly throughout blood supply within 1 min of
absorption

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DRUG ADME - DISTRIBUTION

uneven drug distribution round body due to uneven blood supply

rapid distribution from blood vessels to tissues and organs
(leaky blood vessels)
drug has to enter a cell if the molecular target is within the cell
blood level concentration of drug drops rapidly after absorption
due to distribution, macromolecular binding and storage in fat
tissue (e.g. barbiturates) or bone
blood brain barrier hinders polar drugs from entering brain
- tight fitting cells line the capillaries in the brain
- capillaries have a coating of fat cells
can increase polarity of peripherally acting drugs to reduce CNS
side effects
placental barrier

DRUG ADME - METABOLISM

foreign chemicals are modified by enzyme catalysed reactions,
mostly in liver - detoxification

metabolic reactions also occur in blood, gut wall and other

organs

drug metabolites are products formed from drug metabolism

drug metabolites are usually less active or inactive
(exception - prodrugs)

modification of the structure of drug may interfere or prevent

binding interactions with the target

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DRUG ADME - METABOLISM

orally absorbed compounds pass through the liver before

distribution to the rest of the body
fraction of orally absorbed drug is metabolised in the liver prior
to distribution round the body - first pass effect
drugs absorbed by other routes (injection or inhalation) avoid the
first pass effect and circulate round the body before reaching the
liver
fraction of non-orally absorbed compounds never reaches the
liver due to distribution into fat, cells and tissue

DRUG ADME - METABOLISM

A drug must survive long enough in the body to modify its target
activity.
Medicinal chemists must understand metabolic reactions in order
to design drugs with reduced metabolic rate.
metabolic reactions are defined as phase I or phase II
Phase I Drug Metabolism: most phase I reactions add a polar handle
to the molecule
Oxidation: Cytochrome P450 mixed function oxidases (CYP450)
Flavin containing monooxygenases (FMO)
Monoamine oxidases (MAO) Xanthine oxidase (XO)
Reduction. Aldo-keto reductase, nitro reductase.
Hydrolysis. Esterases, Amidases.
Generate polar metabolites suitable for elimination.

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DRUG ADME - METABOLISM

Phase II Drug Metabolism: phase II reactions are often carried out on
functional groups which have been added by Phase I reactions

Glucuronidation.
Sulfate Conjugation.
Glycine and Glutamine Conjugation.
Glutathione Conjugation.
Acetylation.
Generate water soluble metabolites which are excreted in the
urine.

DRUG ADME METABOLISM

Cytochrome P450 enzymes
located in liver (Fe-heme bound), the most important component
in oxidative metabolism
at least 12 families in human biochemistry
(organic oxidation ~75% of metabolic reactions)
RH + O2 + 2H+ + 2e ROH + H2O
individuals differ in types of Cyt. P450 enzymes present
patient variability in drug metabolism complicates dose levels and
leads to different susceptibilities to drugs
drugs which affect the activity of Cyt. P450 enzymes may affect
the activity of other drugs (drug-drug interactions)

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Background information

cytochrome P450

catalytic cycle

Fe-heme

DRUG ADME - METABOLISM

Phase I Reactions
Oxidations (catalysed by cytochrome P450 enzymes)
Oxidation of exposed alkyl groups
R

CH3

CH2OH

OH

Oxidation of alkenes and aromatic rings

R

epoxidation R

OH

Oxidation of N-alkyl groups (demethylation)

R

R
N Me

N H
R

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Oxidative Metabolism
Aromatic hydroxylation.
Aliphatic hydroxylation.
Epoxidation of alkenes.
Oxidation of alcohols to aldehydes or ketones.
Oxidation of aldehydes to carboxylic acids.
Oxidation of non-basic nitrogen.
Oxidation of sulfides to sulfoxides to sulfones.
Oxidative dealkylation on heteroatoms
Oxidative deamination.

Metabolic Aromatic Hydroxylation

Aromatic hydroxylation takes place on electron rich aromatic rings.
The aromatic ring is epoxidized to arene oxide which undergo the NIH shift
rearrangement to yield the phenol product.
The hydroxyl group ends up at the position para to electron donating groups,
such as hydroxyl, alkoxy, amino and alkyl groups.
When the para position is blocked, some aromatic hydroxylation can takes
place at the ortho position.
Epoxides of polyaromatics are carcinogenic metabolites which have longer
half-life to react with DNA.
Aromatic hydroxylation is inhibited by electron withdrawing groups, such
as fluoro, chloro, bromo, nitrile, carbonyl, sulfoxide and sulfone.
Fluorine at the para position is often used to block aromatic hydroxylation.

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Block Aromatic Hydroxylation

Cholesterol abs. inh.

t1/219-30 hr.

Antiviral
IC1/20.04 ug/ml
Bioavail. F 23%

Antiviral
IC1/20.02 ug/ml
Bioavail. F 9%

Block Aliphatic Hydroxylation

AUC 0.04 ug.h/ml

AUC 1.40 ug.h/ml

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Metabolic Oxidation of Non-Basic Nitrogen

CYP450 oxidizes non-basic nitrogen.
Pyridine is oxidized to pyridine N-oxide.
Introduce steric blocker at the ortho position.
Imine is oxidized to imine oxide.
Aniline is oxidized to N-phenylhydroxylamine to nitrosobenzene which are toxic
and carcinogenic.
Nitrogen heterocycles are not oxidized to nitrosobenzene.

Ritonavir
Norvir (Abbott 1996)
Bioavail. F ~70 %; Half-life ~3 h

HIV protease inhibitor.

Bioavail. F ~20 %; t1/2 ~1.5 h.
Pyridine N-oxidation

Electron deficient thiozole, 20x lower metabolic rate

DRUG ADME - METABOLISM

Phase I Reactions
Reduction of nitro, azo and carbonyl groups
R NO2

R NH2

NH2 + H2N

OH

C
R

C
R

H
R

Hydrolysis of esters and amides

O
C
R

O
OR

+ HO R

C
R

OH

C
R

NR2

+ HNR2

C
R

OH

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