Introduction to cytochrome P450
Contents Introduction Structure of cytochrome P450 Factors involved in drug biotransformation Genetic polymorphism Disease Age Concomitant drugs Induction Inhibition

Introduction

Cytochromes P450 enzymes constitute a large superfamily of haem-thiolate proteins involved in the metabolism of a wide variety of both exogenous and endogenous compounds. (91) They were first discovered in 1955 in rat liver microsomes and they are characterized by an intense absorption band at 450 nm in the presence of carbon monoxide. (92)
 

Spectrum of P450

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The cytochrome P450 (CYP) mixed function monooxygenases are located on the smooth endoplasmic reticulum of cells throughout the body, but the highest concentrations are found in the liver (hepatocytes) and small intestine. (93) These enzymes are responsible for the oxidative (Phase I) metabolism of a wide number of compounds, including many medications. (93) They biotransform lipophilic drugs to more polar compounds that can be excreted by the kidneys. (94) The metabolites are usually less active than the parent compound, although some drugs undergo biotransformation to pharmacologic active agents. In some cases the metabolites can be toxic, carcinogenic or teratogenic. (94)

At least 12 cytochrome P-450 gene families have been identified in humans, although 3 families are involved in the majority of the drug biotransformations; these are the cytochrome P-450 1,2 and 3 (CYP1, CYP2 and CYP3) (94) The enzymes are divided into families based on amino acid sequence similarities, and each family can be further separated into subfamilies, which are designated by capital letters following the family designation (e.g., CYP3A). Individual enzymes are subsequently indicated by arabic numerals (e.g., CYP3A4). (94) An enzyme belongs to a family when the amino acid sequence possesses more then 40% homology, enzymes with more than 55% homology form a subfamily and individual enzymes can have to 97% homology between the sequences. (92) A single hepatocyte can contain a variety of cytochrome P-450 enzymes. An individual enzyme of cytochrome P-450 may be able of metabolizing many different drugs, but a given drug may be primarily metabolized by a single enzyme. (94)

Members of the CYP3A subfamily are the most abundant cytochrome enzymes in humans, accounting for 30% of the cytochrome enzymes in the liver and 70% of those in the gut. CYP3A4 is the major form of cytochrome P-450 in the adult liver and metabolizes the greatest proportions of drugs. (94) This enzyme and CYP3A3, which are 97% identical and cannot be distinguished from each other based on the substrates that they metabolize, are the major enzymes expressed in the small intestine, while CYP3A5 is the major enzyme expressed in the stomach. (95) CYP3A5 is present in only 20%-30% of Caucasians, but being deficient in CYP3A5 poses no problem because the CYP3A4 enzyme is available to assume its functions. (96)

Structure of cytochrome P450 (91)
 

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There exists a widespread division of P450-containing monooxygenases systems into two main types, bacterial/mitochondrial (typeI) and  microsomal (typeII). Mitochondrial P450 systems have three compounds: a FAD containing flavoprotein (NADPH or NADH-dependent reductase), an iron-sulfur protein and P450. The eukaryotic microsomal P450 system contains two components: NADPH:P450 reductase, a flavoprotein containing both FAD and FMN, and P450. CPR appears to be fusion protein consisting of domains which are homologous to ferredoxin: NADP⁺ reductases (FAD domain) and flavodoxin (FMN domain) (97)
All the P450 containing monooxygenase systems described so far share common structural and functional domain architecture. By domain we mean polypeptide existing as an independently folding unit and possessing a certain function. Hence there are no fundamental differences between the protein domain and the individual protein component and all the P450 systems can be considered as three-domain systems:

• NADH- or NADPH-dependent FAD containing reductase (FAD domain)
• an iron-sulfur protein (in a three-component system) or FMN-binding domain (in a two- and one component system)
• P450 domain (haem domain)

Factors involved in drug biotransformation

Genetic polymorphism (94)

Genetic differences are important contributors to the inter individual differences in drug biotransformation seen within a patient population. These differences are called genetic polymorphism's and are linked to inherited autosomal recessive traits. The definition of a polymorphism is the presence within a population of at least 2 groups with distinctly different abilities to metabolize drugs. (98) Individuals can be characterized as extensive (rapid) or poor (slow) metabolizers. Poor metabolizers often have an increased incidence of adverse effects.

Disease (94)

Impaired liver functions can lead to decreased drug biotransformation and is a function of the severity of the disease. Disease state that can impair liver function include hepatitis, alcoholic liver disease, biliary cirrhosis and hepatocarcinoma. Infection can also alter drug biotransformation. There have been reports of impaired drug elimination during viral infections such as influenza, rhino virus, adenovirus, herpes simplex virus and infectious mononucleosis.

Age (94)

Infants do not develop a mature enzyme system until more than 2 weeks after birth. elderly have age related decreases in liver mass, hepatic enzyme activity and hepatic blood flow. In addition the overall metabolic capacity of the liver is decreased, although the considerable inter individual variability in age and disease related changes in organ function makes it difficult to form generalizations.

Concomitant drugs

Two of the most common causes of altered drug biotransformation reactions are induction and inhibition of cytochrome P-450 enzymes. (94)

Induction

Induction is an increased synthesis of enzyme that is associated with exposure to drugs. (94) The induction can occur when a drug stimulates the biotransformation of co-administered drugs either through the same enzyme pathway or via an alternative pathway. (94) However inducers are usually specific for a given cytochrome P-450 family. (94) Sometimes a drug can induce its own biotransformation in addition to that of other agents.
Effects of induction can be seen within the first two days of therapy, but it usually takes more than a week for new enzymes to be synthesized and the maximal effect to occur. (94) The time course of enzyme induction onset and offset is closely related to the plasma concentration of the inducer, as well as the half-life of enzyme production and degradation. (99)

Inhibition

Competitive inhibition is the most common mechanism of inhibition and occurs when 2 or more drugs compete for the same enzyme. The clinical significance of an inhibition interaction depends primarily on the relative concentrations of the drugs, as well as a variety of other patient specific factors.  Some drugs are capable of binding to, and acting as competitive inhibitors of, different P-450 enzymes from the ones that are responsible for the biotransformation of that particular drug. (94)
Drugs can bind irreversibly (mechanism based inhibition) or reversibly with the haem-binding site of the enzyme and inhibit other drugs from binding. Mechanism based inhibition occurs when certain drugs are metabolized by the cytochrome P-450 system to active metabolites that bind to the enzyme and cause irreversible loss of function. Activity can only be restored by synthesis of new enzymes, which may take several days. (94)
More complex mechanisms of inhibition can also occur. Some drugs undergo metabolic activation by the cytochrome P-450 system to inhibitory products. The metabolites may generate relatively stable complexes with cytochrome P-450 so that the cytochrome is held in an inactive state. There can be great clinical significance to this interaction, since it is relatively long in duration. Additionally, when the interaction involves drugs with narrow therapeutic ranges, there is the potential for toxicity. (94)
Unlike induction enzyme inhibition usually begins with the first dose of the inhibitor. (99) Inhibition is maximal when the inhibitor reaches steady state (four to seven half-lives), and the maximum concentration of the inhibited drug occurs when the inhibited drug reaches steady state at its new, longer half-life. Similarly, the time required for the interaction to resolve also depends on the half-lives of the drugs involved. (99)