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95 Cards in this Set
- Front
- Back
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What needs to be added to microsomes to see UGT metabolites? |
UDPGA |
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What needs to be added to microsomes to see CYP metabolites? |
NADPH regenerating system |
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(T or F) Microsomes can be used for long-term studies |
False |
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DME means what |
Drug Metabolizing Enzyme |
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Recombinant CYP's are used in what type of study? |
Reaction phenotyping |
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What is reaction phenotyping? |
finding which CYP is involved in a specific transformation |
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How many time points are used for primary metabolite screening? |
One (usually 30 min) |
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What is the biggest variable in human liver microsome (HLM) studies? |
variability in enzyme activity |
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What is used as a control in HLM studies? |
a standard of a known drug |
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(T or F) Enzyme activity in microsomes may vary from batch to batch |
True |
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DME's that aren't found in microsomes? |
N-acetyl transferase (NAT), sulfotransferase (SULT), glustathione-S-transferase (GST) |
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What needs to be added to see NAT metabolites? |
Acetyl CoA |
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What is S9? |
Supernatant obtained from centrifugation at ~9000 g that contains both microsomes and dissolved DMEs |
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S9 is most useful for? |
test for mutagenicity, finding genotoxic metabolites |
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What can hepatocytes model that S9 can't? |
Transport accross the cell membrane, changes in gene expression (induction) |
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(T or F) Hepatocytes do not require the addition of co-enzymes |
True |
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What is the preferred method for doing in vito to in vivo extrapolation? (Predicting ADME properties in humans) |
Hepatocyte studies |
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What can be introduced to a compound to reduce stability if it is lasting too long? |
"soft" groups capable of metabolic transformation |
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Fixing a metabolite stability problem often leads to problems with _____? |
Absorption |
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What is the most direct way to prevent metabolic degredation? |
Alter the molecule at or very near the site of metabolism |
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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 |
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Metabolism studies are incubated at what temp? |
37 C (body temp) |
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In metabolism studies what is the NCE dissolved in? |
HPLC grade methanol or acetonitrile |
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Microsomes are incubated in what? |
Potassium phosphate buffer (pH = 7.4) |
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What method is used for metabolite analysis after incubation? |
LC/MS/MS |
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(T or F) DMSO can inhibit CYP metabolism |
True |
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What method is used to measure cell viability and concentration before metabolism study can be performed? |
Tryptan Blue Exclusion Method |
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Terfenadine was taken off the market because ...? |
Strong CYP3A4 inhibitors caused serious side effects |
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Post-systemic metabolism occurs in? |
Kidney & Liver |
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Pre-systemic metabolism occurs in? |
Liver & GI tract |
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What conjugation reactions increase polarity? |
Sulfonation, glucuronidation, amino acid conjugation |
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What conjugation reactions do NOT increase polarity? |
Methylation and acetylation |
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What conjugation reaction protects against reactive metabolites? |
Glutathione conjugation |
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What are the 3 most important CYP's? |
3A4 , 2C9, 2D6 |
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What groups are glucuronidated? |
Phenols, Carbox. acids, amine nitrogens, alcohols |
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What groups are sulfonated? |
phenols, alcohols |
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What group is amino acid conjugated? |
Carboxylic acids |
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What group is N-acetylated? |
Amine nitrogens |
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(T or F) At time of NDA submission all clearance pathways (enzymatic and non) and transporters must be known? |
True |
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The requirement of metabolism studies ____ the percentage of IND's failing from unsufficeint PK/bioavailability? |
Decreased (From 39% to 8%) |
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Before clinical studies can be performed, what can be used to predict human metabolites? |
in vivo animal studies and in vitro human |
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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 |
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Olistat (Alli) was found to inhibit..? |
CES2 |
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(T or F) Orlistat inhibits carboxylesterases with intrinsic lipase activity being a determinant factor |
False |
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What are three sites on carboxyesterases |
Active site, Z-site and "side door" |
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Where is CES2? |
Liver, GI, Kidney |
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Where is CES1? |
Liver, small amounts in GI and Kidney |
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Where is CES3? |
Liver, small amount in GI |
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(T or F) CES1 prefers substrates with a large group on the alcohol side |
False |
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(T or F) CES2 prefers substrates with a large group on the alcohol site |
True |
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Clopidogrel is a prodrug metabolized by both CYP's and CES1. Which products the active metabolite? |
CYP's |
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Where are lipases present that CES is not? |
Pancreas |
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Oxidation of amine nitrogens is catalyzed by? |
CYPs or FMO |
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Tertiary amines are oxidized into? |
amine oxides |
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Secondary amines are oxidized into |
Secondary N-hydroxy then Nitrone |
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Primary amines are oxidized into |
Primary N-hydroxy then Nitroso then Nitro |
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A sulfide is oxidized into |
sulfoxide then sulfone |
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A sulfoxide has how many oxygens attached to the sulfur? |
1 |
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A nitroso is what typeof bond? |
Nitrogen double bonded to oxygen |
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A Nitro is what type of bond? |
Positive nitrogen single bonded to a negative oxygen |
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A sulfone has how many oxygens double bonded to the sulfur |
2 |
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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 |
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What enzymes catalyzes steroid aromatization |
CYP19 |
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What happens in steroid aromatization? |
The double bonded oxygen and a methyl group are reomved to create an aromatic ring |
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What happens in 1,4 dihydropyridine oxygenation? |
A hydrogen is removed from a nitrogen |
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What types of groups are hydrolyzed? |
ester, amide, epoxide, carbamate, carbonate, urea |
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What are the two products of amine hydrolysis |
amine and carboxylic acid |
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What is the product of epoxide hydrolyzation |
a trans di-ol |
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What are the three major types of enzymes that catalyze hydrolysis |
esterases, peptidases, epoxide hyrolases |
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What are the three types of esterases |
carboxyesterases, cholinesterases, organophosphatases |
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(T or F) Enzymes allow simultaneous acid and base catalyzed hydrolysis |
True |
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(T or F) histidine acts as a base |
True |
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(T or F) Glutamate acts as a base |
False |
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(T or F) Serine acts as a nucleophile |
True |
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List the 3 steps of esterase catalysis |
1. enzyme-substrate complex 2. Acetylated enzyme intermediate 3. Nucleophilic attack by water |
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Which is hydrolyzed faster, ester or amide? |
Ester |
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Which is hydrolyzed slower, procaine or procainamide? |
ProcainAMIDE |
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Clopidogrel is a prodrug. Its active metabolite binds to what? |
P2Y12 receptor on blood platelets |
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(T or F) Clopidogrel is hydrolyzed by CES1 to form an inactive metabolite |
True |
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(T or F) Clopidogrel competes with Tamiflu for CES1 hydroysis |
True |
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(T or F) Increased hydrolysis of Tamiflu causes behavioral side effect |
False |
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Decreasing hyrolysis of tamiflu causes what two things to increase? |
toxicity and therapy |
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Radiolabeling metabolites allows you to do what? |
Quantify the amount of that metabolite present |
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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 |
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(T or F) Pravastatin is one of the only statins not metabolized by CYP3A4 |
True |
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What is the duration of induction |
The time is takes for the enzyme to be degraded, days |
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What is the duration of an irreversible inhibitor |
The time it takes for the enzyme to be resynthesized, days |
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What is the most common type of CYP2D6 phenotype |
extensive metabolizer |
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Describe a narrow therapeutic window substrate |
exposure-response curve indicates increases in exposure lead to serious safety concerns |
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What 2 CYPs have large binding pockets |
3A4 and 2C8 |
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Which CYP prefers planar molecules |
1A2 |
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What determines the site of CYP3A4 oxidation |
chemical reactivity of each atom in the substrate |
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Describe a sensitive substrate |
plasma AUC increases 5X or more with any known inhibitor of that CYP |
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Four most common oxidation rxns |
Aromatic Carbon hydroxylation, benzylic carbon hydroxylation, N-dealkylation, O-dealkylation |
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What does cyclosporine inhibit |
CYP3A4 and pgp |
