PharmacologyA Selective Overview – Part 2

Carl Rosow, M.D., Ph.D.

Topics for Today

• Pharmacokinetics– Volume of distribution– Clearance– Drug Metabolism

• Pharmacogenomic Variability– Pharmacokinetic– Pharmacodynamic

Pharmacokinetics

• Absorption• Distribution• Metabolism• Excretion

“Clinical” Correlate

What is going on here?

Indians of the Amazon hunt with blowguns and paralyze prey with curare-tipped darts. They eat the animals without suffering ill effects.

1. Absorption: Passive Diffusion

Depends on • Surface area• Concentration gradient• Lipophilicity (oil:water)• Ionizable residues and pKa• Molecular weight

Lipophilicity

• Solubility in lipid relative to water• Oil/water (oil/buffer) partition coefficient

–Drug concentration in oil versus water (adjusted for pH)

Oil

Water

Impact of Ionizable Groups

• Ionization markedly decreases lipophilicity• Drugs with fixed negative or positive charge

not readily transported by passive diffusion

O CH3

CH3C O CH2CH2N CH3

CH3

+

What if the charge is not fixed?

AH A- + H+

BH+ B + H+

low pH high pH

QUESTION: If lidocaine (a weak base) is injected together with sodium bicarbonate, absorption will be

1. Increased2. Decreased3. Unchanged

Active, Carrier-mediated Transport

• Saturable kinetics• Substrate competition• Examples

– P-glycoprotein (MDR-1) “efflux pump”

– OATP “organic anion transport protein”

2. Distribution

DRUG BLOOD

D

D-protein

TARGET

OTHER

TISSUES

DEPOT

D

D

D

D

Tissue Perfusion Rates(ml/min-100g tissue )

HighLung 400Kidney 350Liver 85Brain 55

Intermediate/variableSkeletal muscle 5

LowFat 3

Leaky – Sinusoids of the Liver

Tight – “Blood-Brain Barrier”

“Apparent” Volume of Distribution

Vd =Dose

C0

For Digoxin,

0.25 mg

0.5 ng/mL= 500 Liters ??

HoursHoursP

lasm

a C

once

ntra

tion

Pla

sma

Con

cent

ratio

n((

g/m

l)

C0

DRUG BLOOD

D

D-protein

TARGET

OTHER

TISSUES

DEPOT

D

D

D

D

Sample comes from here Vd

Case

• A 3 year old child has just ingested a bottle of her grandfather’s digoxin tablets. Her mother rushes her to the hospital. Would hemodialysis be a good way to get the drug out of this child’s system?

3. Drug Metabolism(aka, Biotransformation)

• Liver• Plasma• Kidney• Skin• Gut wall

(& bacteria)• Red cell• Lung

Microsomal Enzymes

• A “microsome” is a lab artifact!• Actually pieces of smooth endoplasmic

reticulum• Preparation from liver homogenate

– Centrifugation to remove large debris and mitochondria.

– High speed centrifugation of supernatant gives microsomes

Cytochrome P450 Enzymes

• Metabolize ~75% of all drugs• Heme containing mixed-function oxidases• “P450” from absorption peak of 450 nM• Nomenclature:

CYP 3A4

• 3 – denotes family (>40% a.a. homology)• A – denotes subfamily (>55% a.a. homology)• 4 – specific enzyme

Percentage of Drugs Metabolized by P450

C Y P 1 A 2

C Y P 2 A 6

C Y P 2 C 9 /1 0

C Y P 2 C 1 9

C Y P 2 D 6

C Y P 2 E 1

C Y P 3 A 4 /5

U n k n o w n

CYP3A4/5

CYP2C9/10

All others

CYP1A2

DRUG BLOOD

D

D-protein

TARGET

OTHER

TISSUES

DEPOT

D

D

G.I. TRACT MLIVER

D

D

“First-Pass” Metabolism

Typical Doses

Intravenous Oral

Propranolol 1-2 mg 40-80 mg

Morphine 10 mg 30-60 mg

Lidocaine 100 mg Ineffective

Kinetics of Biotransformation

• Mainly enzyme reactions. • Drug concentrations usually <<Km for the

enzyme, so…– Kinetics are FIRST ORDER– Rate is proportional to drug concentration– Describable by a half-life, T1/2 , which is

independent of dose or concentration.

Significance of Biotransformation

Prodrug Active Drug

Metabolite 1 Metabolite 2 Metabolite 3

(inactive) (active) (toxic)

Clinical Case• A 2 year old has ingested several ounces

of antifreeze (ethylene glycol). The compound itself is not harmful, but it is converted to a kidney toxin, oxalic acid, by the enzyme alcohol dehydrogenase.

How can you protect this child’s kidneys?• Enzyme inhibitor - Fomepizole • Substrate inhibition – Ethanol(!)

4. Clearance

• Definition: Volume of plasma cleared of a compound (or drug) per unit time

• e.g., Renal Clearance of Drug D from 24 hr urine collection:

Expressed as mL/min, mL/hr, etc.

Clrenal =[D]urine x Volumeurine

[D]plasma

Mechanisms of Renal Clearance

Glomerular filtration

Proximal tubular secretion

Reabsorption

Some Important Relationships

Vd = Dose

Co

Co = Dose

Vd

Vd x ln2T1/2 = Cl

Clinical Case

• Patient M.R. weighs 400 lbs. How would this alter her drug therapy with lipophilic drugs?

Clinical Case

Vd x ln2T1/2 = Cl

C0 =Dose

Vd

Patient M.R. weighs 400 lbs. How would this alter her drug therapy with lipophilic drugs?

First-order Process

Number of Half-Lives Amount Completed

1 50%

2 75%

3 87.5%

4 93.8%

5 96.9%

• An 83 year old man is given daily doses of an analgesic that is excreted unchanged in the urine. The half-life of the drug is 50 hr in young adults, but it is ~100 hr in the elderly.

• How long will it be before the drug reaches its maximal concentration in this patient?

Clinical Case

Clinical Case

1. Curare doesn’t affect human cells

2. Curare is cleared by first-pass metabolism

3. The dose in a typical meal is too low

4. Curare is not absorbed from the GI tract

5. They developed tolerance with repeated exposure

Amazon tribesmen routinely killed animals with curare-tipped darts and then ate the meat. Why weren’t they harmed by the poison?

Variability in Drug Response

• Genomic Variability– Pharmacokinetic: Variation in drug ADME– Pharmacodynamis: variation in sensitivity of

drug target• “Idiosyncratic” Variability

– Uncommon, unpredictable drug response– No defined difference in kinetics or target

sensitivity

Clinical Case

A 30 year old female sustains a wrist fracture after a fall on the ice. She is given codeine for her pain, but she gets no relief. She takes 2 more doses during the next hour without detectable effect.

What happened?

Codeine is a Pro-Drug that is O-Demethylated to Morphine

• A minor pathway (<10%) of codeine metabolism, but it accounts for nearly all opioid activity

• The CYP3A4 and glucuronide metabolites (80%) are inactive

CYP2D6 Polymorphism• 78 variants of CYP2D6 identified• Homozygous inactive = “poor metabolizer”

– 5-10% of Caucasians– 1-2% of S.E. Asians

• Gene duplication = “ultrarapid metabolizer”– 25-30% of N. Africans– 1-7% of Caucasians

• Heterozygotes = “intermediate metabolizer” or “extensive metabolizer”

Implications of CYP2D6 Genotype Depend Upon the Activity of Parent Drug (P) vs. Metabolite (M)

• P inactive, M active (codeine)– Poor metabolizer – reduced or no effect– Ultrarapid metabolizer – excessive effect

• P active, M inactive (metoprolol)– Poor metabolizer - excessive effect– Ultrarapid metabolizer – reduced or no effect

• P active, M active (oxycodone)– Metabolizer status – not critical

Question:

• Is it a good thing to hear that your new investigational drug is metabolized by CYP2D6?

6-Mercaptopurine

• A purine analogue “antimetabolite” used for chemotherapy of leukemia.

• Stops rapidly dividing tumor cells in S-phase

• Primary toxicity is to other rapidly-dividing cell populations (bone marrow, GI tract).

Thiopurine S-Methyltransferase Polymorphisms

Relling, 1999

RBC Thioguanine nucleotides during 6-MP therapy

Stopping 6-MP For Toxicity

Relling, 1999

TPMT deficient patient tolerates only 10% of normal dose.

Clinical Correlation

A 32 year old African-American male with a bladder malformation has had repeated urinary tract infections. He is given a prescription for the antibiotic, nitrofurantoin. After the first dose, his urine turns the color of Coca-Cola.

What happened?

1. Glucose 6-PO4 Dehydrogenase (G6PD) DEFICIENCY

• Sex-linked, uncommon in females• Common in Africans (malaria protective)• >300 allelic variants• Not a disease! It is characteristic for

>400 million people world-wide• Hemolysis when exposed to oxidative

compounds

Biochemical Basis forG6PD Deficiency

• Mature RBC lacks Krebs cycle to generate NADPH.

• Glycolysis– 90% of glucose enters here– Generates NADH (and ATP)

• Pentose Phosphate Shunt– 10% of glucose enters here– In first step, G6PD recycles

glucose-6-PO4 to generate NADPH

Detoxification of Hydrogen Peroxide

NADP+ NADPH + H+

glutathione reductase

2GSH GSSG

glutathione peroxidase

H2O2 2H2O

Drugs Causing Hemolysis inG6PD Deficiency

• Primaquine (an antimalarial!)• Nitrofurantoin• Sulfonamides (a few, older ones)• Naphthalene• Fava beans• (Acetaminophen)

Final Thoughts:The Use of Pharmacogenomics in

Drug Research & Development

• Pharmacogenomics May Identify:–New Drug Targets–Patients Most Likely to Benefit–Patients Most Likely to Be Harmed

On the other hand…

• Genetically “enriched” study populations are more likely to show positive results

• Better chance of FDA approval?• Once a drug is approved, prescribing is not

restricted by genetic testing• Cost of tests (if used) added to cost of drug

$

Test Question

Consider the patient we described, who had a pharmacogenetic insensitivity to codeine. What if this patient were given a normal dose of the b-blocker, metoprolol? Would the drug effect be altered? If so, what is the mechanism?