Pharmacokinetics Lecture PDF

Pharmacotherapy Drug Metabolism and Pharmacokinetics Part One Dr. Ahmed Fayez  Learning Objectives To comprehend the purpose of metabolism of drugs To understand phases of metabolism and key metabolising systems Subjects: Metabolism Phase 1 Pha...

Pharmacotherapy Drug Metabolism and Pharmacokinetics Part One Dr. Ahmed Fayez  Learning Objectives To comprehend the purpose of metabolism of drugs To understand phases of metabolism and key metabolising systems Subjects: Metabolism Phase 1 Phase 2 Law of Mass Action T1/2 ionisation First and zero order process Volume of distribution Plasma protein binding Bioavailability First pass effect Phase 1 and 2 metabolism Cytochrome P450 enzymes Glucuronidation Clearance  Pharmacokinetics /metabolism http://www.health.herts.ac.uk/cpd/multimedia/StudyNetPK/Starthere.html Read: Brenner and Stevens ‘Pharmacology’, Elsevier 2012 Chapter 2  Why Metabolism? Components of food and drink Substances from environment absorbed via lungs/skin Drugs (intentional) Metabolism applies equally to all xenobiotics and some endogenous compounds Most metabolic products are less pharmacologically active; important exceptions: Where the metabolite is more active (Prodrugs, e.g. Erythromycin-succinate (less irritation of GI) --> Erythromycin) Where the metabolite is toxic (acetaminophen) Where the metabolite is carcinogenic  Metabolism Close relationship between the biotransformation of drugs and normal biochemical processes occurring in the body: Metabolism of drugs involves many pathways associated with the synthesis of endogenous substrates such as steroid hormones, cholesterol and bile acids Many of the enzymes involved in drug metabolism are principally designed for the metabolism of endogenous compounds These enzymes metabolize drugs only because the drugs resemble the natural compound  Definition Biotransformation of drugs causing two fundamental changes: Reduces lipophilic nature of drugs Reduces biological activity (normally)  Metabolic Phases Divided into two/three categories: Phase I – initial modification Phase II – addition of larger group Phase III – transport processes  Sites of Metabolism Liver – main site, first pass metabolism GI tract Lungs Skin Kidneys  Phase I reactions chemically alter basic structure of a drug Oxidative and Reductive reactions alter and create new functional groups Hydrolytic reactions cleave esters and amides to unmask functional groups Metabolism: Phase II Reactions Conjugation reactions in which the drug or metabolite is coupled to an endogenous substrate such as glucuronic acid Conjugate is usually pharmacologically inactive More water soluble, so eliminated more rapidly  Glucuronic acid derived from glucose Morphine, paracetamol, salicylates Phase I Phase II Cytochrome P450 enzymes H OH O Glucuronic Hydroxylation acid  Phase 1 reactions Convert parent compound into a more polar (=hydrophilic) metabolite by adding or unmasking functional groups (-OH, - SH, -NH2, -COOH, etc.) Often these metabolites are inactive May be sufficiently polar to be excreted readily Eg Oxidation: There are two types of oxidation reactions: Oxygen is incorporated into the drug molecule (e.g. hydroxylation) Oxidation causes the loss of part of the drug molecule (e.g. oxidative deimination, dealkylation)  Paracetamol Metabolized through 3 different pathways: – 42% to 67% undergoes glucuronidation and is excreted in urine – 26% to 36% undergoes sulfation and is excreted in urine – 5% to 8% passes through the cytochrome P-450 pathway producing a potentially hepatotoxic metabolite, N-acetyl-p-benzoquinone imine (NAPQI) In normal dosing, NAPQI is conjugated with the antioxidant, glutathione, and excreted in urine  Paracetamol An example of dose related toxicity, WHY? P450 mediated phase 1 metabolism of paracetamol leads to production of NAPQI, a reactive intermediate which is hepatotoxic Cytochrome P450 (CYP450) family Essential for most drugs eliminated by hepatic metabolism. These enzymes are absorbed at 450nm in the reduced state when carbon monoxide is present; thus the “P450” designation. Based on the homology of amino acid sequences, the CYP450 enzymes have been categorized into families, subfamilies, and individual enzymes  Intracellular Sites Metabolism catalysed by cellular enzymes located in: Endoplasmic reticulum – Majority; isolated as microsomal fraction Mitochondrion Cytosol Lysosomes  Microsomal Mixed Function Oxidases (MFOs) “Microsomes” form in vitro after cell homogenization and fractionation of ER Rough microsomes are primarily associated with protein synthesis Smooth microsomes contain a class of oxidative enzymes called “Mixed Function Oxidases” or “Monooxygenases” These enzymes require a reducing agent (NADPH) and molecular oxygen (one oxygen atom appearing in the product and the other in the form of water)  MFOs Drug metabolising enzymes (MFOs – mixed function oxidases) located in smooth ER. Microsomes are the micro-vesicles formed from fragments of ER liver tissue has been homogenized and centrifuged. Activity dependent on Cytochrome P450 Cytochrome P450 reductase – electron acceptor, contains FMN and FAD NADPH – reducing agent Molecular oxygen – donates one atom to product – other becomes water Low substrate specificity – wide range compounds  Reactions by MFOs Aromatic hydroxylation Lignocaine Aliphatic hydroxylation Pentobarbitone Epoxidation Benzo[a]pyrene N-Dealkylation Diazepam O-Dealkylation Codeine S-Dealkylation 6-Methylthiopurine Oxidative deamination Amphetamine N-Oxidation 3-Methylpyridine S-Oxidation Chlorpromazine Phosphothioionate oxidation Parathion Dehalogenation Halothane Alcohol oxidation Ethanol  CYP450 Isoenzymes More than 30 subtypes identified. Cause differences in the ability of an individual to metabolise drugs. Nomenclature Root: CYP Family: CYP2 Subfamily: CYP2D Gene: CYP2D6 All CYP isoenzymes in same family have at least 40% structural similarity, those in same subfamily have at least 60% structural similarity.  Phase 2 Reactions Conjugation with endogenous substrate to further increase aqueous solubility Conjugation with glucoronide, sulfate, acetate, amino acid Phase I usually precede phase II reactions Liver is principal site of drug metabolism: – Other sites include the gut, lungs, skin and kidneys For orally administered compounds, there is – “First Pass Effect” – intestinal metabolism – Liver metabolism – Enterohepatic recycling – Gut microorganisms - glucuronidases  Phase II Reactions Although Phase I metabolites may be polar – not necessarily excreted readily Undergo subsequent Phase II conjugation to allow more rapid excretion Water soluble product excreted in bile/urine Involve conjugation, includes: – Glucuronidation – Methylation – Acetylation – Glucosidation  Glucuronidation – Phase II Largest capacity enzyme system UDP glucuronosyltransferases (UGT) Enzyme system located in ER in proximity to CYP enzyme Most drugs are changed radically from original structure – no pharmacological effect  UGT Isoforms More than 20 human UGTs Common backbone, isoforms have different N-amino termini Two main families UGT1 and UGT2 Main sites liver and GI tract UGT1A1 (hepatic) utilises wide range of drugs, large aromatic carcinogenic compounds and oestrogenic steroids  Phase II Example Glucuronidation Reactions Morphine Chloramphenicol Salicylic Acid  Paracetamol Phase 2 phase 2 conjugation with hepatic glutathione. In the event of paracetamol overdose the formation of NAPQI exhausts the glutathione stores and hepatic toxicity proceeds.  Phase I and II Reactions Absorption Metabolism Elimination Phase I Phase II Drug Conjugate Inactive metabolite Conjugate Drug Metabolite with modified Conjugate activity Drug LIPOPHILIC HYDROPHILIC  Sequence Phase II sometimes take place before Phase I reactions e.g. isoniazid: Phase Phase I II Isoniazid N-acetyl isonicotinic conjugate acid Hepatotoxic metabolite  End result of metabolism Three possibilities: Drug converted from pharmacologically active to inactive compound (majority) Conversion of drug to metabolite also pharmacologically active Conversion of pharmacologically inactive compound to active drug - PRODRUG Why make a prodrug?  Control of Metabolism Upregulation/downregulation of CYP and UGT isoforms may alter drug metabolism Both working in concordance with each other and Phase III exporter systems Adaptable responses in relation to substrates within cells Pharmacotherapy Drug Metabolism and Pharmacokinetics Part Two  Learning Objectives To comprehend the effects of changes in enzyme systems on drug metabolism To understand the clinical consequences of changes To have an appreciation of the basis of drug- drug interactions  Therapeutic window Increasing Therapeutic Increasing drug window adverse concentration effects Therapeutic Adverse response Effects  THE THERAPEUTIC WINDOW THE LIMITS BETWEEN WHICH A DRUG IS AN EFFECTIVE THERAPEUTIC AGENT Drug conc. in plasma Toxic Concn Therapeutic window Subtherapeutic Concn Time  Maintenance of Drug Control Therapeutic Window achieved when patient responds to treatment and ADRs are minimised Drug Failure – insufficient therapeutic concentrations Drug Toxicity – concentrations elevated beyond the therapeutic window  Enzyme induction: clinical significance Decreased efficacy of drugs – coadministered drug stimulates metabolism oral anticoagulants oral contraceptive pill Osteomalacia – barbiturates and antiepileptics stimulate vitamin D3 metabolism depletes stores of calcium Increased dose requirement of benzodiazepines and analgesics in smokers  Induction of Drug metabolism leads to increased cytochrome P450 synthesis barbiturates industrial chemicals carbamazepine pesticides ethyl alcohol insecticides phenytoin steroids herbicides food preservatives nicotine HME- inducers  HME- inhibitors  Phenobarbitone Cimetidine Phenytoin valproate Phenylbutazone Chloramphenicol Carbamazepine Erythromycin Rifampicin Ciprofloxacin Gresiofulvin Estrogen Testosterone Grapefruit Cortisol Drugs  Hepatic blood flow Tobacco smoking B-Blockers (Propranolol) St john's wort H2-Blockers (Cimetidine) Increase Metabolism of other drugs e.g. Oral anti-coagulants, Oral hypoglycemics & Oral contraceptives decrease their duration of action. Auto-induction → Tolerance  Case Study 63 year old male prescribed simvastatin 10mg daily. Over the course of 3 months the patient did not respond to treatment so dosage increased to 50mg daily. Patient later admitted to hospital with rhabdomyolysis.  Evidence Patient had become depressed and self administered St Johns Wort. He had stopped treatment 10 days before toxicity occurred as he felt in improved mood. What happened?  Characteristics Therapeutic levels of simvastatin only achieved by increasing dose multiple times The presence of another drug (herbal remedy) increased clearance – St Johns Wort induces P450 enzymes – Alterations in plasma levels resulted in loss of efficacy/toxicity Toxic effects appeared gradually (days) Increased drug clearance was reversible  Cause Enzyme Induction: presence of an inducer causes accelerated metabolism Inducer brings about increase in synthesis of specific enzyme isoform The patient had taken SJW, the pateint stopped responding to the statin, so the doctor increased the dosage The patient no longer taking SJW, but still at a higher statin dose, so lead to the toxic effects of rhabdomyolysis  Inducing Agents Anticonvulsants Many CYP isoforms Steroids CYP3A4 Polycyclic aromatic CYP1A1, CYP1A2 hydrocarbons Antibiotics Most CYP isoforms Recreational agents CYP1A2 (nicotine) CYP2E1 (alcohol) Herbal remedies CYP3A4 (St John’s Wort)  Mechanism of Action Inducing agents cause alterations in transcription in the nucleus Results in increased expression of CYP isoforms Consequence – increased clearance of co- administered drugs metabolised by the isoform.  Induction Overview Inducing agent combines to cytoplasmic Inducing agent detected in cell receptor forming complex Receptor-inducer complex migrates to nucleus Interaction of complex with DNA results in transcription mRNA produced mRNA released to cytoplasm Translation via mRNA produces increased levels of CYP Increased CYP levels, increases metabolic rate  Clinical Consequences Induction leads to therapeutic failure Drug levels restored to therapeutic levels by either increasing dose or Removal of inducing agent Effects on patient not serious as efficacy is easily restored  Inhibition of Drug Metabolism Drug interaction - one drug inhibits metabolism of another Two types - 1: inhibition of cytochrome P450, e.g.by binding to haem or protein moieties. 2: inhibition of other specific metabolic pathways Calcium channel blocker 100% 15% 30% 45% 90% CYP3A4 Sinusoid cells CYP3A4 In enterocytes +  Case History A 29 year old healthy male took terfenadine twice daily for allergic rhinitis. He had taken the medication for one year. One day the man took took his tablets after two glasses of grapefruit juice which he drank regularly. He went out to mow his lawn and within one hour he had collapsed and died.  Analysis Post mortem report showed raised levels of terfenadine – usually undetectable The subject did not have any evidence of history of impaired liver function Elevated levels of drug are capable of causing cardiac arrythmias Why did this happen?  Characteristics Patient stabilised on the dosage regimen, indicating that levels of drug already lie in the therapeutic window. GFJ prevented the clearance of the parent drug by interfering with metabolism The toxicity is an intensification of the drug’s usual pharmacological action indicating drug accumulation  Enzyme Inhibition Occurred within hours rather than days Toxic effects appear so rapidly that even the patient was unaware as to what happened Toxic effects may be rapidly reversed once the inhibition is prevented Can be a consequence of ‘self medication’ so physician unable to intervene  Enzyme Inhibition Occurs through four main mechanisms: Competitive Non-competitive Uncompetitive Mechanism based Competitive Inhibition  Competitive Inhibition Competing molecule bears structural similarity to substrate e.g. ketoconazole (anti-fungal agent) competitive inhibitor of CYP3A4 leading to fall in blood testosterone - leads to gynecomastia in males Non-Competitive Inhibition  Non-Competitive Inhibition Inhibitor interacts with a separate allosteric site from the active site. Structural alteration in the active site prevents substrate binding e.g. omeprazole and lansoprazole (proton pump inhibitors) are potent inhibitors of CYP3A4 Uncompetitive Inhibition Inhibitor remains at activebinds Inhibitor site to substrate  Uncompetitive Inhibition As this inhibitor binds to the E-S complex it is relatively rare Caused by some dietary components of citrous fruits e.g. tangeretin acts upon CYP3A4 Mechanism Based Inhibition Inhibitor remains at active site  Mechanism Based Inhibition Irreversibly binds to active site of enzyme ‘suicide inhibition’ as completely deactivates enzyme, causing its destruction. Takes several days for resynthesis. Many examples including GFJ  Inhibition Consequences Irreversible inhibition has serious clinical consequences: Causes elevated levels of drug leading to toxicity Worsens over time if inhibitor not stopped Causes destruction of enzyme – not readily reversible Enzyme induction and inhibition: clinical significance Narrow therapeutic Wide therapeutic window window  Factors Affecting Drug Metabolism Internal Genetic Age Gender Disease External Diet Environment  Gender Menstrual steroid metabolism causes females to handle drugs differently Phases I-III of metabolism is responsible for biotransformation of female steroid hormones Drugs which alter enzymes in Phases I-III are likely to cause more ADRs in females than males Effects of menopause/HRT unknown  Age - Neonates Infants 70 years Polypharmacy (up to 8 medications) Decline may be consequence of drug interactions more than enzyme concentration In vitro studies show similar CYP activity in young and elderly Physiological – oxygen and NADPH limited  First Pass Metabolism Liver blood flow is reduced by 40% in >60yrs Accompanied by decline in renal clearance Problematic for drugs with a high intrinsic clearance (rapid processing by liver) Propranolol – bioavailability is increased with standard doses, leading to ADRs Warfarin – less than half of standard dose required in elderly c.f. younger adults  Diet ‘You are what you eat’ Diet may have severe consequences on drug metabolism Antioxidants Plant toxins Preservatives Polyphenols Polycyclic aromatics  Polyphenols Extend the viability of fruits Toxins to cause death of animals that feed on plant Naringenin and bergottamin in grapefruit juice Polyphenols implicated in altering metabolism of range of Phase I and Phase II reaction enzymes Further work needed  Cruciferous vegetables Broccoli, brussel sprouts, cabbage, cauliflower Induce CYP1A2 and glutathione transferases (GST) in gut resulting in rapid clearance of some compounds CYP1A2 may convert aromatic amines to carcinogens Induction of GST associated with decreased cancers of gut  Barbecued meats Induce CYP1A1 and CYP1A2 Increased clearance of: Paracetamol Amitryptiline Clozaprine Fluvoxamine Naproxen Haloperidol Effect of alcohol on drug metabolism Enzyme inhibition Acute ethanol exposure Longer drug half-lives observed Increases NADH/NAD+ ratio preventing production of co-factor required for glucuronidation Enzyme induction Chronic ethanol exposure Phase I and II metabolism and increases CYP2E1 Enzyme inhibition Alcoholic cirrhosis Structural changes in hepatocytes  Cirrhosis Lliver tissue replaced by fibrous tissue, limiting viable hepatocytes Phase I metabolism appears unaffected but Phase II glucuronidation is impaired e.g. paracetamol glucuronidation is reduced This can also occur in alcoholic liver disease  Environmental Factors Tobacco smoke Motor vehicle exhaust fumes Industrial solvents Industrial pollutants Induce drug metabolism due to presence of many compounds – polycyclic aromatic hydrocarbons Pesticides – inhibition of drug metabolism Malathion and parathion commonly used insecticides induce P450 enzyme systems affecting many drugs  Conclusion Induction and inhibition of drug metabolising enzymes result in modification of drug pharmacokinetics Induction – lowering of drug levels, may lead to therapeutic failure Inhibition – raising of drug levels, often results in adverse reactions  References Rang & Dale's Pharmacology 8th Edition P.116 Section 1.9. Rang, H P, and M M. Dale. Rang & Dale's Pharmacology. Edinburgh: Churchill Livingstone, 2016 8th edition.