Introduction to pharmacology and drug metabolism
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Introduction to pharmacology and drug metabolism Introduction to pharmacology and drug metabolism Presentation Transcript

  • Introduction to Pharmacology and Drug Metabolism Luke Lightning, PhD
  • Outline of topics to be discussed: Introduction Quantitative aspects of drug-receptor interactions Fundamental mechanisms of drug action Drug dose and clinical response Factors modifying effects of drugs ADMEText: B.G. Katzung, Basic & Clinical Pharmacology, chapters 1 & 2
  • Introduction Pharmacology: study of interactions between chemical compounds and biological systems. i.e. - how drugs work - where drugs act - how the body processes drugs, etc. (mechanisms of drug action) The receptor is the cornerstone of pharmacology Explains how the organism interacts with a drug and initiates a chain of biochemical events that results in observed effects An agonist is a drug whose interaction with the receptor stimulates a biological response
  • Purpose of Drug Therapy To produce the characteristic effect(s) of the drug being used. The drug must achieve adequate concentrations at its site(s) of action. To achieve the maximal positive effect of the drug while minimizing undesired effects. No drug will have only one effect (i.e. adverse effects)!
  • Magnitude of Response Following Drug Therapy Dependent on various factors: – amount of drug administered (dose) – concentration at site of action » dependent on rate of absorption and blood flow to the site – amount of time the drug remains at the site of action » dependent on biotransformation (metabolism) and elimination Appropriate dose of a drug: – amount of drug needed at a given time that results in the appropriate concentration at the site of action (where biological effect occurs)
  • Effect of Drugs on Organs and Tissues Drugs only modify cellular function – do not create effects DRUG RECEPTOR RESPONSE – Pharmacodynamics: Drug  Biological Effects – drugs alter the normal biochemical functions of an organ, tissue, or cell e.g. laxatives increase the activity of the GI tract (i.e. stimulation) general anesthetics decrease activity of cells in the CNS (i.e. depression)
  • Drugs, Dose, Receptor, and Response Drugs Dose Target (Receptor/Enzyme) Response Lipitor 10-80 mg HMG-CoA Reductase Decreases LDLSingulair 10 mg Leukotriene Receptors Prevents BronchochonstrictionLexapro 5-20 mg Serotonin Receptors Relieves Anxiety Nexium 20-40 mg Proton Pump Decreases Gastric Secretion Plavix 75 mg Purinergic Receptors Anticoagulation
  • Drug-Receptor Interactions Receptors largely determine the quantitative relationship between dose or concentration of drug and their pharmacological effects. Receptors are responsible for selectivity of drug action – binding to the receptor is dependent on the 3-D characteristics of the drug – size, shape (e.g. stereochemistry), and electrical charge of a drug molecule – changes in the chemical structure of a drug can affect receptor binding – different types of bonds can be formed between drug and receptor (e.g. H-bond) » explore these 2 aspects in more detail in Dr. Dave’s section of MCMP 407
  • Drug-Receptor Interactions (cont.) Receptors mediate the actions of pharmacologic agonists and antagonists – Agonists: drugs that bind to a receptor and stimulate a biological response – Antagonists: » drugs that bind to a receptor but do NOT alter receptor function (i.e. stimulating a response) » alter the interaction of the receptor with another drug » effect depends completely upon its ability to prevent binding of an agonist to its receptor and blocking their biological activity » possess affinity, but lack intrinsic activity
  • Drug-Receptor Interactions LSD is an agonist at the 2-Bromo-LSD is an 5-HT2A receptor antagonist LSD LSD Br CNS effects
  • Effect of Drugs on Organs and Tissues (cont.) site of drug action: where the drug acts to initiate the chain of events leading to a biological effect – extracellular sites: » some drugs do not need to enter the cell to exert their effects » intracellular reactions (i.e. signaling pathways) are responsible » more on these biochemical pathways later – intracellular sites: » usually involve a lipid-soluble drug that is able to cross membranes – sites on the cell surface: » usually involve transmembrane receptors
  • Concentration-Effect Curves and Receptor Binding of Agonists Responses to low concentrations of a drug increase proportionally As the dose increases, the incremental response decreases Finally, concentrations may be reached at which no further increase in response can be achieved with increasing concentration akin to Michaelis-Menten kinetics (principles of Km, Vmax)
  • Concentration-Effect Relationship 100% EC50 = concentration of drug required 75% to produce half-maximal effect Drug Effect 50% At lower concentrations: drug effect is changing rapidly 25% EC50 0% At higher concentrations: 0 200 400 600 800 1000 drug effect is changing slowly Drug Concentration (µM)- difficult to accurately extrapolate quantitative information due to the constantly changing slope of the curve log plot- difficult to compare multiple curves at the low concentrations
  • Concentration-Effect Relationship (cont.)Relatively linear portion in the curve about its central point  more accurate quantitation 100% expansion of scale at lower concentrations 75%Drug compression of scale at higher concentrationsEffect 50% 25% EC50 0% 1 10 100 1000 easier to compare concentration-effect Drug Concentration (µM) (dose-response) curves graphically there is no biological significance to this change in graphical presentation
  • Pharmacological Descriptors of the Receptor KD: – describes the interaction between the drug and receptor – drug concentration where drug binding to the receptor is half-maximal – constant for a given drug-receptor system – The lower the KD, the stronger the interaction Bmax: – total amount of receptor present in a cell or tissue
  • Homer Simpson and KD + beer low KD high affinity very high KD + champagne very low affinity
  • Receptor Binding and Drug Concentrationarithmetic scale the drug-receptor log scale the drug-receptor binding curve is hyperbolic binding curve is sigmoidal Ratio occupied receptor 1.0 1.0 Ratio occupied receptor 0.9 0.9 0.8 0.8 0.7 0.7 50 % occupancy 0.6 0.6 when [Drug] = KD 0.5 50 % occupancy 0.5 0.4 when [Drug] = KD 0.4 0.3 0.3 0.2 0.2 0.1 0.1 0.0 0.0 0 100 200 300 0.01 0.1 1 10 100 1000 Drug concentration (mM) Drug concentration (mM) KD is constant for a drug-receptor system
  • Concept of Affinity affinity: ability of the drug to interact with the receptor KD is a measure of affinity affinity is a determinant of potency – lower KD  higher affinity  more potent a single drug: different affinities for different receptors relative affinities among drugs may change from receptor to receptor
  • Concept of Potency potency: dose of a drug required to produce a particular effect of given intensity compare drug doses that produce the SAME effect (usually at ED50) more potent if less drug is required (higher affinity) higher KD or EC50  less potent potency may be over-rated – imperfect: our world of D + R  DR  response instead determine efficacy
  • Concept of Efficacy efficacy: the biological response resulting from the drug-receptor interaction – not all DR  same amount of response a strong agonist has high affinity and high efficacy maximal efficacy is often limited by toxicity – high doses efficacy is more important than potency as a drug property log dose-response curves good for visual inspection Foye’s: page 90
  • Homer and Agonists Agonists
  • Partial AgonistRemember LMA: conformational change in R  response k1 [D] + [R] [DR] Effect k-1 what about this step? full agonist  full occupancy  maximal effect some agonists  full occupancy  less than maximal effect effects of these agonists are less efficiently coupled to receptor occupancy = “partial agonists”
  • Partial Agonist full agonist A A g o n is t E f f e c t 1 A 0.8 partial agonist B B 0.6 0.6 Drug C partial agonist C 0.4 0.4 Effect 0.2 0 -9 -8 -7 -6 -5 -4 -3 -2 -1 0 1 Log Agonist Concentration log [Drug] does NOT  same maximal effect as a full agonist regardless of the concentration used
  • Partial Agonist (cont.)  reduced response even at 100% receptor occupancy may competitively inhibit the response to a full agonist can have the same affinity for the receptor as full agonists – decreased affinity is not the reason for a less than maximal response mechanisms complex but probably related to drug binding to inactive form of receptor – receptor can take on two forms (active and inactive) – partial agonist can bind to both forms
  • Example of Concepts  Potency A C B  EfficacyResponse  Agonist  Partial Agonist log (Dose)
  • Receptor Antagonism a D-R interaction that inhibits the drug response produced by an agonist binds to the receptor, but does NOT activate it 4 major types of receptor antagonists: – competitive: almost all antagonists in clinical use are of this type – irreversible: these  covalent modifications of the receptor – mixed: we won’t discuss – noncompetitive: we won’t discuss exhibit very different concentration-effect and concentration-binding curves
  • Competitive Antagonist Reversible or equilibrium competitive antagonism: – antagonist combines with the same binding site on the receptor as the agonist – can be reversed by increasing the dose of the agonist – e.g. heroin overdose is treated with competitive antagonist naloxone
  • Competitive Antagonist (cont.) 100% A E ffe c t A : agonist alone B 75% B: (+) competitive antagonist CDrug C: (+) more comp. antagonist D ru g 50%Effect In presence of comp. antag.: 25% Higher [ agonist ] required to: 0% - overcome inhibition 0 100 200 300 400 500 Drug Concentration - produce effect [Drug]
  • Competitive Antagonist (cont.) 100% E ffe c t - increase [ antagonist ]  75% increase EC50 of the agonistDrug D ru g 50% IncreasingEffect [ antagonist ] - magnitude of the shift is proportional to [antagonist ] 25% 0% -6 -5 -4 -3 -2 - potency decreases Log Drug Concentration - efficacy is unchanged log [Drug] EC50
  • Log Dose-Response Curve in the Presence of aCompetitive Antagonist the shape of the log dose-response curve and the maximal response are not altered by the competitive antagonist at very high [antagonist], raising the [agonist] should still  response a competitive antagonist has affinity, but lacks significant intrinsic activity (efficacy)
  • Irreversible Antagonist an irreversible antagonist will usually bind to the same site as the agonist, but will not be readily displaced irreversible inhibition is generally caused by a covalent reaction between antagonist and receptor inhibition persists even after an irreversible antagonist is removed!
  • Irreversible Antagonist (cont.) 100% curve is shifted to the right E ffe c t increasing 75% at high [ irrev. antag. ]: [ antagonist ]Drug - max effect decreases D ru g 50%Effect - covalent bond is formed 25% higher [ agonist ] does not: 0% - overcome inhibition -6 -5 -4 -3 -2 log [Drug] Log Drug Concentration - produce max. effect
  • Time-Action CurveAddresses two main questions for every drug: 1How quickly will the drug act?How long will the drug effect last? 0.8 D ru g E ffe c t 0.6 0.4 0.2 Minimum Effective 0 0 1 2 3 4 5 6 7 8 9 10 Concentration Time (hr) Time to onset Duration of action Time to Peak Effect
  • Residual Effects after the primary effects are terminated, it is possible for a drug to exert a residual effect that is unmasked when another dose of the same drug is given – e.g. impaired psychomotor skills following anesthesia may not be due to the binding at the receptor responsible for the primary effects can only be observed if another dose or a dose of another drug is given – e.g. cognitive decline (sleep disorders, impaired memory, etc.) with chronic MDMA use Can last for long periods of time (months, years) may also occur when another entirely different drug is given and the phenomenon of antagonism or potentiation is manifested – e.g. 2nd drug bind to receptor responsible for primary effects  1st drug released
  • Residual Effects (cont.) Marijuana Use 1 0.8 1 drug effects D ru g E ffe c t 0.6 terminated 0.4 residual effect 0.2 0 0 1 2 3 4 5 6 7 8 9 10 Time (hr) impaired neuropsychology (attention, memory, etc.) women > men
  • Pharmacokinetics (PK) SectionBODILY PROCESSES DRUG Drug Absorption and Transport Pharmacodynamics: Drug  Biological Effects Text: Katzung, Basic & Clinical Pharmacology, chapters 3-4 Foye’s, Principles of Medicinal Chemistry, chapters 7-8
  • PharmacokineticsAbsorption Distribution Biological Effect Drug PharmacodynamicsMetabolism Elimination
  • Pharmacokinetics and Pharmacodynamics ADME: - Absorption - Distribution - Metabolism - Elimination Katzung: page 36
  • PK Curve May Not Correlate with PD Curve  Problem: – PK ≠ PD * » average: 6-8 hr activity, 22 hr t1/2 »  individualized dosing is required – Prescriptions are increasing – contributed to 3,849 deaths in 2004 (790 in 1999) » 82% of those deaths listed as accidental methadone
  • Definitions once thought that the biological response to a drug was due to its pharmacologic activity – it is now apparent that this is NOT the case Absorption: movement of a drug FROM the site of administration  the circulation Distribution: movement of drug FROM circulation  tissues (e.g. plasma  receptor) Metabolism: biotransformation of drugs into metabolites Elimination: removal of unchanged drug and metabolites from the body
  • Introduction in order for a drug  biological activity, it MUST be present at its target site in the body ADME processes occur simultaneously and determine the time course of [drug] at its target in combination with the affinity of the drug for its target site: – ADME processes serve to regulate the pharmacological activity of a drug ADME processes play an important role in the overall drug effect: – drugs are rarely administered directly to the site of action (e.g. topical administration) an understanding of cell membrane properties and structure is required Foye’s: page 145
  • Transport of Drugs: drug transport = movement of a drug molecule across a series of membranes and spaces most often: drug is given into one body compartment and must move to its site of action in another – requires that the drug be absorbed into the blood and distributed to its site of action drug action (time of onset and duration) depends on ALL of the rates of ADME processes elimination can occur by metabolism and/or directly excreted – should occur at a reasonable rate so length of drug effect is appropriate for therapy the rate of uptake/release by a tissue is a function of: – blood flow to that tissue – affinity (partition coefficient) of tissue for drug rates of absorption can depend upon the rate of blood perfusion at the site of absorption
  • Drug Absorption:Routes of Administration
  • Drug Absorption for most routes of administration, drugs must cross epithelial membranes in order to reach the blood – e.g. GI, oral – but NOT injection (sc, im, or iv) therefore, (except for injection) drugs must go through the cells in the membrane – cannot go between cells by bulk flow drug absorption is usually limited by: – the rate the drug can cross cell membranes by drug transport mechanisms: (diffusion, filtration, ion-pairing, endocytosis, facilitated transport, or active transport) – perfusion (i.e. circulation at the site of absorption) and concentration gradient – surface area
  • Routes of Administration choice will have a profound effect upon the rate and efficiency with which the drug acts – enteral = drug placed directly in the GI tract (epithelial barriers – stomach) » oral – swallowing » rectal – absorption through the rectum » sublingual – placed under the tongue – parenteral - BYPASS GI system (endothelial barriers) » injection - sc, im, iv – topical - (epithelial barriers - skin) – inhalation - (epithelial barriers - lung) remember: no single method of drug administration is ideal for all drugs in all situations
  • Bulk Flow (cont.) Absorption Distribution Environment Plasma + + - GI +ORAL Skin - + Lung o - o o - SC, IM o epithelium capillary endothelium (tight junctions) (loose junctions)
  • Enteral Absorption formulation: controls the ability of the active ingredients to dissolve and go into solution – essential 1st step for absorption – especially important at gastric pH (very low) – achieve delayed release into small intestine with pH sensitive coatings – avoid stomach microbial metabolism: – proteolytic and hydrolytic enzymes of intestinal microflora may metabolize drugs  – altered rate of absorption OR – altered biological activity (metabolites)
  • Enteral Absorption (cont.) FOOD (generally decreases absorption) – delays gastric emptying – increases hydrolysis by gastric enzymes – increases intestinal blood flow and subsequent absorption – complexes with drugs to retard absorption » e.g. tetracycline: complexes with Ca2+ in food and milk products  Effect is considerable  can reduce absorption of tetracyclines by 80%  Solution: leave a 2 hour gap between eating and taking tetracycline
  • Routes of Administration: Oral Advantages: – convenient: can be self-administered, pain-free, easy to take – absorption: takes place along the entire GI tract – cheap: compared to parenteral routes Disadvantages: – sometimes inefficient: only part of the drug may be absorbed – 1st pass effect: drugs absorbed orally are initially transported to the liver via the portal vein – irritation to gastric mucosa  nausea and vomiting – destruction of drugs by gastric acid and digestive juices – effect too slow for emergencies – unpleasant taste of some drugs – unable to use in an unconscious patient (patient compliance is a problem)
  • 1st Pass Effect drug is absorbed from the gut and delivered to the liver by the portal circulation enzymes in the liver metabolize the drug to an inactive species before it reaches the systemic circulation – inactive product = metabolite that does not possess the desired pharmacological activity the greater the 1st pass effect: – the less the drug will reach the systemic circulation when administered orally
  • Routes of Administration: Sublingual barrier is oral mucosa (epithelial cells) surface area is limited (< 1 m2), but well perfused cell layer is relatively thin absorption is rapid if lipid/water partition coefficient is high pKa is the major rate limiting factor - saliva pH is 7.0 absorption direct to general circulation - thus bypasses 1st pass metabolism limiting factors: dissolution and transit time in oral cavity – some drugs are taken as smaller tablets which are held in the mouth or under the tongue » advantages: rapid absorption, drug stability, avoid 1st pass effect » disadvantages: incovenient, small doses, unpleasant taste of some drugs
  • GI Absorption size of the absorptive surface of the various parts of the GI tract (in m2): – oral cavity: 0.02 – stomach: 0.1-0.2 – small intestine 100 – large intestine 0.5-l .0 – rectum 0.04-0.07
  • pH in Body Compartments Blood 7 pH 1-3 Mouth 6-7 Colon 8 Cerebral spinal fluid 7 Urine 5-8 5-7 Sweat 4-7 6-7note: stomach pH is variable 7-8 SI and LI pH is near neutral Foye’s: page 144
  • Other Routes of Administration: Advantages Rectal: – Bypasses: » low pH of GI, hydrolytic enzymes in GI, first-pass metabolism » good for drugs affecting the bowel (laxatives) – useful for unconscious or vomiting patients or uncooperative patients (children) Topical: – generally produces only local effects e.g. dermatology: antibacterial, antifungal, sunscreens, antiviral agents Lung: – very highly vascularized and absorption RATE in the lungs is considerably higher than that in the small intestine
  • Parenteral Administration barrier is endothelial cells can bypass epithelial barriers via injection subcutaneous (sc): bypass epidermis - only barrier is dermis intramuscular (im): bypass epidermis and dermis – injected into skeletal muscle – faster absorption than s.c. due to better perfusion and lateral diffusion transdermal: diffusion through intact skin intravenous (iv): bypass ALL barriers (membranes) to absorption – drug injected directly into the blood stream – produces essentially immediate response
  • Advantages of Intravenous Administration absorption phase is bypassed (drug is 100% bioavailable) almost immediate onset of action obtain precise plasma levels; excellent compliance; fairly pain free large quantities can be given good for drugs with narrow therapeutic index (accurate route of administration) useful for rapidly metabolized or labile drugs – bypass 1st pass and absorption phase especially good for drugs which are poorly absorbed by other mechanisms especially good for very large drug molecules (macromolecules that can’t cross membranes)
  • Disadvantages of Intravenous Administration very rapid response  potential for overdose (OOPS! factor is high) non-recoverable – can’t “suck out the poison” requires skilled administration (costly) potential for tissue necrosis potential for embolism – drug or particulate in formulation blocks the flow of blood potential for microbial or viral contamination in preparation
  • IV vs Oral Administration Bioavailability (F) Calculation: – Amount of drug available after oral administration compared to: – Amount of drug available after IV administration (F = 100%) – Tells you: » amount of first pass metabolism » if there were absorption problems  new formulation? » etc.
  • Time-Action Curve (PK) 1 Ideal Situation: Drug Plasma fLevelsPD and PK Time-Action 0.8 D ru g E fe c t CmaxCurves are Correlated 0.6 AUC T1/2 0.4 0.2 0 0 1 2 3 4 5 6 7 8 9 10 Time (hr) Tmax
  • General Scheme of Drug MetabolismLipophilic Hydrophilic Metabolism increase elimination decrease biological activityParent compound Phase I Phase II Metabolites (synthetic) Conjugated (oxidative) Metabolites polarity functionality ionization water solubility
  • Human P450 Isoforms  major drug metabolizing P450s  % of drugs metabolized by P450s Foye’s pages 178-179
  • Clinical Considerations of CYP450 Metabolism Loss of Drug Effect No Toxicities Substrate Oxidation DrugCYP450 CYP450 + Metabolite EliminationCYP450 + Drug + electrons  Activated CYP450  CYP450 + Metabolite (capable of oxidations)
  • NADPH2 P450Oxidations bound molecular oxygen cytoplasmic substrate side endoplasmic reticulum P450 (membrane) luminal side
  • Aromatic Oxidation [O] bioactivation inactivation vs. bioactivation cellular toxicities
  • MDMA and Cytochrome P450 MetabolismMDMA (“Ecstasy”) MINOR H O N CH3 MAJOR CH3 O P450 2D6 P450 1A2 H H N O H HO N CH3 CH3 O CH3 HO
  • CYP450s ISOZYME SUBSTRATES INDUCERS INHIBITORS CYP1A2 Acetaminophen Barbecue Antibiotics (2%) Theophylline Smoking Quinolone CYP2C fam Diazepam Rifampin Fluoxetine (20%) Phenytoin CYP2D6 Codeine None known Quinidine (25%) Imipramine Antidepressants CYP3A4 Quinidine Phenobarbital Antifungals (52%) Warfarin Phenytoin Antibiotics approximate % of drugs metabolized by this CYP450
  • P450-catalyzed reactions: Epoxidation - ring (aromatic)Benzo[a]pyrene – polycyclic aromatic hydrocarbon present in cigarette smoke, smog, charcoal grilled meat P4501A Epoxidation O  known carcinogen in fish, insects, humans, and other animals  epoxide reacts w/ DNA and macromolecules  LC50: cricket = 15mg/g (oral)
  • Clinical Considerations of Cytochrome P450 Inhibition Prolonged or Enhanced Effect Competitive Inhibition Undesirable Toxicities (Drug-Drug Interaction) Drug A Drug B (Inhibitor) (Substrate) P450 Inhibited P450 Drug B slow release of inhibitor Drug-Drug Interaction (DDI)
  • Time-Action Curve – Competitive Inhibitor 1 + inhibitor Drug Plasma Levels 0.8PK and PD D ru g E ffe c tare affected or 0.6 0.4 0.2 0 0 1 2 3 4 5 6 7 8 9 10 Time (hr)
  •  Why are we so interested in DDIs?? FDA: 2006
  • FDA Draft Guidance – Metabolism and DDIs September 2006 – Study design, data analysis methods – Implications for dosing and labeling – Mostly concerned with effects on CYP450 DDIs can be due to metabolism but also: – Changes in PK, transporters, etc. Does not establish legally enforceable responsibilities Describe the FDA’s current thinking View only as recommendations, not required – May be best to be running experiments described to stay ahead of or with the rest of the pack – “Negative findings from early in vitro and early clinical studies can eliminate the need for later clinical investigations.” – i.e. potentially fewer protocols!!
  • Adverse Events Reported to FDA FDA has a website devoted to ADRs: This figure illustrates the patient outcome(s) for reports in AERS since the year 1999 until the end of 2008. Serious outcomes include death, hospitalization, life-threatening, disability, congenital anomaly and/or other serious outcome.
  • Factors Modulating Xenobiotic Metabolism (cont.)DRUG INTERACTIONS (DI’s): competitive inhibition by other drugs and xenobiotics can decrease metabolism of drugs especially important with multiple drug treatments 1 drug 7% 2 drugs 12% potential DI’s with: 4 or more 3 drugs drugs 13% – herbal drugs and illegal drugs relatively unexplored 68% very important with elderly patients who are often taking multiple drugs simultaneously approx. 1000 patients at VA Medical Center, Wichita, KS
  • Steps of the Experiment Combined with tissues of interest and other reaction ingredients Mixture undergoes vigorous shaking for a period of timeTest Articles
  • Purification and Analysis Centrifuged to precipitate protein Injected onto the LC/MS for analysis
  • Data Analysis and Next Steps Go home and let the Process the data LC/MS work overnight disseminate to the Project Team I think we No More should perform Bailouts this experiment or next! DDIs!
  • Competitive Inhibition of Cytochrome P450s (B) coordination to the heme iron (A) lipophilic and H- atom - usually through a nitrogen bonding interactions (esp. imidazole ring) Inhibitor A Inhibitor B N N N Fe N N Fe N N N P450 P450
  • MDMA and Cytochrome P450 Inhibition H O N MDMA CH3 CH3 O Contaminants commonly found: • MDMA structural derivatives: legal, cheaper • caffeine and ephedrine (“herbal ecstasy”): mimic speedy feeling • LSD (very rare) • dextromethorphan (“green triangles”) anti-tussive (cough medicines) raises body temp inhibits sweating
  • Drug-Drug Interaction CH 3 CH3 MDMA N Dextromethorphan N H CH 3 O O CH3 O P450 2D6-Dextromethorphan P450 2D6 cheaper plasma levels of MDMA drugs
  • Drug-Drug Interactions • H2 receptor antagonist (anti-ulcer agent) • general inhibitor of human P450s Cimetidine (Tagamet) • inhibits hepatic elimination of many drugs: H CN warfarin alprazolam N N acenocoumarol triazolam S phenadion theophyllineMeHN N N phenytoin imipramine H carbamazepine caffeine N chlormethiazole propanolol Fe N diazepam labetalol N N chlordiazepoxide metoprolol lidocaine ethanol • imidazole ring able to coordinate to the heme iron atom of several different P450s undesirable toxicities
  • Drug-Drug InteractionsRanitidine (Zantac) HO - N O N Me2 S • H2 receptor antagonistM eHN N O H • replacement of imidazole w/ furan ring: circumvents cimetidine drug interactions Cimetidine (Tagamet) • knowledge of which structural features of a H drug were important for P450 inhibition CN N N SMeHN N N design of a safer drug H
  • Mechanism-Based Inhibition (Irreversible) FDA Draft Guidance Metabolic activity will not be restored until enzyme is re-synthesized
  • Pathways of Mechanism-Based Inhibition of CYP450 MBI* N Fe N N N Fe MBI MBI* N N N N Fe N NN Fe N N N N N Cys
  • Mechanism-based Inactivators of CYP450sRaloxifene (osteoporosis) Phencyclidine (street drug) RU-486 (morning after)Bergamottin (Grapefruit JuiceComponent)
  • Ritonavir “BOOSTER” for + ritonavir other HIV drugs Mechanism-basedinactivator of CYP3A4
  • Experimental Design: Mechanism-based Inactivation ≥ 20-fold dilution AND excess substrate to displace MBI + NADPH (now <<< KD) time time product analysis (0-10 min) (e.g. 7-OH coumarin) • HPLC/fluorescence • LC/MS • GC/MS 1˚ rxn 2˚ rxn• human liver microsomes • CYP450 selective substrate• MBI (e.g. 8-MOP for CYP2A6) (e.g. coumarin at 2X KD)• initiate rxn • initiate rxn with P450 from 1˚ reaction
  • Enzyme-Drug Interaction - Concepts E I [E + I] E time E E [E + I] [E-I] E KI [E + I] S kinact time [E-I] 20X Metabolites [E + S] I [E + S] dilution KD
  • RU486 and CYP2B6 (2008) 0-25 µMcompetitive inhibition 31% remaining kinact KI
  • Esterases > 70 different human esterase genes – Esterases are present in every tissue and blood a/b hydrolase-fold family (>15,000 members) – Carboxylesterases (hCE-1, 2, 3) – broad substrate specificities – Acetylcholinesterase (AChE) – specific for acetylcholine – Butyrylcholinesterase (BChE) – broad substrate specificity Others: – Proteases (Chymotrypsin, Trypsin, etc.) – Albumin – Paraoxonases (hPON-1, 2, 3) – broad substrate specificities
  • Famous Esters heroin aspirin Esther Rolle polyester “Good Times!!”
  • General Esterase Activity esterase H2O ester acid alcohol +
  • Human Carboxylesterases Enzymes known to be involved in drug metabolism – Human carboxylesterases-1 and -2 (hCE-1 and hCE-2) hCE-1 microsomes liver cytosol Two purified hCE-2 enzymes intestine
  • Inhibitors of Esterases: Biological Weapons Sarin Tabun VX AChE inhibitor – developed as a pesticide (1952) most deadly nerve agent in existence 3X more deadly than sarin 300 mg is fatal "Its one of those things we wish we could disinvent." - Stanley Goodspeed, on VX nerve agent
  • Factors Modulating Xenobiotic MetabolismAge and Ontogeny: decreased: – absorption (decreased absorptive surfaces, blood flow, and GI motility) – tissue perfusion – general metabolism and liver function – P450 levels in very young and very old – different P450 are expressed altered drug distribution: – increased % body fat – decreased: serum albumin (plasma protein), muscle mass, total body water
  • Factors Modulating Xenobiotic Metabolism (cont.)PHARMACOGENETICS: sex differences (generally small in humans) ethnic differences (P450) – isoniazid - slow vs. fast acetylators species differences (P450) – MAJOR problem: drug testing in animals and extrapolation to humans individual genetic variability (relative amounts of P450s and Phase II enzymes) organ-specific differences (P450, bioactivation) individualized drug therapy is the goal
  • elimination DRUG Phase II Reactions Glucuronosyl Transferases P450 Sulfotransferases Phase I FMO Glutathione Transferases eliminationReactions ADH esterases Amino Acid Transferases Acetyltransferases amidases Methyltransferases Metabolite elimination
  • Drug Elimination Pharmacological activity of drug can be reduced by: – metabolism – plasma protein binding – redistribution to other compartments (i.e. fat) Elimination: – required to remove the chemical from the body and terminate biological activity » especially if drug is minimally metabolized – necessary to prevent accumulation of xenobiotics in the body
  • Major Routes of Drug Elimination: are highly dependent on metabolism: – KIDNEYS (renal) » represent approx. 1% of of total body weight, » but receive 25% of cardiac output » blood flow rate is approx. 8X more that exercising muscle – Liver – Intestines – Lungs – Sweat, Saliva, Milk – not really significant same physiological mechanisms govern drug elimination as absorption – i.e. cell membranes are the barriers.
  • Methadone  Problems: – PK ≠ PD (average: 6-8 hr activity, 22 hr t1/2) » F = 36-100%, t1/2 = 5-130 hr * »  individualized dosing is required – Lots of interindividual variability – Long t1/2 and high tissue distribution – DDIs – Prescriptions are increasing
  • Methadone Metabolism CYP2B6: S > R CYP3A4: S = R CYP2C19: R >> S Several DDIs possible Methadone EDDP Cmax ~ 0.6 µM (inactive, renally excreted)
  • The End