By Yuchen Lin
When we are ill, we take various kinds of drugs to fight against the disease. But after drugs exert their functions, how do our bodies get rid of them? Lots of drugs are toxic to most of the cells in our body. If we do not detoxify them after they’ve done their job, the drugs will have undesirable functions in other cells or tissues. The pathway by which our bodies break down drugs is called drug metabolism or xenobiotic metabolism. It is part of the natural process that metabolizes endogenous substrates, including hormones, cholesterol, and bile acids, and excretes them.
Liver is the main site for drug metabolism that involves three phases – phase I, phase II and phase III transformation. Drug metabolism converts lipophilic drug compounds into more hydrophilic compounds. Usually, orally administered drugs are dissolved in the gastrointestinal tract and absorbed through the gut lumen, where they enter the bloodstream and go to the liver for circulation and metabolism. Phase I transformations introduce functional groups to make xenobiotic compounds inactive. The most common and essential enzyme in phase I is cytochrome P450 that promotes the oxidation of xenobiotic compounds. This increases the water solubility of the drug compounds, meaning they are more readily excreted as urine. This is a slow conversion (Voet & Voet, 2011). If lipid-soluble non-polar drug compounds are not well-metabolised during phase I, they will remain in our blood and tissues, maintaining their pharmacological functions for a longer period. Then, in phase II transformation, a rapid and complete conversion process will take place to make drug compounds hydrophilic enough to be excreted (Voet & Voet, 2011). These modifications happen on pre-existing functional groups created in phase I. It generates highly polar drug compound derivatives for excretion. Some bigger groups may also be added here to increase the compounds’ polarity. As a result, the metabolites are eliminated by direct intestinal excretion through faeces instead of urine, because metabolites are too big to cross the filter in the kidney. Conjugated xenobiotics can be further processed in phase III before being recognized by efflux transporters that pump them out of cells.
Roles of drug metabolism are not confined to degradation and detoxification of drugs, but are also extended to activation of prodrugs. Some parts of prodrugs can be too toxic for non-target cells or tissues. These parts are masked and phase I transformation activates them after the prodrugs pass through the liver. Take one of the most common drugs in our daily life, Aspirin, as an example. Aspirin is widely used for reducing fever and relieving pain, and is also known as a salicylate and a nonsteroidal anti-inflammatory drug (WebMD, 2021). There are three critical enzymes involved in aspirin metabolism, cytochrome P450 2C9 (CYP2C9), UDP-glucuronosyltransferase 1A6 (UGT1A6), and N-acetyl transferase 2 (NAT2). Weak acid Aspirin is not absorbed by the stomach, which has a pH of 6.5. Its absorption occurs in the upper part of the small intestine through passive diffusion. After absorption, the Aspirin will be metabolized. Notably, CYP2C9 has a significant association with polymorphism in the Aspirin sensitivity phenotype in its DNA. Such genetic polymorphisms are key to drug efficiency and tolerance in our body (Eswara, 2011). Aspirin is a prodrug, and the active form which helps to release pains is salicylic acid. However, the phenolic hydroxyl, -OH, of salicylic acid causes gastric bleeding in the human body. Therefore, an ester is added to mask the phenolic hydroxyl group and form inactive Aspirin – until CYP2C9 activates salicylic acid and enables it to convey its functions. Aspirin brings less toxic side effects to our bodies by preserving salicylic acid in such an inactive form.
CYP2C9 promotes hydroxylation of Aspirin in phase I transformation to increase the polarity of the hydrolysed products of Aspirin, and also increase its water solubility (Palikhe et al., 2011). It catalyses hydroxylation and epoxidation of various substrates. There can be multiple sites available for modification on substrates, which means the metabolised Aspirin can re-enter phase I transformation. However, the catalysis of CYP2C9 is highly specific. If the orientation of the modifiable chemical group is not optimised for the binding site on CYP2C9, further oxidation cannot happen. The oxidation relies on a microsomal mixed-function oxidase system which requires oxygen and a reducing environment. One oxygen atom is transferred to Aspirin and the other oxygen atom undergoes a two-electron reduction followed by a conversion to water. CYP2C9 contains a Heme at its active site as the iron (III) porphyrin cofactor. Molecular oxygen will bind to this cofactor after the reduction of ferric ion to ferrous ion. Then, oxygen is converted to the reactive form that plays a role in oxidizing Aspirin. Therefore, the heme group in CYP2C9 is critical for its function. This group is displayed in figure 1.
Fig 1. Heme structure of CYP2C9. CYP2C9 is shown in grey. The Heme structure is labelled with magenta sticks and the conserved Cys435 which makes contact with ferrous ions is shown with cyan sticks.
For the most part, Aspirin is not hydrophilic enough after phase I transformation, and that is where UGT1A6 and NAT2 come into force. UGT1A6 is responsible for glucuronidation of Aspirin, transferring a sugar moiety onto the hydroxyl group of the phase I product, a D-glucuronic acid, and producing glucuronide. The salicylic acid deacetylated from Aspirin can be converted to various metabolites in phase II, including acyl and phenolic glucuronide conjugates, for excretion. The final products are often excreted in the bile. UGT1A6 catalyses the formation of the conjugates to lower the amount of pharmacologically active salicylic acid present (Kuehl et al., 2005). NAT2 catalyses the acetylation with three enzymatic activities, and co-factor acetyl-CoA is required. These activities involve the O-acetylation of N-hydroxy aryl amines that are generated through oxidation by CYP2C9. Meanwhile, NAT not only has roles in detoxifying drugs but is also implicated in cancer risk because of its role in the activation of carcinogens (McDonagh et al., 2014). Functional alterations in NAT2 gene could lead to asthma (Palikhe et al., 2011) which belongs to the symptoms of acetylsalicylic acid intolerance. Therefore, the various elimination rates of xenobiotics including Aspirin caused by the differential speed of acetylator production is genetically determined.
After processing by these three essential enzymes in phase I and II transformation, Aspirin is ready to be released. However, the specific rates and pathways may vary between individuals as they are affected by many factors such as age, sex, and individuals’ hormone production. In addition, drug metabolism is stereospecific, so two enantiomers can have completely different metabolites and excretion pathways. Toxic metabolites might be produced, or metabolism of one enantiomer may inhibit metabolism of the other.
Drug metabolism detoxifies drugs and increases their hydrophilicity, so our bodies can get rid of them once their aims are accomplished. Apart from the example of Aspirin mentioned in this article, cytochrome P450 also plays important roles in phase I transformation of many other drugs. Although phase II transformation is rapid, it is essential in ensuring the removal of toxic drugs from bodies, especially for prodrugs that utilize phase I transformation for activation. These mechanisms protect our bodies from adverse impacts caused by life-saving drugs.
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Palikhe, N.S. et al. (2011) Polymorphisms of Aspirin-Metabolizing Enzymes CYP2C9, NAT2 and UGT1A6 in Aspirin-Intolerant Urticaria. AAIR. 3(4):273-276. Doi: 10.4168/aair.2011.3.4.273
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WebMD. (2021) Aspirin Oral. Available from: https://www.webmd.com/drugs/2/drug-1082-3/aspirin-oral/aspirin-oral/details [Accessed: 19th May 2021]