Drug Metabolism and Cytochrome P450

Pharmacotherapy Drug Metabolism and Pharmacokinetics

• Topics of Study: Drug metabolism and pharmacokinetics
• Learning Objectives: Comprehend the purpose of drug metabolism, phases of metabolism and key metabolising systems.
• Subjects Discussed: Metabolism, Phase 1 Metabolism, Phase 2 Metabolism

Phases of Metabolism

• Phase I: Initial modification of drugs
• Phase II: Addition of larger groups to metabolites
• Phase III: Transport processes

Processes of Drug Metabolism

• Absorption: First and zero order processes, Ionisation , Volume of distribution, Plasma protein binding, Bioavailability, First pass effect
• Distribution: Movement of drug throughout the body.
• Metabolism: Chemical alteration of drugs to convert them into active or inactive forms, Liver is primary organ of metabolism, Cytochrome P450 (CYP450) enzymes, Glucuronidation, Clearance
• Excretion: Removal of drugs from the body, mainly through urine and bile.

Why Metabolism?

• Metabolism is necessary for food components, environmental substances, and drugs.
• Most metabolites are less pharmacologically active, but some are more active, toxic or carcinogenic.

Mechanisms of Drug Metabolism

• Drugs undergo various types of metabolism
• Important examples include oxidation and reduction, hydrolysis
• Different sites include liver, GI tract, lungs, and kidneys
• Metabolism is a fundamental aspect for many processes in the body

Metabolic Phases

• Phase I reactions: Chemically alter basic drug structure by oxidative and reductive reactions, hydrolytic reactions, cleave esters and amides to unmask functional groups
• Phase II reactions: Couplings in which the drug or metabolite is coupled to an endogenous substrate such as glucuronic acid, conjugate is usually pharmacologically inactive, and is more water soluble so eliminated more rapidly

Sites of Metabolism

• Liver: Main site, first-pass metabolism
• GI Tract: involved in drug metabolism
• Lungs: involved in drug metabolism
• Skin: involved in drug metabolism.
• Kidneys: involved in drug metabolism

Types of Drug Metabolism

• Oxidation: Oxygen is incorporated into the drug molecule.
• Reduction: Loss of oxygen from the drug molecule
• Hydrolysis: Cleavage of chemical bonds in the drug molecule using water

Cytochrome P450 (CYP450) enzymes

• Essential for the majority of drugs eliminated by hepatic metabolism
• Absorbed at 450nM when carbon monoxide is present

Intracellular Sites

• Endoplasmic reticulum: Majority of drug metabolism occurs here, isolated as microsomal fraction
• Mitochondrion
• Cytosol
• Lysosomes

Microsomal Mixed Function Oxidases (MFOs)

• These enzymes require both NADPH and molecular oxygen
• Involved in biotransformation of drugs and other substances

Reactions Catalysed by MFOs

• Aromatic hydroxylation, Aliphatic hydroxylation, Epoxidation, N or O or S-Dealkylation, Oxidative deamination, N-Oxidation, S-Oxidation, Phosphothioinoate oxidation, Dehalogenation, Alcohol oxidation

CYP450 Isoenzymes

• More than 30 subtypes identified
• Cause differences in individual abilities to metabolise drugs
• Nomenclature, Family, and Subfamily based on structural similarity

Paracetamol Metabolism

• Metabolised by 3 different pathways: glucuronidation, sulfation, and cytochrome P450 pathway (producing NAPQI)
• In normal doses, NAPQI is conjugated with glutathione and excreted.
• P450 mediated phase 1 metabolism of paracetamol leads to production of NAPQI, a reactive intermediate which is hepatotoxic.

Drug Metabolism: Phase II

• Conjugation with endogenous substrates for increased aqueous solubility.
• Glucuronidation (adding glucuronic acid), Sulfate conjugation, Amino acid conjugation

UGT Enzymes

• Largest capacity enzyme system, UDP glucuronosyltransferases (UGTs)
• Located in the endoplasmic reticulum
• Involved in the metabolism of many drugs, removing their pharmacological effect

Phase III Example

• Glucuronidation reactions involving morphine, chloramphenicol, and salicylic acid
• UGTs (UDP-Glucuronosyltransferases) are responsible for glucuronidation reactions

Paracetamol Toxicity (and Phase 2)

• In high doses, NAPQI overwhelms the body's glutathione supply, leading to liver damage
• Phase 2 conjugation with hepatic glutathione detoxifies harmful metabolites.

Phase I and II Reactions

• Phase I reactions convert lipophilic drugs to more hydrophilic metabolites.
• Phase II reactions conjugate the metabolites with endogenous substrates like amino acids.
• The end result of metabolism can be the conversion of an active drug to an inactive compound or to another active drug metabolite

Sequence

• Phase II sometimes takes place before Phase I reactions

End Result of Drug Metabolism

• Drug is converted from pharmacologically active to inactive (mostly).
• Conversion of a drug to an active metabolite.
• Conversion of a pharmacologically inactive compound to an active drug – prodrug.

Prodrug Definition

• An inactive drug precursor that is metabolised into an active form in the body.

Control of Metabolism

• Upregulation/downregulation of CYP and UGT isoforms may alter drug metabolism.
• CYP and UGT work in concordance with other systems.
• Adaptable responses within cells.

Therapeutic Window of Drug

• The range of drug concentrations in the plasma needed to cause a therapeutic response without causing adverse effects.

Enzyme Induction: Clinical Significance

• Decreased efficacy of drugs (co-administered drugs stimulate metabolism), Osteomalacia (barbiturates and antiepileptics stimulate vitamin D3 metabolism), Increased dose requirement of benzodiazepines and analgesics in smokers

Induction of Drug Metabolism

• Increased synthesis of cytochrome P450 (CYP450)
• Several classes of drugs, industrial chemicals, and environmental pollutants including barbituates, carbamazepine, and ethyl alcohol

HME- inducers/HME-inhibitors

• Lists several inducers and inhibitors of human microsomal enzymes (HMEs).
• Indicates the influence of various substances on hepatic blood flow.

Case Study(s)

• 63 year-old male prescribed simvastatin 10mg daily, increased to 50mg, presented with rhabdomyolysis
• self-administered St. John's Wort to treat depression, impacting efficacy and toxicity of simvastatin.

Characteristics of Case Study

• Therapeutic levels of simvastatin achieved by multiple dosage increases
• Presence of other drugs increased clearance
• St. John's Wort induced P450 enzymes
• Toxic effects appeared gradually
• Increased drug clearance was reversible.

Cause

• Enzyme induction, presence of inducing agent causes increased metabolism
• Inducing agent brings about increased synthesis of enzyme isoforms.
• Patient had stopped taking inducing agents, requiring higher statin doses
• Patient's subsequent rhabdomyolysis due to increased metabolism resulting from higher than therapeutic doses.

Inducing Agents

• Anticonvulsants, Steroids, Polycyclic aromatic hydrocarbons, Antibiotics, Recreational agents, Herbal remedies (also known as St. John's Wort)

Mechanism of Action of Inducers

• Alterations in transcription within the nucleus, resulting in increases in the expression of CYP isoforms
• Consequence increased clearance of co-administered drugs metabolised by these isoforms

Induction Overview

• Inducing agent combines with cytoplasmic receptor to form complex
• Receptor-inducer complex migrates to the nucleus
• Interaction of complex with DNA results in mRNA production
• mRNA released to cytoplasm, translation produces increased CYP levels, increasing metabolic rate

Clinical Consequences of Enzyme Induction

• Therapeutic failure due to induction
• Drug levels restored to therapeutic levels by increasing doses or removal of inducing agent
• Effects on patients are not serious as effects are readily restored

Inhibition of Drug Metabolism

• Drug interaction whereby one drug inhibits another's metabolism
• Two main types are: inhibition of cytochrome P450 and other specific metabolic pathways

Calcium Channel Blocker Example

• Felodipine, affected by grapefruit juice which inhibits CYP3A4 in enterocytes.
• Reduced bioavailability in presence of grapefruit juice

Case History Example

• 29 year-old healthy male taking terfenadine for allergic rhinitis
• Took grapefruit juice before medication, leading to cardiac arrhythmias within an hour and death
• Post-mortem report showed raised levels of terfenadine

Analysis of Case History

• Post-mortem report revealed elevated terfenadine levels
• No history of impaired liver function
• Higher drug levels can cause cardiac arrhythmias
• Grapefruit juice inhibited the metabolism of terfenadine

Characteristics of the Analysis

• Patient stabilised on dosage indicating drug levels lie within therapeutic window
• Grapefruit juice prevented terfenadine clearance
• Toxicity resulted from intensified drug action due to accumulation

Enzyme Inhibition

• Toxic effects appear rapidly
• Toxic effects are rapidly reversible once inhibition is prevented
• Inhibition can be a result of self-medication

Mechanisms of Enzyme Inhibition

• Competitive inhibition, Noncompetitive Inhibition, Uncompetitive Inhibition, Mechanism-based inhibition

Clinical Consequences of Inhibition

• Irreversible inhibition has severe clinical consequences
• Elevated drug levels can cause toxicity that worsens over time if inhibitor not removed.
• Destruction of enzyme is not readily reversible, making it difficult to reverse the toxic effects rapidly.

Enzyme Induction and Inhibition

• Induction lowers drug levels, leading to failure of therapy
• Inhibition raises drug levels, often leading to adverse reactions
• Graph illustrating the difference between narrow vs wide therapeutic windows

Factors Affecting Drug Metabolism

• Internal factors include genetics, age, gender, and disease
• External factors include diet and environment

Gender

• Menstrual steroids and differences in drug handling between sexes due to drug metabolism variability.
• Phase I-III metabolism handles female steroid hormones with enzymes more affected in females.

Age and Neonates

• CYP450 activity lower in newborns (<4 weeks), requiring smaller doses of certain drugs.
• Developmental differences of metabolising ability can be concerning as differences disappear between 2-6 months for certain drugs

Age and Elderly

• Drug metabolism declines after 70 years
• Polypharmacy or multiple drug use prevalent issues in elderly population
• In vitro studies show similar CYP activities between young and elderly populations.
• Physiological limitations (oxygen and NADPH) may also be crucial factors in metabolism

First-Pass Metabolism

• Hepatic blood flow reduces by 40% in older adults (>60 yrs) accompanied by a related reduction in renal clearance.
• High intrinsic clearance drugs (rapid processing by the liver) are affected by reducing bioavailability with standard doses, creating potential ADR risks for elderly patients.
• Warfarin requires a lower dose in elderly compared to younger adults due to reduced metabolism and clearance

Diet

• Diet significantly impacts drug metabolism
• Antioxidants, plant toxins, and preservatives can impact drug metabolsim

Polyphenols

• Extend viability of fruit
• Toxin to animals consuming plants.
• Naringenin and bergamottin in grapefruit juice
• Polyphenols and altering metabolism of Phase I and II reactions

Cruciferous Vegetables

• Induce CYP1A2 and glutathione transferases, resulting in rapid clearance of certain compounds.

Barbecued Meats

• Induces CYP1A1 and CYP1A2
• Increased clearance of paracetamol, amitriptyline, clozapine, fluvoxamine, naproxen, and haloperidol

Effect of Alcohol on Metabolism

• Acute ethanol exposure – longer drug half-lives due to preventing NADH/NAD+ ratio
• Chronic exposure – phase I and II metabolism and increasing CYP2E1
• Enzyme inhibition due to structural changes in hepatocytes.

Cirrhosis

• Liver tissue replaced by fibrous tissue, reducing viable hepatocytes.
• Phase I metabolism is unaffected, but Phase II glucuronidation is impaired, leading to decreased metabolism of some drugs like paracetamol.
• Can result from alcoholic liver disease.

Environmental Factors

• Tobacco smoke, motor vehicle exhaust fumes, industrial solvents, and industrial pollutants can induce drug metabolism.
• Pesticides such as malathion and parathion induce P450, affect synthesis.

Conclusion

• Induction and inhibition of drug-metabolizing enzymes cause modifications in pharmacokinetics.
• Induction lowers drug levels, possibly leading to therapeutic failure.
• Inhibition raises drug levels, often leading to 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.

Test your knowledge on the role of Cytochrome P450 enzymes in drug metabolism, including Phase I and Phase II reactions. This quiz covers key concepts such as NADPH involvement, glucuronidation, and the first pass effect. Understand the essential functions and characteristics of cytochrome P450 in pharmacology.