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95 Cards in this Set

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What needs to be added to microsomes to see UGT metabolites?

UDPGA

What needs to be added to microsomes to see CYP metabolites?

NADPH regenerating system

(T or F) Microsomes can be used for long-term studies

False

DME means what

Drug Metabolizing Enzyme

Recombinant CYP's are used in what type of study?

Reaction phenotyping

What is reaction phenotyping?

finding which CYP is involved in a specific transformation

How many time points are used for primary metabolite screening?

One (usually 30 min)

What is the biggest variable in human liver microsome (HLM) studies?

variability in enzyme activity

What is used as a control in HLM studies?

a standard of a known drug

(T or F) Enzyme activity in microsomes may vary from batch to batch

True

DME's that aren't found in microsomes?

N-acetyl transferase (NAT), sulfotransferase (SULT), glustathione-S-transferase (GST)

What needs to be added to see NAT metabolites?

Acetyl CoA

What is S9?

Supernatant obtained from centrifugation at ~9000 g that contains both microsomes and dissolved DMEs

S9 is most useful for?

test for mutagenicity, finding genotoxic metabolites

What can hepatocytes model that S9 can't?

Transport accross the cell membrane, changes in gene expression (induction)

(T or F) Hepatocytes do not require the addition of co-enzymes

True

What is the preferred method for doing in vito to in vivo extrapolation? (Predicting ADME properties in humans)

Hepatocyte studies

What can be introduced to a compound to reduce stability if it is lasting too long?

"soft" groups capable of metabolic transformation

Fixing a metabolite stability problem often leads to problems with _____?

Absorption

What is the most direct way to prevent metabolic degredation?

Alter the molecule at or very near the site of metabolism

4 methods for improving metabolic stability

1. Replace atoms at site of instability


2. Alter steric effects to disfavor metabolism


3. Reduce LogP by adding polarity


4. Pre-form an active stable metbolite

Metabolism studies are incubated at what temp?

37 C (body temp)

In metabolism studies what is the NCE dissolved in?

HPLC grade methanol or acetonitrile

Microsomes are incubated in what?

Potassium phosphate buffer (pH = 7.4)

What method is used for metabolite analysis after incubation?

LC/MS/MS

(T or F) DMSO can inhibit CYP metabolism

True

What method is used to measure cell viability and concentration before metabolism study can be performed?

Tryptan Blue Exclusion Method

Terfenadine was taken off the market because ...?

Strong CYP3A4 inhibitors caused serious side effects

Post-systemic metabolism occurs in?

Kidney & Liver

Pre-systemic metabolism occurs in?

Liver & GI tract

What conjugation reactions increase polarity?

Sulfonation, glucuronidation, amino acid conjugation

What conjugation reactions do NOT increase polarity?

Methylation and acetylation

What conjugation reaction protects against reactive metabolites?

Glutathione conjugation

What are the 3 most important CYP's?

3A4 , 2C9, 2D6

What groups are glucuronidated?

Phenols, Carbox. acids, amine nitrogens, alcohols

What groups are sulfonated?

phenols, alcohols

What group is amino acid conjugated?

Carboxylic acids

What group is N-acetylated?

Amine nitrogens

(T or F) At time of NDA submission all clearance pathways (enzymatic and non) and transporters must be known?

True

The requirement of metabolism studies ____ the percentage of IND's failing from unsufficeint PK/bioavailability?

Decreased (From 39% to 8%)

Before clinical studies can be performed, what can be used to predict human metabolites?

in vivo animal studies and in vitro human

What are 4 uses of metabolism studies?

1. Determine which enzymes metabolize a drug


2. Determine if it is an inhibitor


3. Determine if it is an inducer


4. Create a "better" drug design

Olistat (Alli) was found to inhibit..?

CES2

(T or F) Orlistat inhibits carboxylesterases with intrinsic lipase activity being a determinant factor

False

What are three sites on carboxyesterases

Active site, Z-site and "side door"

Where is CES2?

Liver, GI, Kidney

Where is CES1?

Liver, small amounts in GI and Kidney

Where is CES3?

Liver, small amount in GI

(T or F) CES1 prefers substrates with a large group on the alcohol side

False

(T or F) CES2 prefers substrates with a large group on the alcohol site

True

Clopidogrel is a prodrug metabolized by both CYP's and CES1. Which products the active metabolite?

CYP's

Where are lipases present that CES is not?

Pancreas

Oxidation of amine nitrogens is catalyzed by?

CYPs or FMO

Tertiary amines are oxidized into?

amine oxides

Secondary amines are oxidized into

Secondary N-hydroxy then Nitrone

Primary amines are oxidized into

Primary N-hydroxy then Nitroso then Nitro

A sulfide is oxidized into

sulfoxide then sulfone

A sulfoxide has how many oxygens attached to the sulfur?

1

A nitroso is what typeof bond?

Nitrogen double bonded to oxygen

A Nitro is what type of bond?

Positive nitrogen single bonded to a negative oxygen

A sulfone has how many oxygens double bonded to the sulfur

2

Thioridizine is metabolized by CYP2D6, as well as a second pathway which is catalyzed by another CYP or FMO. Which path produces an active

CYP or FMO

What enzymes catalyzes steroid aromatization

CYP19

What happens in steroid aromatization?

The double bonded oxygen and a methyl group are reomved to create an aromatic ring

What happens in 1,4 dihydropyridine oxygenation?

A hydrogen is removed from a nitrogen

What types of groups are hydrolyzed?

ester, amide, epoxide, carbamate, carbonate, urea

What are the two products of amine hydrolysis

amine and carboxylic acid

What is the product of epoxide hydrolyzation

a trans di-ol

What are the three major types of enzymes that catalyze hydrolysis

esterases, peptidases, epoxide hyrolases

What are the three types of esterases

carboxyesterases, cholinesterases, organophosphatases

(T or F) Enzymes allow simultaneous acid and base catalyzed hydrolysis

True

(T or F) histidine acts as a base

True

(T or F) Glutamate acts as a base

False

(T or F) Serine acts as a nucleophile

True

List the 3 steps of esterase catalysis

1. enzyme-substrate complex


2. Acetylated enzyme intermediate


3. Nucleophilic attack by water

Which is hydrolyzed faster, ester or amide?

Ester

Which is hydrolyzed slower, procaine or procainamide?

ProcainAMIDE

Clopidogrel is a prodrug. Its active metabolite binds to what?

P2Y12 receptor on blood platelets

(T or F) Clopidogrel is hydrolyzed by CES1 to form an inactive metabolite

True

(T or F) Clopidogrel competes with Tamiflu for CES1 hydroysis

True

(T or F) Increased hydrolysis of Tamiflu causes behavioral side effect

False

Decreasing hyrolysis of tamiflu causes what two things to increase?

toxicity and therapy

Radiolabeling metabolites allows you to do what?

Quantify the amount of that metabolite present

If a patient takes Rifampin, what will happen to the effectiveness of a drug that is metabolized by CYP3A4?

The drug will be converted into the inactive metabolite too much, making the effectiveness go down

(T or F) Pravastatin is one of the only statins not metabolized by CYP3A4

True

What is the duration of induction

The time is takes for the enzyme to be degraded, days

What is the duration of an irreversible inhibitor

The time it takes for the enzyme to be resynthesized, days

What is the most common type of CYP2D6 phenotype

extensive metabolizer

Describe a narrow therapeutic window substrate

exposure-response curve indicates increases in exposure lead to serious safety concerns

What 2 CYPs have large binding pockets

3A4 and 2C8

Which CYP prefers planar molecules

1A2

What determines the site of CYP3A4 oxidation

chemical reactivity of each atom in the substrate

Describe a sensitive substrate

plasma AUC increases 5X or more with any known inhibitor of that CYP

Four most common oxidation rxns

Aromatic Carbon hydroxylation, benzylic carbon hydroxylation, N-dealkylation, O-dealkylation

What does cyclosporine inhibit

CYP3A4 and pgp