MEDCHEM 570. First Midterm. January 30, 2015

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1 Name MEDCHEM 570 First Midterm January 30, 2015 Instructions: Exam packet totals 7 pages. The last page has a 5 points extra credit question. If you need additional space for a question go to the back of that page and tell me you did so. Write legibly and in complete sentences when indicated. Read the questions carefully and answer the questions you know first. 1

2 1. (20 points) True False (circle the correct answer)-2 points each T F The jejunum is more acidic than the duodenum. T F Glucuronyl transferases are cytosolic enzymes. T F Bioactivation of prodrug Plavix (clopidogrel) requires CYP1A2. T F CYP2D6 IM status is important for warfarin dosing. T F Ritonavir is commonly used for PI boosting. T F Mitochondria contain P450 enzymes but they are not important in drug metabolism. T F Phase II sulfate conjugates are not charged above ph 10. T F Esterases are Phase II enzymes. T F Induction of CYP2E1 limits the toxicity of acetaminophen. T F CYP2D6 PM s have low debrisoquine metabolic ratios. 2) (20 points) Provide short definitions including one major item of relevance for drug metabolism (2-3 sentences total) of the following terms (structures/diagrams are welcome). Answer 4 of 5; cross out the one you don t want graded. 5 points each. If you didn t indicate which one you didn t want answered and you gave an answer to all five, the first four were graded. a) Microsomes Microsomes are donut-shaped vesicles made by homogenizing (lysing and shearing the ER) liver cells and centrifuging at 10k x g and 100k x g. The pellet after the second centrifugation contains the microsomes and is re-suspended. These preparations contain P450 enzymes and glucuronyl transferases, and are used to study drug metabolism in vitro. b) Carbamazepine Carbamazepine is an anti-epileptic drug which is metabolized by and induces CYP3A4. It is important in polytherapy involving other CYP3A4 substrates such as oral contraceptives as it causes increased clearance of these other drugs. Additionally, carbamazepine forms as stable arene oxide upon P450-mediated oxidation. c) Halothane Halothane is an inhaled anesthetic. It is no longer used in the US, but is still widely used in developing countries. It is bioactivated by CYP2E1 into a toxic metabolite that is believed to react with Lys residues. Halothane hepatitis is seen after repeated use and is thought to be an immune-mediated response. d) Carboxylesterases Phase I drug metabolizing enzymes that cleave ester bonds to form an alcohol and a carboxylic acid. They are particularly important in the bioactivation of pro-drugs into their active forms. Some examples of drugs that require carboxylesterase activity are Tamiflu, Fluphenazine esters and Enalapril. 2

3 e) Arene Oxide An arene oxide is an unstable, reactive, and highly electrophilic intermediate formed during P450-mediated oxidation of aromatic rings. They often undergo an NIH shift to form phenols. They can adduct DNA and cause cell damage. 3) (12 points) The Black Box Warning on the label of the anti-androgen flutamide states that serum transaminase levels should be measured before and during therapy. a) Give an example of a transaminase and explain why we measure serum transaminase levels in this case. 3 points ALT or AST ALT and AST are liver enzymes and are released into the bloodstream upon liver damage, thus elevated levels in the blood indicate liver damage. b) Provide the names of two other types of common clinical tests that are often employed and describe briefly why they are also good markers for this type of problem. 6 points BIL bilirubin is processed in the liver and then excreted in the urine. Elevated bilirubin indicates an inability of the liver to perform this function. PT/INR the liver is responsible for releasing clotting factors into the blood stream. Liver damage impairs this function and clotting factor levels go down. This results in a longer clotting time and increased INR. Either AST or ALT was also accepted (whichever one was not mentioned in part 3a) c) Pharmacists can alert patients to signs and symptoms of a developing problem with this drug. Name two. 3 points Any two of nausea, vomiting, abdominal pain, fatigue, anorexia, flu-like symptoms, hyperbilirubinuria, jaundice, upper right quadrant tenderness. 4) (15 points) The magnitude of a particular inhibitory drug-drug interaction depends upon a number of factors that relate to the metabolism and pharmacokinetics the object drug. In some cases the site of administration of the object drug is a major factor. Consider the physiology of blood circulation and answer the following. 5 points each. Scenario for a) and b): A given interactant drug increases the AUC of an iv dose of midazolam by 3-fold. a) What dose adjustment would be required to manage this interaction. What basic PK equation do you use in this analysis. You would LOWER the dose by 3-fold (to 1/3 the original dose). The equation is CL = D/AUC. Variations of that equation were accepted as well; AUC = D/CL, AUC*CL=D, etc. 3

4 b) Would the magnitude of effect on AUC for an oral dose be equal to, greater than, or less than that of the iv dose. Explain your reasoning in a couple of sentences (no equations please). The magnitude of effect on AUC for an oral dose would be GREATER than or EQUAL to the magnitude of effect on AUC for an IV dose. This is because of the first pass effect on an oral dose, specifically the inhibition of enzymes in the enterocytes, which would affect the oral dose but not the IV dose since the intestines and liver are in series for oral drugs. c) Hypothesis: For the detection of the effect of a new drug candidate on CYP1A2 activity in vivo, caffeine is a better object drug than theophylline or clozapine. Explain why this is true (ignore the pharmacological effects of these potential object drugs). An equation would be useful but not required. Caffeine is 90 % metabolized by CYP1A2 whereas theophylline and clozapine are only ~60 % metabolized by CYP1A2. This makes caffeine plasma levels more sensitive to CYP1A2 inhibition and/or induction, thus making it a better object drug than either theophylline or clozapine. 5) (9 points) What evolutionary advantages are conferred by the following facts related to the metabolism of xenobiotics (one or two sentences each). 3 points each a) A P450 enzyme usually will produce multiple metabolites of a given substrate. Producing multiple metabolites prevents any one metabolite concentration from being too high. In cases were toxic metabolites are involved, the final concentration of that metabolite would be lower in cases where other metabolites were also formed than if the toxic metabolite was the only one being made. b) Multiple P450 enzymes will oxidize a given xenobiotic. Having multiple enzymes able to clear a given drug protects against any inter-individual variability as well as inhibition of any given enzyme responsible for clearance. If an individual were deficient for an enzyme needed for the clearance of a drug due to a polymorphism, for example, or were taking another drug that inhibited that enzyme, other routes of clearance of the drug are available. c) The blood from the intestines is directed to the liver. This ensures that anything absorbed into the body through the intestines goes through first-pass metabolism. Any toxins, therefore, have a chance of being cleared before reaching systemic circulation. 6) (24 points) Atomoxetine is an amphetamine analog that is used in the treatment of ADHD (attention deficit hyperactivity disorder). Normally, in pediatric populations, the dose of this drug is determined via individualized dose escalation to desired effect using an algorithm that monitors therapeutic and toxic effects of the drug. The pharmacokinetics of atomoxetine are largely controlled by CYP2D6. Sites of metabolism by CYP2D6 are indicated by the arrows shown below (aromatic hydroxylation, N-dealkylation and O-dealkylation). 4 points each 4

5 Note that partial structures using R groups are OK in your answers. c. (trace) b. (minor) CH 3 O N CH3 Atomoxetine a. (major) a) For metabolite a. show the structure of the unstable intermediate and final metabolic product. Intermediate Product b) Metabolite a is found in urine however the dominant metabolite in urine is the product of a Phase II enzyme that produces a further metabolite and that uses the cofactor shown below. Show the structure of this urinary metabolite, name the enzyme involved and the cofactor shown. Explain why (chemical reason) this type of secondary metabolite would be expected to be found in urine. The cofactor is UDPGA and the enzyme is glucuronyl transferase or GT or UGT. It s expected to be excreted into the urine because it s polar and charged 5

6 c) For metabolite b. show the structure and name the unstable intermediate. Also show the structure of the final metabolic product. Would you expect this metabolite to accumulate in vivo? Why or why not? OH R N H R NH 2 Carbinolamine Hemi-aminal The metabolite would be expected to accumulate because it is not more or less polar than the parent (which accumulates). d) For metabolite c. show the structure of the unstable intermediate and both final metabolic products. Intermediate Products In a very large study of dosing of atomoxetine (roughly 1100 EMs and 100 PMs) prescribers dosed pediatric patients to optimal effect using standard protocols without foreknowledge of metabolizer status and the results of blood concentration measurements. Note that the final mean doses for EMs and PMs (week 10)were not significantly different. The plasma levels are shown below (left). Average AUC for the EMs and PMs were 4 ug*hr/ml and 28 ug*hr/ml respectively. asma atomoxetine exposures in CYP2D6 extensive and poor metabolizers by week on e) Based on the dose and AUC information only estimate the fraction of atomoxetine that is cleared by CYP2D6 and explain your reasoning. f!!! =!"#$!!"#$ è = 6/7 = 85% of atomoxetine is cleared by CYP2D6!"#$ 6

7 f) From the left graph it appears that the time to steady state in the PMs was longer than for the EMs. Why would you expect this to be true? From the graph, we see that PMs have higher AUCs. This would mean that the clearance of atomoxetine is lower and that the half-life of atomoxetine in PMs is higher. Drugs with a higher t 1/2 take longer to reach steady state. g) (5 points extra credit) The conclusion of the study was that foreknowledge of metabolizer status is not required in the dosing of atomoxetine. Consider this conclusion which is fully supported by the data. Does it make sense to you? Why or why not? Hint: Focus on the plasma concentration-effect vs. dose-effect relationships for this drug in your answer. Suggest one plausible reason that might explain these results. Recall from above: prescribers dosed pediatric patients to optimal effect using standard protocols without foreknowledge of metabolizer status and the results of blood concentration measurements and note that the final dose they arrived at for EMs and PMs was the same. Finally the blood levels were 8 fold higher in the PMs. Clearly they dosed to an expected final dose, not effect which confounded the study. A wide therapeutic index for atemoxetine would potentially be a factor. Either that or for some unexplained reason CYP2D6 PM s are more resistant to the effect of the drug. 7